Status of Biomedical Research and Related

Technology for Tropical Diseases

September 1985

NTIS order #PB87-139614
Recommended Citation:

U.S. Congress, Office of Technology Assessment, Status of Biomedical Research and Related

Technology for Tropical Diseases, OTA-H-258 (Washington, DC: U.S. Government Printing Office, September 1985).

Library of Congress Catalog Card Number 85-600508

For sale by the Superintendent of Documents

U.S. Government Printing Office, Washington, DC 20402
Foreword

Billions of people in less developed areas of the world and a small but growing

number of inhabitants of the industrialized countries come into contact with infectious

diseases generally characterized as “tropical. ” Though not necessarily geographically

limited, these diseases are most common in the tropics because of the social and economic, as well as climatic, conditions that prevail there.

In combination with the traditional methods of parasitology and basic research,

the new tools of biotechnology and immunology have opened doors to finding methods for controlling tropical diseases. Scientists using both traditional and recombinant

DNA techniques are pursuing technologies for the control of insects that transmit diseases and the development of vaccines, diagnostic technologies, and new therapies for

tropical diseases. The possibilities for making inroads into the overwhelming morbidity and mortality caused by these diseases have never been so promising.

The United States continues to make significant contributions toward controlling

tropical diseases, even though the resources allotted to these efforts are small: in recent

years, less than $100 million per year has been spent by the U.S. Government on tropical disease research out of a total annual biomedical research budget of $4 to $5 billion.

As one component of U.S. international activities, research in tropical diseases serves

several objectives: social and humanitarian, political, economic, medical (including the

protection of American citizens), and scientific. Opportunities for expanding U.S. influence and contributions in this field are great.

In the spring of 1983, the Senate Appropriations Committee asked OTA to examine the status of biomedical research and technologies for controlling tropical diseases.

The request was precipitated by a question about continued funding of the Gorgas

Memorial Institute of Tropical and Preventive Medicine, Inc. and its operating arm,

the Gorgas Memorial Laboratory in the Republic of Panama. An OTA technical memorandum, Quality and Relevance of Research and Related Activities at the Gorgas

Memorial Laboratory, was released in August 1983 as a response to the immediate question. The larger question is addressed in this full assessment.

An advisory panel, chaired by Dieter Koch-Weser, provided guidance and assistance during the assessment. A large number of individuals from academia, the Federal

Government, the private sector, and the public provided information and reviewed a

draft of the report. Five contractors were essential to the completion of the assessment:

Paul F. Basch surveyed the use of biotechnology in tropical disease research; Veronica

Elliott compiled information about funding; Roger A. Bitar contributed the material

on therapies for tropical diseases; and Myron M. Levine and Lydia Schindler prepared

case studies on oral dehydration therapy for diarrheal diseases and on the development

of a malaria vaccine, respectively.

The final responsibility for the content of the report rests with OTA. Key staff involved in the analysis and writing were Hellen Gelband, Kerry B. Kemp, Susan C. Tripp,

and Steven S. Bjorge.

 JOHN H. GIBBONS

Director

///
Advisory Panel for Status of Biomedical Research and

Related Technology for Tropical Diseases

Dieter Koch-Weser, Pane] Chair

Department of Preventive and Social Medicine

Harvard Medical School

Pedro Acha

Pan American Health Organization

George Alleyne

Pan American Health Organization

Karen Bell

Board on Science and Technology for

International Development

National Academy of Sciences

Richard Cash

Department of Tropical Public Health

Harvard School of Public Health

Barnett Cline

Department of Tropical Medicine

Tulane Medical Center

Joseph Cook

The Edna McConnell Clark Foundation

Robert Goodland

Office of Environmental Affairs

World Bank

Abraham Horwitz

Pan American Health Organization

Francisco Lopez-AntuFiano

Pan American Health Organization

Arnold Monto

School of Public Health

University of Michigan

Ruth Nussenzweig

Division of Parasitology

New York University School of Medicine

Richard Riegelman

Department of Health Care Sciences

George Washington University Medical Center

Gabriel Schmuiiis

Pan American Health Organization

Thomas Simpson

Virginia State Health Department

Ronald Vogel

College of Business and Public Administration

University of Arizona

Kenneth Warren

The Rockefeller Foundation
OTA Project Staff—Status of Biomedical Research and

Related Technology for Tropical Diseases

Roger C. Herdman, Assistant Director, OTA

Health and Life Sciences Division

Clyde J. Behney, Health Program Manager

Hellen Gelband, Project Director

Steven S. Bjorge, Analyst

Kerry Britten Kemp, Health and Life Sciences Division Editor

Susan C. Tripp, Analyst

Virginia Cwalina, Administrative Assistant

Beckie I. Erickson, Word F’rocessor/P. C. Specialist

Carol Guntow, Clerical Assistant

Diann G. Hohenthaner, Secretary/Word Processor Specialist

Contractors

Paul F. Basch, Stanford University School of Medicine

Roger A. Bitar, The Johns Hopkins Hospital

Veronica Elliott, Alexandria, VA

Myron M. Levine, University of Maryland School of Medicine

Lydia Woods Schindler, Darnestown, MD

v
Contents

Chapter

1.

2.

3.

4.

5.

6.

7.

8.

9.

Findings and Options . . . . . . . . . . . . . . . . .

The United States in International Health

Funding of Tropical Disease Research. . .

Description of Selected Tropical Diseases

Page

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59

Strategies and Technologies for Controlling Tropical Diseases . . . . . . . . . . . . . . . . . 99

Vector Control Technologies: Selected Tropical Diseases . . . . . . . . . . . . . . . . . . . . . . 115

Immunization Technologies: Selected Tropical Diseases . . . . . . . . . . . . . . . . . . . . . . . 129

Diagnostic Technologies: Selected Tropical Diseases . . . . . . . . . . . . . . . . . . . . . . . . . . 153

Therapeutic Technologies: Selected Tropical Diseases . . . . . . . . . . . . . . . . . . . . . . . . . 181

Case Studies

A. Oral Dehydration

B. The Developmen!

Appendixes

Therapy for Diarrheal Diseases . . . . . . . . . . . . . . . . . . . . . . . . . . . 201

of a Malaria Vaccine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 225

A. Acknowledgments and Health Program Advisory Committee . . . . . . . . . . . . . . . . . 249

B. Method for Analysis of Tropical Disease Research Funding . . . . . . . . . . . . . . . . . . . 253

C. Glossary of Acronyms and Terms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 254

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 267

Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 287
1 .

Findings and Options
Contents

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . .

Request for the Assessment. . . . . . . . . . . .

Scope of the Report . . . . . . . . . . . . . . . . . .

Structure of the Report . . . . . . . . . . . . . .

Summary of Options Developed in This

Background on Tropical Diseases . . . . . . . .

Health in the Tropics . . . . . . . . . . . . . . . . .

Tropical Diseases in the United States . .

 . . . . . . . . .

. . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . .

Assessment . . . . . .

. . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . .

U.S. Citizens at Risk for Tropical Diseases . . . . . . . . . . . . . . .

Approaches to Controlling Tropical Diseases . . . . . . . . . . . . . . .

Status of Control for Selected Tropical Diseases . . . . . . . . . . . .

Malaria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Schistosomiasis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Trypanosomiasis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Leishmaniasis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Filariasis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Leprosy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Tuberculosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Diarrhea] and Enteric Diseases. . . . . . . . . . . . . . . . . . . . . . . . . .

Acute Respiratory Infections . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Arboviral and Related Viral Infections . . . . . . . . . . . . . . . . . . .

U.S. Efforts in Tropical Disease Research . . . . . . . . . . . . . . . . . .

Rationale for U.S. Efforts in Tropical Disease Research . . . .

U.S. Activities in Tropical Disease Research . . . . . . . . . . . . . .

U.S. Funding of Tropical Disease Research. . . . . . . . . . . . . . .

Funding of U.S. Researchers in Tropical Diseases . . . . . . . . .

Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Development of Medical Technologies for Tropical Diseases

Information for Congressional Decisions . . . . . . . . . . . . . . . . .

Research Funding . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

3445566799

12

12

12

13

14

14

14

15

15

15

16

16

16

17

17

19

19

19

21

23
INTRODUCTION

For most of the world’s population in less developed countries, mainly in the tropics, good

health is not something interrupted by occasional annoying bouts of sickness. One-tenth of the

average person’s life in a developing country is

seriously disrupted by ill health (412). Life expectancy at birth is nearly 20 years shorter in the developing regions than it is in the developed countries, Each year, millions of infants and children

die from enteric and respiratory infections. Several

hundred million people are infected with organisms

that cause chronic disabling and often life-threatening parasitic diseases: malaria, schistosomiasis,

trypanosomiasis, leishmaniasis, and filariasis.

Chronic and debilitating infections with intestinal parasites range, in total, into the billions.

Researchers in all corners of the globe have

aided the efforts against tropical diseases, and the

United States has played a major role. Through

the work of individual U.S. researchers and institutions and through international collaborations, the United States has contributed to the development of oral dehydration therapy (ORT) for

the dehydration that accompanies diarrhea] diseases, the rapidly unfolding progress toward de3
4 ● Status of Biomedical Research and Related Technology for Tropical Diseases

velopment of a genetically engineered malaria

vaccine, and other advances in the prevention,

diagnosis, and treatment of tropical diseases.

ORT and the malaria vaccine fall at opposite

poles of biomedical research and of disease interventions. ORT, developed from research in basic

human physiology, has saved the lives of hundreds of thousands of children by treating the lethal dehydration of diarrhea caused by a plethora of organisms. The hope for a malaria vaccine

is to prevent at least some part of the 300 million

cases that occur worldwide each year. Widespread

application of an effective vaccine, still at best 5

to 10 years away, will be dependent on the use

of recombinant DNA technology, a tool that did

not even exist 20 years ago. These advances are

undeniably valuable contributions, both for

science and for health. But even these achievements pale in the shadow of the massive health

problems that continue to affect the billion or so

who live in less developed areas.

This assessment examines the status of control

for a selected group of diseases that cause extensive morbidity and mortality in developing countries of the tropics and looks at what the United

States is doing in biomedical research to alleviate the human misery caused by these diseases.

OTA found a great deal of recent progress in tropical disease research and a small, but highly motivated, corps of tropical disease researchers. OTA

also found, however, that the U.S. Government

is spending relatively little money on tropical disease research. The small amount of money spent

by the United States on tropical disease research

appears to reflect choices in policy, whether implicit or explicit, and not a lack of promising

avenues for research.

Request for the Assessment

The need for an assessment of research in tropical diseases was focused in the spring of 1983,

when a question arose about continued U.S. funding for Gorgas Memorial Laboratory (GML), a

half-century old research institution located in the

Republic of Panama. The National Institutes of

Health (NIH) requested no funds for GML for fiscal year 1984. If acted upon, the withdrawal of

funding would probably have forced the laboratory’s closing.

The Senate Appropriations Committee asked

OTA to assess the activities of GML and then go

on to examine the overall status of U.S.-funded

biomedical research for tropical diseases. An OTA

technical memorandum fulfilling the first charge,

Quality and Relevance of Research and Related

Activities at the Gorgas Memorial Laboratory,

was released in August 1983 (360). On the basis

of that report, a companion report prepared by

the General Accounting Office (355), and other

information, Congress restored funding for GML

for fiscal year 1984. While recognizing the creditable record of research and service of GML, Congress also required that plans for the future course

of the laboratory’s efforts take account of some

problems identified by OTA and the General Accounting Office.

Scope of the Report

There are many definitions of tropical diseases.

In the strictest sense, tropical diseases are those

that occur only in tropical areas, between the

Tropics of Cancer and Capricorn, and are limited

to those areas by geographic and climatic factors.

In the broadest sense, they can include any health

conditions that occur in the tropics, regardless of

their distribution around the world. A more useful definition includes those diseases or conditions

that occur or could occur in many regions, but

which are considerably more prevalent in tropical areas because of the social, economic, and climatic conditions that characterize many tropical

countries.

In this report, no attempt was made to redefine tropical diseases. For the purpose of this

assessment, OTA focused on a limited group of

conditions that are unquestionably important

causes of morbidity and mortality and that either

occur exclusively in the tropics or have greater

public health implications in the tropics than in

temperate zones. Included in this group are the

six diseases singled out by the Special Program

for Research and Training in Tropical Diseases

(TDR) of the World Health Organization (WHO)/

U.N. Development Program/World Bank: malaria, schistosomiasis, trypanosomiasis, filariasis,
Ch. l—Findings and Options . 5

leishmaniasis, and leprosy. Also considered in this

report are tuberculosis, diarrheal diseases, acute

respiratory infections (ARIs), and arboviral and

related viral infections.

The focus of this assessment is on biomedical

laboratory research pertaining to these diseases

and on some field research in the control of insect and other vectors. In general, this report does

not address the equally important contributions

of entomological research and basic ecological research necessary to improve the understanding of

these diseases. It also does not examine research

in human behavior and societies or the interactions of humans with the environment, elements

that are of great importance in understanding and

controlling disease.

Research, which provides the knowledge and

tools that can be applied to control tropical diseases, is only one arm of the triad that makes up

the U.S. effort in international health. Two other

arms of the triad—application and training—are

not the focus of this assessment but are also deserving of attention.

Application of the knowledge and tools developed through research is aimed at directly improving health conditions. Millions of deaths still occur

as a result of diseases for which control measures

are available, but are not applied.

Training yields the personnel both for research

and for the application of biomedical technologies

in the field. There is widespread concern among

medical and scientific personnel in tropical health

that a crisis exists in the dwindling U.S. capacity

for training in this area. The National Research

Council (NRC) of the National Academy of Sciences (NAS) has recently begun an assessment of

the capacity of U.S. institutions to train personnel and carry out research in tropical medicine.

It will also examine different types of funding

mechanisms and programs for tropical health research. NRC’s report is scheduled for completion

in 1985.

Structure of the Report

This report examines the U.S. role in and contributions to biomedical research in selected tropical diseases, and assesses the status of biomedical technologies for controlling those diseases. The

remainder of this chapter presents the major findings of the assessment along with some background material, identifies issues, and develops

options to address those issues.

Chapter 2 discusses the rationale for U.S. involvement in international health activities and

reviews some important reports and legislation

that have shaped the U.S. role.

Chapter 3 analyzes funding for tropical disease

research, focusing on funding from the U.S. Government, from TDR, and from WHO. It also considers the contributions of foundations and pharmaceutical companies.

Chapters 4 through 9 summarize the status of

biomedical technologies for tropical diseases and

review current advances from research. Chapter

4 briefly describes the diseases or classes of diseases covered—malaria, schistosomiasis, trypanosomiasis, leishmaniasis, filariasis, leprosy, tuberculosis, diarrheal diseases, ARIs, and arboviral

and related diseases—and reports advances in

basic knowledge about the disease organisms.

Chapter 5 describes the overall strategies for

tropical disease control programs. Chapter 6 treats

vector control technologies; chapter 7, immunization technologies; chapter 8, diagnostic technologies; and chapter 9, therapeutic technologies.

Case studies of the development of two technologies are presented following chapter 9. Case

study A is about ORT for diarrheal disease, and

case study B is about the development, up to the

present, of vaccines to prevent malaria.

Summary of Options Developed in

This Assessment

The options that follow are discussed in the

text.

Medical Technology Development (pp. 19-21)

Explicitly include drugs and vaccines for tropical diseases in the definition of “orphan drugs”

under the Orphan Drug Act of 1983 (Public

Law 97-414).

Encourage Federal agencies, such as the U.S.

Agency for International Development (AID),
6 ● Status of Biomedical Research and Related Technology for Tropical Diseases

to examine the possibility of interesting private

companies in developing medical technologies

for tropical diseases by guaranteeing purchases

of products and assisting in field trials.

● Mandate the creation of and authorize funds

for a quasi-governmental nonprofit corporation

to undertake research and development of medical technologies for tropical diseases until the

technologies become economically attractive

enough for private industry to take over with

the right to an exclusive license for the product.

OR

Stimulate the development of an international nonprofit corporation, funded through contributions from the U.S. Government, other

governments, and international bodies, to

undertake such research and development.

OR

Create a nonprofit corporation charged with

ensuring the development and availability for

use in developing countries of prophylactic and

therapeutic agents for which there appears to

be insufficient commercial interest.

Information for Congressional Decisions

(pp. 21-23)

●

●

Hold a special appropriations hearing for tropical disease research with representatives from

NIH, the Centers for Disease Control (CDC),

the Department of Defense (DOD), and AID,

and perhaps invite international agencies and

private foundations to participate.

AND/OR

Require each agency mentioned above to submit a report on the status of its tropical disease

research, providing data specified by the Appropriations Committees for use during appropriations hearings.

Research Funding (p. 23)

● Increase Federal funding for all aspects of tropical disease research.

● Amend the international health mandate of the

Department of Health and Human Services

(DHHS) to remove the limitations on the research DHHS may support in tropical diseases.

BACKGROUND ON TROPICAL DISEASES

Health in the Tropics

In many developing countries of Africa, Asia,

Latin America, and other parts of the world, infectious diseases, which have been eliminated as

major causes of death in the United States and

other developed countries, are the biggest killers.

The victims are often infants and children. In the

United States, only about 11 of every 1,000 live

born babies die in the first year of life, and many

of the deaths are attributable to problems evident

at birth—low birthweight, premature birth, birth

defects, and respiratory problems, for example.

In the developing world, as many as 200 of every

1,000 babies born die before their first birthday.

The causes are almost entirely infectious diseases—especially diarrheal diseases, ARIs, and

malaria. In Latin America and the Caribbean,

diarrheal diseases are the leading cause of death

in children unders years of age (266). Worldwide,

as many children die from diarrheal disease, as

there are total deaths from cancer. In many places

where diarrheal diseases are under better control,

ARIs are the main cause of death in children. Although ARIs are common among children in the

United States, they rarely are fatal, causing a total

of about 5,000 deaths per year (not including

influenza ).

People in the less developed countries of the

tropics are not only afflicted with diseases that

seldom occur in temperate regions—e.g., malaria,

schistosomiasis, trypanosomiasis, leishmaniasis,

and filariasis; they are also afflicted with the diseases of the developed countries in temperate regions. In fact, some types of cancers are more

common in less developed countries than they are

in the United States or Europe. Certain areas of

Central America have the highest known rates of

cervical cancer in the world. Parts of Africa and

Asia have extremely high rates of liver cancer.

Heart disease has become more common in less
Ch. 1—Findlngs and Options ● 7

developed countries as well, particularly in urban

areas. Sexually transmitted diseases are more

prevalent in many developing countries than they

are in the United States.

The presence or absence of good health is determined by forces that interact with the effects

of particular agents of disease. Poor nutrition,

high birth rates, and lack of education, especially

for women, are conditions that exist in the less

developed countries with the poorest health. Poverty itself, if defined in terms of per capita income,

does not necessarily condemn a country to ill

health. In Cuba, for instance, where the average

salary is about $2OO per month, there has been

great emphasis since the 1959 revolution on providing universal access to health care, providing

for adequate nutrition, and providing for universal education. Life expectancy in Cuba is now

close to that in the United States. The infant mortality rate has been cut from over 100 per 1,000

before the revolution to about 17 per 1,000. Malaria and diphtheria have been eliminated from

Cuba. Deaths from diarrheal diseases and tuberculosis, formerly important public health problems, are negligible (316,348).

In many less developed countries, however,

malnutrition is a primary factor in disease. Poorly

nourished people are more susceptible to disease

and often suffer more severe effects when they

contract disease. Where the case fatality rate from

measles is high, the deaths are generally of undernourished children. The body’s immune system

and recuperative powers both are diminished by

the lack of adequate nutrition. Undernourished

pregnant women are of special concern. The

babies born to such women are often of low birthweight, which is the main predictor of infant mortality. Those that survive past infancy may themselves be weak and prone to infection. The mothers

may be unable to produce sufficient milk to nourish the infants, perpetuating the cycle of undernourishment.

Many scientists believe that the high infant mortality rates associated with undernutrition lead to

high birth rates (see case study A). Families who

depend on children as workers and for support

in old age attempt to have enough babies to ensure

the survival of a few. As women undergo pregnancy after pregnancy, they become progressively

weaker themselves and have fewer resources for

the surviving children. The last children born to

them may be the weakest, and most prone to die

in infancy or childhood.

Educating women is an important step toward

better health. Educated women are better able to

learn about the role of nutrition in their own and

their children’s health and may improve the probability of their children’s survival. Though cause

and effect would be hard to prove, for each year

of schooling for women, the World Bank has estimated that the infant mortality rate is reduced by

nearly 1 percent (243). Education also leads to

higher expectations for the women and for their

children, which promote better health.

The cycle of malnutrition, disease, and high

birth rates can be broken in a number of ways.

This report focuses on technologies to address the

problems of disease directly. A 1982 OTA report

(359) considered current and developing technologies for world population and fertility planning.

A more recent OTA study addressed technological alternatives to food aid in Africa (363).

Tropical Diseases in the United States

Although some “tropical diseases” are in large

part restricted to the tropics because the conditions necessary for their existence have been limited by geography or climate, many “tropical diseases” are not limited by natural factors to the

tropics, but also occur in temperate areas, including the temperate United States. The semitropical and tropical areas of the United States are, of

course, vulnerable as well.

Diarrheal diseases and ARIs occur in all parts

of the world. In the United States, virtually all

children survive bouts of diarrhea and colds with

little or no long-term consequences. What makes

diarrheal illnesses and ARIs important as “tropical diseases” is the fact that infants and children

in developing countries commonly die from them.

Tuberculosis was among the leading causes of

death in the United States early in this century.

Improved living conditions for most Americans

decreased transmission of the disease to fairly low
8 . Status of Biomedical Research and Related Technology for Tropical Diseases

levels. Tuberculosis still is widespread in the developing world and wherever substandard, overcrowded living conditions exist. In 1982, more

than 25,000 U.S. cases, undoubtedly an underreporting, were registered by CDC. Tuberculosis is common in the home countries of many recent refugees coming to the United States. In 1982,

at least 300 refugees entered the United States with

active tuberculosis (372).

Malaria was endemic to the United States until its elimination in about 1950, but the conditions for its reestablishment in some parts of the

country stilli exist. In 1982, a total of 1,056 cases

of malaria in the United States were reported to

CDC (372), fewer than the previous 2 years. All

but 17 cases were acquired outside the United

States. The 17 cases acquired in the United States

were either congenital, associated with blood

transfusions, or accidentally acquired in the laboratory. Very recent evidence suggests that malaria caused by Plasmodium vivax is being transmitted among farmworkers by mosquitoes in the

central valley of California.

Chagas’ disease (American trypanosomiasis)

has been limited to the New World tropics and

never was endemic to the United States. However,

the insect vectors of Chagas’ disease (reduviid

bugs) are present across the Southern United

States, and at least some are infected with trypanosomes. A case of Chagas’ disease transmitted

by an insect in northern California was diagnosed

in 1982, the first such case reported in the United

States since 1955, when two infants in Texas were

diagnosed with domestically acquired Chagas’ disease (309).

Four cases of Ieishmaniasis transmitted in the

United States were reported at the 1983 annual

meeting of the American Public Health Association (408).

Some of the traditionally tropical arboviruses

are extending their ranges within the tropics and

to temperate areas, probably aided by the increase

in air travel over the past decade. The principal

vector of dengue fever and yellow fever, the mosquito Aedes aegypti, is common throughout the

world, including the Southern United States. In

1982, there were 45 cases of dengue fever confirmed by CDC, in 14 States. Since 1977, 855 suspected cases have been reported, some from almost all the States. Epidemiologic investigations

have indicated that most of the infections were

contracted outside the United States. More significant though, are the approximately 40 cases

of dengue fever contracted in the United States

in 1980, the first such reports since 1945. Venezuelan equine encephalitis has been recently introduced into the United States from South America. In other parts of the world, arboviruses,

including those that cause Rift Valley fever and

African swine fever, also have demonstrated their

ability to spread.

Leprosy is transmitted in this country. Since

1970, somewhat fewer than 30 cases have been

acquired domestically each year. The total number of cases reported annually in the United States

has been increasing, however, because of an increase in cases acquired outside the country. A

sharp increase in the number of leprosy cases since

the mid-1970s corresponds to the pattern of refugees entering the United States from Southeast

Asia (372).

There are a number of ways for “tropical diseases” to become established or reestablished in

the United States. Such diseases could be spread

into the United States gradually from contiguous

geographic areas (e.g., from Latin America through

Mexico). They also could be introduced from endemic areas by Americans who have contracted

diseases abroad or by foreign visitors or immigrants entering the country. Between fiscal year

1975 and 1982, more than 800,000 refugees, most

from Asian areas where tropical diseases are prevalent, entered the United States and have since

settled in all parts of the country. A large number of other immigrants also come from tropical

areas.

The United States has a relatively strong system of disease surveillance that allows the detection of unusual disease activity in the country and

near U.S. borders. However, the scientific basis

for predicting whether a disease is likely to become established or reestablished is fairly weak.

Climatic conditions do not favor the establishment of some diseases in the United States, and

for other diseases (e.g., African sleeping sickness),

suitable vectors (tsetse flies in the case of sleep-
Ch. l—Findings and Options ● 9

ing sickness) are absent, although for most vectorborne diseases, little is known about the capacity for alternate insects to become vectors.

U.S. Citizens at Risk for

Tropical Diseases

The risks of contracting tropical diseases in the

United States, either from infected individuals or

from insects transmitting disease, are compounded

for Americans who travel to other countries.

The longstanding interest of DOD in tropical

diseases has practical roots. It is said that during

World War II, more U.S. troops in the Philippines

were hospitalized with malaria than were hospitalized with injuries of war. The same was true

of our troops in Vietnam. Currently, more than

half a million American military personnel are

abroad, many located in the tropics (365).

In addition to military personnel, substantial

numbers of Americans reside in less developed

countries as employees of U.S. and international

aid and development agencies or of multinational

corporations. An even greater number annually

travel to the tropics, no longer so far off and inaccessible as they once were. In all, about 5 million Americans each year are at risk of contracting a tropical disease.

APPROACHES TO CONTROLLING TROPICAL DISEASES

The goal of most tropical health programs is

control of disease. In this report, the term control refers to the reduction of morbidity and mortality from disease using any or all of the spectrum of biomedical and environmental tools.

Control is not synonymous with eradication, because eradication implies permanent elimination

of a disease from the face of the earth. While the

possibility exists that certain tropical diseases can

be eradicated, following the example of the spectacular success with smallpox, most of these diseases cannot be either because of biological or

practical constraints. Many tropical diseases can

be eliminated from geographic areas, while existing in other parts of the world. The control of

others can be achieved by maintaining low levels

of incidence and prevalence.

Providing clean water and sanitation systems,

coupled with education and behavioral changes,

removes the sources of infection and the means

of transmission for many diseases that are rife in

developing countries. In the United States, at least

since 1900, long before the antibiotic era, the

spread of public works, particularly sanitation

measures, was leading the country out of the thrall

of infectious diseases. As Lewis Thomas has said,

“Much of the credit [for improved health] should

go to the plumbers and engineers of the Western

World” (413).

The development of more specific disease control measures is heavily dependent on basic and

applied biomedical research. The probability that

researchers will develop successful control measures for tropical diseases has never been greater

than it is today.

The advent and explosive growth in the use of

“biotechnology” —recombinant DNA techniques

and other sophisticated tools relying on the ability to harness and manipulate genetic material—

have given a boost to the study of tropical diseases. Using methods formerly unavailable, immunologists and molecular biologists are beginning to understand the unique biology of the

parasitic organisms that cause malaria, schistosomiasis, trypanosomiasis, filariasis, and 1eishmaniasis. This progress has led to vaccine research

and to a new generation of badly needed diagnostic tools for parasitic diseases, as well as bacterial and viral diseases.

Although there is obviously great progress in

controlling tropical diseases to be made through

the use of biotechnology, there remains a need

to continue more traditional research approaches

in parasitology, infectious disease natural history,

and basic biomedicine. This need is particularly

acute for tropical diseases, because these diseases

are much less well understood than diseases of im-
10 • Status Of Biomedical Research and Related Technology for Tropical Diseases

portance in the United States. Such basic information about tropical diseases as the ranges of

insect vectors that transmit many of them, their

natural history, and their prevalence is incomplete. Although control of one or a few of these

diseases is conceivable without such information,

for most of the diseases, such information will be

required before adequate control measures—

which will of necessity integrate aspects of vector control, prevention, diagnosis, and treatment—can be designed.

The control of mosquitoes and other vectors

(where appropriate) has been a successful route

of disease control in certain geographic areas for

several diseases. The elimination of malaria in the

United States, in most of Europe, the Caribbean

islands (except for Haiti and parts of the Dominican Republic) and other parts of Latin America

was accomplished, at least in part, through control of the mosquitoes that transmit the disease.

Similarly, through control of mosquito vectors,

yellow fever was eliminated from Panama at the

turn of the century, allowing completion of the

Panama Canal.

Before the use of DDT (dichloro-diphenyl-trichloroethane) in the 1940s, vector control programs were based largely on physical measures,

particularly on removing standing water where

larvae could develop. In Africa, early control programs for tsetse flies, the vectors of African sleeping sickness in humans and nagana in livestock,

relied on geographic isolation of flies (one strategy involved burning swathes of vegetation to

prevent migration) and efforts to kill large numbers of flies manually. Although some schemes

were more successful than others, the tsetse control effort on the whole, failed, at least in part

because of a lack of effective coordination among

a number of African nations.

After the advent of DDT, hope for controlling

and eradicating diseases focused on chemical pesticides, The “malaria eradication program, ” announced in the mid-1950s, was WHO’s first global

initiative. WHO’s promotion of a malaria eradication campaign was based on what is perceived

in retrospect as the overly optimistic hope that

a single intervention by insecticide could be effective. An appreciation of the ability of insect

vectors to develop resistance to pesticides and of

the complexity of interactions among vectors, humans, and the environment was the lesson learned

from WHO’s unsuccessful efforts. Pesticides, even

DDT, still have a place in strategies for disease

control, but no single technology is likely to be

able to control most vectors of disease. The integration of chemical and biological approaches

in “integrated pest management” (1PM) may provide the long-term solution, though as yet successes are few in this rather new field.

The application of biotechnology and immunologic tools has yielded a great deal of information about the biology of disease vectors. Specific

arthropod vectors once thought to be homogeneous single species are now known to be “species

complexes.” Different species complexes may vary

subtly or dramatically in their behavior and susceptibility to the organisms that cause disease.

While appearing identical, their roles as disease

vectors may be distinct as well. Knowledge about

the vectors that transmit certain tropical diseases

may become the basis for designing rational control programs.

As a technology, vaccination to prevent disease

has had the greatest impact on health to date and

still holds enormous potential. The principle of

vaccination—the stimulation of the body’s ability to fight off pathogens before they can cause

disease—was established long before the germ theory of disease was developed. As early as 500

years ago, it was common practice in India,

China, and probably Africa to scratch a bit of material from smallpox pustules into the skin of

healthy people. The result of this process, called

variolation, was to provoke a mild case of disease and thus render the person immune to further infection. Today’s familiar vaccines—measles, rubella, mumps, diphtheria, whooping

cough, tetanus, and yellow fever, for example—

have been developed through one of several “conventional” methods. The pathogenic organism itself (attenuated or inactivated) or part of the organism is deliberately introduced into the body to

prime the immune system to combat future infections.

Vaccine development has moved into a new era

with biotechnology (361). Scientists’ ability to de-
Ch. l—Findings and Options . 11

cipher genetic codes and pinpoint the proteins that

trigger an immune response has opened the door

for subunit vaccines that may be safer and more

specific than those derived from whole or partial

disease organisms themselves. The first genetically

engineered human vaccine, for hepatitis B, is now

in early immunogenicity trials in humans. Progress toward a malaria vaccine puts initial trials

in humans perhaps as early as 1986, but development of the vaccine for general use is probably

5 to 10 years off if no major problems occur.

Besides the development of the vaccines themselves, the mode of vaccine delivery occupies the

attention of biomedical researchers and engineers.

The “bifurcated needle, ” nothing more than a double-pronged needle which holds one drop of vaccine between the prongs, is one of the technologies credited with making smallpox eradication

possible. The various vaccination devices used

previously were, for a variety of reasons, not as

well suited to the needs of the worldwide smallpox campaign. The bifurcated needle was developed by industry in response to the needs of the

campaign, which was already under way.

Trials of “aerosolized vaccines, ” which are inhaled, have been going on for the past 10 years.

Just last year, in 1984, Albert Sabin, the oral polio vaccine pioneer, reported successful measles

immunization using an aerosolized vaccine. Delivered in an aerosol form, the vaccine was more

effective than it had been when given by injection (301). This technique may prove important

for a wide range of ARIs.

Just as important as vaccination devices in the

tropics are technologies for the mass production,

storage, preservation, and distribution of vaccines. Concerns about these functions relate to the

stage of development of organized health services

as well as to medical technologies per se. One of

the most critical needs for immunization programs

involving certain vaccines, for instance, is the

maintenance of a “cold chain, ” the means to keep

vaccines cool during transportation from the laboratory to the vaccinee, wherever that person

might be,

Lack of adequate diagnostic technologies has

hampered the study and treatment of many tropical diseases. In addition to benefiting individual

patients, diagnostics are needed to learn about the

ranges of diseases and their prevalence and incidence. Although there is a wide range of conventional diagnostic tests that are adequate for some

tropical diseases, the need for rapid tests that do

not require sophisticated laboratory equipment

remains.

Biotechnology has made significant contributions in diagnostics. As a result, rapid diagnostic

tests, which can be used under field conditions,

are now under development. Diagnostic capability based on the DNA of the disease organisms

has also brought to light within-species differences

in disease organisms. Although the differences are

subtle, they may eventually have significance for

controlling diseases.

Research and development in therapeutic technologies for tropical diseases have lagged behind

research and development in technologies for diseases of U.S. importance. Few new drugs have

been introduced for human tropical diseases in the

past two decades, though there has been a surge

in the development of products for parasitic infections of domestic animals. One reason why

pharmaceutical companies have been reluctant to

invest heavily in drugs for tropical diseases is that

many of the potential beneficiaries of drugs cannot afford to buy them. While a company may

produce an effective drug, there may be no market for it.

Notwithstanding the general slow progress in

the development of therapeutics for tropical diseases, there have also been some new and exciting developments.

The discovery in the 1960s that glucose is actively transported in water into the body through

the intestinal wall even during severe diarrhea

paved the way for the development of ORT. ORT

is a major therapeutic measure for diarrheaI dehydration that has the potential to significantly

alter the mortality statistics of developing countries today. It is not a pharmaceutical in the usual

sense, nor is it even a curative agent. ORT is a

nonspecific treatment for episodes of diarrhea,

particularly prevalent among children, and responsible for at least one-third of all infant deaths

in developing countries.
12 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

ORT was developed as a treatment for cholera, first in adults, then in children and infants.

Its efficacy for a wide spectrum of diarrheal diseases has since been proved. Increasingly, less developed countries are undertaking national diarrheal disease control programs in which ORT

constitutes the keystone of the program. WHO

and AID both are supporting major programs in

ORT.

Praziquantel, a drug marketed in 1980, has revolutionized the treatment of schistosomiasis. Previously, schistosomiasis treatment itself involved

major health risks, and was a long process. Praziquantel has overcome both of these problems and

would probably replace the previous drugs of

choice (which have disadvantages) if it were not

so expensive.

Several new drugs for prophylaxis and treatment of malaria have been introduced in the past

decade. The development and spread of malaria

parasites resistant to chloroquine, for many years

the drug of choice, has made the quest for new

antimalarial drugs imperative.

Activity against a tropical disease by an antiviral drug, ribavirin, has been demonstrated for

the first time. This broad spectrum antiviral was

shown to be effective against Lassa fever in animals by DOD researchers. CDC scientists took

ribavirin to the field in west Africa, where Lassa

fever is a major problem, and the drug proved

effective in humans.

STATUS OF CONTROL FOR SELECTED TROPICAL DISEASES

Control measures for tropical diseases vary in

their availability, safety, and effectiveness. The

status of preventive, diagnostic, and therapeutic

technologies is summarized below for the diseases

considered in this report.

Malaria

Malaria is one of the most widespread diseases

in the world. In the last decade, its world prevalence has increased more than twofold (430).

Worldwide, an estimated 250 to 300 million cases

occur each year. In tropical Africa alone, an estimated 160 to 200 million people are infected every

year, and 1 million people die, mostly infants and

children. Human malaria is caused by four species of protozoan blood parasites of the genus

Plasmodium. Different species are important in

different parts of the world, although there is some

overlap. The parasites have a complex life cycle

and are transmitted to humans by mosquito vectors of the genus Anopheies.

The tools now available to control the mosquitoes that transmit malaria are inadequate to the

task. Varying degrees of resistance have been developed by the mosquitoes to every insecticide

that has been tried, and there is no reason to hope

that a new conventional insecticide will be more

successful on a long-term basis. Work on biological control methods (e.g., the introduction of mosquito predators) is needed.

Another problem in malaria control is the spread

of strains of malaria parasites that are resistant

to chloroquine and other antimalarial drugs.

Screening and testing of new compounds has proceeded at a steady pace since about World War

II, however, and a few of new drugs are now in

various stages of development and testing.

The most exciting development in malaria control is the prospect of a vaccine. Advances in biotechnology have made hopes for a vaccine realistic. With continued success, optimistic estimates

put a malaria vaccine on the market in 5 to 10

years (see case study B).

Schistosomiasis

Schistosomiasis is a chronic, debilitating, parasitic disease caused by trematode worms of the

genus Schistosoma that live in vertebrate host

blood vessels. The three major species of schistosomes that affect humans (S. mansoni, S. haematobium, and S. japonicuzn) have a complex life

cycle, requiring certain freshwater snails as inter-
mediate hosts. These parasites together have a

pantropical distribution. A 1972 survey including 71 countries estimated that 500 million people were exposed to schistosomiasis and 125 million were infected.

In recent years, there has been a drop in the

incidence and prevalence of schistosomiasis in

some areas and an increase in others. Preventing

transmission of schistosomiasis is accomplished

most effectively by reducing human contact with

infected water. Piped water supplies and excreta

disposal are therefore the most effective control

measures. Molluscicides (agents for killing snails)

are available, but all have adverse environmental

side effects and do not provide a permanent solution to control. Biological methods to control

snails are under investigation.

In some areas, large-scale hydroelectric and

agricultural irrigation projects have been responsible for spreading schistosomiasis by providing

year-round breeding sites for snails. The best

known case is the Aswan High Dam in Egypt. According to a 1937 survey, about 5 percent of the

population in upper and middle Egypt was infected with schistosomiasis. Following the construction of the Aswan High Dam, the prevalence

rate rose to 30 percent. Areas where large-scale

development projects are undertaken may require

special surveillance and control measures to prevent human health problems.

Chemotherapy for schistosomiasis, using Praziquantel, along with previously available drugs,

is effective and relatively safe. Work on a schistosomiasis vaccine is still in exploratory stages.

Trypanosomiasis

Trypanosomiasis is a group of clinically different diseases of the blood and tissues caused by

different species of the genus Tzypnosozna. The

important human diseases are African sleeping

sickness and Chagas’ disease.

African Sleeping Sickness

(African Trypanosomiasis)

African sleeping sickness is caused by two varieties of Trypanosoma brucei, which are transmitted by different species of tsetse flies (genus

Ch. l—Findings and Options • 13

Glossina). In west Africa, the disease is caused

by T. brucei gambiense, while in east Africa, it

is caused by T. b. rhodesiense. T. b. gambiense

causes a chronic, debilitating disease in humans

that saps the energy and eventually kills. T. b.

rhodesiense infects both humans and animals.

People become infected when they enter hunting

grounds and grazing areas where there are infected

animals. The disease is rapidly fatal to humans

and destructive to domestic livestock.

Control of tsetse fly vectors of sleeping sickness has, by and large, been unsuccessful. Consequently, large areas of land have been abandoned or left unsettled because of the threat of

disease.

Therapy for African sleeping sickness requires

hospitalization because of the use of toxic, intravenously administered, and frequently only partially effective drugs over an extended period of

time. Safe, effective, short course chemotherapy

is needed.

Chagas’ Disease (American Trypanosomiasis)

Chagas’ disease occurs in almost every country of Latin America. The disease is caused by

Trypanosoma cruzi,. a protozoan parasite that

lives in the blood and tissues. It is transmitted to

humans and about 150 other species of mammals

by reduviid bugs, blood-sucking insects found

throughout the Americas. Reduviid bugs are harbored in the mud and thatch of substandard housing, and the transmission of Chagas’ disease is

especially high in rural areas. In 1974, WHO estimated that out of 50 million exposed, 12 million

people were infected with T. cruzi.

The acute phase of Chagas’ disease, which may

cause heart and nervous system damage, can be

fatal, but it usually passes into a chronic stage.

Heart failure and grotesque enlargement of the digestive tract are among the long-term sequelae

which lead to death.

There is no effective treatment beyond the acute

phase and no vaccine. Control measures for Chagas’

disease concentrate on periodic insecticide spraying of houses. Improved house construction,

which could eliminate breeding sites for aduviid

bugs, could provide a permanent control solution.
14 ● Status of Biomedical Redearch and Related Technology for Tropical Diseases

Leishmaniasis

Leishmaniasis is the collective term for a spectrum of parasitic diseases caused by several species of the protozoan genus Leishmania. All species, which segregate largely into Old World and

New World forms, are transmitted by blood-sucking phlebotomine sandflies. In 1977, an estimated

400,000 new cases of leishmaniasis occurred

worldwide, and in some countries, the number

is increasing. Many wild animals in jungle areas

that are being cleared for agricultural development

are infected with leishmanial parasites, putting

new settlers at risk.

Depending on the infecting species, leishmaniasis takes several clinical forms. In the least severe form, cutaneous leishmaniasis, self-resolving

skin lesions appear at the site of the insect bite.

In the mucocutaneous form, sores may spread into

the nasal and pharyngeal mucous membranes, disfiguring the face, nose, and throat. The most destructive form, called “kala azar, ” attacks internal organs—spleen, liver, bone marrow, and lymph

glands—and in epidemics kills thousands of people. Recent epidemics in Asia have resulted from

reemergence of the sandfly vectors after spray

programs were discontinued.

There is no vaccine against leishmaniasis. Therapeutic drugs for treating leishmaniasis, mainly

antimony compounds, are not always effective

and have serious side effects. Rapid field diagnostic techniques are being developed through biotechnology. These techniques could both help patients by detecting disease earlier in its course and

facilitate field epidemiologic studies of the natural history of the disease. Overall, control of leishmaniasis is poor.

Filariasis

At least eight species, in several genera, of filarial nematode worms, threadlike in form, inhabit

the skin, other tissues, or the lymphatic system,

causing filariasis in humans. These parasites are

transmitted by blood-sucking insects.

Of the many filarial worms that infect humans,

three are of major public health importance on

a global scale: Wuchereria bancrofti and Brugia

malayi, both of which can result in elephantiasis;

and Onchocerca volvulus, which is the agent of

onchocerciasis (river blindness). W. bancrofti and

O. volvulus are widespread in Africa, and B.

malayi in parts of Southeast Asia. All three of

these species have become established to lesser degrees in the New World.

There are no vaccines to prevent filariasis.

Treatment for all forms of filariasis is similar.

There is effective therapy for the microfilariae that

cause the symptoms of disease, but no nontoxic

therapy against adult worms. All available drugs

have side effects, some serious. There is a great

need for improved filaricidal agents and immunopotentiating agents, drugs able to stimulate the

natural defenses of the body.

Leprosy (Hansen’s Disease)

Worldwide, an estimated 15 million people

have leprosy, most of them in the Old World. The

disease ranges from “tuberculoid” leprosy, with

localized skin lesions and minor nerve involvement, to the severe “lepromatous” leprosy, with

spreading, disfiguring lesions, resulting in destruction of the nose, involvement of the vocal cords

and eyes, and severe nerve damage.

The agent that causes leprosy, Mycobacteriuzn

leprae, is closely related to the bacterium that

causes tuberculosis. Although leprosy has been

recognized and feared for centuries, frustratingly

little is known of its natural history. Even the way

in which leprosy is transmitted is not clear. The

most likely means is through the respiratory tract,

infection being acquired by direct contact with infected individuals.

Treatment of leprosy involves months or years

to a lifetime of treatment with a combination of

drugs. There are only a handful of useful drugs,

most of which do not actually kill the bacteria,

but only arrest its spread. Resistance has developed even to the most effective drug. Considerable progress has been made toward a leprosy vaccine, but it is difficult to predict whether a vaccine

will be successful as a control measure for the

majority of those at risk. Technical problems in

studying leprosy, particularly scientists’ limited

ability to culture M. leprae in the laboratory, have

hampered research in all areas of leprosy control.
Ch. l—Findings and Options ● 1 5

Tuberculosis

Tuberculosis is a major public health problem

in most developing countries and is a resurgent

problem in some crowded, poor inner cities in the

United States. It is also the most common serious infectious disease among Indochinese refugees

to the United States. Tuberculosis is caused by

the bacterium Mycobacterium tuberculosis and

is transmitted from person to person by airborne

droplets. The lungs are infected first, but infection can subsequently spread to all parts of the

body. Infections are chronic and debilitating if

untreated.

The vaccine against tuberculosis is known as

BCG (Bacillus Calmette-Guerin) vaccine. BCG

vaccine is used mainly in areas of high transmission rates, but its effectiveness has shown great

variability in field trials in different parts of the

world. There is adequate treatment now for most

cases of tuberculosis, though there are drug-resistant strains. A minimum of 6 to 9 months treatment is necessary with the best drugs in combination. In many tropical countries that use less

expensive and less effective drugs, much longer

treatment is necessary. New antibacterial agents

are needed.

Diarrheal and Enteric Diseases

Diarrheal diseases occur throughout the world.

Research since the early 1970s has uncovered an

array of organisms that cause diarrheal diseases,

and there is a continuing need to identify and

characterize these organisms. The agents of diarrheal diseases include viruses, bacteria, and protozoa. These agents are transmitted to humans by

fecal contamination of food or water.

The greatest danger in diarrheal disease, regardless of the specific cause, is severe, life-threatening

dehydration and shock. In adults, diarrheal diseases other than cholera are generally not lifethreatening. In children under 5 in many developing countries, however, diarrheal diseases are

the leading cause of illness and death, Infants and

young children in developing countries may experience four to eight separate episodes of diarrhea

per year. One out of every 150 to 200 episodes

results in severe, life-threatening dehydration.

One way to prevent diarrheal diseases is to

break the transmission cycle by providing clean

water and sewage disposal. The encouragement

of breast feeding, proper weaning practices, and

health education could also help. Vaccines against

rotaviruses, Shigella, and Sahnonella, the agents

of a large percentage of childhood diarrhea, are

undergoing development.

From World War II through the 1970s, the only

accepted means of treating diarrheal dehydration

was through intravenous replacement of body water and salts. In the late 1960s and continuing into

the 1970s, a therapy for diarrheal dehydration was

developed that has been called the most important therapeutic advance in tropical medicine.

That therapy, ORT, involves the oral administration of a simple solution of water, salt, and sugar

and thus does not require hospital facilities (see

case study B). ORT has revolutionized the treatment of adults with cholera and, even more importantly, is effective in infants and children with

a wide range of diarrheal infections.

Acute Respiratory Infections (ARIs)

ARIs are among the most important causes of

preventable deaths in the world. These infections

include both lower respiratory tract infections

(e.g., pneumonia and bronchitis) and upper respiratory tract infections (e.g., influenza, measles,

diphtheria, and whooping cough). The organisms

that cause ARIs are transmitted by airborne droplets and include viruses, bacteria, and mycoplasma.

In developed countries, the toll from ARIs

(other than influenza in pandemic years) is relatively low and stable. ARIs are exacerbated by

malnutrition and substandard living conditions,

however, and in developing countries, these infections take a large annual toll. Some 12 percent

of all deaths of children living in Africa, Central

America, and Asia are attributed to ARIs. ARIs

not only pose a serious mortality risk for the very

young and very old in developing countries, but

reduce productivity in all age groups and impose

tremendous demands on health care systems.

Effective vaccines are available for measles, pertussis (whooping cough), and diphtheria, but a

large percentage of the world’s children have not
76 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

been vaccinated. Immunization rates have increased in some countries as a result of WHO’s

Expanded Program on Immunization (EPI), but

EPI’s goal of universal vaccination of all children

is still far from being realized. Vaccines for pneumococcal pneumonia and influenza are available,

but have limited use in developing countries.

The lack of adequate rapid diagnostic methods

that can be used in less developed areas of the

world hampers effective treatment of ARIs. Some

researchers are attempting to develop new diagnostics with the methods of biotechnology. Effective treatments exist for many of the bacterial and

mycoplasmal ARIs, but the development of antiviral chemotherapy is still in its infancy. For bacterial ARIs, drug resistance to penicillin and other

antimicrobial is an increasing problem.

Arboviral and Related Viral Infections

The name “arbovirus” is derived from the descriptor “arthropod-borne virus. ” The unifying

characteristic of arboviruses is that they replicate

in and are transmitted by arthropods (predominantly mosquitoes, but also ticks, sandflies,

midges, and gnats). Arboviruses occur worldwide, infecting humans and other animals. The

80 or so human types have varying distributions

and exhibit an apparent trend toward spread rather than containment. Many of the viruses that

principally infect animal populations also cause

occasional severe outbreaks in human populations.

Important arboviral infections in terms of severity and prevalence are yellow fever, dengue fever,

oropouche fever, chikungunya, Japanese encephalitis, other viral encephalitides, and hemorrhagic

fevers. Clinically, arbovirus infection can cause

either mild disease (e.g., acute benign fevers or

self-limiting arthritic symptoms), severe central

nervous system disease with brain inflammation

which can be fatal, or hemorrhagic fevers with

high case-fatality rates. Yellow fever also causes

liver damage and jaundice.

Effective vaccines exist for yellow fever and Japanese encephalitis, and work is proceeding on vaccines for dengue fever and a few others. Vector

control, at present, provides the most promising

avenue for control. Development of therapeutic

drugs for arboviral infections is still in its infancy.

Treatment for such infections now consists mainly

of relieving symptoms.

U.S. EFFORTS IN TROPICAL DISEASE RESEARCH

Rationale for U.S. Efforts in 1.

Tropical Disease Research

The reasons for U.S. involvement in tropical

disease research, and in the whole area of international health, have not changed since the end

of World War II. It was at that time that the

United States began to provide substantial

amounts of aid to encourage the development of 2.

less developed nations. The Interdepartmental

Committee on International Health Policy, a

group set up in 1960 as one attempt at coordination among the interested agencies, identified four

basic objectives of international health policy.

Restated here, they are (75):

Social and humanitarian objectives. The

ideals and the humanitarian and philosophical beliefs basic to the American democratic

tradition should be reflected in U.S. international health activities. The desire for the

world’s people to enjoy good health is an extension of the attitude of Americans toward

their own health.

Political objectives. Involvement in international health contributes to the U.S. position

of world leadership. The United States is not

alone in supplying aid in international health.

In addition to European nations, Cuba, for

instance, is extremely active, with health

personnel now in about 25 countries (348).
Ch. l—Findings and Options ● 1 7

3.

4.

Health programs can contribute to the political stability of developing nations.

Economic objectives. Improved health in developing countries can have economic benefits for those countries as well as for the

United States. For the countries themselves,

increased productivity can accompany gains

in health. For the United States, markets for

U.S. goods and the atmosphere for U.S. investments may improve with increased involvement in the health sector.

Medical objectives, including self-protection

against diseases. The self-interest of protecting Americans from tropical diseases is increasing in importance as the world community grows smaller. Americans abroad, as

well as at home, may be exposed to the risks

of tropical diseases. In addition, there may

be spinoffs in understanding or controlling

diseases of importance in the United States.

For example, techniques developed during

the campaign against smallpox have been

successfully applied to measles control in this

country (161).

To these four may be added a fifth objective:

5. Scientific objectives. Tropical diseases pose

challenging problems for scientists who are

interested in advancing knowledge and improving health.

U.S. Activities in Tropical

Disease Research

The major U.S. Government supporters of

tropical disease research are: NIH, mainly the

National Institute of Allergy and Infectious Diseases (NIAID), and CDC (both in DHHS); DOD;

and AID. Each U.S. Government agency supporting tropical disease research acts in accordance

with its own specific mandates, which by and

large do not overlap with the mandates of other

agencies. NIH funds research of scientific merit

that will advance the state of knowledge about

causative agents, the response of humans to the

disease agent, and the disease itself. CDC responds

to specific problems around the world in the control of diseases as they affect the health of U.S.

citizens, and also maintains applied research programs in diseases of public health concern (e.g.,

malaria and helminthic diseases) (27). DOD focuses on health problems of direct relevance to

U.S. military personnel in different parts of the

world. Most AID-funded research has relevance

to development assistance and political objectives.

The Federal agencies that fund research do so

under various legislative mandates, which for the

most part allow, but do not necessarily encourage,

delving into tropical health issues. To quote a 1978

report of the White House Office of Science and

Technology Policy (27):

At best, current authorization is passive and

certainly does not act as a stimulus. Other legal

restrictions further limit the use of agency authorizations to support international health research.

The situation has not changed appreciably since

that report was issued.

The U.S. Government also contributes to research through programs supported by WHO and

through TDR (the Special Program for Research

and Training in Tropical Diseases) which is

administered by WHO, cosponsored by the U.N.

Development Program and the World Bank, and

funded by international contributions).

Private sector supporters of tropical disease research include foundations, pharmaceutical companies, volunteer agencies, and religious organizations. Private U.S. foundations have programs

that have often served to highlight problems and

identify opportunities not exploited by Federal

agencies. The Rockefeller Foundation’s Great Neglected Diseases program and the Edna McConnell

Clark Foundation’s focus on schistosomiasis are

examples.

U.S. Funding of Tropical

Disease Research

It is difficult to pin down a dollar figure that

represents U.S. research in tropical diseases, because information is not standardized across agencies, and definitions of tropical diseases differ. On

the basis of analyses of information developed for

this assessment, OTA estimates that, in recent

years, the U.S. Government has annually contributed less than $100 million toward research in

tropical diseases, out of a total annual biomedi-
18 ● Status of Biomedical Research and Related Technology for Tropical Diseases

cal research budget of well over $4 billion. Contributions of private foundations are of lesser magnitude. Figures on tropical disease research

funding by pharmaceutical companies are undocumented.

U.S. Department of Health and

Human Services

NIAID’s tropical medicine funding covers research in three general areas: 1) traditional tropical diseases (malaria, schistosomiasis, trypanosomiasis, leishmaniasis, filariasis, and leprosy);

2) general parasitology (cestodes, nematodes,

trematodes, and protozoa); and 3) general tropical medicine (rickettsia, bacteriology, mycology,

virology, and vector pathogens). In fiscal year

1983, NIAID funding for tropical medicine research was about $33 million, about 12 percent

of NIAID’s budget (less than 1 percent of the total NIH budget). About one-quarter of this funding supported researchers working at NIH (intramural research), and three-quarters went to

researchers outside NIH (extramural research). In

addition to the $33 million for tropical medicine

research, NIAID spent approximately $3 million

on projects in diarrheal diseases and ARIs (a large

proportion of ARI funding is for influenza research, directed at domestic problems).

The Fogarty International Center of NIH does

not have a tropical disease research program, but

approximately $2 million annually is channeled

through the Fogarty budget to the Gorgas Memorial Institute, for the operation of its laboratory

in Panama. Fogarty also holds international conferences and provides fellowships to foreign scientists, some of which contribute to the overall effort in tropical disease research.

CDC conducts tropical disease research primarily through the Center for Infectious Diseases and

its Division of Parasitic Diseases. In fiscal year

1983, CDC spent an estimated $5 million on tropical disease research. This includes support of

CDC’S Medical Entomology Research and Training Unit in Guatemala.

U.S. Agency for International Development

Most AID funding for tropical disease research

comes from the Office of Health. In fiscal year

1982, this amounted to about $14 million. About

$5 million of this was the U.S. contribution to

TDR, about $5.8 million was spent on research

in malaria immunology and vaccine development,

and $1.9 million was part of the core support for

the International Center for Diarrheal Disease Research in Bangladesh. AID’s Africa Bureau spent

approximately $1.3 million on the biomedical research components of three projects: onchocerciasis control, combatting communicable childhood diseases, and schistosomiasis activities in

Cameroon and the Sudan.

AID’s Office of the Science Advisor supports

additional research at the level of about $1 million in grants through the Program in Science and

Technology Cooperation (PSTC) and through the

National Research Council’s Board on Science

and Technology for International Development

(BOSTID). Most of these funds go to researchers

in developing countries.

U.S. Department of Defense

In fiscal year 1982, DOD spent about $13.5 million for biomedical research in tropical infectious

diseases, representing about 6 percent of all DOD

biomedical research funding. The Army and Navy

conduct a certain amount of research themselves,

as well as contracting out research both in the

United States and in the tropics. The U.S. Army

currently maintains the only large-scale antimalarial drug screening program in the world, It also

maintains medical research units in eight developing countries, the largest such program of any

U.S. Government agency.

U.S. Contributions to International Programs

The Special Program for Research and Training in Tropical Diseases (TDR), administered by

WHO, is cosponsored by WHO, the World Bank,

and the U.N. Development Program. TDR was

initiated in 1975 and has two main objectives: 1)

to strengthen the biomedical research capabilities

of the developing countries; and 2) to develop

tools to control six tropical diseases (malaria,

schistosomiasis, trypanosomiasis, leishmaniasis,

filariasis, and leprosy).

By the end of 1981, TDR had received more

than $95 million in contributions from 25 gov-
Ch. l—Findings and Options ● 1 9

ernments, other organizations, and the sponsoring agencies. The U.S. annual contribution to

TDR was $4 million in 1980 and 1981 and just

over $5 million in 1982.

WHO supports some tropical disease research

in addition to its contributions to TDR, Under the

general classification of communicable disease

prevention and control, about $2.4 million of

WHO funds went for research in tropical diseases

during the 2-year period 1980-81.

WHO also supports two separate programs relevant to tropical diseases: one in ARIs and one

in diarrheal diseases. Over the 2-year period 1980-

81, WHO spent $381,000 for ARI research and

just over $1 million for diarrhea] disease research.

WHO’s 1982-83 diarrheal disease research budget

was estimated to be about $3.8 million.

Precisely determining the U.S. contribution to

WHO research programs is difficult, because

funding for each set of projects comes both from

the “regular budget” and from a spectrum of “extrabudgetary sources, “ in varying amounts. For

the period 1980-81, the annual U.S. monetary

contribution is estimated by OTA to be between

$250,000 and $500,000.

Funding of U.S. Researchers in

Tropical Diseases

Most of the tropical disease research funded by

U.S. institutions is carried out by U.S. researchers. In fiscal year 1982, for example, NIAID

awarded 96 percent of its tropical medicine funds

to U.S. institutions and 4 percent to researchers

in other industrialized countries. Often, however,

OPTIONS

Development of Medical Technologies

for Tropical Diseases

To encourage research and development of

medical technologies for tropical diseases, Congress might consider the following options.

OPTION 1: Explicitly include drugs and vaccines for tropical diseases in the definition of “orphan drugs” under the Orphan Drug Act of 1983

(Public Law 97-414).

investigations undertaken by U.S. institutions are

carried out in close collaboration with institutions

in less developed countries.

AID’s Bureau of Science and Technology allocates almost 75 percent of its biomedical research

funds to American institutions. Many of these institutions collaborate with organizations in the less

developed countries. AID’s Research Grants Program, administered by the Science Advisor’s Office and by BOSTID (NAS), has so far allocated

about 45 percent of its resources to American institutions, though the BOSTID portion of the program makes grants only to institutions in developing countries.

The DOD biomedical research program allocates about 75 percent of its extramural funds to

organizations in the United States, 18 percent to

institutions in less developed countries, and 5

percent to organizations in other industrialized

countries.

The majority of biomedical research funds both

of the Rockefeller Foundation (52 percent) and of

the Edna McConnell Clark Foundation (77 percent) were awarded to American institutions.

U.S. researchers also receive funds from international organizations in excess of the U.S. contributions to such organizations’ research budgets.

In 1981, TDR awarded about one-third of its research funds to Americans, nearly equal to the

U.S. contribution. The same year, WHO’s program in diarrheal diseases awarded one-quarter

of its research funds to American institutions, but

the U.S. contribution represented 1 percent of the

program’s total budget.

OPTION 2: Encourage Federal agencies, such

as AID, to examine the possibility of interesting

private companies in developing medical technologies for tropical diseases by guaranteeing purchases of products and assisting in field trials.

OPTION 3: Mandate the creation of and authorize funds for a quasi-governmental nonprofit

corporation to undertake research and development of medical technologies for tropical diseases

until the technologies become economically attrac-
20 ● Status of Biomedical Research and Related Technology for Tropical Diseases

tive enough for private industry to take over with

the right to an exclusive license for the product.

OR

Stimulate the development of an international

nonprofit corporation, funded through contributions from the U.S. Government, other governments, and international bodies, to undertake

such research and development.

OR

Create a nonprofit corporation charged with

ensuring the development and availability of prophylactic and therapeutic agents for use in developing countries for which there appears to be insufficient commercial interest.

Relatively few U.S. pharmaceutical companies

are pursuing the development of drugs and vaccines for tropical diseases. The few that are often

have entered the field because research for tropical parasitic diseases of humans is a spinoff of research on parasitic diseases of domestic agricultural animals. There is, at present, little financial

incentive for pharmaceutical company activity in

most human tropical diseases. The 1978 Office of

Science and Technology Policy report states (27):

There is almost no domestic U.S. market for

vaccines, drugs, or pesticides used against tropical diseases. The main potential purchasers of

these products are developing countries or international assistance organizations acting on their

behalf. At present, these markets are unprofitably small and offer no realistic incentive for industry research in this area. . . .

We conclude that underutilization of existing

drugs and vaccines, researched and developed at

considerable expense by industry, is a major disincentive to new investment in tropical medicine

research and development by pharmaceutical

firms.

It is unfortunate that financial incentives are

lacking to develop drugs for some of the most

widespread diseases of humankind. The lack of

profitable markets does not correlate with a lack

of patients, but with the high price of newly developed drugs (necessary to recoup research and

development costs); the relatively small health

budgets of developing country governments and

the inability of most people in developing countries to pay for the drugs themselves; and unstable political conditions which make it risky for

U.S. companies to rely on trade with specific developing countries.

The new medical products developed with biotechnology, largely vaccines and diagnostics,

might provide a stimulus for activity by the pharmaceutical industry. With much of the developmental work on these products coming from

publicly funded research, the research and development costs incurred by industry should be

somewhat less. Much of the expense of production, particularly for vaccines, however, is incurred in scaling up production and in clinical

trials.

As the malaria vaccine now under development

in the United States illustrates, there are also issues to be addressed in the commercialization of

publicly funded research. Funding for the malaria

vaccine’s development has come from various

U.S. Government agencies, private foundations,

and WHO. WHO’s policy is that the fruits of

WHO research funding should be available to the

world at large. This policy means that a commercial company desiring to produce the malaria vaccine, should it move to the production stage,

would not be allowed to secure a patent. Because

substantial costs are involved in developing a

process for large-scale production of the vaccine,

many companies would be unlikely to undertake

the project without patent protection.

The Orphan Drug Act of 1983 charges the U.S.

Government with the task of identifying and promoting “orphan products, ” defined as drugs and

devices for rare diseases or conditions. The act

defines rare diseases and conditions as ones that

occur so infrequently that there is no reasonable

expectation that the cost of development can be

recouped by sales within the United States.

Four kinds of support for orphan drugs are authorized by the 1983 act:

1. A 50-percent tax credit on all clinical testing expenses associated with the drug.

2. Award of an exclusive 7-year right to market a drug that is unpatentable (through the

Food and Drug Administration’s new drug

approval authority).
Ch. l—Findings and Options ● 2 1

3.

4.

In

crue

.

Technical assistance in the development of

clinical testing protocols.

Award of grants and contracts for clinical

testing expenses associated with an orphan

drug.

addition, public relations benefits may acto pharmaceutical companies developing orphan products.

The Orphan Drug Act is aimed at diseases that

are rare in the United States, certainly the case

with most tropical diseases. It does not address

the prevalence of diseases in other parts of the

world. The Food and Drug Administration (FDA)

is planning to publish for public comment proposed regulations under the act. None now exist, though FDA is operating under interim guidelines and is designating orphan products. The

regulations, when final, could encourage research

and development in drugs for tropical diseases.

Even with the incentives of the Orphan Drug

Act, there is little reason to expect a major increase

in the activities of U.S. pharmaceutical companies in tropical disease research. One alternative

is some form of public funding expressly for pharmaceuticals for tropical diseases. Although setting up a nonprofit corporation is not the only

means of injecting public funds into the process,

there might be advantages to setting up a corporation, rather than simply adding money to TDR

or increasing grants through NIH. The nonprofit

corporation’s research could be expressly aimed

at eventual commercialization, and rules could be

set up for the transfer of projects to for-profit

pharmaceutical companies at the appropriate

time.

Information for Congressional

Decisions

OPTION 4: Hold a special appropriations

hearing for tropical disease research with representatives from NIH, CDC, DOD, and AID, and

perhaps invite international agencies and private

foundations to participate.

AND/OR

OPTION 5: Require each agency mentioned

above to submit a short report on the status of

its tropical disease research, providing data specified by the Appropriations Committees for use

during appropriations hearings.

Evaluating the adequacy of U.S. research efforts in the area of tropical diseases is difficult,

because no specific goals have been enunciated

except in the case of U.S. contributions to the

TDR and WHO programs, By and large, each

U.S. Government agency involved in funding or

carrying out tropical disease research does so

according to its own needs, and, in most cases,

through legislation that passively allows tropical

disease research activities rather than promotes

them.

In the past, suggestions have been made to improve coordination among the Federal agencies

involved with tropical disease research (27,252).

Two very basic aims of such coordination would

be: 1) to avoid duplication of effort, and 2) to

identify gaps in the overall research agenda.

OTA found no evidence that the tropical disease research efforts supported by different Federal agencies are duplicative. The professional

community in international health is relatively

small and closeknit, and within that community,

there is a high degree of awareness of the various

activities taking place. Undetected duplication of

effort is relatively unlikely also because the total

amount of money spent on tropical disease research is small.

There do appear to be gaps in current tropical

disease research activities, however, and some of

them are identified in this assessment. It is possible that a coordinating committee of some sort,

with representatives from the relevant agencies,

could identify gaps and devise plans to fill them.

However, the options cited above would allow

the Appropriations Committees to see the range

of related activities in tropical disease research and

might serve as well as or better than a coordinating committee.

Looking at just one component of tropical disease research does not allow an assessment of the

adequacy of the effort. A look at the mix of Federal funding for malaria research, the most heavily

funded disease-specific research, illustrates the

value of examining information across agencies.
22 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Of the funds NIAID spends for research on the

six TDR tropical diseases, about 20 percent goes

to malaria research. (The highest percentage of

those NIAID funds, about 25 percent, goes to

trypanosomiasis research. ) The focus of NIAID’s

malaria work is on immunology and vaccine

work. Of the money CDC spends for research on

the six TDR diseases, 60 percent goes for malaria

research. CDC maintains a colony of monkeys,

large colonies of mosquitoes, and laboratory facilities for studies of malaria transmission, drug

testing, and other biological characterizations.

DOD is heavily involved in the screening and testing of potential antimalarial drugs. Most of the

drugs currently available for chemoprophylaxis

and treatment of malaria either have been developed by DOD researchers or their contractors or

have benefited from DOD research. About 60 percent of DOD’s budget for the TDR diseases goes

for malaria research. AID’s research activities in

tropical diseases have focused on malaria immunology and vaccine development,

TDR itself allocates about 30 percent of its annual budget for malaria research. Most of TDR’s

funded projects concern immunology, including

vaccine development, and chemotherapy of malaria. More than one-third of the WHO budget

for parasitic disease research is spent on malaria,

To obtain basic information comparable across

Federal agencies, the Appropriations Committees

could provide a framework for the presentation

of information at hearings or in a report from

agencies.

The following types of information might be

included in the reports from U.S. Government

agencies:

1. A summary of funded activities for each disease from a core list of tropical diseases (e.g.,

the diseases covered in this report). The information listed below might be required, including, where appropriate, indications of

which research is strictly associated with

tropical diseases and which only partially:

—type of research—basic or technologyoriented;

—if basic, category (immunologic, physiologic, etc.); and

—if technology-oriented, category (preventive, diagnostic, therapeutic).

2.

3.

4.

5.

6.

The same information as in (1) for other diseases at the discretion of each agency:

—number of projects; and

—total funding levels.

A rationale for the allocation of funds.

An indication of how much research is

funded at institutions in the tropics, and how

much in the United States, and acknowledgment of funded American researchers collaborating with researchers in the tropics.

An indication of U.S. contributions to international research efforts (principally WHO

and TDR).

An assessment of gaps in research as perceived by the agencies.

Private foundations and international agencies

funding tropical disease research could provide

similar information about their activities and

could provide additional perspective on research

needs.

Even given specific categories, Federal agencies

will not be able to clearly identify all research

directed at tropical diseases. There are two basic

reasons.

First, some research will address health problems that are important both domestically and in

developing countries. A clear example is research

on ARIs, which are prevalent all over the world.

Most of the ARI research funded by U.S. institutions is directed at the U.S. problem, particularly

influenza, but progress may have spinoffs for the

tropics.

ARIs are caused by a range of diverse and unrelated organisms, and one important research

problem is to gauge the importance of different

causal agents. The spectrum of ARI agents in the

United States is probably quite different from that

in the tropics, in which case, advances in diagnosis and treatment may benefit the United States

without having an appreciable effect on conditions

in developing countries. Some ARI research, however, is directed specifically at conditions in the

tropics. In examining funding for tropical diseases,

either including or excluding all general ARI research would be misleading. In cases such as ARI,

the agencies could be asked to submit total funding amounts and estimates of the proportion related to conditions in the tropics, with a narrative describing how the breakdown was made.
Ch. l—Findings and Options ● 2 3

The second reason for the difficulty in assigning research to the tropical disease category is that

many tropical disease parasites, trypanosomes for

instance, have become subjects of heuristic study

in basic science laboratories. Because these parasites are interesting organisms, researchers have

begun using them to study basic functions, such

as the transport of substances across membranes

and, in many cases, basic immunology, gene transcription, and gene regulation. These studies are

not necessarily directed toward the control of disease and may be successful without ever contributing to a diagnostic, preventive, or therapeutic

technology.

Research Funding

OPTION 6: Increase Federal funding for all aspects of tropical disease research.

OPTION 7: Amend the international health

mandate of the U.S. Department of Health and

Human Services (DHHS) to remove the limitations on the research DHHS may support in tropical diseases.

U.S. Government funding for tropical disease

research in recent years has been less than $100

million annually, out of an annual Federal biomedical research budget of $4 to $5 billion. It is

understandable that the disease problems of importance in the United States consume most of

the resources, but there are perhaps a billion people in other countries suffering from or at risk for

diseases which most U.S. citizens will never encounter.

Since the focus of this assessment has been tropical disease research itself, OTA has not identified specific funding programs or institutional

bases for increases in funding. An NAS study now

in progress is developing information that will be

of direct value in formulating such programs and

identifying the institutional resources that exist

in this country. In the short term, however, it appears that the funding mechanisms that now exist could productively absorb funding increases.

Current authority for international health research within DHHS derives primarily from the

International Health Research Act of 1960 (Public Law 86-61 o), the purpose of which is “to advance the status of the health sciences in the

United States and thereby the health of the American people through cooperative endeavors with

other countries in health research and research

training.” The authority to undertake research for

the good of people outside the United States rests

with the President.

Congress could transfer the authority to make

decisions about all aspects of tropical disease research to DHHS itself, the Department which includes NIH and CDC. It could do this by amending the language of the International Health

Research Act to give DHHS the authority to “advance the status of the health sciences in the

United States and thereby the health of all

people. ”

Such a change would explicitly provide greater

justification for DHHS research on diseases that

are not major public health problems in the United

States. Giving NIH, in particular, more flexibility to fund research on tropical diseases could

stimulate the submission of a greater range of

grant proposals and might provide the basis for

additional contracts in the area of tropical

diseases.
2 .

The United States in

International Health
Contents

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Rationale for U.S. Involvement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Political Objectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Economic Objectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Scientific Objectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

U.S. Support of International Health Activities . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

The International Health Research Act of 1960 . . . . . . . . . . . . . . . . . . . . . . . . . . .

Support of Multilateral Efforts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

U.S. Reports on Tropical Disease Research and Training . . . . . . . . . . . . . . . . . . . . .

1962 NAS/NRC Report on Tropical Health . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Current NAS Study of U.S. Capacity To Address

Tropical Disease Problems . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Tropical Disease Research in the Context of U.S. International Health

Activities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

27

27

27

28

29

30

30

31

31

31

32

33
2 ■

The United States in International Health

INTRODUCTION

Over the course of the 20th century, the U.S. ter describes the reasons for U.S. participation in

Government and U.S. scientists and medical international health activities, and then discusses

workers have sustained an interest in the health some of the important influences on those activities.

of people in other parts of the world. This chapRATIONALE FOR U.S. INVOLVEMENT

In 1983, the United States spent more than $14

billion in foreign aid to less developed countries.

Although much of this aid went for defense, the

United States increasingly has recognized the

value of assisting developing countries with health

services and building health infrastructure. The

health budget of the U.S. Agency for International

Development (AID) for 1983 was about $123

million.

Five basic objectives of U.S. activities in international health were discussed in chapter 1. In

general, the reasons for U.S. involvement in international health activities apply to the subset

of tropical disease research. Briefly, the motives

for U.S. involvement are:

social and humanitarian;

political;

economic;

medical, including protection of U.S. citizens

against disease; and

scientific.

The social and humanitarian motive is either

convincing or not to an individual or a society,

and is something that can shift in importance over

time.

The medical motive, particularly to protect

U.S. citizens, may be gaining strength, as it is becoming more apparent that opportunities for

worldwide disease transmission have been and are

continuing to increase. A still small but growing

number of U.S. citizens are exposed to tropical

diseases both abroad and at home (see ch. 1).

Furthermore, the opportunities for worldwide

transmission of disease have recently been demonstrated by the emergence of acquired immunodeficiency syndrome (AIDS) as a public health

problem in the United States, particularly among

the gay community. There is convincing evidence

that the syndrome originated in Africa and is

present in the heterosexual community there. Its

appearance in the United States probably came

through migrations of Africans to Haiti, where

vacationing Americans came in contact with it

(362).

The political, economic, and scientific objectives of U.S. involvement in international health

activities are discussed further below.

Political Objectives

One aim of all U.S. international aid is to win

friends for the United States, and thus sustain the

U.S. role as a world leader. Contributions in

health, particularly toward primary health care,

may also be one paving stone on the road to social harmony and political stability. Furthermore,

there is an opportunity to raise grassroots support for the United States by having Americans

participate at local levels in developing countries.

The United States is not the only country providing health aid. One of the more active countries

in recent years has been Cuba. Cuban medical

personnel are present in more than 25 countries

(348). Certainly in relation to size, Cuba accords

tropical medicine much greater importance than

27
28 ● Status of Biomedical Research and Related Technology for Tropical Diseases

does the United States. Cuba has virtually eliminated its classical tropical disease problems, so its

present emphasis is not mainly a response to

domestic needs, though surveillance to prevent

reintroductions of once prevalent diseases is a

priority.

In the summer of 1983, when issues concerning continued funding of the Gorgas Memorial

Laboratory in Panama were addressed by OTA

and the General Accounting Office, the role of

Cuba was raised by an official of the U.S. State

Department. It was feared that “closure of the

Laboratory would give Cuba the opportunity to

attract undue attention to its institute [the Cuban

Institute of Tropical Medicine] which could be disadvantageous to the United States” (355). Going

beyond the specific case of Gorgas, countries may

be inclined to accept health aid from those best

equipped and most willing to provide it.

Economic Objectives

There is intuitive appeal to the notion that improvement in the health of a population will lead

to economic improvement. Solid proof of this intuition has been elusive, but some evidence now

exists to support it. Belief in the concept that

“health pays” is reflected in the actions of the

World Bank over the past years.

The World Bank is an institution which, like

commercial banks, requires that projects for

which loans are given be economically viable. The

Bank first began making loans for health components of development projects in 1975. The Bank’s

experience from 1975 through 1978—involvement

in 70 health components of development projects

in 44 countries—prompted the Bank in 1980 to

begin lending directly for health projects.

The economic reasons for supporting health

projects are set forth in the second edition of the

World Bank’s Health: Sector Policy Paper (412).

The economic effects of ill health, on which the

Bank bases its policy, are summarized below.

1. Reducing Availability of Labor. The effective labor force is reduced by premature death and

by illness. The death of workers may not have

a national impact on productivity in developing

countries, where unemployment is high and there

is a steady supply of replacements. Premature

death may, however, impose some costs on society when dependents are left.

Morbidity of workers, resulting in absenteeism,

has a more direct effect on overall productivity.

That effect has been measured in a few studies,

though it is generally difficult to prove. Antimalarial programs in the Philippines and parts of

Africa, and yaws control in Haiti have substantially reduced absenteeism (412).

2. Impairing Productivity of Labor. The ability of a sick worker is impaired not only in terms

of physical capacity, but also in terms of the capacity to concentrate and think. The World Bank

studied construction and rubber plantation workers in Indonesia, 85 percent of whom had hookworm infestations. After the workers were given

iron supplements to correct the anemia caused by

the worms, productivity increased 19 percent, for

a cost of only 13 cents per worker over 12 months

(17).

At a more elemental level, the ability to learn,

at all levels of education and training, is impaired

by ill health. Absence from school compounds the

negative effect of sickness.

3. Wasting Current Resources. Resources are

used up in often ineffectual treatments of diseases.

Even when effective, many treatments afe costly

and pull disproportionate amounts from the overall health budget. Many illnesses also cause an increase in calorie use by the body, using up scarce

food resources, and in turn contributing to malnutrition.

4. Impeding Development of Resources. Some

areas of the world, particularly parts of Africa,

are effectively closed to development because of

disease threats. Some areas have been settled only

to be abandoned because of the lack of effective

control measures for diseases. African sleeping

sickness (African trypanosomiasis) and river blindness (onchocerciasis) render parts of Africa uninhabitable by humans and/or domestic animals.

The World Bank and the World Health Organization (WHO) have undertaken vector control programs to open some areas. Making areas safe for

inhabitants also can increase the attractiveness of

a country for tourists, a definite economic

advantage.
Ch. 2—The United States in International Health ● 2 9

What of the cost of the 10-year campaign? Approximately $83 million has been spent in international assistance for the smallpox eradication

program since 1967. The endemic countries themselves have spent roughly twice that amount, but

few of them have spent much more than they

were already spending on smallpox control. The

total amount of money spent in international

assistance is little more than half what was computed in 1968 to be the yearly expenditure for

smallpox control in the U.S. alone; worldwide

expenditures for smallpox vaccination and quarantine measures have been estimated as being in

the range of from $1 billion to $2 billion a year.

With the eradication of the disease, smallpox

vaccination will no longer be required, nor will

international certificates of smallpox vaccination.

Apart from the alleviation of human suffering,

the savings have already repaid the small investment many times over.

In addition to directly saving money, improved

health can lead to a generally higher quality of

life and, ultimately, to improvements in the economic development of a country.

Scientific Objectives

The tropical diseases of particular scientific interest today are the parasitic diseases, mainly

those caused by protozoa (e.g., malaria, trypanosomiasis, leishmaniasis) and helminths (e.g., filariasis, schistosomiasis). While U.S. research on

these diseases is still at a very low level in comparison with research on diseases of domestic importance, scientific interest has been heightened

by advances in immunology and biotechnology.

The parasites that cause tropical diseases are more

complex immunologically than the organisms that

cause infectious diseases common in the “United

States, and these parasites have captured the attention of both basic and applied scientists. Their

research activities are highlighted in this report,

particularly in chapters 6, 7, and 8.

In a more general sense, scientists want their

work to have an effect, and such desires do not

stop at political boundaries. Advancing the control of diseases in developing countries is a great

scientific challenge, and success in meeting it

would be richly rewarding in terms of bettering

the human condition.
30 ● Status of Biomedical Research and Related Technology for Tropical Diseases

U.S. SUPPORT OF INTERNATIONAL HEALTH ACTIVITIES

For the past four decades, U.S. Presidents and

Members of Congress have shown sporadic enthusiasm for international health activities. Nearly

every President since Truman has spoken out for

a U.S. role in international science and international health. From President Truman’s “Point

Four Program” through President Carter’s assertion that “Health is a basic human right, ” improvements in health have been seen as part of

the solution to social, political, and economic

problems. Congress has acted similarly, but the

pronouncements and legislation have not been

matched by funding for new programs.

One observer summarized the history of the interactions as follows (75):

. . . the process over a period of time has become

cyclical. The Legislative and Executive branches

have not been able to agree on a course of action because the enthusiasm of one branch of

Government has been asynchronous with that

of the other. Furthermore, the Office of Management and Budget has deterred any Departmentor Agency-generated initiatives which would entail increased expenditures abroad.

The International Health

Research Act of 1960

In 1958, Senator Lister Hill introduced a bill to

institute broad international programs in health

research. Congressman John Fogarty introduced

an identical bill in the U.S. House of Representatives. The specific measures called for: 1) a National Advisory Council for International Medical

Research; 2) a National Institute for International

Medical Research as part of the National Institutes of Health (NIH); and 3) a $50 million annual budget authority for international medical

research.

Senator Hill stressed the need not only for research, but for information exchange and for

training of research personnel. He contrasted his

$50 million budget authority for international

health research with the $400 million then being

spent on medical research in the United States and

the $40 billion for defense that had just been authorized.

The bill passed the Senate, but the House made

changes consistent with the wishes of the Eisenhower Administration and removed many of the

specific provisions. The objections to the Senate

version, according to Corning, were as follows

(75):

. . . too much money; a new institute at the NIH

for its administration was unnecessary; the international program authorized by the bill was considered to be a foreign policy matter; and the

proposed program should be linked with the Department of State and the International Cooperation Administration and executed under the

immediate supervision of the President.

The House version passed and became the International Health Research Act of 1960. An important distinction was made in the revision to

separate international health activities that benefit the United States from those designed to advance the status of international health as a whole.

Authority for activities benefiting the United

States was delegated to the Secretary of Health,

Education, and Welfare (now Health and Human

Services). Authority to engage in activities for the

purpose of advancing international health rests

with the President and extends only to research

and training for research, not operational health

programs.

Some research conducted by NIH under the

broader authority of the Public Health Service Act

(the legislation that provides overall direction to

Public Health Service activities) is of great benefit to other countries. Current work in the broad

field of immunology is a case in point. The International Health Research Act, however, still provides the basis for most U.S. Department of

Health and Human Services activities in international health and places limits on the initiatives

that may be taken by NIH and the Centers for

Disease Control (CDC).

The most recent forceful assertion of the importance of international health came under President Carter. In 1978, the White House Office of

Science and Technology Policy produced the report New Directions in International Health Cooperation (27). Federal agencies responded to that

report by proposing several initiatives for domes-
Ch. 2—The United States in International Health . 31

tic and multilateral programs in international

health. The proposals included research in tropical

medicine, establishing a global epidemic intelligence service, supporting the U.N. Development

Program/World Bank/WHO Special Program for

Research and Training in Tropical Diseases

(TDR), developing a worldwide immunization

program, establishing a health unit to respond to

disasters around the world, encouraging the pharmaceutical industry to be more responsive to the

needs of the developing world, and establishing

nutrition surveillance and research programs. In

spite of the enthusiasm, no new funds were allocated for these purposes, and progress was made

only to the extent that existing budgets could support it, which for the most part was relatively

little.

According to one observer, “the United States

has been sensitive to international health needs,

has had continuing concern, but has not always

addressed the issue of availability of resources and

responded with active programs” (75). That observation continues to be accurate (see ch. 3).

Support of Multilateral Efforts

The United States has an admirable record in

supporting multilateral international health efforts. Since the Pan American Sanitary Bureau

was established in 1902, the United States has provided major financial support (about 65 percent

of the regular annual budget) and technical assistance to it and to the later Pan American Health

Organization. The United States energetically

promoted and participated in the formation and

funding of WHO beginning in the mid-1940s, The

United States is currently a major supporter of

TDR (see ch. 3).

U.S. REPORTS ON TROPICAL DISEASE RESEARCH AND TRAINING

The most comprehensive look at tropical disease research was a 1962 report by the National

Academy of Sciences/National Research Council (NAS/NRC). A current NAS study is addressing the U.S. capacity to address tropical disease

problems. These reports are discussed further

below.

1962 NAS/NRC Report on

Tropical Health

The most comprehensive look at tropical disease research, training, and practice, and the resources put toward those ends was taken more

than 20 years ago in an NAS/NRC report entitled

Tropical Health: A Report on a Study of Needs

and Resources (252).

In the mid-195&, the American Society for

Tropical Medicine and Hygiene (ASTMH) recognized that interest in tropical medicine had begun

to decline. The boost given by World War II, interest generated from practical necessity, had

ceased to have effect. Money for research was drying up, and medical schools were cutting back the

teaching of tropical medicine. At the same time,

the United States was developing its foreign aid

programs to include health. Without continuing

support for research and training for the health

problems of recipient countries, mainly those in

which tropical diseases are endemic, the professionals necessary to implement programs would

not be available.

Dr. Albert Sabin, who chaired an ASTMH

committee to consider these problems, was largely

responsible for enlisting NRC to carry out its

study. Funding was provided by NIH, the U.S.

Army Research and Development Command, and

the Rockefeller Foundation.

The 1962 NRC report is encyclopedic in scope.

Geographically, the five major regions covered

are: 1) the Caribbean and Central and South

America, 2) Africa, 3) Southwest Asia, 4) South

Central and Southeast Asia, and 5) Oceania. In

20 chapters, the report discusses the major human

and animal diseases, including what is known of

their incidence, prevalence, and mortality rates

by region, and the status of prevention and treatment; resources for health and medical care; impacts of tropical health and disease on the United

States; U.S.-based and foreign research programs
32 . Status of Bjomedical Research and Related Technology for Tropical Diseases

on tropical diseases; research grant and fellowship programs in tropical health; the teaching of

tropical medicine and hygiene and facilities for

training; and career opportunities and future manpower needs.

Research recommendations for each disease are

presented as an “assembly of informed opinion, ”

gathered by polling experts worldwide. Very specific suggestions are included, covering all aspects

of laboratory, field, and clinical research.

The committee responsible for the report also

drew up several broad resolutions addressed to

various public and private entities in the United

States, specifically (252):

1.

2.

3.

4.

5.

6.

7.

Pave the way to increased U.S. participation

in international health activities, if necessary

through legislation. Solutions to disease

problems of the tropics require systematic

research and development programs. The

United States, by virtue of its position of

world leadership has an obligation to contribute to the fullest extent to research and

training activities. Participation should not

be tied to direct benefits to the United States.

Explore creation of an advisory group within

NAS/NRC to organize support for a “National Program for Research in Tropical

Health, ” with involvement of the relevant

government agencies and private organizations.

Seek authority to make direct grants to foreign institutions for support of research, development, or training in tropical health.

Strengthen undergraduate and postgraduate

medical training in tropical diseases. This

could include opportunities for training in

the tropics.

Seek to improve statistics relating to tropical diseases, by working through WHO and

by offering WHO increased U.S. support to

accomplish this aim.

Encourage studies to document economic

loss due to disease and gains accruing from

disease control.

Encourage and provide additional support

for international veterinary health organizations.

The 1962 NAS/NRC study was, and still is, of

inestimable value to people in the field, giving a

clear picture of the state of knowledge at the time.

That study is now the benchmark against which

progress can be measured. But it does not appear

to have had a significant impact on U.S. Government policies or practices.

Legislation to foster U.S. involvement in tropical health has not been passed. The advisory

group suggested in resolution 2 was not formed.

Funding for research, and particularly for developing country institutions, is still passively allowed in most cases, rather than actively encouraged. There is widespread agreement that the U.S.

capacity to train tropical health professionals has

eroded, if anything, in recent years.

On the positive side, statistics regarding health

conditions in the tropics have gradually become

more reliable. The major improvements have been

in birth and death reporting, and not so much in

the inherently more difficult task of recording

nonfatal cases of disease. The number of studies

of the type recommended in resolution 6 that have

been carried out could probably be counted on

one hand.

Current NAS Study of U.S. Capacity To

Address Tropical Disease Problems

Leaders in tropical medicine and tropical public health have expressed alarm at the current erosion of U.S. support for training in those fields,

The capacity of the United States to train people

in parasitology, vector control, tropical medicine,

and related fields in tropical public health is reaching a low ebb. This crisis is the subject of a current study by NAS.

The NAS study, U.S. Capacity TO Address

Tropical Disease Problems, is being carried out

by NRC’s Board on Science and Technology for

International Development (BOSTID), in cooperation with the Institute of Medicine (IOM). IOM

was approached about the need for this study by

an ad hoc committee of ASTMH.

The NAS study has several objectives:

●

●

to examine current capabilities of U.S. centers

to conduct research and postgraduate training in tropical medicine and tropical public

health;

to estimate future manpower needs in applied

research and training; and
Ch. 2—The United States in International Health ● 3 3

● to assess the adequacy of mechanisms for

providing research and training support in

the field.

Funding for the study is being provided by the

U.S. Army Research and Development Command,

the National Institute for Allergy and Infectious

Diseases, CDC, and AID. The Rockefeller Foundation will cosponsor and support an international workshop in connection with the study. A

report from the study is expected in late 1985. This

report should provide information on which to

make decisions about the types of funding mechanisms and programs that could be emphasized

or mounted in future funding decisions. There will

still be a need to examine the opportunities for

application of available technologies and the constraints to their application.

TROPICAL DISEASE RESEARCH IN THE CONTEXT OF

U.S. INTERNATIONAL HEALTH ACTIVITIES

Efforts to improve international health consist

of a great deal more than research, and in fact,

research is a small component in terms of money

spent and personnel deployed. Most U.S. international health assistance is directed toward operational activities—particularly in providing primary health care, environmental services, and

nutrition and population programs. These activities are closely tied to U.S. foreign policy. Guidelines for operational activities are established by

the Department of State, and the activities themselves are largely carried out by AID. Programs

and resources are provided not solely on the basis of need, but in accordance with U.S. foreign

policy goals.

Much of the published analysis of the U.S. international health effort relates more closely to

operational activities than to research. Tropical

disease research shares some but not all of the

problems of the larger field of international health

and is also subject to other influences.

One concern of many observers of international

health is the lack of a central Federal policy stating U.S. goals for international health activities.

There is a statement of policy for research, which

is stated in the International Health Act of 1960—

but that policy is fairly weak (see discussion

above). The lack of an overall U.S. policy subordinates international health aims to the general

mandates and strictures of each agency as it carries out its activities.

Furthermore, lack of a clear-cut international

health policy results in tropical disease research

remaining low in status in relation to domestic

biomedical research. Operational programs can

use the leverage of foreign policy benefit to support their activities, but research programs, which

may have less direct benefits, may be more difficult to justify.

A strong central policy supporting an aggressive U.S. role in international health could have

a salutary effect on all aspects of the field, including tropical disease research. However, the effect

of such a policy would probably be greater in expanding operational programs than in expanding

research. The reason is that operational programs

have a more immediate, tangible, foreign policy

impact than do research programs. A program in

a particular country in need of health assistance

is a more dramatic display of U.S. interest than

is a researcher at NIH searching for long-term solutions to disease problems. For this reason, if a

policy statement is considered by the U.S. Government, the intent of the policy concerning research, as distinct from operational aims, should

probably be spelled out.
3 .

Funding of

Tropical Disease Research
Page

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . 37

Funding Sources . . . . . . . . . . . . . . . . . . . . . . . . 37

Special Program for Research and

Training in Tropical Diseases . . . . . . . . 37

World Health Organization . . . . . . . . . . . . 38

U.S. Department of Health and Human

Services . . . . . . . . . . . . . . . . . . . . . . . . . . . 41

U.S. Agency for International

Development . . . . . . . . . . . . . . . . . . . . . . 44

U.S. Department of Defense . . . . . . . . . . . 45

Private U.S. Foundations. . . . .

Pharmaceutical Companies . . .

Types of Research Funded . . . . . .

Types of Tropical Diseases . . .

Types of Research Objectives .

Recipients of Research Funding. .

Summary . . . . . . . . . . . . . . . . . . . . .

Table

3-1.

3-2.

3-3.

3-4.

3-5.

3-6.

3-7.

LIST OF TABLES

No.

. . . . . . . . . 46

. . . . . . . . . 47

. . . . . . . . . 49

. . . . . . . . . 49

. . . . . . . . . 50

. . . . . ... 52

. . . . . . . . . 55

Page

TDR’s Research and Development “

Program Area: Distribution of Projects

and Budget Amounts for Specific

Tropical Diseases, 1977-81. . . . . . . . . . 39

WHO Funding for Selected Biomedical

Research Pertaining to Tropical

Diseases, 1980-81 Biennium . . . . . . . . . 40

WHO Program for the Control of

Diarrheal Diseases: Funding for

Biomedical Research Projects,

1980-83 . . . . . . . . . . . . . . . . . . . . . . . . . . 40

NIAID Funding for Tropical

Disease Research, Fiscal Years

1981-83 . . . . . . . . . . . . . . . . . . . . . . . . . . 41

NIAID Funding Specifically for

Research on the Six TDR Diseases,

Fiscal Years 1981-83 . . . . . . . . . . . . . . . 42

NIAID Funding for Research on

Diarrheal and Enteric Infections

and Acute Respiratory Infections,

Fiscal Years 1981-83 . . . . . . . . . . . . . . . 43

CDC Funding for Tropical Disease

Research, Fiscal Years 1981-83 . . . . . . 43

Table No. Page

3-8.

3-9.

3-10<

3-11.

3-12.

3-13.

3-14.

3-15.

3-16.

3-17.

3-18.

CDC Funding for Research on the

Six TDR Diseases, Fiscal Years

1981-83 . . . . . . . . . . . . . . . . . . . . . . . . . .

AID’s Bureau for Science and

Technology: Funding for Disease

Control Research Projects,

Fiscal Years 1979-84 . . . . . . . . . . . . . . .

Total AID Funds Committed Since

1981 for Research on Selected

Diseases by the Programin Science

and Technology Cooperation and

the Board on Science and Technology

for International Development . . . . . .

DOD Funding for Tropical Disease

Research, Fiscal Years 1981-83 . . . . . .

DOD Funding for Research on the

Six TDR Diseases, Fiscal Years

1981-83 . . . . . . . . . . . . . . . . . . . . . . . . . .

Summary of Annual Funding by

U.S. Sources and WHO/TDR Sources

for Biomedical Research unselected

Tropical Diseases, Various Years . . . .

Distribution offending for

Biomedical Research on the Six

TDR Diseases by Research

Objective, 1981 . . . . . . . . . . . . . . . . . . .

NIAID Funding for Biomedical

Research on Diarrheal and Enteric

Infections and ARIs by Research

Objective, Fiscal Year 1981.. . . . . . . .

Distribution of Extramural

Biomedical Research Funds by

Various Funding Sources to Recipient

Organizations in the United States,

Other Industrialized Countries, and

Less Developed Countries,

Various Years.. . . . . . . . . . . . . . . . . . . .

NIAID Funding for Tropical

Medicine and International Health

by Type of Recipient, Fiscal Years

1981-83 . . . . . . . . . . . . . . . . . . . . . . . . . .

DOD Funding for Biomedical

Research on Tropical Diseases by

Type of Recipient, Fiscal Years

1981-83 . . . . . . . . . . . . . . . . . . . . . . . . . .

44

44

45

45

46

50

51

53

54

54

55
3.

Funding of Tropical Disease Research

INTRODUCTION

This chapter presents an analysis of funding of

biomedical research in the tropical diseases of

interest in this assessment-namely, malaria,

schistosomiasis, trypanosomiasis, leishmaniasis,

filariasis, leprosy, tuberculosis, diarrheal diseases,

acute respiratory infections (ARIs), and diseases

caused by arboviruses.

The chapter describes the major funding sources

and levels of funding for tropical disease research.

It also discusses the types of research (by disease

and by research goals) that the funding sources

support. The concluding section of the chapter

presents information about the types of organizations that actually conduct the research.

FUNDING SOURCES

The sources of tropical disease research funding considered in this analysis are as follows:

●

●

●

the Special Program for Research and Training in Tropical Diseases (TDR) and the

World Health Organization (WHO);

U.S. Government agencies, specifically, the

Department of Health and Human Services

(DHHS), the Agency for International Development (AID), and the Department of Defense (DOD); and

U.S. private foundations.

Private pharmaceutical companies are discussed

separately.

Each source of tropical disease research funding considered in this analysis is profiled below.

Data made available by the funding sources themselves are analyzed to show the level of funding,

historical trends, and the allocation of funding

across the tropical diseases of interest.

The analysis in this chapter represents the first

attempt in over 20 years to review all major funding sources for tropical diseases. The information

in this analysis was obtained from various sources

that do not report according to a common set of

definitions or in a common format. Every attempt

has been made to take account of variations and

to identify assumptions and accommodations that

have been made. Even so, the data presented in

this chapter should be regarded as indicative

rather than definitive. Additional information

about the limitations of the data is presented in

appendix B.

Special Program for Research and

Training in Tropical Diseases (TDR)

TDR was planned and initiated by WHO, with

the assistance and cosponsorship of the U.N.

Development Program (UNDP) and the World

Bank. The program was initiated in 1974, and between that date and the end of 1982, nearly $120

million had been contributed to the program by

25 governments, other organizations, the World

Bank, UNDP, and WHO. The U.S. contribution

alone was about $15 million, and by fiscal year

1983, the United States had contributed a total

of about $20 million over a 7-year period

(351,353).

TDR has two interdependent objectives (353):

● to develop new and improved tools to control six tropical diseases: malaria, schistosomiasis, trypanosomiasis, leishmaniasis, filariasis, and leprosy; and

37
38 . Status of Biomedical Research and Related Technology for Tropical Diseases

● to strengthen the biomedical research capabilities of developing countries.

TDR is organized into four program areas: 1)

Administrative and Technical Bodies; 2) Research

and Development; 3) Research Capability Strengthening; and 4) Program Management.

TDR’s Research and Development program

area has 10 components: one component for each

of the six specific tropical diseases and four additional components that cut across disease lines.

The four transdisease components of TDR’s Research and Development program area are biomedical sciences, vector control and biology, epidemiology, and social and economic research.

Scientific working groups make recommendations

about program direction and the allocation of research funding.

Table 3-1 presents information about the overall growth of the disease-specific components of

TDR’s Research and Development program area

from 1977 through 1981. (These components of

the program area comprised about 85 percent of

the program area’s total expenditures from 1977

to 1981. ) The data in table 3-I show that the

disease-specific components of TDR’s Research

and Development program area grew from 128

projects amounting to $2,791,000 in 1977 to 374

projects amounting to $10,377,000 during 1981.

From 1977 through 1981, TDR allocated a total of some $43,767,000 to investigations into the

six diseases (351). The percentage of this research

funding allocated to specific diseases from 1977

to 1981 was as follows: malaria, 30 percent; schistosomiasis, 14 percent; trypanosomiasis, 22 percent; leishmaniasis, 6 percent; filariasis, 16 percent; and leprosy, 13 percent. The percentage of

funding for research on specific diseases remained

fairly constant each year, with the exceptions of

a decline for schistosomiasis from 16 percent in

1979 to 10 percent in 1981 and an increase for

leprosy from 11 percent in 1979 to 16 percent in

1981.

World Health Organization (WHO)

Research on two groups of diseases discussed

in this report —ARIs and diarrheal diseases-is

funded through separate WHO programs.

In the biennium 1980-81, WHO allocated a total of $2,389,000 for biomedical reseach pertaining to tropical diseases of interest in this assessment (see table 3-2). This research falls in the

broad category of communicable disease prevention

and control. The data in table 3-2 were collected

by a combination of budget review and personal

interviews and should be regarded as approximations (108). Sixty-six percent of the funding

for such research came from regular budgetary

sources.

The United States contributes 25 percent of the

regular WHO budget, just under $400,000 for the

1980-81 period. The United States (as do other

countries and organizations) makes additional allocations on a program-by-program basis (extrabudgetary contributions), bringing the total annual contribution of the United States to between

$250,000 and $500,000 for 1980-81. An example

of other extrabudgetary WHO funds in 1980-81

was $600,000 for research into leprosy, much of

which was a gift from the Japan Shipbuilders

Association.

WHO Program in Acute Respiratory

Infections

In 1980-81, WHO spent a total of $381,000 for

research in ARIs and tuberculosis, 45 percent of

which was from regular budgetary sources (see

table 3-2). In 1983, ARI was identified as a distinct WHO program area. Since then, while ARIs

have been receiving greater attention within the

WHO regular budget, support from extrabudgetary sources has been relatively modest. The Pan

American Health Organization (PAHO) reports

that in 1983 its program for ARIs received only

$31,000 in direct support funds from WHO, and

a further $45,000 for tuberculosis (207).

WHO Program in Diarrheal Diseases

WHO’s Program for Control of Diarrheal Diseases (CDD) was initiated in 1978. Its objectives

are to reduce the mortality and morbidity caused

by acute diarrheal diseases and to promote the

self-reliance of individual countries in the delivery of health and social services for the control

of diarrheal diseases (427).
Table 3.1.—TDR’s Research and Development Program Area: Distribution of Projectsa b and

Budget Amounts for Specific Tropical Diseases, 1977-81
40 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

Table 3-2.—WHO Funding for Selected Biomedical Research

Pertaining to Tropical Diseases, 1980=81 Biennium

Amount of funding (000s)

Regular Extra- Total

Type of research budget budgetary budget

Malaria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ... ... ..$ 222 $ 11 $ 233

Other parasitic diseases . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 335 0 335

Bacterial diseases, plague, meningitis and mycotic diseases. . . . . 105 0 105

Leprosy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66 600 666

Tuberculosis and acute respiratory infections (ARIs) . . . . . . . . . 170 211 381

Viral diseases . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 250 0 250

Vector biology and control . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 419 0 419

Total . . . . . . . . .......:. . . . . . . . . . . . . . . . . .. .. .. .. ... ... .$1,567 $822 $2,389

SOURCE: Data presented in V. Elliott and P. Contacts, ’’A Profile of Selected Biomedical Research Efforts Into Diseases of

Major Public Health importance to People of Developing Countries;’ typescript, prepared for the US. Agency for

International Development Washington, DC, November 1982.

Support for CDD is predominantly from extrabudgetary WHO sources. In the biennium 1980-

81, CDD resources totaled $6,879,018, of which

$5,004,840(73percent) came from extrabudgetary

sources. In 1982, total resources were $7,857,429

of which $6,932,352 ($8 percent) were extrabudgetary. The contribution of the United States was

$157,400 in 1980-81 and $78,800 in 1982.

CD Dobligateda total of $5,654,423 during l980-

81 and unestimated $14,373,600 during 1982-83.

Of these obligations, 33 percent in 1980-81 and 39

percent in 1982-83 were set aside for research. The

research component of CDD includes both biomedical and operational activities. In 1980-81,$1,060,095

(56percent) ofCDD’s resarch budget was allocated

to biomedical research; estimates for 1982-83 are

$3,814,700 (67percent) (427). These figures include

funds for planning, coordination, and support of

collaborating centers as well as direct support of research projects.

Table 3-3 shows that between 1980 and 1983,

CDD provided $3,035,000 in funds for a total of

149 biomedical research projects. Bacterial enteric

infections were the focus of 63 of these research

projects and accounted for $l,150,000 (38percent)

of research project funds over the period. Drug

development and the management of acute diarrheas accounted for 25 percent of these funds

and viral diarrheas for somewhat less (23 percent)

(425).

Table 3-3.—WHO Program for the Control of Diarrheal Diseases (CDD) Funding for

Biomedical Research Projects, 1980-83

Total Percent

1980-83 of total

Number of

Budget amount (OOOs)

budget 1980-83

projects amount budget

Type of research 1980-83 1980-81 1982 1983 (000s) amount

Bacterial enteric

infections. . . . . . . . . . . 63 $222 $312 $ 616 $1,150 38%

Parasitic diarrheas . . . . . 8 0 10 111 121 4

Viral diarrheas . . . . . . . . 36 61 256 390 707 23

Drug development and

acute diarrhea

management . . . . . . . . 31 109 159 497 765 25

Other . . . . . . . . . . . . . . . . 11 25 119 148 292 10

Total by year. . . . . . . . 149 $417 $856 $1,762 $3,035 100%

SOURCE: Derived from World Health Organization, Program for Control of Diarreal Diseases, DCC/TAG/84.2A (Geneva: WHO,

February 1984).
Ch. 3—Funding of Tropical Disease Research ● 4 1

U.S. Department of Health and

Human Services (DHHS)

Three organizational units within DHHS fund

research in the tropical diseases of interest in this

assessment:

● the National Institute of Allergy and Infectious Diseases (NIAID) of the National Institutes of Health (NIH);

● the Centers for Disease Control (CDC); and

● the Fogarty International Center for Advanced Study in the Health Sciences of NIH.

The activities of these organizations are discussed below.

National Institute of Allergy and

Infectious Diseases

NIAID has primary responsibility for tropical

medicine research at NIH. In fiscal year 1981, the

total NIAID budget accounted for $232 million

(or 6 percent) of the total NIH obligations of

$3,572 million (383). That year, as shown in table

3-4, NIAID awarded about $26,865,000 in extramural and intramural grants for research activities in tropical diseases, general parasitology, and

general tropical medicine (379). These grants represented about 12 percent of NIAID funds and less

than 1 percent of all NIH funds in fiscal year 1981.

Table 3-4 shows that overall NIAID funding for

tropical disease research increased from a level of

almost $27 million in fiscal year 1981 to a little

more than $33 million in fiscal year 1983 (379,

380,381). This NIAID-funded research can be classified under three general headings: 1) tropical diseases (malaria, schistosomiasis, trypanosomiasis,

leishmaniasis, filariasis, and leprosy); 2) general

parasitology (cestodes, nematodes, protozoa, and

trematodes); and 3) general tropical medicine

(rickettsia, bacteriology, mycology, virology, and

vector pathogens). As shown in table 3-4, the percentage of funds allocated to research under the

Table 3.4.—NIAID Funding for Tropical Disease Research, Fiscal Years 1981-83

Combined extramural and

intramural funding

Fiscal year and Amount of funding (000s ) Amoun t

program are a Extramural Intramural (000s) Percen t

Fiscal year 1981:

Tropical diseases . . . . . . . . . . .

General parasitology . . . . . . . .

General tropical medicine . . . .

Total . . . . . . . . . . . . . . . . . . . .

Percent of combined

funding amount. . . . . . .

Fiscal year 1982:

Tropical diseases . . . . . . . . . . .

General parasitology . . . . . . . .

General tropical medicine. . . .

Total . . . . . . . . . . . . . . . . . . . .

Percent of combined

funding amount. . . . . . .

Fiscal year 1983:

Tropical diseases . . . . . . . . . . .

General parasitology . . . . . . . .

General tropical medicine. . . .

Total . . . . . . . . . . . . . . . . . . . .

Percent of combined

funding amount. . . . . . .

$ 9,101

3,551

7,660

$20,312

$4,187

1,163

1.203

$13,288

4,714

8,863

$26,865 a

490/0

18

33

$6,553 100%0

760/o 240/o 100 ”/0

$10,042

3,502

7,659

$5,627

1,854

1,991

$15,670

5,356

9,650

51 0/0

17

31

$21,203 $9,472 $30,676 100 ”/0 b

690/o 31 % 100 ”/0

$16,211

2,908

6,182

$5,182

1,245

1,430

$21,393

4,153

7,612

650/o

13

23

$25,301 $7,857 $33,158C 100 ”/0

760/o 240/o 100%
42 ● Status of Biomedical Research and Related Technology for Tropical Diseases

tropical diseases heading rose from 49 percent in

fiscal year 1981 to 65 percent in fiscal year 1983,

while the percentage allocated to general tropical medicine declined from 33 percent in fiscal year

1983 to 23 percent in fiscal year 1983. About

three-quarters of NIAID funding for tropical disease research was awarded through extramural

grants and contracts in fiscal years 1981 and 1983

and a slightly smaller proportion in fiscal year

1982. There were no major differences in the proportion of extramural awards between the three

categories of research.

Table 3-s shows the allocation of NIAID funds

for research under the heading of tropical

diseases—i.e., on the six TDR diseases—for fiscal years 1981, 1982, and 1983. Funds for research

into each of these diseases increased over the 3-

year period, with the exception of funds for leprosy research, which were substantially reduced

in fiscal year 1982, but increased again in fiscal

year 1983. Over the 3-year period, trypanosomiasis research accounted for 26 percent of NIAID

funding for research on the six TDR diseases;

schistosomiasis research, 22 percent; malaria research, 20 percent; leishmaniasis research, 18 percent; filariasis research, 9 percent; and leprosy research, 5 percent. The same relative level of

NIAID research funding among the six diseases

is seen for each year for which data are presented,

with the exception of fiscal year 1983, when funding for leishmaniasis research slightly exceeded

that for research in malaria.

Table 3-6 shows NIAID funding for research

on diarrheal and enteric diseases and ARIs, through

grants awarded under the headings of general

parasitology and general tropical medicine, for

fiscal years 1981, 1982, and 1983. Most NIAID

funding for research in diarrheal and enteric diseases and ARIs is considered domestic research

for reporting purposes, but the share allocated to

tropical medicine has increased. In fiscal year

1981, only 1 percent of NIAID funds for research

in diarrhea] and enteric diseases and ARIs came

under the heading of tropical medicine; in 1982,

16 percent; and in 1983, 15 percent (263).

The data presented in table 3-6 show that NIAID

funded research on ARIs at more than twice the

funding level for research on diarrhea] and enteric

infections during the 3-year period 1981-83. The

distribution of funding between extramural and

intramural research for these two disease classes

varied during the 3-year period, ending with a

slight shift toward extramural research in fiscal

year 1983.

Centers for Disease Control

CDC conducts research in all of the categories

of tropical diseases of interest in this assessment.

Table 3-7 shows the levels of CDC funding of such

research during fiscal years 1981, 1982, and 1983.

Overall, CDC’S tropical disease research funding

grew from $3,877,000 in fiscal year 1981 to

$4,929,000 in fiscal year 1983 (288). This growth

is reflected in each of the categories of disease

shown in the table. Overall, about 44 percent of

CDC’S tropical disease research funding from 1981

to 1983 was allocated to research on the six TDR

diseases; 22 percent was allocated to research on

diarrheal diseases; 29 percent to research on ARIs;

and 4 percent to research on arboviral infections.

Table 3-5.—NIAID Funding Specifically for Research on the Six TDR Diseases, Fiscal Years 1981.83

Fiscal year 1981 Fiscal year 1982 Fiscal year 1983 Fiscal years 1981-83

Amount a

Amount Amount Amount

Disease (000s) Percent (000s) Percent (000s) Percent (000s) Percent

Malaria. . . . . . . . . . . . . . . $ 2,426 180/0 $3,567 230/o $3,950 180/0 $9,943

Schistosomiasis. . . . . . . 2,984 22 3,550 23 4,598 21 11,132

Trypanosomiasis . . . . . . 3,783 28 3,889 25 5,308 25 12,980

Leishmaniasis. . . . . . . . . 1,927 15 2,883 18 4,107 19 8,917

Filariasis . . . . . . . . . . . . . 1,189 9 1,452 9 2,051 10 4,692

Leprosy . . . . . . . . . . . . . . 979 7 330 2 1,379 6 2,688

Total by year. . . . . . . . $13,288 100% $15,670 1000/0 $21,393 100% $50,351

20%

22

26

18

9

5

100%

SOURCE U S Department of Health and Human Services, National Institutes of Health, “lnternational Cooperation by the National Institute of Allergy and Infectious

Diseases, FY 1981, FY 1982, FY 1983, ” mimeo, Bethesda, MD, no date
Ch. 3—Funding of Tropical Disease Research ● 4 3

Table 3-6.—NIAID Funding for Research on Diarrheal and Enteric Infections and

Acute Respiratory Infections (ARIs), Fiscal Years 1981-83

Extramural Intramural Combined

funding funding funding

amount amount amount

(000s) (000s) (000s)

Fiscal year 1981:

Diarrheal and enteric infections . . . . . . . . . . . . . . . . . . . . . . . $4,459 $2,497 $6,956

ARIs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11,819 4,442 16,261

Total . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . $16,278 $6,939 $23,217

Percent of combined funding amount . . . . . . . . . . . . . . 70 ”/0 30 ”/0 100 ”/0

Fiscal year 1982:

Diarrheal and enteric infections . . . . . . . . . . . . . . . . . . . . . . . $4,195 $2,984 $7,179

ARIs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10,194 4,293 14,487

Total . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . $14,389 $7,277 $21,666

Percent of combined funding amount . . . . . . . . . . . . . . 66% 34% 100%

Fiscal year 1983:

Diarrheal and enteric infections . . . . . . . . . . . . . . . . . . . . . . . $5,452 $2,731 $8,183

ARCS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14,689 3,944 18,633

Total . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . $20,141 $6,675 $26,816

Percent of combined funding amount . . . . . . . . . . . . . . 75% 25% 100%

SOURCE J E Nutter, Chief, Office of Program Planning and Evaluation, National lnstitute of Allergy and infectious Diseases, National Institutes of Health, Bethesda,

MD, personal communication, April 1984.

Table 3-7.–CDC Funding for Tropical Disease Research, Fiscal Years 1981-83

Fiscal year 1981 Fiscal year 1982 Fiscal year 1983 Fiscal years 1981-83

Amount Amount Amount Amount

(000s) Percent (000s) Percent (000s) Percent (000s) Percent

TDR diseasesa 

. . . . . . . . $1,722 44% $1,927 45% $2,160 44% $ 5,809 44%

Diarrheal diseases . . . . . 859 22 976 23 1,113 23 2,948 22

ARIs . . . . . . . . . . . . . . . . . 1,130 29 1,261 29 1,397 28 3,788 29

Arboviral infections . 166 4 166 4 259 5 591 4

Total by year. . . . . . . . $3,877 100% $4,330 100% $4,929 100% $13,136 100%

aMalaria, scillstosomiasis, trypanosomiasis, filariasis, Ieishmaniasis, and leprosy

SOURCE J G Randolf, Budget Analyst Financial Management Office, Centers for Disease Control, Atlanta, GA, personal communication, April 1984

Table 3-8 shows the amount and distribution

of CDC funds among the six TDR diseases for fiscal years 1981-83. Over that 3-year period, research in malaria received more than 60 percent

of this funding. Schistosomiasis research funding

increased from 15 percent of the funds in 1981 to

21 percent in 1983, while filariasis research funding declined from 6 percent of the funds in 1981

to 4 percent in 1983.

Fogarty International Center for Advanced

Study in the Health Sciences

FIC is the focal point within NIH for scientific

exchange and collaboration at the international

level. The FIC budget was $5.1 million in fiscal

year 1979, $4.5 million in fiscal year 1980, and

$4.3 million in fiscal year 1981 (382). Most of the

activities of FIC fall outside the biomedical research concerns that are the subject of this

analysis.

A major exception, however, is FIC’S responsibility for transmitting the U.S. Government’s

care support to the Gorgas Memorial Institute of

Tropical and Preventive Medicine. The operating unit of the institute is the Gorgas Memorial

Laboratory, located in Panama. Neither FIC nor

any other U.S. agency has programmatic control

over the activities at the laboratory. Gorgas received $1.7 million in core support through FIC

in fiscal year 1980, $1.8 million in fiscal year 1981,

$1.692 million in fiscal year 1982, and $1.8 million in fiscal year 1983 (360). These amounts con-
44 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Table 3-8.—CDC Funding for Research on the Six TDR Diseases, Fiscal Years 1981-83

Fiscal year 1981 Fiscal year 1982 Fiscal year 1983 Fiscal years 1981-83

Amount Amount Amount Amount

Disease (000s) Percent (000s) Percent (000s) Percent (000s) Percent

Malaria . . . . . . . . . . . . . . . $1,127 650/o $1,208 630/o $1,290 600/0 $3,625 620/o

Schistosomiasis. . . . . . . 250 15 352 18 461 21 1,063 18

Trypanosomiasis . . . . . . 40 2 48 2 58 3 146 3

Leishmaniasis. . . . . . . . . 66 4 66 3 98 5 230 4

Filariasis . . . . . . . . . . . . . 101 6 101 5 86 4 288 5

Leprosy . . . . . . . . . . . . . . 138 8 152 8 167 8 457 8

Total by year. . . . . . . . $1,722 100 ”/0 $1,927 100 ”/0 $2,160 100 ”/0 $5,809 100 ”/0

SOURCE J G Randolf, Budget Analyst, Financial Management Office, Centers for Disease Control, Atlanta, GA, personal communication, April 1984

stitute between 64 and 74 percent of all support

received by the institute. Other agencies of the

U.S. Government also provide funds to Gorgas

Memorial Institute through grants and contracts.

Further information about Gorgas is presented in

OTA’S technical memorandum entitled Quality

and Relevance of Research and Related Activities

at the Gorgas Memorial Laboratory (360), which

was published in August 1983 as part of this

assessment.

U.S. Agency for International

Development (AID)

AID’s Bureau for Science and Technology supports a variety of biomedical research activities

pertaining to tropical diseases. Table 3-9 presents

information for fiscal years 1979 through 1984

about projects that are classified by AID as “Disease Control—Research” (152). This category includes AID’s support for TDR and for the International Center for Diarrheal Disease Research,

Bangladesh. In addition to providing this institutional support, AID funds research through grants

and contracts. The research activities funded by

AID in the past have been primarily concerned

with malaria immunology and vaccine development, but AID is planning to expand its tropical

disease research agenda (110).

Since 1981, the Office of the Science Advisor

of AID has also been supporting biomedical research through AID’s Program in Science and

Technology Cooperation (PSTC) and through the

complementary Research Grants Program of the

National Academy of Sciences’ Board on Science

and Technology for International Development

(BOSTID). These programs administer grant

funds which are intended to stimulate investiga-
Ch. 3—Funding of Tropical Disease Research ● 4 5

tion of problems that confront developing countries. The Research Grants Program administered

by BOSTID provides funds only to developing

country institutions. PSTC gives priority to proposals received from developing countries, but

also funds activities in the United States.

PSTC has concentrated on five areas of research. One of these relates to tropical diseases—

namely, biotechnology/immunology and biological control of human schistosomes and associated

snail vectors (73).

The BOSTID program concentrates on six areas

of research. Three of these areas-field studies of

the mosquito vector, rapid epidemiologic assessment, and the diagnosis and epidemiology of ARIs

in children—fall within the broad rubric of biomedical research in tropical diseases.

Table 3-10 shows AID funds committed to

PSTC and the BOSTID program for research relevant to this analysis. The data show that AID has

committed about $8 million of research funding

for investigations into these areas of interest since

the AID-funded programs began in 1981. The duration of any single project may be from 1 to 4

years (73,255).

U.S. Department of Defense (DOD)

In fiscal year 1982, DOD obligated $233 million for biomedical research and development

(383). Of this amount, $21 million (9 percent) was

for research on about 30 infectious diseases.

Almost $13.5 million of the latter amount was for

research pertaining to the tropical diseases of interest in this analysis (140). That $13.5 million represents 64 percent of DOD’s infectious disease

research funding and 6 percent of DOD’s total biomedical research funding.

Table 3-11 shows the levels of DOD funding

for research pertaining to the six TDR diseases,

diarrheal diseases, and arboviral diseases for fiscal years 1981, 1982, and 1983. DOD does only

a small amount of research in ARIs. The figures

in table 3-11 are for activities of the Army Medical Research and Development Command, which

Table 3.1 l.— DODa Funding for Tropical Disease Research, Fiscal Years 1981-83

Fiscal year 1981 Fiscal year 1982 Fiscal year 1983 Fiscal years 1981-83

Amount Amount Amount Amount

Disease (000s) Percent (000s) Percent (000s) Percent (000s) Percent

TDR diseasesb . . . . . . . . $ 7,115 57 ”/0 $ 7,797 580/o $ 8,19 0 580/o $23,10 2 580/o

Diarrheal diseases . . . . . 1,55 8 1 2 1,63 5 12 1,71 7 1 2 4,91 0 1 2

Arboviral infections . . . . 3,81 5 31 4,00 4 3 0 4,20 4 3 0 12,02 3 3 0

Total bv year. . . . . . . . $12,488 1000/0 $13,43 6 100 % $1411 1 100% $40,03 5 100 %
46 ● Status of Biomedial Research and Related Technology for Tropical Diseases

manages almost all DOD tropical disease research

efforts. The percentage of DOD funds for research

in the specific categories of diseases remained

almost constant from fiscal year 1981 to fiscal year

1983, with 58 percent of funds allocated to the

six TDR diseases, 12 percent allocated to diarrheal

diseases, and 30 percent allocated to arboviral diseases. The actual amounts available rose by approximately $1 million each year.

Table 3-12 shows the level and distribution of

DOD funding for investigations in the six TDR

diseases for fiscal years 1981 through 1983. As

shown in that table, DOD conducts no research

in filariasis or leprosy. About 58 percent of the

DOD funding shown in table 3-12 is concerned

with malaria; 19 percent with leishmaniasis; 17

percent with trypanosomiasis; and 7 percent with

schistosomiasis.

Private U.S. Foundations

A number of private foundations in the United

States fund activities in health, but do not report

supporting biomedical research in the tropical diseases of interest in this analysis. These include the

National Science Foundation, the Lasker Foundation, the Kellogg Foundation, the Ford Foundation, and the Carnegie Foundation. The paragraphs below describe the objectives and funding

levels of the three U.S. foundations that do support these activities: the Edna McConnell Clark

Foundation, the Rockefeller Foundation, and the

MacArthur Foundation.

Edna McConnell Clark Foundation

The Edna McConnell Clark Foundation reports

assets of $225 million in 1982 and makes grants

of approximately $14 million annually (105).

Grantmaking procedures suggest that applicants

write a brief letter describing the program, which

is reviewed by the appropriate program officer.

If the request seems to fit the program’s goals,

more information and a proposal is requested. In

reviewing proposals, the Clark Foundation’s

officers look for sound strategy and staff with the

skills to accomplish work central to the aims of

a particular program. Proposals are reviewed by

trustees and a committee of outside experts and

acted upon by the trustees.

The Clark Foundation makes grants in four

program areas: 1) children in foster or institutional

care, 2) jobs for the disadvantaged, 3) improvements in the criminal justice system, and 4) tropical disease research. Recently, the foundation

made a decision to broaden support and include

research on preventable causes of blindness. The

tropical disease research program is currently concerned mainly with schistosomiasis, supporting

research in immunology for vaccine development,

the metabolism and biochemistry of schistosomes,

and studies of the epidemiology of schistosomiasis.

The grants payable by the Clark Foundation’s

tropical disease program were $3,174,651 on September 30, 1980; $2,433,047 on the same date in

1981; and $2,238,369 in 1982 (106), These amounts

Table 3-12.– DODa Funding for Research on the Six TDR Diseases, Fiscal Years 1981.83

Fiscal year 1981 Fiscal year 1982 Fiscal year 1983 Fiscal years 1981-83

Amount Amount Amount Amount

Disease (000s) Percent (000s) Percent (000s) Percent (000s) Percent

Malaria. . . . . . . . . . . . . . . $4,259 600/0 $4,471 57 ”/0 $4,694 57% $13,424 580/o

Schistosomiasisb . . . . . . 270 4 611 8 644 8 1,525 7

Trypanosomiasis . . . . . . 1,224 17 1,285 16 1,350 16 3,859 17

Leishmaniasis. . . . . . . . . 1,362 19 1,430 18 1,502 18 4,294 19

Filariasis . . . . . . . . . . . . . 0 0 0 0 0 0 0 0

Leprosy . . . . . . . . . . . . . . 0 0 0 0 0 0 0 0

Total by year. . . . . . . . $7,115 100 ”/0 $7,797 1000/0 $8,190 100 ”/0 $23,102 100 ”/0
Ch. 3—Funding of Tropical Disease Research ● 4 7

constitute about 30 percent, 25 percent, and 20

percent of total grants awarded by the foundation in each of these years.

Rockefeller Foundation

The Rockefeller Foundation’s Health Sciences

program began in 1977 and has three components:

1) the Great Neglected Diseases of Mankind, 2)

the Health of Populations, and 3) Coping With

Biomedical and Health Information. Since these

components were put in place, the foundation has

allocated about $12 million, $3 million, and

$676,000 to the respective components (293).

The Rockefeller Foundation’s Great Neglected

Diseases program seeks to improve the knowledge, the means of treatment, and the control of

these diseases by attracting outstanding scientists.

The diseases included under the program are those

such as malaria, schistosomiasis, hookworm, and

diarrheal diseases. The objectives of the Rockefeller Foundation’s efforts have been to establish

and support research units of excellence, to establish collaboration in clinical investigation and field

research among research institutions in developed

and developing countries, and to maximize collaboration among these researchers through an

annual meeting.

Funds for the Great Neglected Diseases program

were $1.6 million in 1980, $1.9 million in 1981,

and $1.9 million in 1982. By 1980, there were 14

research units established, 3 of which are located

in the developing world, and collaboration had

been established with 22 developing countries.

Foundation funding for each research unit is

$150,000 at most, and this support is about 30

percent of the support received by the units from

all sources.

In addition to supporting these research units,

the Rockefeller Foundation awards some grants

related to tropical disease research to institutions

not included under its program of institutional

support. In 1981, such grants supported investigations in malaria, schistosomiasis, trypanosomiasis, and leprosy, as well as an investigation

by the Center for Public Resources of ways in

which the pharmaceutical industry might be encouraged to expand its role in improving health

in developing countries.

MacArthur Foundation

In 1983, the board of the MacArthur Foundation decided to devote $20 million over the next

5 years to the support of centers of excellence to

apply the techniques of modern biology to parasitic diseases (282). The MacArthur Foundation

is particularly interested in supporting research

by scientists who have not previously studied

parasitic diseases and in fostering collaboration

among scientific disciplines. Accordingly, it has

invited proposals from a selected group of individuals and institutions and made its first

awards under this program in late 1984. The

MacArthur Foundation has also recently made a

grant of $1 million to TDR.

Pharmaceutical Companies

Detailed information about the money spent by

U.S. pharmaceutical companies to conduct biomedical research in tropical diseases is not available. Neither individual firms nor the industry

publish comprehensive data about research expenditures because such information is considered

proprietary. The discussion below summarizes

relevant facts and conjectures about research on

tropical diseases being undertaken by private

companies.

Overview

The Pharmaceutical Manufacturers Association (PMA) reported in 1979 that its 132 U.S.

members spent $1.3 billion each year on research.

About one-fifth of the companies conduct 80 percent of the research and development in drugs.

A PMA survey published in 1979 found that 21

companies had done, or were doing, research relevant to tropical diseases. PMA estimated that this

research accounted for about 5 percent of the

overall research and development effort of the industry (306).

Another report indicates that the U.S. pharmaceutical industry spends more than 50 percent of

its net income on research and development (165).

The report notes, however, that these activities

are extremely concentrated within the industry.

Only 14 of the 26 companies that spend more than

$1 million a year on research and development
48 ● status of Biomedical Research and Related Technology for Tropical Diseases

actually spend at the industry average rate of 50

percent. The four most research-intensive companies account for 37 percent of the industry’s

research and development, but produce only 21

percent of industry sales. The next four most research-intensive companies account for 23 percent

of the industry’s research and development and

24 percent of its sales. The next 12 companies represent 33 percent of total research and development, but 47 percent of industry sales (165).

A survey of 15 research-oriented European

pharmaceutical companies in 1977 found that 7

of these firms were engaged in tropical disease research. These companies allocated approximately

$40 million to tropical disease research (387).

An analysis of the leading U.S. and European

pharmaceutical companies in the area of research

in tropical diseases suggests that seven companies

spent a total of $22,327,000 on research in parasitology during fiscal year 1979 (73).

The data cited above should be reviewed in relation to the costs of developing new drugs. One

study found that the average cost to a U.S.-owned

pharmaceutical firm for developing a new chemical entity (NCE) to the point of marketing approval in the United States was $54 million in 1976

dollars. The average length of time from initiation of clinical testing to market approval for all

NCES approved in 1976 was more than 6 years.

During the period 1963-76, approximately 900

NCES were studied in humans by U.S.-owned

firms. Of these 900, 20 (2 percent) were candidates primarily for tropical or parasitic diseases.

These NCES came from 11 U.S.-owned pharmaceutical firms (95).

Examples of Pharmaceutical Company

Activities

In the past 25 years, a number of pharmaceutical companies have conducted biomedical research related to malaria, often in collaboration

with the public sector. During the period 1960-

69, for example, Parke-Davis invested about $16

million in antiparasite research, much of which

was concerned with malaria (a disease for which

Parke-Davis developed seven drugs). Furthermore, Parke-Davis reported in 1979 that, with

AID support, it was engaged in studies of an

immunological approach to malaria (20). Research in malaria by Roche led to the development of Fansidar, a prophylactic and chemotherapeutic drug marketed in 1970 (20). Since then,

Roche has been collaborating with TDR and

Walter Reed Army Institute of Research to conduct clinical trials of derivatives of the natural

alkaloid quinine (20). A 1979 PMA survey found

14 U.S. companies and 4 of the 7 European firms

surveyed were conducting research in malaria

(306,387).

The PMA survey reported 11 American companies and 4 of the 7 European firms were engaged

in schistosomiask research (306,387). Pfizer’s major work has been in schistosomiasis. Roche has

been engaged in schistosomiasis research for more

than 20 years and, after screening tests, has focused attention on two compounds, one of which

has now been selected for clinical trials (20). Wellcome has worked on schistosomiasis for 30 years,

but without commercial success (20). Bayer and

E. Merck/Darmstadt began intensive research in

schistosomiasis about 10 years ago (20).

Three of the 7 European firms surveyed by the

PMA work in trypartosomiasis (387). Roche has

given high priority to investigations into Chagas’

disease (American trypanosomiasis), and reports

that once the company succeeds, it will emphasize investigations pertaining to African trypanosomiasis (20). Bayer discovered the first effective

drug against African trypanosomiasis in 1916, and

after more than 30 years of research, Bayer introduced the first drug to treat the acute and chronic

phases of Chagas’ disease in 1972 (20). E. Merck/

Darnstadt also has an active program in trypanosomiasis (20).

Research in leishmaniasis was stopped by many

companies about 10 years ago because the market was small and a drug was available (20). However, TDR has provided some stimulus to new efforts, and six U.S. companies and two European

companies are engaged in leishmaniasis research

(306,387). Among these, Squibb was reported to

be collaborating with TDR in supplying a compound to be tested in Africa (306). BurroughsWellcome is currently testing allopurinol, an existing drug used for gout, which was found to

have antileishmanial activity.
Ch. 3—Funding of Tropical Disease Research ● 4 9

Various companies have spent some $20 million on filariasis research, and there is as yet no

satisfactory drug. Roche, Bayer, and Ciba-Geigy

have been particularly active in this research effort, and Janssen has also been having good results (20). Seven members of the PMA reported

research projects in filariasis, including ParkeDavis which was under contract to WHO for the

synthesis of antifilarial drugs (306). Four of the

seven European firms questioned in the PMA survey reported research in filariasis (387).

Leprosy is under investigation by two of the

European firms and six members of the PMA

(306,387). Ciba-Geigy and Lepetit have drugs on

the market and are doing further research to try

to reduce treatment costs by requiring a less frequent application (20).

Information about research into other diseases

of interest to this analysis is not readily available.

However, Lederle, Sterling, and Merck are reported to be conducting research in tuberculosis

(306). The WHO program concerned with diarrheal diseases reports that during 1981-82, it colTYPES OF RESEARCH FUNDED

Types of Tropical Diseases

Table 3-13 summarizes annual funding from

WHO, TDR, and from U.S. Government and private foundation sources for research pertaining

to the six TDR diseases, diarrhea] diseases, ARIs,

and arboviral diseases, for roughly the period

1979-81. Combined WHO/TDR and U.S. funding for biomedical research on these diseases was

a little over $109 million per year. Less than $100

million came from U.S. sources.

Combined funding for research in the six TDR

diseases—malaria, schistosomiasis, trypanosomiasis, leishmaniasis, filariasis, and leprosy—was

almost $55 million, or about 50 percent of the total $109 million per year. Malaria research alone

was funded at $22.8 million per year (21 percent

of the total $109 million). TDR, AID’s Bureau of

Science and Technology, and DOD allocated a

greater portion of their research funding for malaria research than for research in any of the other

laborated with eight pharmaceutical companies,

none of which were American (427).

The Rockefeller Foundation has sought to expand the role of the pharmaceutical industry in

developing products for tropical diseases by making a series of grants to the Center for Public Resources. The center organized a task force of

leaders from the pharmaceutical industry, the developing countries, academe, and assistance agencies following a 1979 conference at the Institute

of Medicine. This task force designed a number

of projects, but in 1981 and 1982, the pharmaceutical industry determined that it did not wish

to match the Rockefeller Foundation’s commitment to the cooperative effort and the individual

projects, and the project was terminated. The

Rockefeller Foundation points out that the outspoken support of Senators Jacob Javits, Richard

Schweiker, and Edward Kennedy during the 1979

conference was not reflected in the political atmosphere of 1980, and suggests that industry cooperation may have been discouraged as a result

(293).

categories of tropical diseases considered in OTA’S

analysis. As shown in table 3-13, the distribution

of combined annual funding for biomedical research on the other TDR diseases was as foIlows:

schistosomiasis, $9.2 million (8 percent of the total

$109 million); trypanosomiasis, $9 million (8 percent); leishmaniasis, $6.3 million (6 percent); filariasis $3.7 million (3 percent); and leprosy, $3.8

million (4 percent).

Research in diarrheal diseases was funded at an

annual level of $14.5 million (or about 13 percent

of the total $109 million per year); research in

ARIs was funded at $20.7 million (I9 percent of

the total); and research in arboviral diseases was

funded at $7.5 million (7 percent of the total).

Combined annual funding for research that is related to the diseases of interest in this analysis but

for which data were not available in sufficient detail to allocate the funding by disease (the “unspecified category in table 3-13) amounted to

$11.5 million (11 percent of the total).
50 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

Table 3.13.—Summary of Annual Funding by U.S. Sourcesa and WHO/TDR Sourcesb

for Biomedical Research on Selected Tropical Diseases, Various Yearsc

Combined WHO/TDR and

U.S. sources

U.S. sourcesb WHO/TDRc

Amount

Disease (000s) (000s) (000s) Percent

Malaria . . . . . . . . . . . . . . . . . . . . $19,354 .$ 3,435 $22,789 21 “/0

Schistosomiasis . . . . . . . . . . . . 8,136 1,044 9,180 8

Trypanosomiasis . . . . . . . . . . . . 6,716 2,335 9,051 8

Leishmaniasis . . . . . . . . . . . . . . 5,707 614 6,321 6

Filariasis . . . . . . . . . . . . . . . . . . 2,137 1,548 3,685 3

Leprosy. . . . . . . . . . . . . . . . . . . . 1,546 2,299 3,845 4

Subtotal . . . . . . . . . . . . . . . . . $43,596 $11,275 $54,871 500%

Diarrheal diseases . . . . . . . . . . $14,081 $ 417 $ 14,498 130%

Acute respiratory infections , . 20,301 381 20,682 19

Arboviral diseases . . . . . . . . . . 7,482 NA 7,482 7

Subtotal . . . . . . . . . . . . . . . . . $41,864 $ 798 $ 42,662 39 %

Unspecified . . . . . . . . . . . . . . . . $10,359 $ 1,109 $ 11,468 11 %

Total by source . . . . . . . . . $95,819 $13,182 $109,001 100%

Percent of combined

funding amount. . . . . 880% 12 ”/0

and the Edna McConnell Clark Foundation

These data are affected by the fact that not all

agencies define diarrheal diseases and ARIs as

tropical diseases. Research funds for these diseases

from NIAID sources other than those that fall under tropical medicine are included, but it is likely

that WHO and other institutes of NIH, in particular, fund biomedical research in these diseases

which is not included in this analysis.

Furthermore, information about the contribution of pharmaceutical companies to research in

these diseases is not available. Although the PMA

suggests that tropical disease research receives

somes percent of the $1.3 billion spent by all U.S.

industry on biomedical research and development, this figure cannot be substantiated.

Types of Research Objectives

Recently, AID attempted to classify TDR research projects according to four research objectives: diagnostic methods, chemotherapy, immunology and vaccination, and vector control (108).

AID compiled a matrix of research projects by

looking at a list of TDR project titles and, using

the opinion of a small number of experienced

scientists, allocated each research project to one

disease category and one research objective category. The matrix that was developed by AID cannot be construed as comprehensive or definitive,

but it does give some indication of the relative

attention paid to each objective as research funds

were allocated among diseases and disease types.

Table 3-14 is an expansion of AID’s matrix for

the six TDR diseases. The figures in table 3-14 include projects funded by NIAID, AID, DOD, the

Rockefeller Foundation, and the Edna McConnell

Clark Foundation, as well as those funded by

TDR.

In the data presented in table 3-14, the research

projects included under TDR include relevant

projects funded under both the disease-specific

and transdisease components of TDR’s Research

and Development program area. Thus, in table

3-14 (unlike in table 3-1 concerned with TDR as

a funding source), a research project funded under

one of the four transdisease components of the

program area (i.e., biomedical sciences, vector

control and biology, epidemiology, or social and

economic research) was included if it seemed to

relate directly to one of the six TDR diseases.
Table 3-14.—Distribution of Funding for Biomedical Research on the Six TDR Diseasesa

.

Diagnostic methods Chemotherapy immunology/vaccination Vector control

Number of Amount Number of Amount Number of Amount Number of Amount

Disease projects (000s) projects (000s) protects (000s) projects (000s)

by Research Objective, 1981

Other Total

Number of Amount Number of Amount

projects (000s) projects (000s)

Malaria 2 $ 118 47 $3.819 67 $8.12 8 16 $1 166

Schistosomiasis 6 310 35 1,748 48 2,800 15 446

Trypanosomiasis 4 245 15 1 027 37 2,397 6 156

Leishmaniasis 6 409 11 728 11 917 4 106

Filariasis 3 156 18 1 237 15 971 3 136

Leorosv 5 164 22 785 19 693 NA NA

27 $ 2,797 159 $16,028

31 2,400 135 7704

40 3,145 102 6,970

13 1,532 45 3,692

10 716 49 3,216

r , 15 1,052 61 2694

Total by research objective 26 $1.402 148 $9,344 197 $15,906 44 $2,010 136 $11,642 551 $40,304

Percent’ of total number

of projects 5 % 2 7 % 3 6 % 8 % 2 5 % 10070

Percent of total funding 3 % 23% 39% 5% 29% 100%

DC November 1982 M Groves U S Army Medical Research and Development Command Ff

April 1984 and J Erickson Division Chief Office of Health U S Agency for International

Detrick MD personal communication March 1984: J.E. Nutter Chief Off Ice of Program Planning and Evaluation NIAID personal communication

Development personal communication March 1984
52 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

Although the data in table 3-14 are not comprehensive and are only an indication of research objectives, they do show that immunology/vaccination was the objective of 39 percent of the total

$40.3 million funding and 35 percent of the 551

projects. Research in malaria immunology and

vaccination accounted for $8.1 million (20 percent of the total funds shown). Research in chemotherapy amounted to $9.3 million (27 percent of

the total); vector control, $2 million (8 percent);

and diagnostic methods, $1.4 million (3 percent).

Biomedical research in other aspects of the TDR

diseases amounted to $11.6 million, which is 29

percent of the total $40.3 million.

Table 3-15, presenting data on NIAID-funded

research concerned with diarrheal and enteric infections and ARIs, shows that in fiscal year 1981,

research projects with the objective of immunology/vaccination accounted for over $9 million,

or 39 percent of the $23.2 million funding NIAID

provided. Research projects with objectives other

than diagnostic methods, chemotherapy, or immunology and vaccination accounted for $12.8

million, or 55 percent of the total $23.2 million.

RECIPIENTS OF RESEARCH FUNDING

The organizations receiving funds for biomedical research in tropical diseases are predominantly

universities and research institutes. Table 3-16

presents information about the distribution of extramural biomedical research funds from various

sources to institutions in the United States, in

other industrialized countries, and in less developed countries.

Table 3-16 suggests that about one-third of

TDR biomedical research funding in 1981 was

awarded to institutions in the United States. The

Country Profile: USA, published by TDR, reports

that between 1975 and mid-1982, $15,103,228 (20

percent) of the $77.3 million obligated by the program was in support of research and training by

U.S. institutions. During this same period, the

United States contributed $15,372,912 to TDR

(352).

CDD, the WHO program in diarrheal diseases,

awarded $214,000, or one-quarter of its biomedical research funds, to U.S. institutions during 1982

(see table 3-16). In 1982, the United States contributed less than $80,000 to CDD, about 1 percent of the total CDD budget (427).

NIAID research in tropical medicine and international health is concentrated at U.S. institutions

(see table 3-16), though these institutions often

work in close collaboration with scientists in the

developing world. In addition, NIAID has a role

in bilateral programs for scientific exchange and

collaboration with Egypt, Israel, and India, which

are funded by other agencies. On occasion, NIAID

makes grant and contract awards to institutes outside the United States, usually in other industrialized countries. The distribution of NIAID funding for intramural and extramural (U.S. and

foreign) research in tropical diseases during fiscal years 1981, 1982, and 1983 is shown in table

3-17.

In fiscal year 1983, AID’s Bureau of Science and

Technology allocated almost three-quarters of its

biomedical research funds to U.S. institutions (see

table 3-16) (110). Many of these institutions collaborate with organizations in less developed

countries. The remaining biomedical research

funding is in the form of support to the International Center for Diarrheal Disease Research in

Bangladesh. AID’s Research Grants Program administered by the Science Advisor’s Office and by

BOSTID has allocated 45 percent of its resources

to American institutions (11,255). The BOSTID

portion of this program makes grants only to institutions in developing countries.

The DOD biomedical research program allocates about three-quarters of its extramural funds

to organizations in the United States (see table 3-

16). A further 18 percent is allocated to organizations in less developed countries and 5 percent

to organizations in industrialized countries other
Table 3-15.—NIAID Funding for Biomedical Research on Diarrheal and Enteric Infections and ARIs

by Research Objective, Fiscal Year 1981

Immunology

Diagnostic methods Chemotherapy vaccination Other Total

Number of Amount Number of Amount Number of Amount Number of Amount Amount

projects (000s) projects (000s) projects (000s) projects (000s) (000s)

Diarrheal and enteric infections . . . . O $ 0 0 $ 0 17 $1,657 51 $5,299 $6,956

ARCS . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 749 6 487 48 7,443 76 7,582 16,261

Total by research objective . . . . . . 7 $749 6 $487 65 $9,100 127 $12,881 $23,217

Percent of total amount for

all research objectives . . . . . . 30/0 2% 39 ”/0 550/0 100 ”/0

SOURCE J E. Nutter, Chief, Office of Program Planning and Evaluation, NIAID, National Institutes of Health, Bethesda, MD, personal communication, April 1984

0

●
54 ● Status of biomedical Research and Related Technology or Tropical Diseases

Table 3-16.-Distribution of Extramural Biomedical Research Funds by Various Funding Sources to

Recipient Organizations in the United States, Other Industrialized Countries, and Less Developed Countries,

Various Years

Other

industrialized

United States countries

Amount Amount

Funding source (000s) Percent (000s) Percent

Less developed

countries Total

Amount

(000s) Percent

Amount

(000s)

TDR (1981) . . . . . . . . . . . . . . . . . . $ 4,003

WHOICDD (1982) . . . . . . . . . . . . . 214

NIAID (fiscal year 1982)a . . . . . . 24,346

Al D/Bureau of Science and

Technology b (fiscal year 1983) 5,987

AlD/Science Advisorc . . . . . . . . . 3,608

DOD (fiscal year 1983) . . . . . . . . 3,984

Edna McConnell Clark

Foundation (1981) . . . . . . . . . . 1,667

Rockefeller Foundation

(1981) . . . . . . . . . . . . . . . . . . . . . 995

320/o

25

96

$4,313

205

1,039

340/0

24

4

73

45

76

77

52

0

0

285

436

579

0

0

5

20

30

$4,386 350/0

438 51

0 0

2,217 27

4,433 55

952 18

76 3

345 18

$12,702

857

25,385

8,204

8,041

5,221

2,179

1,919

Total by recipient country . . . $44,804 $6,857 $12,847 $64,508

Percent of total for

Table 3-17.—NIAID Funding for Tropical Medicine and International Health by Type of Recipient,

Fiscal Years 1981-83

than the United States. The distribution of DOD The comparable figures for the Edna McConnell

funding, both U.S. and foreign, in fiscal years Clark Foundation are 77 percent, 20 percent, and

1981, 1982, and 1983 is shown in table 3-18. 3 percent (105).

In 1981, the majority of research funds of both Institutions in the United States received $44.8

the Rockefeller Foundation and the Edna McCon- million (69 percent) of biomedical research funds

nell Clark Foundation was awarded to U.S. insti- from the funding sources identified in table 3-16.

tutions (see table 3-16). The Rockefeller Founda- Institutions located in other industrialized countion awarded 52 percent of grants in the United tries received $6.7 million (11 percent); and those

States, 30 percent to other industrialized countries, in less developed countries received $12.8 million

and 18 percent to less developed countries (293). (20 percent).
Ch. 3—Funding of Tropical Disease Research . 55

Table 3-18.—DODa Funding for Biomedical Research on Tropical Diseases by Type of Recipient,

Fiscal Years 1981-83 -

-.

Fiscal year 1981 Fiscal year 1982 Fiscal year 1983

Amount Amount Amount

(000s) Percent (000s) Percent (000s) Percent

Fiscal years 1981-83

Amount

(000s) Percent

Intramural . . . . . . . . . . . . $ 7,867 630/o $8,431 630/o $8,890 63 0/0

Extramural U. S.. . . . . . . 3,244 26 3,554 26 2,984 28

Extramural foreignb . . . . 1,377 11 1,450 11 ”/0 1,237 9

Total by year. . . . . . . . $12,488 100 ”/0 $13,436 100 ”/0 $14,111 100 ”/0 $40,034 100 ”/0

$25,188 63

10,782 27

4,064 10

SUMMARY

Several departments and agencies of the U.S.

Government support biomedical research on tropical diseases. The important players are DHHS,

AID, and DOD. The U.S. Government also supports international programs in tropical disease

research. Of greatest consequence are TDR and

WHO’s various research programs. A small number of U.S. foundations support research in tropical diseases. The Rockefeller Foundation and the

Edna McConnell Clark Foundation have a long

history of involvement in this field, and the

MacArthur Foundation has become active more

recently. A handful of pharmaceutical companies

also invest in research to develop products for

tropical diseases, but funding levels are undocumented.

Actual dollar amounts for funding of tropical

disease research are difficult to assemble for a variety of reasons. The figures presented in this report represent the best efforts of the public and

private bodies to provide this information, and

OTA believes that the totals are sufficiently accurate to give a rough estimate of U.S. spending

for tropical disease research. OTA estimates that

U.S. public and private organizations (excluding

pharmaceutical companies) have spent somewhat

less than $100 million per year on tropical disease research. This figure includes contributions

to international programs.

A high proportion of U.S. tropical disease research funds is awarded to U.S. investigators.

About 96 percent of NIAID extramural research

funds for this purpose goes to Americans. Perhaps more surprising, U.S. research institutions

and investigators have been awarded at least as

much as the U.S. Government has contributed to

TDR and the WHO programs. Only one program—

the AID-supported Research Grants Program administered by the National Academy of Sciences’

BOSTID—is specifically designed to make grants

only to institutions in developing countries.
4

Description of Selected

Tropical Diseases
Contents

Page

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . 59

Malaria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59

Aspects of Natural History . . . . . . . . . . . . 59

Malaria Control . . . . . . . . . . . . . . . . . . . . . . 61

Recent Progress . . . . . . . . . . . . . . . . . . . . 64

Research Needs . . . . . . . . . . . . . . . . . . . . . . 65

Schistosomiasis. . . . . . . . . . . . . . . . . . . . . . . . . 65

Aspects of Natural History . . . . . . . . . . . . 65

Interventions. . . . . . . . . . . . . . . . . . . . . . . 67

Recent Progress . . . . . . . . . . . . . . . . . . . . . . 67

Trypanosomiasis . . . . . . . . . . . . . . . . . . . . . . 67

African Sleeping Sickness . . . . . . . . . . . . . 69

Chagas’ Disease . . . . . . . . . . . . . . . . . . . . . . 71

Leishmaniasis . . . . . . . . . . . . . . . . . . . . . . . . . 74

Aspects of Natural History . . . . . . . . . . . . 74

Interventions. . . . . . . . . . . . . . . . . . . . . . 78

Recent Progress . . . . . . . . . . . . . . . . . . . . . . 78

Filariasis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78

Aspects of Natural History . . . . . . . . . . . . 78

Obstacles to Research . . . . . . . . . . . . . . . . . 80

Leprosy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80

Aspects of Natural History . . . . . . . . . . . . 80

Interventions. . . . . . . . . . . . . . . . . . . . . . . . . 83

Recent Progress . . . . . . . . . . . . . . . . . . . . . . 83

Tuberculosis . . . . . . . . . . . . . . . . . . . . . . . . . . . 84

Aspects of Natural History . . . . . . . . . . . . 84

Interventions. . . . . . . . . . . . . . . . . . . . . . . . . 84

Recent Progress . . . . . . . . . . . . . . . . . . . . . . 85

Diarrhea] and Enteric Diseases . . . . . . . . . . . 85

Aspects of Natural History . . . . . . . . . . 85

Viruses. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86

Bacteria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86

Protozoa and Other Agents.. . . . . . . . . . 88

Acute Respiratory Infections. . . . . . . . . . . . . 88

Aspects of Natural History . . . . . . . . . . . . 88

Viruses. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89

Bacteria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89

Other ARI Agents. . . . . . . . . . . . . . . . . . . . 90

Interventions. . . . . . . . . . . . . . . . . . . . . . . . . 90

Biotechnology and ARIs . . . . . . . . . . . . . . 91

Arboviral and Related Viral Infections . . . . 91

Aspects of Natural History . . . . . . . . . . . . 91

Research Needs . . . . . . . . . . . . . . . . . . . . . . 94

TABLE

Table No, Page

4-1. Location in This Report of lnformation

About Selected Tropical Diseases . . . . . 60

Figure

4-1.

4-2.

4-3.

4-4.

4-5.

4-6.

4-7.

4-8.

4-9.

4-1o

4-11.

4-12.

4-13.

4-14.

4-15.

4-16.

4-17.

LIST OF FIGURES

No. Page

Generalized Life Cycle of Plasmodium,

the Cause of Malaria . . . . . . . . . . . . . .

Geographic Distribution of Malariaas

of 1982 . . . . . . . . . . . . . . . . . . . . . . . , .

Geographic Distribution of

Chloroquine-Resistant Malaria as

of 1983 . . . . . . . . . . . . . . . . . . . . . . . . . .

Generalized Life Cycle of Schistosorna,

the Cause of Schistosomiasis. . . . . . . .

Geographic Distribution of

Schistosomiasis: Schistosoma

haematobium, S. mansoni and S.

japonicum . . . . . . . . . . . . . . . . . . . . . . . .

Generalized Life Cycle of Trypanosoma

bruceigambiense and T.b. rhodesiense,

the Causes of African Sleeping Sickness

Geographic Distribution of African

Sleeping Sickness . . . . . . . . . . . . . . . . . .

Life Cycle of Trypanosoma cruzi the

Cause of Chagas’ Disease . . . . . . . . . . .

Geographic Distribution of

Chagas’ Disease . . . . . . . . . . . . . . . . . . .

Generalized Life Cycle of leishmania,

the Cause of Leishmaniasis . . . . . . . . .

Geographic Distribution of

Leishmaniasis Caused by Four Species

of Leishmania . . . . . . . . . . . . . . . . . . . . .

Generalized Life Cycle of Two

Important Filarial Worms: Wuchereria

bancrofti and Brugia malayi . . . . . . . .

Geographic Distribution of Major

Filarial Diseases: Infection With

Onchocerca volvulus, Brugia malayi,

and Wuchereria bancrofti . . . . . . . . . .

Geographic Distribution of Leprosy..

Generalized Transmission Cycle of

an Arbovirus . . . , . . . , . . . . . . . . . . . .

Geographic Distribution of

Yellow Fever. . . . . . . . . . . . . . . . . . . . . .

Geographic Distribution of

Dengue Fever . . . . . . . . . . . . . . . . . . . . .

62

63

64

66

68

70

71

73

74

76

77

79

81

83

92

93

94
4.

Description of Selected Tropical Disease

INTRODUCTION

This chapter describes the major tropical diseases that are discussed in this report. The diseases considered are a rather diverse group.

Representing the diseases traditionally considered “tropical” in this report are malaria, schistosomiasis, trypanosomiasis, leishmaniasis, filariasis, and leprosy. These are the six diseases

singled out for attention by the Special Program

for Research and Training in Tropical Diseases

(TDR), which is sponsored jointly by the U.N.

Development Program, the World Bank, and the

World Health Organization (WHO). The six TDR

diseases together affect between 700 million and

800 million people worldwide. Malaria, schistosomiasis, and filariasis each affects more than 200

million people.

Other diseases discussed in this report—tuberculosis, diarrheal diseases, acute respiratory infections (ARIs), and arboviral infections—occur

in nontropical countries as well as tropical ones.

The toll these diseases take in developing countries is much greater than the toll they take in the

developed world because of developing countries’

higher incidence rates and poorer diagnostic and

therapeutic methods.

Tuberculosis was once a public health problem

worldwide. It now persists mainly in developing

countries, where it infects large numbers of

people.

Diarrheal diseases and ARIs are the leading

causes of death among infants and children in developing countries. In countries where the infant

mortality rate is more than 100 per 1,000 live

births, at least one-third of infant deaths are from

diarrheal diseases.

Arboviral infections have a worldwide distribution, but are concentrated in the tropics. With

no specific treatments available and a vaccine

against only two (yellow fever and Japanese encephalitis), arboviral infections are a potential

threat to the U.S. population.

These are far from the only health problems in

tropical developing countries. In addition to being affected by the often debilitating infectious diseases discussed here, people in the developing

world are increasingly affected by the chronic diseases that cause so much sickness and death in

the developed world: heart disease, cancer, stroke,

and diabetes, for instance. Ironically, these

chronic diseases increase in incidence as life expectancy increases and as the probability of dying from infectious diseases decreases.

This chapter provides basic information about

each of the diseases considered in the later chapters of this report. Chapter 5 discusses strategies

for controlling tropical diseases, and chapters 6

through 9 examine the status of disease control

measures: vector control technologies, immunization technologies, diagnostic technologies, and

therapeutic technologies. Table 4-1 summarizes

some basic information about each disease and

guides the reader to the relevant sections, for each

disease, in the remainder of the report.

MALARIA

Aspects of Natural History is no question that malaria is the most important of all infectious diseases.

In 1940, the Nobel Laureate Sir Macfarlane

Burnet wrote (37):

His statement still holds true. Malaria is one of

the most studied of all tropical diseases. The disIf we take as our standard of importance the ease is caused by various species of the protozoan

greatest harm to the greatest number, then there genus Plasmodium . Malaria parasites have a com59
60 ● Status Of Biomedical Research and Related Technology for Tropical Diseases
Ch. 4—Description of Selected Tropical Diseases Ž 61

plex life cycle, alternating between vertebrate

hosts and mosquito vectors of the genus Anopheles.

Four species of Plasmodium cause malaria in

humans: Plasmodium falciparum, P. vivax, P.

ovale, and P. malariae. Other species of Plasmodium infect a wide variety of other vertebrates.

The four species that infect humans have somewhat different clinical effects. P. falciparum can

cause severe anemia, kidney failure, and brain

damage and is often fatal, especially in children.

P. vivax and P. ovale infections are seldom fatal, but relapses of the symptoms (chills, high fever, nausea, headache, etc. ) can occur periodically

for up to 3 years. P. malariae infections can persist in the blood for years without causing any

symptoms.

Life Cycle

All species of Plasmodium progress through a

similar life cycle, though each species differs in

some of the details. All human malaria infection

begins with the bite of an infected female Anopheles

mosquito (see fig. 4-l). As the mosquito ingests

a blood meal to nourish her eggs, she incidentally

injects saliva containing plasmodial sporozoites

(which have been clustered in the mosquito’s salivary glands) into the human bloodstream.

Within about an hour, the threadlike sporozoites leave the bloodstream and move to the human liver. Over the next week or two, depending on the species of Plasmodium, each sporozoite

that has invaded a liver cell becomes a schizont,

a developmental stage that contains thousands of

merozoites, the next stage in the life cycle of the

parasite. When the schizont matures, it ruptures

out of the infected liver cell and discharges

merozoites into the human host’s bloodstream.

In P. vivax and P. ovale malaria, some sporozoites, instead of developing into shizonts, become

hypnozoites, forms that can remain dormant in

the liver for months or years before they start to

proliferate.

Merozoites invade red blood cells (erythrocytes)

and there undergo a second round of asexual reproduction, similar to that in the liver. In 2 or

3 days, the merozoites develop into trophozoites,

then into a second dividing schizont form. When

the schizonts mature, they rupture and release

another round of merozoites, perpetuating the cycle of infection. It is at the time of this rupture

that clinical symptoms of malaria appear. The cycle repeats every 2 or 3 days, depending on the

species of Plasmodium.

Some merozoites, instead of developing into

schizonts, differentiate into sexual forms, gametocytes. Mature gametocytes remain in the host’s

red blood cells, and can be ingested by female

Anopheles when they bite. Gametocytes develop

further into male and female gametes, which

undergo sexual reproduction in the mosquito’s

gut. Eventually a new generation of sporozoites

develops in the mosquito and migrates to the

mosquito’s salivary glands, ready to infect another

human.

Incidence and Prevalence

Malaria is one of the most widespread and destructive of diseases, having doubled in world

prevalence in the last decade (430). Worldwide,

an estimated 250 million to 300 million cases of

malaria occur each year. In tropical Africa alone,

an estimated 160 million to 200 million people are

infected every year, and 1 million people die,

mostly infants and small children (193).

About 1.5 billion people live in areas of the

world where the risk of malaria ranges from moderate to high (see fig. 4-2). The countries of highest malaria incidence are Haiti, Guatemala, Honduras, El Salvador, Colombia, Bolivia, Brazil,

India, Sri Lanka, Pakistan, and many parts of

Africa and Asia. In Europe, the Caribbean, North

America, and parts of South America, and Australia, the gains against malaria brought about by

the WHO eradication program (see discussion below) have been maintained. However, developed

nations such as England and the United States

have experienced an increase in the number of imported malaria cases.

Malaria Control

Before World War II, approximately two-thirds

of the worlds population was at risk for malaria.

DDT (dichloro-diphenyl-trichloro-ethane) was initially highly successful at controlling the Anopheles
62 ● Status of Biomedical Research and Related Technology for Tropical Diseases

0

o

c
Ch. 4—Description of Selected Tropical Diseases ● 6 3

Figure 4“2.–Geographic Distribution of Malaria as of 1982

Key:

O Areas in which malaria has disappeared, been eradicated, or never existed

o Areas with limited risk

~ Areas where malaria transmission occurs

SOURCE U S Department of Health and Human Services, Centers for Disease Control, Hea/th /nforrnat/orr for /n fernaf/ona/ Trave/. HHS Publlcatlon No

(CDC)840-8280 (Washington, DC U S Government Prlntlng office, 1984)

mosquito vector of malaria, however, and because of this, WHO began a worldwide malaria

eradication program in the late 1950s. Large-scale

spraying efforts kept the mosquito population

down, eliminating the risk of disease for some 400

million people, mostly in temperate regions.

By the mid-1960s, malaria had been eliminated

from nearly all of Europe, most of the Asian part

of the U. S. S. R., several countries of the Near East,

most of North America including all of the United

States, most of the Caribbean, large areas of the

northern and southern parts of South America,

Australia, Japan, Singapore, Korea, and Taiwan.

Eighty percent of the originally malarious area

was almost free of malaria.

By 1969, however, there had been little additional progress. The worldwide eradication campaign failed for several reasons. Increasing

mosquito resistance to DDT and P. falciparum

resistance to chloroquine, the most widely used

malaria drug, were two of the most important

(33). Other factors in the increased incidence of

malaria were behavioral changes in the mosquito

vector species and the migration of workers who

unintentionally brought the vectors with them to

new areas.

While efforts to keep the Anopheles vector under control have been successful in some areas,

the cost of applying traditional malaria interventions has greatly restricted the ability of developing countries to protect the populations at risk.

Furthermore, few efforts to slow the spread of

drug-resistant malaria have worked. Resistance

of P. falciparum to chloroquine has been present

in Panama, parts of some South American countries, India, Southeast Asia, Indonesia, China, the

Republic of the Philippines, and other Pacific Islands for several years (see fig. 4-3). In Latin
64 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 4-3.—Geographic Distribution of Chloroquine-Resistant Malaria as of 1983

 Areas where chlorquine -

resistant malaria has

America, resistant strains of the organism are appearing in most parts of Bolivia, Venezuela,

French Guyana, and northern Peru, but serious

drug-resistance has still not moved north of the

Panama Canal. There is growing evidence of resistance in east Africa, which includes Kenya,

Tanzania, eastern Zaire, Burundi, Uganda, Rwanda,

Malawi, Zambia, northern Sudan, Madagascar,

and the Comoro Islands (376).

Resistance to chloroquine is not the only problem. Some malaria parasites have also developed

resistance to newer drugs. Regular monitoring of

local parasite strains is necessary as an indicator

of therapeutic changes that may be needed, both

in type of drug and dose. Renewed efforts to develop new drugs are producing results, but moving new drugs from laboratory screening through

animal testing to human trials takes years.

Recent Progress

Since 1976, with the development of methods

allowing for the continuous cultivation of malaria

parasites in the laboratory, research on the immunology of malaria has moved ahead considerably. All life cycle stages of the malaria parasite can now be grown in culture. An important

recent discovery is that the proteins on the surface of the parasite (surface antigens) vary not

only among the species of Plasmodium but also

among strains of a single species. Perhaps of even

greater interest is the finding that malaria parasites can change their surface antigens during the

course of an infection. The human (or animal)

host, in turn, must play “catch-up” in order to

destroy the new forms of the parasite. This finding has direct implications for the development

of vaccines and diagnostic tests (231).
Ch. 4—Description of Selected Tropical Diseases . 65

Basic malaria research has advanced considerably. Metabolic studies have identified parasitespecific enzyme pathways that can be exploited

to kill the malaria parasite without harming the

human host. Membrane research has revealed

how the parasite finds, attaches to, and invades

the human host’s red blood cells, yielding important clues for drug therapy and vaccine research.

Recent clinical studies have suggested better ways

of preventing and treating cerebral malaria, an

often fatal complication of severe malaria infection.

A vaccine against the sporozoite stage of malaria may be available for testing by 1986. Vaccines against the merozoite and the gamete stages

are behind this in development. If animal testing

in humans confirms the feasibility of immunization against the malaria parasite, extensive human

and field trials will be required before the vaccine can be widely used. Furthermore, given the

difficulties that have plagued other disease immunization campaigns there is some debate about the

SCHISTOSOMIASIS

Aspects of Natural History

Schistosomiasis is a debilitating disease caused

by trematode worms of the genus Schistosorna.

There are three major species affecting humans,

Schistosoma mansoni, S. haematobium, and S.

japonicum, all of which originated in the Old

World but now occur worldwide. S. mansoni

originated in Africa, but now has become established in the Americas. A fourth species, S.

mekongi, was discovered in the late 1960s and is

endemic in areas of Southeast Asia.

Life Cycle

The adult parasitic worms that cause schistosomiasis live in pairs in the human host’s bloodstream, sometimes for many years. Eggs (from a

few hundred to several thousand per day) are

produced by the female schistosome and are deposited in the host’s blood vessels. The eggs escape into the host’,s bowel (S. mansoni, S. japonicum) or bladder (S. haematobium), and then are

excreted in feces or urine. If the eggs become

usefulness of a malaria vaccine (see Case Study

B: The Development of a Malaria Vaccine for a

more comp

Research

ete discussion).

Needs

Field- and community-based studies are needed

to assess the impact of antimalarial interventions.

The emergence of insecticide-resistant mosquito

vectors of malaria has seriously handicapped current malaria control efforts, making studies of the

ecological impact of future interventions critically

important. The effect of antimalarial activities on

the actual immunity levels of populations needs

clarification. Studies in the past have clearly documented the immediate impact of antimalarial

projects on morbidity and mortality, but not the

long-term consequences when projects cease or

fail. Other studies are needed to evaluate the importance of sociological and human behavioral

factors and the usefulness of health education,

community self-help, and volunteer collaborators.

trapped in the human host’s bladder or liver, however, granulomas (masses of small blood vessels

and connective tissue) form around them, and the

eggs eventually die and calcify, producing inflammation and scars. The damage done by schistosomiasis is primarily due to the human body’s reaction to accumulated eggs and their associated

granulomas.

The schistosome eggs that are excreted by the

host into freshwater lakes or streams hatch into

ciliated larval worms (miracidia). Miracidia then

enter an intermediate snail host, in which they

proliferate. Immature worms (cercariae) are released from the snail and can rapidly penetrate

the unbroken skin of persons who enter infected

water (see fig. 4-4). The tailless immature worms

(now called shistosomules) enter the human circulatory system and proceed to the liver, where

they mature. As they mature, they coat their surfaces with proteins acquired from their human

hosts, which enable them to fool the hosts’ immune system into tolerating their presence (130).

After emerging from the liver, mature worms
66 . Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 4-4.—Generalized Life Cycle of Schistosoma, the Cause of Schistosomiasis

Excreted

in feces

l \

4

Excreted

1. Microscopic cercariae are released from snail intermediate host.

2. Cercariae penetrate skin of human host.

3. Tailless cercariae, now called schistosomules, migrate to the human host’s small blood vessels, are carried to lungs, then

through the heart into the liver to mature; paired (male and female) mature schistosomes lodge in the human host’s veins

(S. mansoni shown).

4. Female schistosomes continually produce eggs (up to thousands each day), which penetrate the human host’s intestine

(S. mansoni and S. japonicum) or the urinary bladder (S. haernatobium), from which they are excreted in feces or urine,

respectively.

5. Eggs hatch in fresh water into ciliated miracidia.

6. Miracidia penetrate intermediate snail host, lose cilia, and become sporocysts. Sporocysts reproduce asexually in the snail,

proliferating greatly, eventually giving rise to cercariae.

SOURCES Adapted from T C Cheng. Symbiosis (New York Pegasus, 1970), and E R Noble and G A Noble Parasitology. 3d ed (philadelphia Lea & Febiger. 1973)
Ch. 4—Description of Selected Tropical Diseases . 67

mate for life within small veins around either the

bowel or bladder area, thus completing the parasite’s life cycle.

Incidence

A worldwide survey in 1972 (including 71 countries) estimated that 500 million people were exposed to infection by schistosomiasis and 125 million were infected (169). The various forms of this

disease occur in parts of Africa, the Caribbean,

South America, and the Orient (see fig. 4-5). In

Latin America, recorded schistosomiasis incidence

is highest in Surinam, where 385 of every 100,000

inhabitants were infected in 1980 (265). S. mansoni is also established in suitable snail hosts in

more than half of Brazil, where 10 million people are believed infected, and in parts of Venezuela, where 10,000 more people are thought to

have the disease. Foci in the Caribbean occur in

the Dominican Republic, Guadaloupe, Martinique, and St. Martin. A few cases have been

detected in Montserrat.

Interventions

Over the last 10 years, the incidence and prevalence of schistosomiasis have dropped considerably in several countries. In Japan, the prevalence

dropped from 25 percent in 1950 to less than 1

percent in 1973 (169). This formidable decrease

was brought about by the concrete lining of irrigation ditches, snail control, land reclamation,

environmental sanitation, chemotherapy, and

health education. Similar decreases reported for

some parts of Egypt, Iran, Puerto Rico, Tunisia,

and Venezuela were brought about by more or

TRYPANOSOMIASIS

Trypanosomiasis is a general term for two separate diseases caused by protozoan parasites of

the genus Trypanosoma. Trypanosoma brucei infections cause African sleeping sickness (also

called African trypanosomiasis), and T. cruzi infections cause Chagas’ disease (also called American trypanosomiasis). The two diseases have

less the same combination of control methods and

improved socioeconomic conditions (169). On the

minus side, schistosomiasis has spread to new

areas as a result of water impoundment and irrigation projects which create and expand suitable

environmental conditions for snail hosts and increase human-snail contact. Areas where large

hydroelectric dams are being built, especially in

South America, may require special surveillance

and assessment.

Chemotherapy against schistosomiasis with the

newest generation of chemotherapeutic agents is

effective and relatively safe. However, total and

complete control in endemic areas is difficult to

achieve, since it requires attention to other measures, particularly water supply and sanitation,

and treatment of snail breeding sites (mollusciciding).

Recent Progress

A fairly large corps of American researchers

studies schistosomiasis, and about a dozen laboratories use the techniques of biotechnology. The

life cycle of the schistosomiasis parasite is readily adaptable to the laboratory, and small rodents

are easily infected. The immunology of schistosomiasis has been studied extensively (39). Most

workers are attempting to identify and isolate relevant protective antigens, and many are actively

engaged in gene cloning experiments. A live irradiated larval vaccine has been used in cattle in

the Sudan with encouraging results (338), but its

use in humans is not feasible, because the larvae

are alive (though unable to continue their life

cycle) when injected.

completely different transmission cycles, different vectors, and cause a different pathology. Chagas’ disease begins as a blood infection but ultimately attacks various body organs, principally

the heart, African sleeping sickness also begins as

a blood infection but progresses to central nervous system disease and death,
68 . Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 4-5.—Geographic Distribution of Schistosomiasis:

Schistosoma haematobium, S. mansoni, and S. japonicum

SOURCES: V Zaman and L Keong, Handbook of Medical Parasitology (Australia: ADIS Health Science Press, 1982); and

H. W. Brown and F A. Neva, Basic Clincal Parasitology (East Norwalk, CT: Appleton-Century-Crofts, 1983), used

by permission
Ch. 4—Description of Selected Tropical Diseases ● 6 9

African Sleeping Sickness

(African Trypanosomiasis)

Aspects of Natural History

T. brucei, the agent of African sleeping sickness in humans and nagana in livestock, is endemic to large areas of the African continent but

does not occur naturally outside of Africa. In the

United States, there have been six published reports of imported cases of African sleeping sickness since 1967. All the infected travelers had been

on vacation in game parks in eastern or southern

Africa, where they were exposed to the tsetse fly

vector. The tsetse fly vector of African sleeping

sickness is not found in the United States, so there

is little danger that the disease will become established in this country (368).

Life Cycle.—There are actually two kinds of

African sleeping sickness, each caused by a different variety of T. brucei and occurring in its own

environmental niche. Both forms of the disease

have an early stage involving the blood and lymphatic system and a late stage involving the brain.

In west Africa, sleeping sickness is caused by

the parasite T. brucei gambiense, which is transmitted by tsetse flies (Glossina palpalis) that feed

only on humans. The disease is transmitted near

streams where tsetse flies breed. T. b. gambiense

infection usually results in a chronic condition that

slowly leads to death.

In east Africa, sleeping sickness is actually a

zoonosis (a disease of animals that can be transmitted to humans) caused by T. b. rhodesiense

and transmitted by a different species of tsetse fly

(G. morsitans). Humans acquire the disease when

they venture into areas, such as hunting grounds

and grazing lands, where animals are infected.

The disease is fatal to humans within weeks or

months and very destructive to livestock. Large

expanses of land have become totally unusable

because of the disease risk to humans entering

them.

Either form of African sleeping sickness begins

when an infected tsetse fly feeds on a human or

animal host, depositing trypanosomes in the

wound (see fig. 4-6). The trypanosomes invade

the host’s blood, lymph, and tissue fluids. In cyclical waves, the trypanosomes wane, then reappear, as the body’s immune system and the

parasites interact. These parasites can undergo frequent changes in a specific surface protein, known

as variant surface glycoprotein (VSG) or variant

surface antigen. Located on the blood form of the

trypanosome, VSGS stimulate the production by

the host of a particular responsive and protective

antibody.

Within any individual human or animal host,

the VSGS of the trypanosome population shift

over time through a repertoire of changes numbering potentially over 100, each having no exposed determinants in common with other past

or future populations. Two distinct mechanisms

appear to be responsible for the VSG changes,

which are called antigenic switching. Each mechanism generates cohorts of parasites to which the

mammalian host has not yet made antibody.

Since antibodies raised against one variant are

generally not effective against other variants, the

trypanosome is able to evade the host’s immune

response.

Until the host can mount another antibody response, symptoms of fever, headache, and joint

pain occur. As the infection progresses, lesions

develop in the brain, heart, and small blood vessels. Ultimately, nervous system involvement

develops, first with signs of insomnia and excitability, then coma and death. Symptoms and outcome are similar in both the acute and chronic

forms, but the duration differs.

Incidence.—Recent estimates put 45 million

people at risk of sleeping sickness infection in

Africa. Until 1979, about 10,000 new cases were

recorded annually, but since then, serious outbreaks have resulted in more than 20,000 new

cases recorded per year (353). Because of the difficulties of diagnosing chronic cases and the usual

occurrence of the disease in rural areas, these

figures are probably underestimates. Although

widespread across Africa (see fig. 4-7), sleeping

sicknesss occurs in well-defined endemic foci because the tsetse fly vectors breed in rivers and

streams. Unfortunately, those same water sources

are essential to humans and grazing animals.
70 . Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 4-6.—Generalized Life Cycle of Trypanosoma brucei gambiense and T. b. rhodesiense, the Causes of

African Sleeping Sickness (African Trypanosomiasis)

v

Trypanosomes develop in tsetse fly vector to a form infective to humans and other mammals.

Humans and animals are infected through bite of tsetse fly. Enlarged lymph nodes are an early sign of trypanosome

infection.

Trypanosomes eventually invade the human host’s central nervous system, causing classical symptoms of sleeping

sickness, leading to coma and death.

Forms of trypanosome infective to tsetse fly vector are released into blood circulation.

Tsetse fly picks up infective form of trypanosome while biting infected human or other infected mammal.

SOURCE” Office of Technology Assessment, 1985. photos from H. W. Brown and F A Neva, Basic Clinical Parasitology (East Norwalk CT Appleton-Century-Crofts,

1983), used by permission

Interventions there is a risk (2 to 5 percent) of succumbing to

Treatment for African sleeping sickness requires the treatment itself.

hospitalization because of the dangerous side ef- Control of the tsetse fly vectors of sleeping sickfects and toxicity of the drugs, as well as the need ness is currently the only feasible means of interfor their intravenous administration. Although the vention. Because of the focal nature of transmisinfection is uniformly fatal without treatment, sion, insecticiding is feasible (no insecticide
Ch. 4—Description of Selected Tropical Diseases ● 7 1

Figure 4-7.—Geographic Distribution of African

Sleeping Sickness (African Trypanosomiasis)

resistance exists, as yet, in the vectors) and has

been used successfully, though it is expensive, contaminates the environment, and is labor-intensive.

In some cases, clearing vegetation near river

breeding sites has worked. Overall, vector control has met with only limited, sporadic success.

Recent efforts have focused on the use of insecticide-impregnated traps that attract the vector flies.

Recent Progress

Since about 1979, trypanosomes have become

increasingly popular for studies in molecular biology, The organisms of the T. brucei complex

are becoming the Drosophila (fruit flies) of modern molecular biologists. The purpose of most of

the studies using these trypanosomes has been to

work out the intricacies of gene coding, transcription, translation, and expression, rather than to

control African sleeping sickness. These studies

have made many contributions to the fundamental understanding of gene function, however, and

there is hope that useful spinoffs can be applied

to controlling sleeping sickness.

More disease-focused work by American investigators is centered largely on the molecular biology of antigen switching of VSGS. Monoclinal

antibodies (MAbs) to different VSGS have been

produced in several laboratories (43). One laboratory deals exclusively with the nonvariant antigens and regulatory proteins, studying the stagespecific expression of membrane and internal proteins and host reactions to them. In this laboratory, an attempt is being made to find proteins

that could be used in immunodiagnosis and metabolic targets for chemotherapy (16).

Chagas’ Disease

(American Trypanosomiasis)

Aspects of Natural History

Chagas’ disease is caused by the protozoan

parasite T. cruzi. Though sometimes congenital

and occasionally transmitted through blood transfusion (308), Chagas’ disease is primarily transmitted to humans (and other mammals) by reduviid

bugs, blood-sucking insects found throughout

Central and South America. Thus, it is primarily a disease of poor rural areas, where adobe

brick houses and thatch roofs provide harboring

sites for reduviid bugs to live and breed.

Several strains of T. cruzi exist in different parts

of Latin America. The strains vary considerably

in the pathology they produce in humans because

of the range of immune system reactions they can

evoke. About 150 species of mammals, including

dogs, cats, guinea pigs, opossums, rats, and other

rodents, are thought to be reservoir hosts of T.

cruzi.

There is no effective cure for Chagas’ disease.

In the acute phase of the disease, when the parasites are invading internal organs, headaches, fever, anemia, and exhaustion may occur. The

severity of this phase varies with the age of the

patient. The younger the patient, the more severe

the disease. Thus, children under the age of 2 may

die, while adults may exhibit no symptoms. The
72 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Photo credit: H. W. Brown and F. A. Neva, “Basic Clinical Parasitology, “AppletonCentury-Crofts, 1983. Reprinted by permission,

Megacolon, one effect of Chagas’ disease.

acute stage of Chagas’ disease may resolve completely in a few weeks or months or instead may

pass into a subacute or chronic stage.

Long-term sequelae of Chagas’ disease include

grotesque enlargement of the digestive tract

(megaesophagus and megacolon), circulatory

problems, and central nervous system damages

and most seriously, damage to the heart muscle,

sometimes leading to death from heart failure.

Life Cycle. -T. cruzi infection in humans results when an infected reduviid bug bites a person, usually around the eye while the person is

sleeping, and deposits feces containing 7’. cruzi

parasites into the bite wound (see fig. 4-8). T. cruzi

has two life stages in the mammalian host, one

that circulates in blood and another that proliferates intracellularly within the tissues. Forms of the

parasites infective to reduviid bugs are released

into the mammalian host’s circulation, ready to

be picked up by reduviid bugs in the course of

another insect bite.

Incidence.—Chagas’ disease occurs in almost

every country of Latin America: Brazil, Peru,

Venezuela, Chile, Bolivia, Paraguay, Uruguay,

Argentina, Colombia, Mexico, Costa Rica, and

Panama (see fig. 4-9) (265). Although the vectors

and reservoir hosts are also present across the

Southern United States, only three indigenous

cases of Chagas’ disease have been reported. Two

were infants reported to have contracted the disease in Texas in 1955, and the third was a woman

from the Sacramento Valley area in California in

1982. None of the three had previously had contact with pets carrying the disease or had recent

blood transfusions or had been outside the country (309).

It has been estimated that about 12 million of

the 50 million exposed people living in endemic

areas are infected with T. cruzi (229). Since notification of authorities regarding the presence of

Chagas’ disease is not compulsory, there are no

reliable morbidity data. Studies in Brazil have

shown Chagas’ disease to be a significant cause

of mortality in people under 45 years of age (287)

and a heavy burden to society because of the need

for hospitalization and disability assistance (268).

Interventions

Control measures for Chagas’ disease concentrate on insecticide spraying of houses and upgrading of housing construction. In one area of

Venezuela, the use of insecticides was believed to

account for a significant decline in the percentage of the population infected with T. cruzi during the 1970s (29). Vector bionomics remains an

important research topic for defining transmission

areas, vector behavioral characteristics, and improved control measures.

Recent Progress

In an attempt to understand why the human

host’s immune system is not effective in controlling Chagas’ disease, some investigators have examined the proteins produced by T. cruzi. There

is no evidence for VSGS in T. cruzi (322), although

a large number of local genetic strains do exist.

Some other mechanism must be responsible for

T. cruzi’s ability to evade the human immune

system.
Ch. 4—Description of Selected Tropical

Figure 4-8.— Life Cycle of Trypanosoma cruzi, the Cause of Chagas’ Disease

\

Diseases ● 7 3

1

1. Trypanosomes develop in the reduviid bug vector to a form infective to humans and other mammals.

2. Parasites deposited on human skin in insect feces at time of insect bite invade human through the bite wound. “Romana’s

sign,” the swelling of one eye, is a sign of a reduviid bite.

3. Trypanosomes enter and multiply in muscle tissue, including the heart.

4. Forms of trypanosomes infective to reduviid bug vector are released into blood circulation.

5. Reduviid bug picks up infective form of trypanosome while biting infected human or other infected mammal.

SOURCE Office of Technology Assessment, 1985

Major surface proteins isolated from T. cruzi system raises antibodies against both insect and

organisms resembling the insect stages of the blood stages of the parasite. MAbs to culture and

parasite (grown in culture) and from mammalian intracellular stages of T. cruzi have been made

blood stages were recognized by immune sera by several workers, and Snary and colleagues

from naturally infected humans and experimen- (323) have demonstrated that the antigenic detertally infected mice (259). This means the immune minants are strain-specific. The ideal vaccine must
74 . Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 4-9.—Geographic Distribution of Chagas’

Disease (American Trypanosomiasis)

SOURCES: World Health Organization, Geneva, Switzerland, and London School

of Hygiene and Tropical Medicine.

LEISHMANIASIS

Aspects of Natural History

Leishmaniasis is the collective term for the diseases caused by several species of the protozoal

genus Leishmania. Among the protozoan diseases,

leishmaniasis is commonly considered second in

importance, following malaria.

Depending on the infecting species, leishmaniasis may appear in different forms. Cutaneous

leishmaniasis, which appears as self-limiting and

usually self-resolving sores located at the point

of infection, is caused by L. mexicana, L. braziliensis, or L. tropica. Mucocutaneous leishmaniasis, which also begins as a sore, is caused by certain geographic strains of L. brazdiensis that

produce antibodies that do not cross-react with

heart and nerve tissue and that are effective

against all strains and stages of T. cruzi.

The development of an effective therapeutic

drug for Chagas’ disease is a critical research need.

With a therapeutic drug in hand, a simple effective test for early diagnosis of the disease would

be essential, for once the long-term effects appear

they are irreversible. Vaccine research is under

way, but because the long-term pathology seems

to result from the body’s immune response against

the parasite and cross-reacting with its own heart

and nerve tissue, prospects for an effective vaccine are uncertain.

Fewer than 10 laboratories in the United States

focus on T. cruzi. Most are in academic institutions, with one or two each in government and

in industry. Significant research is taking place

in South America, as well as in a few European

laboratories. In part because of this dearth of effort, work on Chagas’ disease has not advanced

as much as that on some other diseases (16).

commonly metastasize and proliferate in the nasal and pharyngeal mucous membranes. Gross destructive disfigurement of the face, nose, and

throat results.

A third type of this leishmaniasis is a visceral

form, caused by L. donovani. In this form, called

“kala azar, ” the spleen, liver, bone marrow, and

lymph glands are the sites of parasite proliferation. Fatal outcome in children is common. Kala

azar occurs sporadically in many tropical areas,

but it has also appeared in epidemics, killing many

thousands of people at a time in southern Asia.

Recent epidemics of kala azar have resulted from

the reemergence of the sandfly vector after spray

programs were discontinued.
Photo credit Dr Roberf Edelrnan, National Institutes of Health

A lesion of cutaneous Ieishmaniasis.

Photo credit” Office of Technology Assessment

Destruction of tissue resulting from mucocutaneous

Ieishmaniasis.

Ch. 4—Description of Selected Tropical Diseases ● 7 5

Life Cycle

All types of leishmanial parasites are transmitted by blood-sucking phlebotomine sandflies,

in which the parasitic organism undergoes part

of its complex life cycle (see fig. 4-10).

Leishmania invade and multiply within the

host’s macrophages, cells that are part of the immune system. Since macrophages are specialized

for ingestion and destruction of most foreign organisms, the ability of leishmanial organisms to live

in them is paradoxical. A key to controlling leishmaniasis may be to identify a mechanism that will

activate macrophages to kill Leishrnania and then

to attempt to activate this mechanism with a vaccine or drug.

Incidence

The various forms of leishmaniasis are distributed widely, along the U.S. Texas/Mexico border, in Latin America, in the Mediterranean, and

in Africa, India, and China (see fig. 4-11). Incidence rates range from a low in Guatemala of 1.2

per 100,000 in 1980, to a high of 60.5 per 100,000

in Costa Rica for the same year. A total of about

400,000 new cases were reported worldwide in

1977, 100,000 of which were in Bihar, a province

of India where a severe epidemic of kala azar was

occurring (341).

In some countries, the number of cases of leishmaniasis is increasing because of agricultural

colonization of jungle areas. Most forms of the

disease are transmitted to humans (via sandflies)

from animals native to the jungle where the disease occurs. This makes living or working in areas

in or near jungles a major health hazard. In the

late 1970s, cutaneous leishmaniasis seriously impeded a Bolivian scheme to relocate people outside the overcrowded altiplano. Many of the

colonists abandoned their land. More than 60 percent of the people who did said that leishmanial

disease was their reason for returning to the

mountains. Oil exploration and roadbuilding in

several Andean countries have also been significantly hampered (265).

A seroepidemiologic survey in Panama revealed

an apparent focus of leishmanial transmission,

without detecting the presence of clinical infec-
76 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 4.10.—GeneraIized Life Cycle of Leishmania, the Cause of Leishmaniasis

J

1. Parasites develop to “promastigote”

2. Parasites are transmitted to humans

stage infective to humans and other mammals in sandfly vector.

and other mammals through bite of sandfly.

3. Parasites transform to “amastigote” stage in mammalian host, multiplying within certain cells of the host’s immune

system.

4. Some amastigotes infect new cells in the mammalian host.

5. Some amastigotes are picked up by sandflies during bite of infected mammalian host.

SOURCE: Off Ice of Technology Assessment, 1985. Photo from H. W. Brown and F. A. Neva, Basic Clinical Parasitology (East Norwalk, CT: Appleton-Century-Crofts,

1983), used by permission.
Ch. 4—Description of Selected Tropical Diseases • 77

Figure 4-11 .—Geographic Distribution of Leishmaniasis Caused by Four Species of Leishmania

Key: ● Sporadic human cases High endemic to epidemi c

Low to moderate endemi c ---- Presumed limits endemic zones

SOURCE. U.N Development Program/World Bank/WHO, Special Program for Research and Training in Tropical Diseases, SIxth Programme Report 7 Ju/y 1981—31

December 1982 (Geneva: WHO, 1983).
78 . Status of Biomedica/lResearch and Related Technology for Tropical Diseases

tion. Completely subclinical leishmaniasis was

previously unknown and may be an important

clue to vaccine development.

Interventions

For disease caused by most species of Lekhmania, no effective prevention is known. However, immunity to Old World cutaneous leishmaniasis (also called Oriental sore) arising from L.

tropica infection can be induced by inoculation

of a susceptible person with organisms from an

active lesion, a procedure apparently known from

ancient times in endemic regions.

Specific treatment for leishmaniasis is now limited to antimony compounds. These compounds

are not always effective and often have adverse

toxic side effects. Another disadvantage of these

compounds is that they require daily injections

for 10 to 20 days, making them impractical for

patients living in remote and inaccessible areas.

Hospitalization for such a period is not only expensive but also a major inconvenience to patients

who cannot afford to leave work or their farms

for an extended period. For these reasons, improved treatment of the tens of thousands of existing cases is a priority research goal. The Pan

American Health Organization/WHO is attempting to foster development of new therapeutic

drugs. One, allopurinol, in combination with

other drugs, is a promising new treatment (393).

FILARIASIS

Aspects of Natural History

Filariasis is a collective term for several distinct

parasitic infections of tissue-dwelling, threadlike

nematodes, which are transmitted by mosquitoes

and other insects. There are at least eight different types of human filarial infections, among them

infection by the infamous guinea worm, which

before modern chemotherapy evolved was removed by gradually winding the protruding

worm around a stick. Probably the most impor-.

tant of the filarial parasites, in terms of worldwide prevalence and severity of disease, are WuRecent Progress

Fewer than 10 laboratories in the United States

study leishmanial organisms. The difficulty in distinguishing between Leishmania species (necessary

to properly predict clinical outcomes and select

treatment) has been a persistent problem. The solution, however, may be aided by biotechnology.

In recent years, MAbs have been prepared against

a variety of antigenic determinants in Leishmania

spp. (16) and used to probe morphologic and taxonomic differences. Rapid identification of Leishmania spp. may soon be possible with a recently

published technique of DNA hybridization (40s).

A variation of the enzyme-linked immunosorbent assay (ELISA), called the “DOT-ELISA”

method (267), is also a major advance, allowing

for rapid easy field diagnosis. These and other

methods would permit early treatment of the destructive mucocutaneous form of the disease and

would also facilitate precise epidemiologic field

studies.

The possibility of developing a vaccine against

the promastigote form of the leishmaniasis parasite (the stage transferred through the bite of the

sandfly) remains. Such a vaccine would have

limited usefulness because it would be ineffective

against the amastigote form of the parasite (the

stage which lives within the microphage). The

transformation from promastigote to amastigote

occurs rapidly within the host, before a vaccineinitiated antibody response could be effective.

chereria bancrofti and Brugia malayi, which cause

lymphatic forms of filariasis; and Onchocerca

volvulus, the agent of onchocerciasis (river

blindness).

Life Cycles and Interventions

The life cycles of W. bancrofti and B. malayi

are fairly similar (see fig. 4-12). These organisms

are transmitted to humans by several species of

mosquitoes, including common household pest

species. The adult filarial worms live in the human host’s lymphatic system and cause pathol-
Ch. 4—Description of Selected Tropical Diseases ● 7 9

Figure 4-12.-Generalized Life Cycle of Two Important Filarial Worms:

Wuchereria bancrofti and Brugk malayi

v &40 x 0.11 mm

3 83x 0.24 m m

1. Infective larvae develop, but do not multiply, in mosquito vector.

2. Larvae deposited on skin of human or other mammalian host (B. malayi only) through mosquito proboscis at time of bite.

3. Larvae penetrate skin, migrate to host’s lymphatic system, and mature.

4. Masses of adult worms can block lymph vessels, causing accumulation of lymph fluid and growth of lymph tissue, in a

manifestation called elephantiasis.

5. Adult worms release immature microfilariae into blood circulation, usually in a circadian rhythm.

6. Mosquitoes ingest microfilariae during bite of infected human or other mammal.

SOURCE: Office of Technology Assessment, 1985. Photo from F. W. Brown and F. A. Neva, Basic Clinical Parasitology (East Norwalk, CT: Appleton-Century-Crofts,

1983), used by permission.
80 . Status of Biomedical Research and Related Technology for Tropical Diseases

ogy that differs with the host’s immune response.

Inflammation and gross obstruction results in

varying degrees of swelling of the lymph glands,

which may result grotesque enlargement (elephantiasis) of the legs, breasts, or scrotum. Adult

worms release immature forms (microfilariae) that

circulate in the human blood and then infect feeding mosquitoes to complete the transmission cycle. Drugs are available to kill the microfilariae,

but the means to kill mature worms are poor.

0. volvulus parasites are transmitted to humans

by blackflies of the genus Simulium. These blackflies require running water to complete their life

cycles, limiting their habitat mainly to areas

around rivers (hence the name river blindness for

onchocerciasis). The adult parasitic worms live

in the tissues of the human body and often form

large nodules where an intertwined clump of

worms localizes. Microfilariae released by the

adult worms migrate through the human host’s

body in subcutaneous tissues where they can be

picked up by feeding blackflies. When microfilariae reach the human eye, blindness can result. The

prevention of blindness is imperfectly achieved

by chemotherapy and surgical removal of the

nodules. Few preventive measures are available,

and larvicides used to control the blackfly vector of onchocerciasis are subject to resistance and

have only transient effects.

Incidence and Prevalence

Filariasis due to W. bancrofti has a wide but

focal urban distribution throughout the Pacific reLEPROSY (HANSEN’S DISEASE)

Aspects of Natural History

Leprosy is a chronic bacterial infection that continues to be an important public health problem

in many countries. The disease is caused by

Mycobacterium leprae, a bacterium similar to the

one that causes tuberculosis. Leprosy is mainly

a disease of the skin and peripheral nerves, but

it is characterized by a wide array of clinical

presentations.

gion, Asia, Africa, and Latin America (see fig. 4-

13) (406). Lymphatic filariasis caused by B. malayi

is primarily found in rural foci in Sri Lanka, Thailand, Malaysia, Vietnam, China, South Korea, Borneo, and Indonesia. Onchocerciasis is found in

central and western Africa, North Yemen, Saudi

Arabia, Mexico, Venezuela, Colombia, Brazil,

Ecuador, and Central America.

It is estimated that bancroftian filariasis and

onchocerciasis are more prevalent today than they

were more than 100 years ago (256). More than

300 million people are exposed to mosquito-transmitted lymphatic filariasis, and more than 30 million are infected. The main endemic areas in India remain, and there is little control of the disease

in Africa, where in Savannah areas it is estimated

that more than 15 percent of the adults are infected.

Obstacles to Research

Considering the great number of people affected

or at risk, their widespread geographical distribution, and the severity of their pathology, the

filarial diseases are relatively neglected by American researchers. A major obstacle to research on

filariasis is the difficulty in maintaining filarial

organisms for laboratory study. Their complicated life cycles and the unavailability of suitable

animal models make these parasites among the

most frustrating to work with.

Much of the pathology of leprosy is associated

with a defective cell-mediated immune response

in certain individuals. Depending on the host’s immunologic response, leprosy ranges from benign

tuberculoid leprosy, with localized skin lesions

and nerve involvement (sometimes severe peripheral neuropathy) and the presence of few M.

leprae bacteria, to lepromatous leprosy, with

spreading lesions that become nodular and dis-
Ch. 4—Description of Selected Tropical Diseases ● 8 1

Figure 4.13. -Geographic Distribution of Major Filarial Diseases: Infection With Onchocerca volvulus,

Brugia malayi, and Wuchereda bancrofti

SOURCE V Zaman and L. Keong, Handbook of Medical Parasitology (Australia: ADIS Health Science Press, 1982), used by permission.
82 • Status of Biomedical Research and Related Technology for Tropical Diseases

figuring, resulting in destruction of the nose, involvement of the vocal cords and eyes, and often

severe nerve damage, with a heavy infection of

M. leprae.

Questions about why leprosy has such varied

effects on its victims have not been fully answered.

Researchers are not even sure how the disease is

transmitted. The latent period, the time between

infection and the actual appearance of symptoms,

often lasts for many years. The individual is capable of unknowingly infecting others during this

time. Leprosy is more likely to be spread by

chronic exposure to dried skin lesion matter and

nasal secretions from the lepromatous and more

severe borderline patients. The general consensus seems to be that M. leprae bacteria enter the

human body through the respiratory system, although some researchers suspect entry may also

be through the skin (406).

Immunologic diagnostic techniques have shown

that people with tuberculoid leprosy have a strong

and effective cell-mediated immune response (via

lymphocytes) that controls the infection, whereas

lepromatous leprosy patients do not. There is

some evidence to suggest a genetic basis for this

difference. The nerve damage in patients with

lepromatous leprosy seems to result from an absence of cellular immune response. Characterization of this defect in Iepromatous leprosy patients

is being investigated with MAbs that can identify lymphocyte subsets and also by analysis of

the patients’ genetic type.

Incidence and Prevalence

Worldwide there are an estimated 15 million

leprosy cases (307). The prevalence of leprosy has

been reduced in many places, but the overall incidence (i.e., the number of new cases per year)

has not changed with advances in science (see fig.

4-14). In some very small, isolated communities

in parts of Africa and Australia, the prevalence

of leprosy may be as high as 1 out of every 50

inhabitants. The disease is also common in southern Asia, especially India (210). In China, an estimated 500,000 cases of leprosy occurred in the

early 1950s, but fewer than 200,000 cases were

reported in 1984 (307). As of 1984, there were 2.5

million cases in Southeast Asia alone (135).

In the Americas, there are about 400,000 cases

of leprosy, 80 percent of the new cases occurring

in Argentina, Colombia, and the Amazon area

of Brazil (265). The recorded incidence of leprosy

in the Americas has almost doubled in the last 10

years, but the increase is thought to reflect improvements in case finding and notification of authorities, rather than to be a true change. Over

half the clinical cases are of the more severe

lepromatous form. However, as many as fourfifths of those infected with M. leprae never get

sick, though they may transmit the disease to

those with more susceptible immune systems

(210).

About 20 cases of leprosy acquired in the

United States are diagnosed each year, occurring

in Texas, Louisiana, California, Florida, and Hawaii. Of the 250 new cases of leprosy diagnosed

in the United States in 1982, 233 were immigrants
Ch. 4—Description of Selected Tropical Diseases ● 8 3

Figure 4-14.—Geographic Distribution of Leprosy

Key:

Incidence rate per

1,000 population

SOURCE: L. M. Bechelli and V. Martinez Dominguez, “The Leprosy Problem in the World, ” Bull. W.H.O. 34:811-826, 1966

who had contracted the disease in their homelands. In the past, most cases have come from

Mexico and the Philippines, but in recent years

immigrants from Cuba, Haiti, El Salvador, Nicaragua, and Southeast Asia have entered the

United States with leprosy (135).

Why leprosy occurs more in some parts of the

world than in others is not fully understood.

Interventions

A number of useful drugs are now available for

leprosy treatment. The most effective drug is dapsone. Unfortunately, however, strains of M. leprae

have developed resistance to dapsone (217). This

situation has resulted in new recommendations

for combination chemotherapy for leprosy that

will shorten the treatment period and increase the

likelihood of effective control of the disease. If

organized and administered well, combination

chemotherapy will lighten the workload of health

services, improve patient compliance, and result

in better prognosis.

Vaccination against leprosy is in the human trial

stage, but still years from proof of its effectiveness and routine use. Research on the epidemiology of leprosy is still needed to improve intervention strategies.

Recent Progress

A small number of laboratories in the United

States, probably fewer than 10 (including the U.S.

Public Health Service in Carville, LA) specialize

in leprosy research. Past leprosy research efforts

were hampered by the inability of researchers to

grow M. leprae in culture and also by the lack

of a suitable animal model. The introduction of

the armadillo model of infection in the early 1970s

has helped both in the characterization of the disease and in the development of serodiagnostic procedures and a potential vaccine (307). The fairly

recent discovery of the susceptibility of mangabey

monkeys (217) may also help research efforts. The

problems at this point are the slow growth rate

of M. leprae in culture and the inability of arma-
84 ● Status of Biomedical Research and Related Technology for Tropical Diseases

dillos to breed in captivity, necessitating their continual trapping from the wild.

Despite the paucity of workers, the tools of biotechnology are being applied to M. Ieprae. At least

TUBERCULOSIS

Aspects of Natural History

Tuberculosis is caused by the bacterium Mycobacterium tuberculosis, transmitted mainly by airborne droplets from person to person. There is

no insect vector or animal reservoir. Tuberculosis

remains a major threat to health in many parts

of the world, causing several million deaths annually. Though basically a chronic respiratory disease, tuberculosis can spread into the cardiovascular, endocrine, and genital systems, and into

the lymph nodes, bone, brain, kidneys, and other

organs (406).

Incidence

A 1982 worldwide estimate put the number of

tuberculosis cases at 11 million (265), with some

3.5 million new cases occurring each year (92).

Tuberculosis is still a serious problem in most

countries of the world. About 500,000 deaths are

attributed to the disease each year.

Even in countries such as Canada and the United

States, with highly developed coverage for diagnosis and treatment of tuberculosis available, a

significant number of cases of tuberculosis are encountered each year. Often the disease is present

among immigrants arriving from tuberculosisprevalent tropical areas. From 1962 to 1975, prior

to the current wave of immigrants to the United

States from Indochina, the tuberculosis case rate

among children under 14 in the United States decreased from 10.4 to 3.7 per 100,000, representing a 9-percent decline per year. But this downward trend has not continued, and the rate in 1984

was about the same as that of 1975 (281). In 1980,

some 30,000 new cases of pulmonary tuberculosis

were reported in the United States (70).

three investigators are preparing MAbs as part of

studies on the immunochemistry or antigenic

structure of the bacterium (16).

Interventions

In the past, control of tuberculosis was based

on the identification, isolation, and treatment of

patients with active disease, since these patients

are the source of continued transmission and

spread. More recently, with the advent of effective chemotherapy and reduction of active cases,

developed countries with lower prevalence of

tuberculosis have emphasized identification of

newly infected persons via the tuberculin skin test

and treatment of these individuals, in addition to

identification and treatment of persons with active

disease. Developing countries continue to emphasize identifying and treating patients with active

disease, who account for most of the transmission. Tuberculosis is generally diagnosed by sputum microscopy or culture.

Treatment of tuberculosis patients consists of

daily administration of one or more drugs in combination for 6 to 12 months. The drugs most commonly used include isoniazid (INH), streptomycin, Para-aminosaliqlic acid (PAS), pyrazi.namide,

and rifampicin. If the course of treatment is followed properly, cure rates can be as high as 100

percent (406). Because of the practical problems

of long-term treatment, however, a 100-percent

cure rate is seldom realized.

Unfortunately, some strains of M. tuberculosis

have developed resistance to INH, the most effective drug available. Cases of INH-resistant

tuberculosis are commonly reported in tropical

areas from Asia to Latin America. Patients infected with resistant strains must take several

chemotherapeutic drugs, and for a longer time

period, before the infection is arrested.
Ch. 4—Description of Selected Tropical Diseases ● 8 5

A factor strongly contributing to the resistance

problem is the high cost of rifampicin. This drug,

when used with INH, is highly effective in controlling tuberculosis, but it costs about 400 times

as much as INH (see ch. 9). Developing countries

that cannot afford to use rifampicin in their treatment regimes often opt for other less expensive

drugs. The short-term savings, are lost, however,

because patients often do not complete long-term

regimens. Partial treatment that does not eliminate the infection encourages the proliferation of

drug-resistant organisms, as the most susceptible

are kiIIed off even with an incomplete course of

therapy.

Despite the availability of a vaccine for tuberculosis, Bacillus Calmette-Guerin (BCG), there is

considerable controversy as to its effectiveness.

BCG vaccine is derived from a live, attenuated

bovine tubercle bacillus, isolated from a single

strain by the Pasteur Institute 50 years ago. BCG

vaccination of uninfected persons can produce

high resistance to tubercle bacilli, but the protection against tuberculosis has varied greatly in field

trials. Since the vaccine was made years ago

against only one isolated strain, it may not be capable of immunizing individuals against all currently active strains of M. tuberculosis. Nevertheless, some trials have shown very effective

protection, and BCG vaccine is still recommended

in high-risk areas.

Recent Progress

The tuberculin skin test is not an adequate

predicter of infection or active tuberculosis, particularly for research purposes. The use of antigen probes and ELISA methods may be helpful

in more accurately characterizing the state of an

infection (16).

DIARRHEAL AND ENTERIC DISEASES

Aspects of Natural History

Diarrheal diseases, all of which are transmitted

by fecal contamination of food and water, constitute a clinical syndrome of varied etiology. Such

diseases are caused by a variety of viruses (primarily rotaviruses), bacteria (Shigella, Sahnonella, Cryptosporidium, Escherichia coli, Campylobacter, and Yersinia), protozoa (Entamoeba

and Giardia), and worms (Ascaris, Ancylostoma,

and Necator), interacting in a complex fashion

within the susceptible host. Diarrheal diseases are

distributed worldwide.

Diarrhea is primarily a disease of infants and

children. The great danger in diarrheal disease is

the dehydration and subsequent shock caused by

tremendous losses of fluids and electrolytes (salts).

Severe dehydration is the most frequent cause of

diarrheal deaths. Although treatment directed at

the disease-causing organisms may or may not be

effective, there now exists effective treatment for

most cases of dehydration caused by diarrhea.

Oral dehydration therapy (ORT) for diarrheal diseases is considered one of the most significant therapeutic advance in the past several decades (see

Case Study A: Oral Dehydration Therapy for

Diarrheal Diseases).

In 1980, WHO conservatively estimated that

among children under the age of 5, 750 million

to 1 billion diarrheal episodes occur yearly in

Africa, Asia (excluding the People’s Republic of

China), and Latin America (324). Data from several developing countries indicate that children

under 5 years of age in these countries typically

experience four to eight diarrheal episodes annually (23,215,216). In contrast, infants in the United

States and other developed countries experience

one or two diarrheaI episodes yearly. In some

countries, up to 45 percent of all hospital visits

during the months of highest diarrhea prevalence

are due to childhood diarrhea, and case fatality

rates as high as 40 percent have been recorded (249).

A comparison of death rates between children

under 1 year of age in Latin America and North

America is startling. For the United States and

Canada, the mortality rate due to diarrheal disease among infants in 1979 was 21.9 deaths per
86 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

100,000 infants; for Latin America, it was 914.6

per 100,000. The Latin American figure is 40 times

higher, which means almost 1 in 100 infants born

there dies of diarrheal dehydration. Wherever the

infant mortality rate exceeds 100 per 1,000 births,

at least one-third of the deaths can be attributed

to diarrhea (216,257).

Viruses

The complex of diarrheal diseases caused by

viruses-rotaviruses, Norwalk-like agents, adenoviruses, astroviruses, enteroviruses, coronaviruses, calciviruses, and others, perhaps not yet

identified—present extreme difficulties in diagnosis (15). Viral agents cause a significant amount

of diarrheal illnesses in the tropics, but very little

is known about any of them except rotaviruses.

Rotaviruses, the most frequently isolated group

of viruses, have a worldwide distribution. They

were first detected in humans in 1973. Rotaviruses, which are believed to cause one-third of

all diarrheal disease in the world, may cause up

to 40 percent of diarrheal disease in children in

developed countries. Serologic studies have shown

that by the age of 2, nearly all children have been

infected with rotaviruses.

Photo credit: S. S. Raphael, “Lynch’s Medical Laboratory Technology, ”

W. B. Saunders Co., 1983. Reprinted by permission.

Electron micrograph of a rotavirus, a major cause of

diarrheal disease in children.

In Australia, Canada, the United Kingdom, and

the United States, a large percentage (40 to 60 percent) of all children hospitalized with diarrhea are

infected with rotaviruses; the percentage in countries like Guatemala and El Salvador is smaller

(20 to 40 percent) (371,420). However, since the

incidence of all diarrhea] disease is much higher

in developing countries than in developed countries, the toll due to rotaviral infection in these

developing countries is actually much greater than

the percentages indicate.

Growing recognition of the importance of rotaviruses emphasizes the need for further epidemiologic, clinical, and basic research. Rotavirus biology currently is studied at the National Institutes

of Health, where each of the rotavirus genes has

been cloned into bacteria and identified (117). Immunologically based diagnostic testing can be used

for field studies of prevalence and incidence. Since

a rotavirus vaccine exists for animals, a major objective now is to develop one for humans.

Bacteria

Numerous bacteria are known to cause diarrheal illness, and the list continues to grow as laboratory identification methods improve. In many

areas of developing countries, the facilities needed

to identify the agents responsible for diarrhea are

not available. The question of causality is complicated by the wide variety of bacteria which live

harmlessly in the intestine. This large “intestinal

flora” confounds efforts to isolate and identify the

important pathogens, among them enterotoxigenic

(toxin-producing) E. coli, Shigella spp. (which

cause dysentery), Salmonella spp. (which cause

food poisoning and typhoid fever), Vibrio

cholerae (which causes cholera), and Campylobacter.

At present, there is a great deal of concern

about the apparent rise in antibiotic-resistant bacteria. Two separate factors may be responsible for

the increase. One factor is the widespread misuse of antibiotics both in the United States and

abroad, due to ineffective dosing of individuals,

indiscriminate prescribing of drugs (regardless of

the etiologic agent), and the availability of antibiotic drugs over-the-counter in many developing countries. As a result of this misuse, many
Ch. 4—Description of Selected Tropical Diseases ● 8 7

intestinal bacteria, both pathogenic and nonpathogenic, are being exposed to antibiotics. Most

intestinal bacteria are killed by antibiotics, but

those bacteria that have some sort of resistance

mechanism, often genetically determined, survive

and are passed on.

The second factor is the ability of many intestinal bacteria to exchange genetic material among

different strains, species, and even genera through

the transfer of DNA. Bacteria that survive antibiotic exposure may transfer the genes that code for

their resistance to previously susceptible bacteria, and these bacteria in turn incorporate them

into their genetic material and pass them on to

both pathogenic and nonpathogenic bacteria.

Thus, in the past, a cholera infection might have

been successfully treated with a few highly effective doses of tetracycline, but now larger doses

of two or more other antibiotics may be required

to combat the disease. Strains of E. coli, Shigella,

and Vibrio cholerae have all been shown to be

antibiotic resistant.

Escherichia coli. —E. coli is second to rotaviruses as an important cause of diarrheal disease.

For the American traveling abroad (or the foreign

visitor touring the United States), E. coli is frequently the source of the infamous travelers’

diarrhea.

Ironically, although E. coli is one of the most

intensely studied of all organisms in the laboratory, little is known about its disease-causing abilities. Most strains are harmless inhabitants of the

intestine, and only certain strains cause disease:

enterotoxigenic E. coli produces toxins that result

in excessive fluid production; enteroinvasive E.

coli invades the cells of the intestinal wall; and

enteropathogenic E. coli produces a toxin similar to that of Shigella, which causes diarrhea in

infants. Investigators are currently studying the

genetic basis for disease-causing properties of different E. coli strains.

Shigella. —In 1981, there were 19,859 cases of

diarrhea due to Shigella reported in the United

States (366). Infections by Shigella dysertteriae result in bacillary dysentery, a serious and sometimes fatal disease. Most Shigella infections, however, are by species other than S. dysenteriae (e.g.,

S. sonnei and S. flexneri), and the symptoms are

not as severe. Shigella is difficult to study because

it colonizes only primates; thus, research on its

pathology is very expensive. The genes coding for

the attachment factors (which allow the organism

to adhere to the intestinal wall) have been cloned

and inserted into E. coli.

Salmonella. —This genus is frequently divided

into Salmonella typhi, the cause of typhoid fever, and the nontyphi species. In general, the nontyphi species of Salmonella cause self-limited

gastrointestinal distress, although some species

may cause bacteremia bacteria in the blood. The

wide variety of symptoms associated with Salmonella infection, ranging from mild flu-like

stomach upsets to severe food poisoning to typhoid fever, make the collection of accurate incidence and prevalence statistics almost impossible.

In 1981, there were 39,990 cases of salmonellosis

food poisoning reported in the United States (367).

Salmonellosis is usually associated with the consumption of contaminated livestock or poultry

products. In that year, there were approximately

half of the 510 cases of typhoid fever reported in

the United States, about half of which were acquired during travel abroad. During the period

1970-80 while the incidence of typhoid fever in

Latin America increased, the number of cases remained stable in North America and the Caribbean region (265).

Sahnonella spp. usually cause endemic diarrhea, but they may cause epidemics. The need for

improved sanitation and new antibiotics was

brought forcefully home during a nontyphoid epidemic in Mexico in 1972-73, when it was discovered that the strain causing the epidemic was resistant to chloramphenicol, the drug of choice, and

to other antibiotics (265).

Recently, researchers at the Walter Reed Army

Medical Research and Development Command

cloned Shigella genes into an attenuated live typhoid vaccine developed by a group of researchers in Switzerland. The result was a vaccine that

produced immunity to both Shigella and Salmonella (126,423).

Vibrio cholerae. -Cholera occurs in both endemic (in parts of China and India) and epidemic

forms (in Asia and Africa). Because of improved
88 ● Status of Biomedica/ Research and Related Technology for Tropical Diseases

sanitation in many countries, cholera epidemics

are not as common as in the past, although one

reached the U.S.S.R. as recently as 1970 (406).

For unknown reasons, cholera epidemics have

spared the Western Hemisphere.

The severe diarrhea characteristic of cholera is

caused by the action of a bacterial toxin on the

gut wall. One of the factors that makes cholera

so virulent is the number and variety of transmissible genetic elements, which vary from one strain

to the next. Researchers have been collecting cholera strains from endemic areas around the world,

and recombinant DNA libraries, consisting of segments of genes from wild-type organisms and

laboratory-grown strains, have been created. By

studying and comparing the gene segments of toxigenic and nontoxigenic strains, scientists hope to

pinpoint exactly which genes are responsible for

cholera’s virulence (16).

Campylobacter. —Various species of Campylobacter are commonly the cause of sporadic

diarrhea in developing and developed countries

(45). Following rotavirus and enterotoxigenic E’.

coli, Campylobacter is the third most common

cause of diarrhea in developing countries (297).

Campylobacter also is a “frequent, cosmopolitan

risk to travelers” (278). Epidemics sometimes affecting thousands of people, have been caused by

Campylobacter contaminating unpasteurized

milk, chicken carcasses, and water (194).

Protozoa and Other Agents

There are several protozoal diarrheal pathogens, among them Entamoeba spp., Giardia Jamblia, and Cryptosporidium spp.

Giardiasis is now recognized as significant and

ubiquitous throughout the United States, but is

more prevalent in tropical countries. Outbreaks,

such as occurred in Aspen, CO, and Rome, NY,

have been well publicized (313). G. lamblia is a

well-known hazard to camping and backpacking

enthusiasts, who are becoming aware that wilderness water may not be as pristine as it looks. Many

other less publicized outbreaks have occurred in preschool day care centers (400).

Cryptosporidium spp. are common diarrheacausing agents in individuals whose immune systems are compromised, and they have also been

found in some otherwise healthy individuals.

Cryptosporidium was first described in 1907, but

its oocysts (spherical egglike cells) were not recognized in animal feces until 1978. The importance of Cryptosporidium spp. has been appreciated only recently. Investigators using a

simple diagnostic procedure developed in the last

few years have estimated that the organism accounts for 1 to 4 percent of all cases of diarrhea

in human beings (400).

Not much is known about chronic low-level enteric infections by bacteria such as Yersinia (the

agent of plague). Multiple infections by several

different pathogens frequently occur, making

diagnosis difficult. One study of people living in

a poor rural area of Panama with substandard

sanitation showed that 90 percent of the 202 people examined were infected by one or more parasites, the majority of which were either Ascaris

lumbricoides (roundworm), E. histolytica, or G.

lamblia (88). WHO estimates there are at least 650

million people in the world with roundworm (ascariasis), 450 million people with hookworm (ancylostomiasis), 350 million people with amebiasis,

and 350 million people with whipworm (trichuriasis) infections (318).

ACUTE RESPIRATORY INFECTIONS (ARIs)

Aspects of Natural History the elderly, sometimes exceeding the mortality

rate due to diarrheal diseases. The ultimate conARIs are among the most important causes of sequences of an ARI depend on the organism(s)

preventable deaths in the world. They are a ma-, responsible for the infection and the patient’s nujor cause of mortality among children under 5 and tritional status and age.
Ch. 4—Description of Selected Tropical Diseases . 89

All of the ARIs are aggravated by malnutrition

and substandard living conditions, and the presence of other infectious diseases. A principal epidemiologic factor of ARI transmission is close,

overcrowded conditions that promote inhalation

of pathogens aerosolized by coughs, sneezes, and

personal contact.

ARI agents include viral, bacterial, chlamydial,

and mycoplasmal organisms, all transmitted to

humans by airborne droplets. For medical purposes, ARIs are usually characterized as upper or

lower respiratory tract infections (URTIS or

LRTIs); then as community- or hospital-acquired;

and then grouped by etiologic agent. Many of the

same organisms cause infections in both the upper and lower respiratory tracts, however.

Incidence and Prevalence

Data about the frequency of ARIs are not generally available, making a discussion of incidence

and prevalence of this important group of diseases

difficult. Surveys in India, Guatemala, the United

States, and the United Kingdom suggest that the

incidence of ARIs is similar throughout the developing and developed countries, averaging four

to eight separate episodes per year (175). However, mortality rates for ARIs in India are thought

to be 30 to 75 times higher than those in the United

Kingdom or the United States (175). What might

cause inconvenient days out of work or school

in a developed country might well kill a person

in a developing country.

Some 12 percent of all deaths of children living in Africa, Central America, and Asia are attributed to ARI (289), and most of these children

die of either pneumonia, bronchiolitis, or acute

obstructive laryngitis (croup) (370). Among infants, mortality from ARI can range as high as

1,500 per 100,000 in areas of Egypt, Paraguay,

and Mexico—a figure 30 times higher than in the

United States and Canada (57)—or even higher

to the 4,000 to 4,400 per 100,000 observed in areas

of Bolivia and Brazil (370). The incidence of pneumonia, 70 to 100 cases per 1,000 per year, in children under 5 in developing countries, is double

that of the United States. Among children who

are suffering from malnutrition, almost half will

also contract pneumonia during any given year

(370).

WHO data, collected from 88 countries on five

continents, representing a quarter of the world’s

population, reported over 660,000 deaths from

ARIs in 1978 (35). Extrapolating this figure to the

world population suggests that there are more

than 2.2 million deaths from ARIs per year, a significant number of which could have been prevented. In addition to presenting a serious mortality risk for the very young and the very old,

ARIs impose a tremendous social burden in terms

of lost productivity and demands on the health

care system by all age groups.

Viruses

Viruses cause most URTIS and some important

infections of the lower respiratory tract. Most viral infections of the upper respiratory tract—infections with respiratory syncytial virus (RSV),

adenoviruses, rhinoviruses, coronaviruses, and influenza, parainfluenza, measles, and Epstein-Barr

viruses, for example—are self-limiting, eliminated

by a healthy immune system. In weakened individuals, particularly children, who may be malnourished and have other infections, however, even

normally benign URTIS can be life-threatening.

Measles is one of the few URTIS against which

a highly effective vaccine exists. The vaccine’s use

is not universal, however, and measles remains

a major cause of death among children in the

developing world. Worldwide, 900,000 people,

mainly children, died from measles in 1979 (392).

The high death rate may be due to concurrent infections, or overcrowded living conditions, which

result in heavy exposure of the susceptible individual to the virus whenever several members of

the same household are infected (118).

In general, LRTIs cause more serious health

problems than do URTIS, though they are caused

by many of the same viruses. RSV, measles, and

influenza are major causes of pneumonias, which

often lead to death in developing countries.

Bacteria

Bacteria cause a host of infections in and around

the upper respiratory tract including the sinuses,

throat, tonsils, epiglottis, larynx, and trachea.

Four types of bacteria are particularly significant
90 ● Status of Biomedical Research and Related Technology for Tropical Diseases

causes of URTIS. Corynebacterium diphtheria,

is the cause of diphtheria, an infection of the pharynx. A second, Bordetella pertussis, is the cause

of pertussis (whooping cough). Effective vaccines

to prevent diphtheria and whooping cough are

routinely given to newborns in developed countries and are included in WHO’s Expanded Program on Immunization. Immunization is much

more effective than treatment for these two

diseases.

A third bacterium, Streptococcus pyogenes,

commonly infects the pharynx, but is also the

organism responsible for rheumatic fever. The

fourth is Hemophilus influenzae, which causes ear

infections and the more serious condition of meningitis in children.

Bacterial pneumonias (LRTIs) area major cause

of morbidity and mortality in developing countries, as they are in developed countries. Streptococcus pneumoniae is one of the most common

causes of bacterial pneumonia affecting all ages.

Other causes of bacterial pneumonia are S. pyogenes, Staphylococcus aureus, H. influenza,

Klebsiella pnemonia, E. coli, and Pseudomonas

pseudomallei.

Most bacterial ARIs respond to treatment with

antibiotics. Important exceptions are many hospital-acquired pneumonias, which are often caused

by drug-resistant organisms, a problem in developed as well as developing countries.

Other ARI Agents

Other less known ARI agents include Chlamydia spp. (congenital) and Mycoplasma spp.,

usually responsible for URTIS and atypical pneumonia among adolescents and adults. Chlamydia

spp. appear similar to bacteria, but their size is

close to that of viruses. Mycoplasma spp., a group

of organisms which lack a rigid cell wall, are considered by some to be primitive bacteria.

Interventions

For most viral ARIs, there is no treatment other

than symptomatic and supportive relief, though

bacterial infections can be effectively treated with

antibiotics. Vaccinations for measles, whooping

cough, and diphtheria are effective and are promoted for vaccinating children under the Expanded Program on Immunization of WHO.

However, use of the vaccines for these diseases

is frequently limited by the cost of vaccines, the

ability of the health infrastructure to maintain a

vaccination program, and the problems inherent

in maintaining a “cold chain” for certain vaccines.

(Cold chain is the name given to the means for

continuous refrigeration of vaccines from production to vaccinee. )

Measles is an example of an ARI which could,

like smallpox, be eradicated by vaccination. Prior

to the development of the measles vaccine in 1962,

481,530 cases were reported in the United States,

46 resulting in deaths. In 1979, after several years

of widespread use of the vaccine, there were only

13,597 cases and only 1 death (366). In many developing countries, however, vaccinations against

measles have been infrequent. Among children

under 1 year of age in 1982, only 8.3 percent in

Mexico, 15.9 percent in Bolivia, and 22.4 percent

in Colombia (265) were immunized. Measles vaccine must be administered after maternal antibodies lose their effectiveness, but prior to the child’s

first exposure to a virus, in order for it to work—a

critically short time span.

Vaccines against pneumococcal pneumonia

(due to S. pneumonia) and influenzas are available, but their usefulness in developing countries,

is limited because pneumococcal vaccine is not

very effective in children under 2 years old and

influenza vaccines must be renewed periodically

according to the currently prevalent strain. Because of the high cost of annually manufacturing

a different strain of influenza vaccines, influenza

vaccines are generally available only to high-risk

groups.

Improved living conditions and access to health

care are critical factors in controlling ARI, but

field-based epidemiologic studies are also needed.

ARI control is largely ignored in most developing countries. This situation is a function of several factors: difficulty in identifying the etiologic

agents of ARIs, lack of effective treatment for

many ARIs, and failure to define ARIs as tropical diseases, or to recognize ARIs as worthy of

focused research.
Ch. 4—Description of Selected Tropical Diseases Ž 91

The prevention and treatment of ARIs could

be simplified through studies precisely identifying the important etiologic agents in different geographic areas and determining the risk factors that

make ARI mortality so high. Practical management of ARI depends on differential diagnosis of

viral from other bacterial, chlamydial, and mycoplasmal agents for which specific treatments are

effective.

Biotechnology and ARIs

The tools of biotechnology are slowly being applied to the many viruses that cause ARIs in humans, but bacterial agents are being largely

ignored.

ARBOVIRAL AND RELATED VIRAL

Aspects of Natural History

Arboviral infections constitute a large group of

diseases caused by viruses (currently about 80

known in humans) defined by ecologic, epidemiologic, and clinical, rather than taxonomic, characteristics. The term “arbovirus” is a contraction

of “arthropod-borne virus. ” Strictly speaking,

arboviruses replicate in and are transmitted by

arthropods (predominantly mosquitoes, but also

ticks, sandflies, midges, and gnats) (a generalized

arbovirus life cycle is shown in fig. 4-15). However, there are some arbovirus-like diseases whose

vector is still unidentified (e.g., those caused by

the Arenaviridae family) and some whose early

epidemiologic profile incorrectly suggested arthropod transmission (e.g., Argentinean and Bolivian

hemorrhagic fevers). These exceptions are noted,

but for lack of a better characterization system,

they are discussed here.

Arboviruses are widely distributed throughout

all areas of the world and cause significant endemic and epidemic disease. Most arboviral diseases are infections of animals accidentally transmitted to humans (zoonoses), although epidemics

of human-to-human transmission, via insect vectors, can occur.

The number of known arboviruses has grown

rapidly as the etiologic agents of many fevers and

ARI viruses are being isolated, identified, and

characterized. A few laboratories are working on

the molecular biology of influenza are studying

mechanisms of immunity (76) as well as the effectiveness of immunization (271). All of the influenza genes have been identified and sequenced.

Recently, some of the influenza genes have been

cloned into the vaccinia virus used to inoculate

against smallpox. Hamsters were inoculated with

the vaccine and immunity was produced against

both smallpox and influenza (319). There is hope

that all the genes from the various types of influenza can eventually be cloned into the vaccinia

virus to produce a single vaccine effective against

every strain.

INFECTIONS

brain inflammations (encephalitis) have been identified and their transmission cycles elucidated. The

major groups of arboviruses are classified by their

biochemical and physical properties into the

Togaviridae, Bunyaviridae, and Arenaviridae

families.

The Togaviridae family is divided into two

groups. There are about two dozen “alphaviruses”

(formerly known as Group A), including the agents

of Eastern, Western, and Venezuelan equine encephalitis frequently found in North and South

America, Chikungunya of Africa and the Far East,

and Sindbis and Semliki Forest viruses of Africa;

and three dozen or so “flaviviruses” (formerly

Group B), including the agents of yellow fever,

dengue fever, Japanese B encephalitis and Saint

Louis encephalitis (found in the United States).

About a third of the flaviviruses are tick-borne

and cause various febrile and hemorrhagic illnesses.

The Bunyaviridae family includes several hundred distinct viruses, among them the agents of

California encephalitis, Oropouche fever of Brazil, Crimean hemorrhagic fever, and Rift Valley

fever of east Africa.

The Arenaviridae are not arboviruses, although

the possibility remains that arthropod vectors will

someday be identified for the group. Included here
92 . Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 4-15.—Generalized Transmission Cycle of an Arbovirus

1

Rodents, bats,

birds, humans

.

SUMMER/RAINY SEASON:

EPIDEMIC CYCLE

 ‘ \

Introduction of virulent viral subtypes by:

, \

? selection or mutation from endemic cycle

in humans and/or animals t \

? bird or bat migrations J

? movement of viremic equines

? improperly inactivated vaccines

DRY SEASON:

LITTLE OR NO

DISEASE ACTIVITY

known part of cycle

--- speculative part of cycl e

SOURCE: Adapted from G. T. Strickland, Hunter’s Tropical Medicine (Philadelphia: W, B. Saunders Co., 1984), used by permission

are some of the most deadly infectious agents

known, such as the Lassa fever agent, as well as

the more common lymphocytic choriomeningitis

and various South American viruses, such as Junin (Argentine hemorrhagic fever) and Machupo

(Bolivian hemorrhagic fever). Lassa and Machupo

fevers are carried by rodents, but the rodent-tohuman connection has not been made. Work with

the more dangerous arenaviruses is restricted to

the very few laboratories in the world with adequate containment facilities, among them the Fort

Detrick facility of the U.S. Army in Frederick, MD.

Four basic types of clinical conditions are

caused by arboviral infections. Two are generally

benign and self-limited: 1) fevers of short duration, with or without a rash; and 2) painful joints

and rashes of a short duration. Complications can

develop from either of these two conditions, but

they are the exception. The two much more serious clinical syndromes caused by arboviral infections are: 1) acute central nervous system

disease usually with inflammation of the brain

(encephalitis), ranging in severity from mild aseptic meningitis to coma, paralysis, and death; and

2) hemorrhagic fevers, with extensive hemorrhaging, associated with shock and high case fatality

rates (liver damage and jaundice accompany these

symptoms in yellow fever).

Yellow fever was the first arboviral disease of

the tropics to be recognized, by Walter Reed, as

a mosquito-borne disease. William Gorgas led the

campaign to eliminate the disease from the Panama

Canal Zone and from Cuba in the early 1900s.

Further success occurred throughout Latin America. Although the virus is maintained in monkeys

in the wild, yellow fever has not occurred in urban areas of Latin America since the 1920s.

Symptoms of yellow fever include rapid onset,

high fever (l03oF), headache, nausea, vomiting,

and muscle pain. The disease in a population occurs in periodic cycles stretching over several

years. The cycles depend on the buildup of nonimmune individuals in a population, who are then
Ch. 4—Description of Selected Tropical Diseases ● 9 3

swept by an epidemic of the virus, leaving an immune population of survivors. Safe and effective

vaccination of human populations near endemic

jungle areas is one control strategy and provides

immunity for at least 10 years. Surveillance of

monkey populations and jungle mosquitoes by

sampling for virus isolation is an important monitor, providing early warning of high levels of infection.

Yellow fever still remains a major threat in tropical America and Africa (see fig. 4-16), because

the virus is maintained by transmission through

a number of jungle mosquitoes, with monkeys

and possibly certain marsupials serving as reservoirs. Recent research has demonstrated that passage of the virus from the female mosquito to the

egg (transovarial transmission) occurs among the

vectors of yellow fever. Thus, the mosquito may

function not only as a vector, but also as a reservoir (100).

Cases of yellow fever in humans are associated

with humans invading the jungle habitat. In recent years, however, an outbreak appeared in CoPhoto credit: K.S. Warren and A.A.F. Mahmoud “Tropical and Geographical

Medicine, ” McGraw-Hill Book Co., 1984. Reprinted by permission.

Electron micrograph of the dengue virus, agent

of dengue fever.

lombia where there were no apparent known vectors or reservoirs, and in Trinidad where no cases

had been detected for almost 20 years. The possibility of unknown reservoirs is of concern, as

Aedes aegypti, the vector of yellow fever in the

urban setting, remains abundant throughout the

Figure 4-16.—Geographic Distribution of Yellow Fever

SOURCE: U.S. Department of Health and Human Services, Centers for Disease Control, Health Informat/in for International Travel, HHS Publication No.

(CDC)840-8280 (Washington, DC: U.S. Government Printing Office, 1984).
94 . Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 4-17.—Geographic Distribution of Dengue Fever

b m - —

SOURCE: World Health Organization, Geneva, Switzerland.

world (including the United States), posing a persistent threat of epidemic outbreaks in large population centers.

Dengue fever is a disease of worldwide distribution (see fig. 4-17) caused by four serotypes of

the dengue flavivirus. The disease is usually mild,

characterized by a rash resembling measles or

scarlet fever, accompanied by generalized swollen

lymph glands. Convalescence is long and distressful (406). Serious, sometimes fatal complications

of dengue fever are dengue hemorrhagic fever and

dengue shock syndrome. Dengue hemorrhagic fever usually affects children. In recent years, large

epidemics of this virus have swept the Caribbean

and Central America. In 1981, the first indigenous

cases in the United States since the 1940s occurred.

The virus is endemic to the Caribbean and is transmitted by mosquitoes of the genus Aedes, including the common urban mosquito A. aegypti,

which is found as far north as St. Louis, MO.

Oropouche fever, found in Trinidad and Brazil, is e-merging as an important arboviral disease

because of its debilitating symptoms, which include anorexia, rash, and joint and muscle pain.

The virus is transmitted by biting midges (Culicoides spp. ) in urban and suburban areas. There

is probably also a transmission cycle in forest animals away from populated areas, as in the case

of yellow fever.

Rift Valley fever was first characterized in the

Rift Valley of Kenya in 1931. Sandflies and mosquitoes are the suspected vectors, and while they

have been shown capable in the laboratory, transmission has not been established in the field. Until 1977, Rift Valley fever was geographically limited to sub-Saharan Africa. Human fatalities were

not known until the 1975 epizootic (epidemic

among animals) in South Africa (KM). Currently,

the disease is present throughout Africa, but has

not yet spread to other continents. In 1977-78, a
Ch. 4—Description of Selected Tropical Diseases ● 9 5

widespread epizootic and epidemic of Rift Valley fever occurred in Egypt. Some 18,000 human

cases were officially reported, and almost 600 people died of the disease (209). The disease in animals has a serious economic effect, causing cattle to abort and high fatality rates among newborn

lambs. In humans, infection usually results only

in mild dengue-like febrile illness, although it can

result in severe ocular (inflammation of the retina and blindness), encephalitic, or hemorrhagic

complications.

Lassa fever is endemic to some regions of West

Africa. Symptoms include prostration, severe sore

throat, tonsillitis, chest pain, and pneumonitis (inflammation of the walls of the air sacs in the lung).

There is evidence that mild or subclinical infections occur, but among those hospitalized with

Lassa fever, fatality rates range from 20 to 40

percent.

Research Needs

Laboratory techniques can identify arboviruses

and define antigenically similar groups, but there

is great geographic and climatic diversity in each

serologic grouping. This situation emphasizes the

complexity and challenge of arbovirus research

and control.

There is no cure for arboviral diseases, only

symptomatic and supportive relief. Early diagnosis of the agents responsible for potential outbreaks and epidemics identified down to the finest

level possible, has three important values: 1) to

differentiate ambiguous presenting symptoms; 2)

to anticipate life-threatening complications, as in

yellow fever and dengue fever; and 3) to target

the vector, which then determines control strategies. Current control efforts rely on early identification of epidemic outbreaks of arboviraI infections, allowing the institution of vector-control

measures such as insecticide fogging. In many

tropical countries, however, disease surveillance

is not well developed.
5 .

Strategies and Technologies for

Controlling Tropical Diseases
Contents

Introduction . . . . . . . . . . . . . . . .

Strategies for Controlling Trop

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

cal Diseases . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Tools__for Controlling Tropical Diseases . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

General Measures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Vector Control Technologies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Immunization Technologies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Diagnostic Technologies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Therapeutic Technologies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Genetic Tools... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

TABLE

Table No.

5-I. Considerations for Use of a Vaccine for a Tropical Disease . . . . . . . . . . . . .

LIST OF FIGURES

Figure No.

5-1. Structure of

5-2. preparation

Antibody Molecules. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

of Monoclinal Antibodies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Page

99

100

102

102

103

103

105

105

106

Page

104

Page

109

110
5 ■

Strategies and Technologies for

Controlling Tropical Diseases

INTRODUCTION

Throughout history, diseases have been brought

under control by a range of direct and indirect

measures. In most cases, disease control has been

an unplanned bonus of economic and social development. The opportunities for controlling a

disease either directly or indirectly depend on the

relationships among the disease-producting organism, the environment, and humans. The more

complex the relations, the more points for intervention, but not necessarily for success in controlling the disease. Experience shows that the

most complex infectious diseases are, in general,

the most poorly controlled.

In the developed world, the control of infectious diseases has come about largely as a result

of improvements in living conditions. Because of

better sanitation, uncontaminated potable water,

uncrowded housing, and an adequate diet, exposures to pathogens are less frequent and less intense, and people are better able to fight off severe infection.

Many recent declines in U.S. mortality rates are

due to specific interventions, such as new vaccines

and drugs, that are targeted at specific diseases

(e.g., vaccines and antiviral drugs for hepatitis B).

Three types of biomedical tools are used to control many of the infectious diseases of the developed world: 1) immunization techniques to prevent disease; 2) diagnostic technologies; and 3)

therapy, including surgical and medical treatment.

A fourth tool important to controlling many tropical diseases is vector control—i.e, control of the

insects and other organisms that transmit diseaseproducing organisms to humans (e.g., mosquito

vectors of malaria) or are otherwise necessary for

the disease-producing organism to complete its life

cyle (e. g., aquatic snails that serve as intermediate hosts for schistosomes).

These four tools are the primary direct measures that can be used to control diseases in individuals. Along with disease surveillance (which

may involve screening segments of the population for signs of disease or immunity) and keeping track of vector populations and their infection rates, these tools are also the means for

controlling infectious diseases in populations.

The remaining chapters of this report review

the status of vector control (ch. 6), immunization

(ch. 7), diagnosis (ch. 8), and treatment (ch. 9)

for the tropical diseases of interest in this report:

malaria, schistosomiasis, trypanosomiasis, leishmaniasis, filariasis, leprosy, tuberculosis, diarrheal diseases, acute respiratory infections (ARIs),

and arboviral and related viral infections. Current research and promising developments are

highlighted for each. This chapter provides background for the chapters that follow by laying out

the strategies, using single and multiple interventions, for controlling tropical diseases.

Some gains in understanding tropical diseases

have yet to be applied to any specific control

measure. The methods of biotechnology (e.g., the

use of recombinant DNA technology), for example, have led to new knowledge about the immunology of parasitic disease organisms. Although

some of this knowledge may eventually be applied

to the development of vector control, vaccines,

diagnostic or therapeutic measures, it currently

has no practical application. Biotechnology’s contributions to advance in research toward disease

control are brought to light in several chapters

of this report. In the latter part of this chapter,

some of the methods basic to biotechnology are

described.

99
100 ● Status of Biomedical Research and Related Technology for Tropical Diseases

STRATEGIES FOR CONTROLLING TROPICAL DISEASES

Most developing countries of the tropics are still

at the stage where increased investment in sanitation and clean water would have a tremendous

benefit in the control of disease. Extending clean

water and sanitation facilities to as many of the

world’s people as possible continues to be a goal

of development agencies and international health

groups. Although this report does not evaluate

the success of such efforts, it is clear that large

portions of the world’s poor remain unserved. The

tropical disease control measures in the biomedical spectrum, which are the focus of this report,

will not solve the underlying problem of the lack

of sanitation and clean water, which in many

cases contributes to disease transmission. In many

cases, however, these measures can protect against

contracting disease and reduce morbidity and

mortality due to disease.

Worldwide eradication is possible for very few

diseases. Smallpox was eradicated through a

worldwide vaccination program, and the eradication of measles may also be possible. For most

of the diseases considered in this report, however,

the most realistic goal at present is not eradication but control. Control means lowering morbidity and mortality to tolerable levels and containing the spread of disease.

Certain features of smallpox were essential to

its eradication: 1) smallpox is transmitted directly

from person to person, with no vector; 2) smallpox

is transmitted only through fairly close contact

with an infected person; 3) there are no animal

reservoirs for smallpox; 4) there are no subclinical cases of smallpox (virtually everyone who is

infected breaks out in the smallpox rash); and 5)

smallpox develops within a relatively short time

after exposure.

Most other major parasitic diseases—certainly

the five included in the Special Program for Research and Training in Tropical Diseases (TDR)—

depart from these characteristics. The parasites

that cause malaria, trypanosomiasis, leishmaniasis, and filariasis are transmitted by vectors, and

the parasites that cause schistosomiasis live part

of their lives in intermediate snail hosts. The probability of eradicating vector-borne diseases by killing off all vector species or intermediate hosts appears slim. The story of the worldwide “malaria

eradication campaign” waged by the World Health

Organization (WHO) beginning in the 1950s has

taught important lessons in that regard (see ch. 6).

Diseases that affect both humans and other animals pose special problems for control. The existence of animal reservoirs of disease means that

even if all human cases are eliminated at any one

time, the disease will still exist in animals and will

eventually pass into humans once again. One

strain of African trypanosomiasis is actually a disease of livestock, called nagana, which can be

transmitted to humans. Yellow fever is an arboviral disease that exists in monkeys as well as humans. A highly effective, long-lived vaccine for

yellow fever has existed for decades, and in many

areas, transmission to humans has been largely

curtailed. Because of the monkey reservoir, however, the absence of cases in humans does not allow discontinuing vaccination for yellow fever as

was possible in the case of smallpox. Workers in

development projects in Brazil, in which jungle

is cleared for roads or agriculture, have been the

victims of a type of yellow fever (“jungle yellow

fever”) in which the virus picked up by mosquitoes from monkeys is transmitted to humans.

The control of yellow fever is quite good, but realistically, there will be no eradication.

The control of most of the other tropical diseases considered in this report is not as successful as that of yellow fever. The development of

successful malaria vaccines, however, could put

malaria control on a similar footing.

If eradication is not the goal, the options for

disease control are varied, but it has been difficult to discover the most effective and most costeffective mix of control measures. The “St. Lucia

Experiment, ” for the control of schistosomiasis on

the West Indian Island of St. Lucia, is one example of a successful research effort to compare the

effectiveness and costs of several control measures (see box 5-A).
Ch. 5—Strategies and Technologies for Controlling Tropical Diseases •101
102 ● Status of Biomedical Research and Related Technology for Tropical Diseases

TOOLS FOR CONTROLLING TROPICAL DISEASES

General Measures

The infectious diseases that killed most Americans at the turn of the century are now largely

under control in the United States. Improved sanitation, the presence of adequate nutrition, and

higher levels of education claim much of the

credit. Those three conditions are not met in most

developing countries.

Providing a secure, clean water supply for

drinking and washing, and successfully educating people in how to use it, can reduce the transmission of diseases such as schistosomiasis (which

is transmitted to humans through contact with

water in which infected snails live) and diarrheal

diseases caused by water-borne and food-borne

pathogens. Sanitary disposal of human and animal waste further reduces the chance of spreading disease. In southern Africa, south Asia, east

Asia, and the Pacific, less than half the population has access to a secured water supply or excreta disposal. In general, access is much greater

in urban than in rural areas. In India, for instance,

80 percent of the urban population, but only 18

percent of the rural population, has water supplied, In Ethiopia, the corresponding figures are

58 and 1 percent (412).

Adequate nutrition buoys functioning of the

immune system, increasing the individual’s resistance to contracting diseases and his or her ability to fight a disease once

is particularly critical for

infants.

contracted. Nutrition

pregnant women and

Education, particularly of women, has beneficial effects on maternal and child health. The

World Bank estimates that for every year of maternal education, infant mortality rates drop by

nearly 1 percent (412). Although the exact sequence of events has varied around the world, in

general, birth rates drop as the level of health increases. Fewer births lead to healthier children and

women, and, in the long run, can help to improve

nutrition.

Chagas’ disease (American trypanosomiasis)

could be controlled through the application of

present-day knowledge. In Panama, for example,

the Corozal palm (Schelia spp. ), which provides

thatch for rural dwellings, has been found to be

a mini-ecosystem, harboring both reduviid bug

vectors and mammalian reservoirs of Trypanosoma cruzi. If houses were constructed with corrugated metal or adobe tile roofs rather than

thatch, then human contact with the reduviid bug

vector of the disease could be limited considerably. Any attempts to control the reduviid bugs

by insecticide only must be continuous, because

the mammalian reservoirs of Chagas’ disease provide a steady supply of T. cruzi parasites ready

to infect the next generation of reduviid bugs that

feed on them.
Ch. 5—Strategies and Technologies for Controlling Tropical Diseases ● 103

Vector Control Technologies

(see ch. 6)

Many important tropical diseases, including

malaria, trypanosomiasis, leishmaniasis, filariasis, and arboviral infections, are transmitted to

humans by mosquitoes or other arthropods (e.g.,

ticks, sandflies, midges, and gnats). One, schistosomiasis, requires a second host, a freshwater

snail, for the parasite to complete its life cycle.

Control of vectors and snails in endemic areas is

one way to control the transmission of these

diseases.

Before about the middle of this century, most

vector control efforts used mechanical approaches

such as catching insects, removing breeding sites,

or setting up physical barriers to the vectors’

migration (e.g., by clearing strips of forest). Most

vector control activities today use pesticides. The

pesticide era began with the introduction of DDT

(dichloro-diphenyl-trichloro-ethane) after World

War II and has had some significant accomplishments. It also has taught that insects have a great

capacity to evolve pesticide-resistant forms. In

many parts of the tropics, stronger and stronger

pesticides have been used against ever more resistant insects.

Other vector control strategies include biological control measures (e.g., the introduction

of predators or pathogens of vectors) and environmental control measures (i. e., altering the environment to eliminate conditions necessary for

the vectors’ survival). The use of a combination

of measures to control vectors is generally referred

to as “integrated pest management” (1PM).

Immunization Technologies (see ch. 7)

The purpose of immunization is to prevent an

individual from getting a disease if exposed to the

agent that causes the disease. Vaccines are the

most important tools of immunization. The functional components of vaccines are “antigens,” substances that cause the body to marshal its own

Photo credit: Dr. Robert Edelman, National Institutes of Health

Irrigation ditches are frequently infested with the snail intermediate hosts of schistosomes.
104 ● Status of Biomedical Research and Related Technology for Tropical Diseases

immune system by activating one or more types

of immune system cells. Some cells may be programmed to physically attack disease organisms

and others are primed to produce “antibodies,”

substances that act to neutralize or kill the disease organism should the organism invade the

body. Less important than the active immunity

induced by vaccines is a passive immunity produced in an individual by the transfer of antibodies in blood serum from other, actively immune

individuals.

All the human vaccines in use today are vaccines for bacterial and viral diseases. There are

currently no vaccines for human parasitic diseases, but there is a great deal of research activity toward their development. The furthest along

is a vaccine against one stage of the malaria parasite Plasmodium (see Case Study B: The Development of a Malaria Vaccine).

Vaccines benefit both the individuals immunized by them, and the community as a whole.

Even if every person in the community is not vaccinated, if a high proportion are vaccinated or

have acquired immunity from having had the disease, the nonimmune members will also be protected because there are not enough susceptible

people to sustain disease transmission. This

phenomenon is called “herd immunity.”

In the United States most, but not all, children

are immunized against measles, mumps, rubella,

poliomyelitis, tetanus, diphtheria, and pertussis.

In general, immunization levels are high enough

to produce herd immunity. Outbreaks do occur,

however, particularly outbreaks of measles. Investigations usually reveal that a critical mass of

unvaccinated children existed in the community,

and at least one was exposed to a case of measles. Although there are seldom deaths associated

with measles in the United States, that is not the

case in the developing world.

Immunization programs are potentially the

most effective means of disease control for most

infectious diseases, but they require a long-term

commitment of public health resources to be successful. The initiation of vaccine programs requires careful analysis of the health, social, political, and economic conditions of the population

to be immunized. (Table 5-1 lists the biologic considerations for using a vaccine against a tropical

disease. ) Since 1974, WHO has been the major

promoter of childhood immunization, through its

Table 5.1.- Considerations for Use of a Vaccine

for a Tropical Disease

Regarding the target organism:

. Complexity of antigenic structure

● Multiple stages of the organism’s life cycle

● Accessibility of infection site within the host

Ž Occurrence of different species, geographic strains, local

and temporal variants

Ž Inherent genetic variability of organisms

● Possibility of genetic selection for vaccine resistance

Regarding the vaccine:

Ž Immunogenicity of antigen: efficacy and effectiveness

● Use and type of adjuvant or diluent

● Possible undesirable side effects

● Targeted at one or several variants of the disease organism

● Separate administration or combined with other target

vaccines

● Storage requirements (e. g., refrigeration and shelf life)

Regarding the vaccinee:

●

●

●

●

●

●

●

●

A g e

Health status, including pregnancy

Degree of possible resistance

—due to genetic or ethnic factors

—due to prior exposure or acquired immunity

—due to current infection

Protocol; type, number, and time of other immunizations

Access to prophylactic and therapeutic drugs

Route, mode, dosage, and number of inoculations

Measurement of immune response

—comparison with age-matched groups in developed

countries

—monitoring for side effects or adverse reactions

Degree and duration of protection

—from infection

—from pathologic effects of disease

Regarding the vaccine trial:

●

●

●

●

●

●

●

●

●

●

●

●

●

Randomized selection of vaccinee and control groups

Characteristics of placebo used in control group

Availability of baseline disease incidence and prevalence

data

Ethical aspects

Training of field teams and standardization of procedures

Site selection: number and distribution of trial populations

Identifying populations at risk but not already infected or

immune

Determining exposure to and risk of infection by target organism

Statistical validation of data; required number of participants

Presence of related or competing organisms

Length of followup

–for safety

—for efficacy

Coordination with other public health programs—effect of

vector control or other simultaneous projects

Source and adequacy of funding

SOURCE: P. Basch, “The Role of Biotechnology in Tropical Disease Research,”

contract report prepared for the Office of Technology Assessment, U.S.

Congress, Washington, DC, 1984.
Ch. 5—Strategies and Technologies for Controlling Tropical Diseases ● 105

Expanded Program on Immunization (see box

5-B).

Diagnostic Technologies (see ch. 8)

Diagnostic technologies serve several purposes:

● to determine pertinent characteristics of an

individual’s disease, so that appropriate treatment can be given;

● for public health reasons, to determine the

prevalence of pathogenic agents in populations; if control measures (e.g., a vaccine) are

begun, determining the prevalence of such

agents is important to allow evaluation of the

success of the measures; and

● for research purposes, to find out about the

ranges of diseases affecting a population and

the immune status of populations, for instance.

A wide range of biochemical tests supplements

the observational abilities of medical personnel

in diagnosis. Many of these diagnostic tests rely

on reactions between antigens and antibodies and

have radioactive or color markers that signify the

test result. Recombinant DNA technologies are

contributing to diagnostics in several ways, including the production of pure reagents for tests

and the production of “DNA probes, ” which recognize the genetic material of disease-producing

organisms.

Therapeutic Technologies (see ch. 9)

The object of therapy is to improve the lot of

a person with disease. In the best case, therapy

involves ridding the person of disease, but for

many conditions, symptomatic relief is equally

important. This report concentrates on drug treat-
106 ● Status of Biomedical Research and Related Technology for Tropical Diseases

ment; it does not consider surgical treatment, because surgery plays a relatively minor role in the

treatment of most tropical diseases.

Drugs to treat most bacterial infections exist,

though bacteria in many cases are resistant to one

or many drugs. Drugs to treat helminthic (worm)

infections also exist, but these are generally less

effective and less safe and require longer periods

of treatment than do drugs against bacterial infections. The development of drugs to treat viral

infections is just beginning. The antiviral drugs

currently available are not commonly used against

tropical diseases.

Genetic Tools

The science and industry of present-day biotechnology—the use of recombinant DNA techniques and other sophisticated genetic tools—are

outgrowths of the ideas and experiments of a 19th

century Austrian monk. In 1865, Gregor Mendel

first described the concept that traits of higher

organisms are transmitted from one generation

to the next as discrete units. Although Mendel’s

work was unrecognized and unappreciated for

half a century, the science of genetics had been

born. For decades, the units of heredity, genes,

were considered to be vaguely defined units or

particles riding in some sort of linear order upon

the chromosomes of animals and plants. Beads

on a string were used as a popular analogy.

In 1953, Watson and Crick suggested a structure for the precise chemical makeup of Mendel’s

hypothetical hereditary units. The now familiar

double-helix structure of deoxyribonucleic acid,

or DNA, is at the core of modern molecular genetics, for it provides the framework of our understanding of the functioning of the genetic material and the basis for manipulating it.

The DNA molecule consists in part of an invariant “backbone” made up of two coiled strands

of repeating sugar (deoxyribose)-phosphate components. Each sugar has attached to it one of four

chemical bases. These four bases—adenine (A),

cytosine (C), guanine (G), and thymine (T)—in

certain combinations form bridges, something like

the rungs of a ladder, across the paired helically

coiled deoxyribose-phosphate strands. An A must

pair with a T, in either order (A-T or T-A); and

a C must pair with a G. Therefore, given a sequence of bases on one strand of a DNA molecule, say A-T-A-C-G-T, one can deduce the complementary sequence, in this case T-A-T-G-C-A,

which must be attached to the other strand.

In the synthesis of new DNA from existing

DNA, a process known as “replication,” the two

strands of the DNA molecule separate, each serving as a template. In the process of “transcription,”

the DNA strand serves as a template for a strand

of a slightly different nucleic acid known as RNA,

which in turn codes for the synthesis of proteins

(see 357 and 361).

Recombinant DNA Technology

In 1973, Cohen, Chang, Boyer, and Helling reported the first in vitro production of DNA

molecules that were “replicated” when transferred

into living organisms. The details of recombinant

DNA work are extremely complex and constitute

the subject of many thousands of technical publications (e.g., 187, 132, 224, and 397).

Recombinant DNA technology is the term used

for methods to transfer segments of genetic material from one organism to another, to replicate

it, and to use it to make chemicals (see box s-C).

Recombinant DNA technology represents an extremely powerful and precise set of tools with unknown, but certainly great, implications not only

for tropical medicine but for many aspects of life

in the future. A widely acclaimed use of recombinant DNA technology is for the production, in

commercial quantities, of materials such as human insulin, interferon, or growth hormone. An

application of cloned complementary DNA or

RNA (see box s-C) is in locating a viral, bacterial,

or parasitic DNA sequence within a host organism. These nucleic acid hybridization probes

promise to be useful in molecular diagnosis of

disease.

Monoclinal Antibodies (MAbs)

MAbs are homogeneous antibodies derived

from clones of a single cell. To understand MAbs

and the hybridomas that produce them, it is necessary to go back to the work of early immunologists.
Ch. 5—Strategies and Technologies for Controlling Tropical Diseases ● 107

Box K.—Recombinant DNA Technology-.

Recombinant DNA technology is a process

whereby pieces of DNA from different sources,

as different as humans and bacteria, are joined

together. The basic method for recombining

DNA follows.

1. Start with a, b, or c:

a.

b.

c.

DNA is extracted from an organism.

Within the total DNA (the genome) is the

genetic information for making ill of the

components and characteristics of that organism. The tick is to pull out from all that

DNA the gene of interest, which probably

constitutes only l/1,000th or l/10,000th of

the total.

A reversal of the transcription process (in

which DNA is a template for RNA) is used

to make a particular piece of DNA. RNA

is usually extracted from cells that are actively producing the desired material.

Through the use of a special enzyme called

reverse transcriptase, the strand of RNA is

used as a template on which a segment of

single-stranded DNA is synthesized. Because the structure of this new DNA strand

is complementary (like the relationship between a photographic negative and a print

made from it) to the structure of the RNA

template, the newly formed DNA is called

“complementary” DNA. The DNA and

RNA strands of the “hybrid” molecule are

separated and the DNA strand used as a

template with the enzyme “DNA polymerase I“ to synthesize a double-stranded

DNA configuration.

If the sequence of bases in the desired DNA

is known, chemical methods are used for

the direct synthesis of DNA. These methods are limited to “shorter” pieces.

Z. The extracted DNA (from la) orcomplementary DNA (from lb) is cut into pieces by use

of restriction enzymes (restriction endonucleases) that cleave the DNA or complementary DNA molecule at certain specific sites.

Restriction enzymes have been extractedfrom

more than 200 different strains of bacteria and

cleave the DNA molecule at any of more than

90 known sites, unique for any particular erkzyme. The cleavage site is determined by the

sequence of constituent base pairs. One of the

most commonly used restriction enzymes,

called “ECO RI” and derived from Escherichia
Investigators had observed around the turn of

the century that in response to the natural or intentional introduction of foreign materials (“antigens,” animals produce specific substances which

circulate in the blood. These responsive substances

became known as “antibodies,” and early researchers such as Paul Ehrlich believed that the

antibody and its inducing antigen fit together

physically like a lock and a key.

Various theories of antibody formation and action were proposed by many workers, but it was

not until 1959 that the Australian immunologist

Sir Macfarlane Burnet conceived the theory of

clonal selection. Burnet’s idea, which led to a

Nobel Prize, was that a great variety of quiescent

but potentially antibody-producing cells exist in

the normal human or animal, each with the inherent capability of secreting one specific kind of

antibody. These quiescent but potentially antibody-producing cells are now known to be a certain type of white blood cells, which are called

B-lymphocytes. According to Burnet’s clonal

selection theory, the introduction of any particular antigen into the body causes only those cells

preadapted to it to become stimulated and to proliferate and differentiate into plasma cells that release specific antibody in measurable amounts.

The many cells created by the repeated division

of the originally stimulated cell are “clones,”

whose secreted antibody mixes in the bloodstream

with the products of all other clones of antibodyproducing cells.

The structure of antibodies is now known with

great precision. Although there are several subtypes of antibodies, their basic composition and

behavior is quite similar. As shown in figure 5-1,

an antibody molecule has two symmetrical halves,

each containing one “heavy” and one “light” protein chain. Each of the four chains contains a constant region (for certain generalized functions),

and a variable region (on which are located the

antigen-binding sites that determine the antibody’s

specificity).

For decades, immunologists had obtained antibodies for their research by the inoculation of animals, usually rabbits, with antigen. The antigenic

dose could be as simple as a single purified protein, or complex as a whole-organ extract. After

some time, the rabbit was bled, its blood allowed

to clot, and the clear serum, containing the antibodies, was collected. At that point, the problem

was always how to separate the particular antibody of interest from the mixture of very similar

materials in the rabbit’s blood serum. Scientists

devised various chemical precipitations and immunologic binding methods to try to fish the

desired antibody out of the complex “polyclonal”

pool. Many problems were encountered in these

procedures.
Ch. 5—Strategies and Technologies for Controlling Tropical Diseases “ 109

Figure 5.1 .—Structure of Antibody Molecules

I

A.

A. The basic unit of all antibodies is a protein molecule made

up of two “light” and two “heavy” chains joined by disulfide

bonds. Foreign antigens are recognized by regions at one end

of the molecule (variable regions). Other parts of the molecule

are specialized for “effecter functions” that aid in deactivating and removing the foreign antigen. Antibodies in the

immunoglobulin classes lgG, lgD, and lgE exist as single

monomers.

B. Antibodies belonging to immunoglobulin classes lgM and

lgA exist as molecules with multiples of the basic unit. lgM

(in diagram) is a pentamer (5 units). Different types of lgA exist

in monomers, dimers (2 units), and trimers (3 units). The units

are joined by disulfide bonds.

SOURCE Office of Technology Assessment, 1985.

One problem was that complex antigens, as

large molecules, usually bear several different

structural antigenic determinants called “epitomes, ” each of which is recognized by distinct

antibody-producing B-lymphocytes. The inoculation of a complex antigen results in the production by the immune system of several different

antibodies, each of which is specific to a portion

of the same antigen. Conversely, several distinct

antibodies may combine with the same portion

of the antigen. An additional complication is that

rabbits, even from closely inbred lines, vary in

their individual ability to recognize and respond

to particular antigens: the antibodies produced are

sometimes heterogeneous and vary from rabbit

to rabbit or even within the same animal over

time.

Scientists’ inability to obtain a uniform, reproducible, and above all, specific product hindered

biomedical research until the invention of hybridoma technology in the mid-1970s. This invention, by Georges Kohler and Cesar Milstein at

Cambridge University in England, was a landmark in the history of immunology.

For some time, immunologists had attempted’

to isolate and clone antibody-forming B-lymphocytes in artificial culture media, but these cells normally have a limited life span and clones cannot

be established. Malignant tumors of the immune

system called “myelomas” are capable of continuous proliferation in cultures, but cultured myeloma cells produce little or no antibody. Kohler

and Milstein devised a method for forcing the

recalcitrant myeloma cells to produce specific

antibodies in quantity. Their work, published in

1975, has led to an explosion of subsequent

studies. What Kohler and Milstein did was to

combine an immortal but nonsecreting myeloma

cell line with a secreting but nonculturable Blymphocyte in order to produce “chimeras” having the desirable properties of both parental types.

This is the “hybridoma” ( =hybrid myeloma) so

commonly used today. Hybridomas and MAbs

have been reviewed several times (e.g., 6), and

step-by-step instruction manuals for their preparation are available (206,437).

The method for preparing MAbs is shown in

figure 5-2. First, a mouse is immunized with an

antigen and permitted some time to respond.
110 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 5-2.–Preparation of Monoclinal Antibodies (MAbs)

Mouse i s

immunized with

a foreign

substanc e

or ‘antigen. ”

Spleen is

removed and

minced to

release antibodyproducing cells-

(B-lyrnphocytes).

Mouse myeloma

(tumor) cells

are removed

and placed in

tissue culture.

Cells divide in

liquid medium.

The products of this

Myeloma cells which are both “immortal” and contain genes

from the antibodyproducing cells survive,

These are called

“hybridomas. ”

Hybridomas are cloned

and the resulting cells

are screened for antibody production.

Those few cells that

produce the antibodies

being sought are grown

in large quantities for

production of

monoclinal antibodies

(MAbs).

Lymph nodes or spleens of immunized animals

are then removed, and their cells are dispersed and

added to cultures of certain types of mouse myeloma cells. The mixed cells are incubated so that

their cell membranes are allowed to fuse together,

producing hybrid cells with two nuclei. After a

period, the hybrid cell nuclei stabilize, and the cultures, consisting of thousands of cells, are placed

into a medium in which the growth of the original myeloma cell line is prevented. Only the hybrid cells are now capable of proliferation in the

new culture conditions, and the task at this point

is to identify and separate those cells that are active secretors of a desired antibody. This is accomplished through any of several types of screening assay, in which the overlying culture medium

is tested for the presence of antibody by incubation with a particular antigen labeled (with a

radioactive or fluorescent marker) to permit the

ready identification of resultant antigen-antibody

complexes. Those cell populations demonstrating

antibody production are then diluted, and each

surviving hybridoma cell is isolated to start a

clone of genetically identical cells. The result of

this process is the development of large colonies,

each derived from a single hybrid cell, and each

secreting one specific—hence “monoclonal”-

antibody.
Ch. 5—Strategies and Technologies for Controlling Tropical Diseases • 111

Milstein has written (223):

A monoclinal antibody is a well-defined

chemical reagent that can be reproduced at will,

in contrast to a conventional antiserum, which

is a variable mixture of reagents and can never

be reproduced once the original supply is exhausted.

All progeny cells from a single hybridoma clone

secrete antibody molecules whose variable regions

are identical—the variation is only between, not

within, clones.

A single experiment may yield a large number

of hybridoma clones, about 10 percent of which

secrete a monomolecular antibody specific to that

particular cell line. Some of these maybe of little

or no interest to the investigator, who now faces

the lengthy and arduous task of determining the

specificity of each secreting hybridoma clone and

deciding on its relevance. In some experiments,

it may be the goal to generate a variety of different hybridomas. More commonly, an investigator

wants to obtain a highly specific antibody. Therefore, he or she attempts to inoculate a correspondingly purified antigen into the B-lymphocyte-contributing mouse in order to reduce the “noise” of

unwanted hybridomas.

Hybridoma-derived clones of secreting cells

may be propagated in culture media, from which

the specific antibody may be separated. Since

these cells are derived in part from mouse myelomas, they will induce tumors in mice, with the

production of relatively large volumes of antibody-containing ascites fluid, This fluid can be

harvested for use, and the hybridoma line can be

perpetuated by transfer from mouse to mouse.

A particularly intriguing application of MAbs

is in the production of what are referred to as

“anti-idiotypic” antibodies. In this technique, animals are immunized with a previously prepared

MAb made against a disease-causing organism.

The idea is to stimulate the production of a complementary “anti-antibody” which may substitute

for the antigen against which the original monoclinal was raised. In an early application of this

method to a tropical disease pathogen, Sacks and

Sher (303) found that several specific anti-idiotypic antibodies were able to protect mice against

challenge with Trypanosoma rhodesiense, while

others were ineffective. Other investigators have

recently reported successful immunization of mice

against the bacterium Streptococcus pneumonia

with an anti-idiotype vaccine (22). The technique

is certain to have wider application, although its

eventual usefulness is unknown.

Applications of Genetic Tools

to Tropical Diseases

Applications of genetic tools to tropical medicine have been discussed extensively by Falkow

(112) and Cross (81), who indicated some uses and

advantages of cloned genes in medical and veterinary science:

●

●

●

●

elucidation of genetic basis of normal and abnormal cell function;

diagnosis of disease and disorders;

synthesis of biologically active proteins; and

production of subunit vaccines, which may

provide:

—improvements in existing vaccines,

—production of new vaccines,

—reduction in adverse side effects as a result of increased purity,

—safety in manufacture and use, and

—consistency in manufacture and use.

The many applications of MAbs to parasitic

tropical diseases have been discussed by Mitchell and Cruise (237), Mitchell (235,236), McBride

(218), and Rowe (295). In brief, the uses of MAbs

in tropical diseases caused by parasites are:

●

●

●

●

●

●

●

for diagnosis of infection and identification

of the particular type or strain of parasite,

for information about the genetics and relationships among the organisms,

as “probes” for defining the antigenic composition of the parasites and of their

products,

for identification of the parasite components

likely to stimulate host protective responses,

with possible vaccine production,

for standardization of diagnostic reagents and

vaccines,

possibly for drug targeting or other means

of immune-based treatment of infected hosts,

and

to effect host protection in vivo.
712 ● Status of Biomedical Research and Related Technology For Tropical Diseases

MAbs can also stimulate production of antiidiotypic antibodies and are widely used as probes

and reagents in molecular biology laboratories to

detect expression of cloned parasite DNA by host

cells.

In the area of tropical medicine, the genetic

tools find their greatest potential application in

the search for protective vaccines. If a parasitic

organism contains a particular antigenic portion

that can stimulate protective immunity in humans, it would be desirable to obtain large

amounts of that specific antigen (isolated from the

remainder of the organism) to use as a vaccine.

One way to obtain such material might be to grow

the parasite either in animal hosts or in laboratory cultures, and then extract the desired antigen by chemical means. Because of the time and

expense involved, this approach is feasible only

for experimental work in the laboratory. However, identification, isolation, replication, and

production of the antigen by the expression of the

DNA sequence responsible for antigen production

could provide access to an essentially unlimited

and relatively inexpensive source of the immunizing material.

Biotechnology research with agents and vectors

of tropical diseases requires a laboratory equipped

with a wide range of modern apparatus, including optical and photographic equipment, gel electrophoresis devices and power supplies, centrifuges, refrigerators and freezers, sterilization

facilities for culture media, biological safety cabinets, and myriad smaller devices. A great number of chemicals and reagents are also necessary.

Facilities for growing the disease organism must

be maintained or the organism must be obtained

from elsewhere. Trained technical personnel are

indispensable. The work is demanding of effort,

skill, and imagination, and requires a certain level

of financial support. Most developing countries

do not have the capacity to do this type of research, so nearly all of it is done in the United

States and other developed countries.
6 .

Vector Control Technologies:

Selected Tropical Diseases
Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Vector Bionomics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Control of Arthropod Vectors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Insecticides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Biological Control Measures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Detection of Species Complexes. . . . . . . .

Identification of Disease Organisms With

Control of Trypanosomiasis Vectors . . .

. . . . . . . . . . . . . . . . . . . . . .

n Vectors , . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . .

Control of Snail Intermediate Hosts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Molluscicides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Biological Control Measures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Detection of Species Complexes. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Integrated Pest Management. . . . . . . . . . . . . . . . . . . . . . . . . . . ........: . . . . . .

Research Needs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Insecticides and Molluscicides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Vector Bionomic Studies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Page

115

115

117

117

120

120

121

121

122

122

123

123

123

124

124

124

124
6 ■

Vector Control Technologies:

Selected Tropical Diseases

INTRODUCTION

The discovery at the turn of the century that

some tropical diseases were transmitted by mosquitoes was a major breakthrough in understanding these diseases. The control of mosquito vectors was the earliest and often the only method

of intervention against many tropical diseases.

Early public health workers, such as Gorgas in

Cuba and Panama, and Watson in Malaya, were

able to use vector control methods such as drainage and filling, or larviciding standing water with

oil and the dye “Paris green, ” with spectacular success in controlling the carriers of malaria and yellow fever. These methods continued to be used

until the advent of the DDT era in the mid-1940s.

This chapter reviews the current status of control technologies for the insect and other arthropod vectors of diseases such as malaria, filariasis, leishmaniasis, and arboviral infections (e. g.,

yellow fever). It also reviews the status of control technologies for certain mollusks (e.g., snails)

that are essential in the life cycles of trematode

parasites that cause diseases such as schistosomiasis. The terms vector biology and control have

VECTOR BIONOMICS

Progress in vector research tends to come in

small increments that relate to better understanding of vector bionomics (the study of vectors’ habits and their interactions with the environment).

Such research is often narrowly focused research,

from which extrapolations can only infrequently

be made to other vector control situations. Studies

of mating habits, host preferences, oviposition

(egg laying) behavior, larval habitats, flight densities, and natural infection rates with pathogenic

organisms provide insights that add to our abiltraditionally applied only to insects and other arthropods that carry diseases. Currently, however,

these terms also apply to snail intermediate hosts.

Control of vector-borne diseases is possible

only through detailed and accurate understanding of the factors involved in transmission. Such

understanding is gained through meticulous and

tedious observation and experimentation wherever the disease exists. Understanding the role of

the vector necessarily overlaps with field studies

of the human host’s interaction with the environment. Cultural practices, such as storing barrels

of drinking water without lids (which provide perfect breeding sites for mosquitoes), or poor sanitary habits need to be studied to best design social intervention techniques.

Prospects for the future, including the relatively

new concept of integrated pest management (1PM),

are summarized in this chapter. Vector bionomics,

the study of vectors and their interactions with

the environment, provides the scientific rationale for interventions against vectors and is described briefly below.

ity to understand transmission dynamics and to

target interventions against susceptible vector

species.

Information about the resting habits of mosquito vectors of malaria has provided the rationale for, and accounted for the success of, the indoor residual spraying of insecticides in many

tropical areas. Because some mosquitoes rest on

a surface before and after the blood meal, often

near the person they have bitten, spraying all

115
116 ● Status of Biomedical Research and Related Technology for Tropical Diseases

available resting surfaces (the walls, ceilings, undersides of furniture, etc. ) with a long-lasting insecticide (e.g., DDT) has been the method of attacking the vector species.

In many cases, this approach to malaria control has led to great reductions of malaria prevalence. (In some areas of antimalarial spraying,

sandflies, vectors of leishmaniasis, have also been

controlled. ) Unfortunately, in many cases, lack

of information about the behavior of specific populations of mosquitoes has led to failure. Some

species, or subpopulations within species, exhibit

a behavior pattern of entering a house to feed,

but resting outside, or actually avoiding sprayed

surfaces. For example, the malaria vector Anopheles nuneztovari in parts of Colombia and Venezuela enters houses, obtains a blood meal, and

then leaves immediately to rest outside, without

acquiring a lethal dose of DDT (107). A. minimus flavirostris, an important vector of Wuchereria bancrofti (filariasis) in Luzon and Palawan,

the Philippines,

only during the

filarial stage of

enters houses to feed on humans

middle of night, when the microthe parasite is at its highest con-

. .

centration in the blood. This particular mosquito

also leaves the house to rest outside, and is unaffected by indoor spraying of DDT.

Control of the primary vector of a tropical disease does not always result in successful control

of the disease. The disease may continue to be

prevalent and a second previously unsuspected

vector may be discovered. In one case, the elimination of stagnant groundwater as a breeding site

for malarial vectors was followed by the discovery of another vector breeding in tiny pools of

water trapped in the fronds of palm trees.

Work in the field of vector bionomics is progressing at a productive, though undramatic,

pace. This area is grossly understaffed and underfunded, and there are critically few new scientists

entering the field (253). Some progress of a fundamental nature has occurred in the laboratory.

For example, recent work has provided information about the importance of mosquito saliva,

which functions as an anticoagulant, in the mosquito’s proboscis (hollow tongue). The mosquito’s

saliva promotes the formation of a blood gradient

in tissues as it probes, helping the insect to find

a blood-filled capillary. Malarial sporozoites in

the mosquito damage the mosquito’s salivary

glands, reducing the production of saliva; the reduced production of saliva, in turn, causes an increase in probing time, and thus promotes greater

transfer of sporozoites to humans being bitten

(329).

Research in both the field and laboratory aspects of vector bionomics is time- and labor-intensive, but is a necessary activity for the eventual

control of vectors. As noted in a study of pest control by the National Research Council (253):

The control of arthropod vectors of disease or

other pests of public health should be attempted

only with recognition, and insofar as possible,

an intimate knowledge, of the significance of the

ecology and behavior of the target species and

the epidemiology of the disease and with appreciation for environmental values that may be depreciated.
Ch. 6—Vector Control Technologies: Selected Tropical Diseases ● 117

CONTROL OF ARTHROPOD

Insecticides

Insecticides, classified as either chemical

VECTORS

or biological, remain the primary vector control intervention. Two strategies are involved in the use

of insecticides: 1) reducing the population size of

the target vector (e.g., by treatment of a breeding site to kill immature forms); and 2) reducing

the longevity of adult forms of the vector, so that

they die before transmitting the disease (e.g., by

residual house-spraying to intercept only those

adult forms that feed on humans). Regardless of

the strategy, all insecticides act as a selective

mechanism against disease vectors, resulting in

the development of resistant insect populations

that have lost their susceptibility to the particular agent in use.

Chemical Insecticides

Chemical insecticides began to dominate the

field of vector control around the end of World

War II, with the widespread use of DDT. It was

at that time that the World Health Organization’s

(WHO) worldwide “malaria eradication campaign” began, and through that experience that

the world began to learn about the amazing capacity of insect populations to evolve that are

resistant to pesticides (see box 6-A). The development of resistance called for using even more

lethal chemicals.

Most insecticide development today is directed

at pests of agricultural importance rather than at

medically important vectors of diseases. Although

many new chemical insecticides are produced and

tested annually, these new chemicals are not necessarily appropriate for specific disease vectors.

Also, because of research and development costs,

new chemicals are expensive. The development

of insects that are resistant to new chemical insecticides is a virtual certainty. The only question

is simply how quickly the emergence of resistance

will cause the insecticide to be abandoned.

In spite of the gloomy prospects for the complete success of chemical pesticides, such pesticides

still form the backbone of vector control programs

and are responsible for controlling disease in many

Photo credit: Robert Edeman, National Institutes of Health

An anopheline mosquito, vector of malaria, after

a blood meal.

areas. Even DDT, which has been banned for agricultural use in the United States and many other

countries, is still a useful chemical.

DDT was not banned for public health use

(which is far more circumscribed than agricultural

use), however, and it continues to be used in many

countries worldwide for residual house-spraying

against vectors, especially those of malaria. The

only constraint on its use is resistance in local vector species.

The emergence of DDT-resistant strains of mosquitoes and the need to decrease the use of DDT

have spurred the development of new insecticides.

The principal compounds that have been substituted for DDT are malathion, fenitrothion, and

propoxur. Another insecticide, pirimiphos methyl,

has been field tested as a substitute for malathion

in several sites where local mosquitoes have developed malathion tolerance (e.g., in Turkey and

in Pakistan).

All of the newer chemical insecticides have greater immediate toxicity to humans and animals than

DDT has. The irony of the replacement of DDT

by other, more quickly degrading chemicals is that

in order to be effective within a limited time span,

the new chemicals must also be highly toxic. Thus,

although the long-term environmental effects of

DDT’s persistence have presumably been reduced,

the short-term hazards to users (i.e., workers in-
[page omitted]

This page was originally printed on a dark gray background.

The scanned version of the page was almost entirely black and not usable.
Ch. 6—Vector Control Technologies: Selected Tropical Diseases • 119

It turned a subtle and vital science dedicated to

understanding and managing a complicated natural system-mosquitoes, malarial parasites and

people-into a spraygun war.

The goal of malaria eradication has now been

laid aside, and current efforts to control the disvolved in the manufacture and application process, and people and wildlife inhabiting the areas

being sprayed) have probably increased. The new

insecticides are also many times more expensive

than DDT, a problem that has greatly limited their

use in the endemic countries unable to afford

them.

Biological Insecticides

Various agents called biological control measures (see below) are in many cases more accurately

described as biological insecticides. The biological insecticides include pathogenic bacteria, fungi

and their products, and growth hormones. In

most cases, the function and application of biological insecticides are completely analogous to

those of synthetically manufactured insecticides.

Often, the two types of insecticides differ only in

that the former are produced by a micro-organism

and the latter are produced by chemical synthesis. Most biological insecticides are directed

against larval forms of arthropods, which means

they are suitable only where conventional larviciding would also be feasible (e.g., where the

breeding sites are easily found and treated).

Despite the appeal of using naturally occurring

agents, it is not safe to assume that biological insecticides will be nonpathogenic to humans, nor

that because these agents occur naturally, the target organism is less likely to develop early resistance.

Pathogenic Bacteria .—The cultivation and application to vectors of infectious or toxic agents

such as viruses, bacteria, fungi, and helminths is

an area of active research. So far, however, there

is only one practical application in use, Bacillus

thuringiensis israeliensis (BTI), of which the most

common strain is called B.t. H-14.

BTI was first isolated in Israel in 1976, and extensive research has revealed that the bacterium

produces a toxic crystalline compound (delta-toxin) fatal to virtually all insects when they ingest

it. Now commercially produced, this toxin is used

on farms and gardens, as well as in public health

projects, in the United States and around the

world. B.t. H-14 is now applied in the control of

the Simulium (blackfly) vectors of onchocerciasis in West Africa. A number of field trials using

B.t. H-14 against mosquito species are in progress.

BTI differs in its mode of action from other insecticides. It is not readily susceptible to enzymatic

degradation or other biological processing within

the insect. Although there is no reason to believe

that BTI will differ from other insecticides in placing selective pressure on heavily exposed vector

species, resulting in the eventual emergence of resistant strains, development of resistance is not

expected to emerge quickly.

Following from the success of B.t. H-14, the

search for other pathogenic bacteria has intensified. Several promising strains of Bacillus sphaericus have been isolated and are undergoing evaluation as larvicides.

Although biological insecticides are often assumed to have few, if any, adverse effects on humans, a recent report (305) described a corneal

ulcer in a farmer following splashing of the eye

with a biological agent. BTI was subsequently isolated from the ulcer. This may be an extremely

rare occurrence or early warning of a potential

problem.

Fungi.—Several species of fungi are being applied to vectors of disease in the hope of producing

a lethal fungal infection. Those in the development and field testing stage include Tolypocladium cylindrosporum, Culicinomyces clavor-
120 ● Status of Biomedical Research and Related Technology for Tropical Diseases

sporus, and Coelomornyces spp. An earlier promising agent, Metarhizium anisopliae, has been

found ineffective in the field.

Growth Hormones. —The identification and

testing of insect hormones that alter growth and

reproductive behavior is another area of research.

The most notable example is juvenile hormone,

an essential and naturally produced mediator of

the growth process, which regulates insect development during pre-adult stages.

When applied in excess to insect breeding sites,

juvenile hormone can prevent maturation of larvae and thus function as a biological larvicide.

Safety and efficacy arguments regarding the use

of this hormone as a larvicide are similar to those

surrounding BTI. A drawback to the use of juvenile hormone is that resistance has now been demonstrated in some species, contrary to all earlier

expectations. The high cost of juvenile hormone

has limited its use in public health projects. Furthermore, juvenile hormone is useful only where

Iarviciding is a feasible intervention.

Biological Control Measures

To control arthropod vectors of tropical diseases, various ingenious genetic and biological

control procedures have been attempted. Many

are based on the concept of introducing either sterile males or predators to a vector population, or

by breeding arthropods that are resistant to infection by disease pathogens.

Introduction of Sterile Males

Large numbers of sterile hybrid males or of

males sterilized by irradiation or chemicals can

be released into the vector population. To the extent that sterile males outcompete normal males

for females, the vector population is reduced.

The release of sterile males to control screwworm flies has been a spectacular veterinary success. Unfortunately, however, methods involving the release of sterile males are not practical

for the control of mosquito vectors, nor indeed

for most other medically important arthropods.

Tsetse flies or other vectors with low density or

reproductive potential may be controlled in this

way.

Introduction of Predators and Competitors

The introduction of predators or competitors

in vector populations has been tested a number

of times, with variable results. These methods

cannot be used in a broad-brush manner because

of the many variations in the ecology and epidemiology of vector-borne disease. Only vectors

breeding or existing in habitats suitable to the

predator are susceptible to these methods.

The efficacy of using larvae-eating fish, such

as Gambusia spp. and Aplocheilus spp., to control mosquitoes that transmit malaria is under

study in a number of trials. This method works

well against vectors that breed in large ponds or

rice paddies, and a successful trial in northern

Somalia is being extended (353).

A different tactic involves the use of cannibalistic larvae of the mosquito genus Taeneorhynchus.

These larvae eat other larvae present in their

breeding waters. The method is limited, however,

to those species which share the same breeding

habits as the predator.

The use of nematodes, such as Romanomermis

adicivorax and R. iyengari, is under study. These

worms prey on the immature forms of some insects.

Detection of Species Complexes

Scientists are investigating improved arthropod

vector control using powerful new methods available for assessing genetic differences within vector species complexes. The elucidation of species

complexes allows seemingly identical species to

be differentiated according to their capacity as disease vectors. Once differentiated, highly specific

field interventions may be initiated against the

most capable vector species.

The concept of a species is theoretically simple, centering on reproductive isolation. In the

case of large plants or animals, morphological and

behavioral differences can be fairly easily observed and correlated to infer uniqueness as a species. In the case of many vector species, however,

visually identical specimens may exhibit diametrically opposed behavioral habits (e.g., visually

identical specimens of mosquitoes appear to breed

in both freshwater and saltwater).
Ch. 6—Vector Control Technologies: Selected Tropical Diseases ● 121

Past questions of species identity were frequently resolved through analysis of all stages of the

life cycle (egg, larvae, pupae, adult) using classical taxonomic procedures, which were labor- and

talent-intensive. Early attempts sorted out conflicting behavior patterns by tedious cross-breeding experiments which demonstrated reproductively incompatible populations (populations

whose matings produced either no or sterile

progeny).

Several alternative technologies are now available which, in conjunction with classical taxonomic and ecologic studies, can answer questions

of vector identity. Cytogenetic studies may be performed using photomicrographs of the chromosomes in the salivary glands of insects. (Chromosomes in the salivary glands exist as multiple

intertwined copies which are large enough for isolation, preparation, staining, and photomicography, allowing for mapping and comparison of

banding patterns of the genes.) In addition, isoenzyme electrophoresis has been used to investigate

questions of insect species and strain similarity.

Certain enzymes, common to all related species,

are functionally identical but differ in their chemical composition. The differences can be detected

and used to identify separate species or strains.

This method is being assessed on wild-caught mosquitoes in Tanzania (353). More recently, DNA

hybridization techniques have been developed to

demonstrate whether or not a match exists between the chromosomes of insects whose species

identity is uncertain (94).

Identification of Disease Organisms

Within Vectors

In addition to problems of controlling insect

vectors, there are difficulties in determining

whether and to what extent a vector is harboring

a disease-producing organism.

Immunologic work on malarial sporozoite vaccines (see ch. 7 and case study B) has resulted in

the development of two methods to detect sporozoite forms of the malaria parasite in mosquitoes.

Each species of the malaria parasite (P]asnmfiurn spp. ) has one predominant, unique surface antigen, and monoclinal antibodies (MAbs) against

each of the different species’ antigens have been

produced. Using these MAbs, investigators developed a radioimmunoassay (RIA) and an enzymelinked immunosorbent assay (ELISA) (36,435)

that can sensitively and specifically identify the

presence of sporozoite forms of the malaria parasite in crushed preparations of field-captured mosquitoes (69). These immunoassay replace the previous method of detecting sporozoites, which was

manual dissection of the salivary glands from individual mosquitoes, followed by microscopic examination. (See ch. 8 for a full discussion of these

diagnostic tests. )

The RIA and the ELISA greatly facilitate several types of field investigations and may lead to

better understanding of malaria transmission dynamics and vector control. They will allow field

researchers to: 1) identify new vector species; 2)

determine the relative prevalence of the various

malaria species, specifically those extremely uncommon or clinically or microscopically unrecognized; 3) correlate vector species with transmission of particular malaria species; and 4) quantitate

each vector species’ role in transmission (vectorial

capacity) by its infection rate.

Similar technologies will probably be developed

for the diagnosis of other vector-borne pathogens,

and this will greatly facilitate field research and

surveillance of diseases such as onchocerciasis and

filariasis (155).

Control of Trypanosomiasis Vectors

Two insect vectors against which some progress

has been made are the tsetse fly, conveyor of African sleeping sickness (African trypanosomiasis,

and the reduviid (or kissing) bug, which spreads

Chagas’ disease (American trypanosomiasis).

Tsetse Fly Control

Field research on the life cycle and bionomics

of tsetse flies (Glossina spp. ) has demonstrated differential susceptibility and vectorial capacity

among fly populations. Various techniques of

tsetse fly control have been used over the decades,

including the placement of sticky patches on the

backs of plantation workers, clearing of fly habitat vegetation along river banks, and ground and

air spraying of DDT and other insecticides. As
122 . Status of Biomedical Research and Related Technology for Tropical Diseases

yet, no tsetse fly resistance to insecticides exists,

and in fact, the insecticides are highly effective,

though expensive.

Field trials of tsetse fly traps are under way in

various locations. A trial in Burkina Faso (formerly Upper Volta) achieved excellent success in

reduction of Glossina numbers using a biconical

trap (195,353). A cheaper monoconical trap has

also been developed, which is insecticide-impregnated and attracts flies by odor. Insecticide-impregnated screens and car tires have also been

tested (353).

Reduviid Bug Control

Currently, control of the reduviid bug vector

of Chagas’ disease is the only practical measure

against this disease. Over the long term, improvement of rural housing is the single most important intervention against Chagas’ disease. Poorly

constructed, substandard housing provides both

breeding and resting sites for the reduviid bugs.

Periodic spraying of residences with residual inPhoto credit: Dr. Robert Edelman, National institutes of Health

Reduviid bugs, the vectors of Chagas’ disease, live in

the thatched roofs of houses such as this one.

secticides is one method of control. A recent innovation has been the application of paints and

plasters containing slow-release malathion, which

controlled the vector for up to 1 year.

CONTROL OF SNAIL INTERMEDIATE HOSTS

Although mosquitoes, biting flies, and other

blood-feeding insects are the usual emphasis of

vector control programs, one of the most important tropical diseases, schistosomiasis, involves

snails in the transmission cycle. The snail is actually an intermediate host rather than a true vector, as the parasite is released by the snail into

a body of water and enters through the skin of

a human host when the person comes in contact

with it through bathing or wading.

Controlling snail populations, like controlling

insect vectors of diseases like malaria, is one way

of controlling the spread of schistosomiasis. Snail

control methods are analogous to those used in

insect control, namely alteration of snail habitats

and mollusciciding (direct poisoning of snails).

Artificial lakes and irrigation projects can lead

to the spread of schistosomiasis, through the expansion of habitats for the snail intermediate host

(390). Various environmental or engineering measures have been used successfully for snail control,

such as irrigation canals lined with cement in Japan, and reclamation of swamps in the Philippines. Mollusciciding was one of the interventions

used to reduce the incidence of schistosomiasis on

the island of St. Lucia (see ch. 5). Each approach

is potentially effective, but requires careful management and is expensive.

Molluscicides

Attempts have been made to control snail hosts

since the early 1900s, when the role of snails in

the schistosomiasis transmission cycle was elucidated. A number of chemical compounds have

been used over the decades. Independent of the

agent used, an important concept in snail control

has been the use of “focal mollusciciding” which

concentrates effort on the often localized areas of

actual transmission (58,284). Since only those

areas of known transmission are molluscicided,

focal mollusciciding saves chemicals and personnel. However, good diagnostics and surveillance
Ch. 6—Vector Control Technologies: Selected Tropical Diseases . 123

mechanisms are needed for focal mollusciciding

to be effective. For molluscicides, as for insecticides, both biological and chemical agents have

been used.

Chemical Molluscicides

Two molluscicides are commercially available:

copper sulfate, available since 1920, and niclosamide, available since 1959. Scientists have recently

investigated slow-release compounds, which would

reduce the logistical demands of frequent, periodic applications to snail habitats (176). Organotin compounds, very cheap and shown to be highly effective in laboratory and field trials, are

undergoing research and development (176).

The development of chemical means of controlling snails has been hindered by a lack of information about the biochemistry and metabolism

of the snail intermediate hosts and about the mode

of action of molluscicides. B-2 (a dichlorobromophenol) is a molluscicide used extensively in Japan (353), It has been shown to affect several important glycolytic metabolic steps in Oncomelania

and Biomphalaria, two important genera of schistosomiasis vectors.

Plant-Derived Molluscicides

Plant-derived molluscicides have been investigated as possible alternatives to chemicals for a

number of years, because plant-derived compounds could be less costly and would probably

be less toxic to forms of life other than snails. The

assumption is that plant-derived compounds will

pose less of an environmental hazard than chemicals, and it is possible they could be developed

through village level, self-help schemes (399).

Screening for potential molluscicides has concentrated on plants with known medicinal properties. Many of these have been identified as containing active molluscicidal compounds. The most

thoroughly studied is Phytolacca dodecandra,

whose berry extract (ended) has been used in Ethiopia. Other promising species are Ambrosia maritima and other species of that genus, and Anacardium occidentals.

The search for effective plant-derived molluscicides has been impeded by the lack of knowledge about snail metabolism and physiology. Currently, the Special Program for Research and

Training in Tropical Diseases (TDR) is promoting the development of screening methodology

and developing goals for the use of plants (353).

Biological Control Measures

The most promising biological agents for controlling the snails involved in the transmission of

schistosomiasis appear to be competitor snails,

such as Marisa cornuarietis, which has been used

in Puerto Rico. In the course of browsing over

vegetation, M. cornuarietis eats all stages of the

snail Biomphalaria spp. Other competitor snails

under study are Pomacea, Thiara, and Helisoma

Spp. (176,353).

Detection of Species Complexes

As is true in the case of many insects, susceptibility to infection in the case of snails varies between species and strains. Isoenzyme electrophoresis

has been developed for the detection of species

complexes in various snail populations (173,174)

and used experimentally to identify snail populations with differing susceptibility to schistosome

infection (227). Knowledge of susceptibility will

increase our understanding of the epidemiology

of schistosomiasis transmission.

INTEGRATED PEST MANAGEMENT (1PM)

A new emphasis is being placed on an inte- their environment—with biological and chemical

grated approach to vector control based on 1PM control agents. In contrast to single-tactic prostrategies. 1PM strategies emphasize the need for grams based on chemical insecticides, 1PM stresses

combining basic field studies of vector bionom- multifaceted environmental measures reduce disics—the study of the interaction of vectors with ease transmission. In the Panama Canal Zone, an
124 . Status of Biomedical Research and Related Technology for Tropical Diseases

IPM-type program eliminated malaria; then DDT

was added to the control program, and screening and drainage practices were neglected. The

single-tactic approach failed when the mosquitoes

became resistant to DDT, and malaria returned.

Theoretically, 1PM requires an intimate knowledge of all factors relating to vector biology and

disease transmission. Critical factors are selected

and monitored for change, and tailored intervention techniques are applied as needed to control

the vector. Successful 1PM models are based on

the analysis of factors such as weather/climate,

vector density, pathogen infection rate, and other

RESEARCH NEEDS

Insecticides and Molluscicides

The search for cheap, safe, and effective insecticides and molluscicides, whether of chemical or

biological derivation, characterizes research in all

vector-borne diseases. The lack of an obvious

profit element greatly inhibits commercial development of any insecticide having only public

health applications. However, agricultural pests

remain a subject of intense research, and there will

quite likely be benefits for public health problems.

Two methods are used in developing new insecticides. In the first case, knowledge about toxic

substances is used to test known and newly synthesized compounds against a range of target species. In the second case, knowledge is developed

about the physiology and biochemistry of target

species in order to identify critical enzyme pathindicators that are of predictive importance for

disease transmission. In practice, 1PM has sometimes meant nothing more than using several insecticides in combination, instead of just one.

Paradoxically, in many vector control projects,

intuitive intervention may lead to counterproductive results (329). For example, insecticide spraying may reduce the population of larvae in a

breeding site, but have the unexpected result of

producing more robust adults, better able to transmit disease, because of reduced competition for

food.

ways or other life functions against which insecticides could be made. Further research is needed

in either instance.

Vector Bionomic Studies

There is a continuing need for better understanding of the biology of all disease vectors: identification of new vectors, vectorial capacity, physiology, genetics, insecticide susceptibility, behavioral

characteristics such as biting, resting, and breeding habits, and any other factors that contribute

to the maintenance and transmission of disease.

Improved means of detecting disease-causing

agents within vectors is an important area in need

of research and development. There is a vital need

for increased numbers of trained personnel at all

levels of research (253).

SUMMARY

Attempts to control arthropod vectors and in- behavior and natural history of vector species was

termediate hosts such as snails are relatively re- encouraged by the requirements of physical concent, proceeding from the discovery of their role trol methods. After World War II, the old methin human disease transmission around the turn ods were largely abandoned in favor of synthetic

of this century. For several decades, physical chemical insecticides, whose promise was to eradmethods were the only means available for con- icate vector species, particularly the mosquito vectrolling vectors of tropical diseases. Study of the tors of malaria.
Ch. 6—Vector Control Technologies: Selected Tropical Diseases ● 125

The field of vector control is slowly emerging cine, will be sufficient by itself to reduce signififrom total reliance on chemical control methods cantly the prevalence of vector-borne tropical

toward 1PM (integrated pest management). 1PM, diseases. To control vector-borne tropical disat least theoretically, includes chemical, biologi- eases, several methods adapted to local conditions

cal, and physical control methods, once again re- are probably necessary.

quiring more intimate knowledge of the lives of

vectors, now including knowledge of their genetic

and biochemical characteristics. 1PM is as yet a

new field, but one in which there is a great potential to influence the occurrence of disease in

developing countries.

Public health authorities are beginning to emphasize basic vector control engineering measures,

such as drainage, filling, and control of water bodies. However, these physical measures alone are

not sufficient for disease

methods are developed,

It is unlikely that any new medical interven- practical alternatives to

tion, whether a chemotherapeutic drug or a vaccontrol, and unless new

there will be few if any

insecticides.
7.

Immunization Technologies:

Selected Tropical Diseases
Contents

Introduction . .

Immunity and

Immunity . .

Vaccines . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Vaccines: Background . . . . . . . . . . . .

. . . . . . . . .

. . . . . . . . .

Vaccination and Vaccine

. . . . . . . . .

Research;

Selected Tropical Diseases . .

Malaria . . . . . . . . . . . . .

Schistosomiasis . . . . .

Trypanosomiasis . . . .

Leishmaniasis . . . . . . .

Filariasis . . . . . . . . . . .

Leprosy . . . . . . . . . . . .

Tuberculosis . . . . . . . .

Diarrheal and Enteric

. . . . . . . .

. . . . . . . .

. . . . . . . .

. . . . . . . .

. . . . . . . .

. . . . . . . .

. . . . . . . .

Diseases

Acute Respiratory Infections

Arboviral and Related Viral

Summary . . . . . . . . . . . . . . . . . .

. .

. .

. .

. .

. .

. .

,.

. .

. .

. .

. .

. . . . . . . . . . . . . .

. . . . . . . . . . . . . .

Current Status

. . . . .

. . . . .

. . . . .

. . . . .

. . . . .

. . . . .

. . . . .

. . . . .

. . . . .

. . . . .

Infections

. . . . . . . . .

. . . . . . . . .

. . . . . . . . .

. . . . . . . . .

. . . . . . . . .

. . . . . . . . .

. . . . . . . . .

. . . . . . . . .

. . . . . . . . .

. . . . . . . . .

... . . . . .

. . . . . . . . .

. . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

f o r

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . ..,,, .

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . ,.

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . .

FIGURE

Figure No.

7-I. Methods Used To PreDare . Subunit Vaccines Against Viral Diseases:

The Recombinant DNA Method and Chemical Synthesis Method . . . . . . . .

Page

129

129

129

131

134

134

137

138

140

140

140

142

142

144

148

149

Page

133
7.

Immunization Technologies:

Selected Tropical Diseases

INTRODUCTION

Vaccines exist for many of the viral and bacterial diseases that are important in developing

countries, but there are no vaccines against the

parasitic diseases that are generally associated

with the tropics. Antiparasite vaccines are more

difficult to formulate than vaccines for viruses and

bacteria, the reason being that protozoan and helminthic parasites are more complex in biochemical and physical composition, life cycle stages, and

interactions with their hosts. The immunology of

parasitic diseases is in a vigorous phase of research

(66,164,213,241,273) and forms the underpinning

for attempts at vaccine development.

The available vaccines vary in the protection

against disease that they induce and the risks of

adverse effects in individuals who are vaccinated.

Vaccines against yellow fever, measles, rubella,

and mumps, for example, are almost always effective and provide for long-term protection.

Others—influenza, pneumococcal pneumonia,

cholera, and typhoid, for instance—do not induce

immunity in as high a proportion of those vaccinated and may last only a matter of months or

a few years. Both new vaccines against diseases

currently not immunizable and improvements in

existing vaccines are needed.

There is great optimism in vaccine research today. Large strides in biotechnology, particularly

in the use of monoclinal antibodies (MAbs) and

recombinant DNA, and a rapidly growing body

of knowledge in immunology are aiding the field

of immunization technology. The development of

replacement vaccines for viral and bacterial diseases that are safer, more effective, and possibly

less expensive than vaccines currently available,

and the development of new vaccines against

some tropical diseases, including parasitic diseases

for which no vaccines now exist, are advances that

now depend more on research with the tools already at hand than on major technological breakthroughs (310).

The single most visible, exciting development

in vaccines today is that a vaccine against malaria

may be ready for widespread use in a matter of

a few years. In 1984, progress could be followed

almost weekly, as the immunologic properties of

various life-stages of malaria parasites were characterized in laboratories in different parts of the

world and the information was processed for practical significance to a vaccine.

IMMUNITY AND VACCINES: BACKGROUND

Immunity

The term “immunity” refers to the capacity of

an organism to resist a particular disease. Diseasecausing organisms themselves require certain conditions for survival, and only certain potential

host organisms will provide those conditions.

Thus, as a species, humans are susceptible to a

unique set of pathogens that differs from the set

of pathogens that cause disease in other species,

although there are areas of overlap (zoonoses).

“Species immunity” does not stem from a response

of the potential host’s immune system against the

disease, but simply from an inability of the disease organism to become established. From an

evolutionary point of view, such immunity is important, but from the standpoint of preventing

diseases and developing vaccines against them,

129
130 • Status of Biomedical Research and Related Technology for Tropical Diseases

the important kind of immunity is that which individuals have or can develop against diseases that

do occur in human beings.

The immune status of an individual is built

from a complex of mechanisms, some genetically

determined and some acquired during life, which

we are just beginning to understand. A person

with complete immunity will not get a disease

when exposed to the causative agent. The converse is complete susceptibility, which means that

anytime the person comes in contact with the

causative organism, the disease will develop. Between solid resistance and complete susceptibility, a range of states of partial immunity and partial susceptibility exist.

“Innate immunity” is genetically determined at

birth. Many people of African descent are resistant to Plasmodium vivax malaria because they

lack a protein on their red blood cell surface that

normally allows the malaria parasite to bind to

the red blood cell. Many people in endemic malaria regions are resistant to P. falciparum malaria because they carry the sickle-cell or thalassemia trait. Genetically determined aberrant types

of red blood cells cause sickle-cell anemia in individual who inherit the aberrant trait from both

parents (homozygotes), but prevent malarial infection in individuals who have inherited the trait

from only one parent (heterozygotes).

“Acquired immunity” is immunity that results

either from the body’s response to exposure to disease or from vaccination (“active immunity”) or

from the transfer of antibodies from another person or from animals (“passive immunity”). Active and passive immunity are discussed further

below.

Active Immunity

“Active immunity” is immunity acquired by an

originally susceptible individual by effective contact with an infectious organism (or a closely related species) or its products. Such immunity results when a biochemical entity known as an

antigen (which may be on, in, or produced by the

infectious organism) stimulates a response by the

immune system of the host.

Active immunity is the result of two general

types of stimulation of an individual’s immune

system:

● humoral immunity: immunity resulting from

the production by antigenically stimulated BIymphocytes (specialized white blood cells)

of nonliving proteins called “antibodies” (or

immunoglobulins) that circulate in the blood;

and

● cell-mediated immunity: immunity resulting

from increased activity by living cells in the

blood and other tissues (e.g., T-lymphocytes,

macrophages/monocy tes, eosinophils, mast

cells, and natural killer cells) that directly and

nonspecifically destroy foreign material in

conjunction with antibodies.

Active immunity often is a result of natural exposure to an antigen that results in subclinical or

clinical infection (e.g., chickenpox infection), but

it may be induced by vaccination (e. g., measles

or polio vaccination).

Vaccines usually stimulate humoral immunity,

i.e., antibodies against a specific antigen, but some

immunizing agents strengthen cell-mediated immunity. An example of the latter is Bacillus Calmette-Guerin (BCG) vaccine, a weakened or attenuated tuberculosis vaccine. Whether stimulated

by natural infection or vaccination, active immunity usually takes some weeks to develop and may

thereafter be lifelong, as with yellow fever or

smallpox immunization, or may wane after a variable period, as with typhoid fever.

Passive Immunity

“Passive immunity” is immunity conferred on

an individual by the direct transfer to that individual of antibodies produced by another individual. It may result from direct transfer of immune serum that is obtained from individuals with

acquired immunity against a specific infectious

agent. Large injections of immune serum globulin against hepatitis (“gamma globulin”) are sometimes given to Western travelers to the tropics.

Newborns acquire certain antibodies passively

through the placenta from their mothers and also

receive some protection through antibodies in
Ch. 7—immunization Technologies: Selected Tropical Diseases ● 131

breast milk. The protection offered by passive immunity is relatively short-lived, usually disappearing within a period of weeks. The transfer of antibodies does nothing to stimulate the body’s

immune system to produce its own antibodies.

The use of MAbs to confer passive immunity

against experimentally attempted infection (“challenge”) is important for laboratory investigations

and for establishing the theoretical possibility of

inducing immunity, but probably will not become

a major tool for disease control.

Vaccines

Vaccines are used to produce active immunity.

In all cases, a vaccine contains some biochemical

compound whose structure is the same as, or similar to, some part or product of the infecting organism. Introducing that foreign material (antigen)

stimulates the host’s immune system to produce

humoral and cell-mediated immunity. Following

vaccination, the host’s immune system is primed

to attack the organism more quickly and efficiently should it ever reinvade the host’s body.

Most tropical diseases are chronic infections

(e.g., malaria, schistosomiasis, leprosy) in which

the proliferation of the pathogen does not exceed

a certain level and an equilibrium of sorts is established between pathogen and host. The immune

system of the host reacts to the parasite, yet the

parasite is not completely eliminated because it

has various methods of evading the immune system. The equilibrium reached is “beneficial” to the

parasite: the host’s body is not overwhelmed or

killed by the infection. Parasites evade or subvert

the immune system in many different ways. They

can lose the surface antigen by which they are recognized by the host, take up host proteins as a

surface coat, shed and generate altered surface

antigen over time, or internalize in the host’s cells,

The challenge of current immunology is to make

the immune system produce an effective response

against the parasite.

A further complexity is that the immune response itself often causes the pathology of the disease. For example, the immune response to schistosome eggs in the host’s tissue causes the clinical

symptoms of schistosomiasis. In eliciting a protective response by the host’s immune system, it

is important to avoid stimulating a response that

causes immunopathology.

Types of Vaccines

The production and composition of vaccines

differ depending on the disease-producing organism and the available knowledge about it. It is

no accident that most currently available vaccines

are for diseases caused by viruses and bacteria,

which have limited variability in their structure,

biochemistry, and genetic makeup. Most of these

vaccines were created through empirical work,

often with very imperfect understanding of the

underlying biochemistry and molecular biology.

Conventional Vaccines. —The commonly produced vaccines are derived from actual disease

organisms (or closely related organisms) grown

in a culture medium, embryonated eggs, or in cell

culture. Despite great efforts at quality control,

such biological products are always subject to

some variation. Potency can vary, improper inactivation may make a virulent vaccine, and organisms may change or mutate in culture, reverting

to or developing pathogenicity. Another potential problem is contamination. For example, some

early batches of polio vaccine grown on monkey

kidney cell cultures were contaminated by a monkey virus. In some vaccines, there maybe latent,

undetected viruses whose effects may not be evident for years.

The major types of conventional vaccines are

the following.

1. Live infectious organisms: The infectious

agent is obtained from someone who has the disease. For instance, scrapings from lesions of cutaneous Ieishmaniasis have long been used in endemic areas for deliberate inoculation of children

to prevent the possible appearance of disfiguring

facial lesions later in life (168).

2. Closely related organisms that stimulate

cross-immunity: Jenner’s discovery in the 18th

century that inoculation with cowpox could prevent smallpox marked the beginning of modern

vaccination.

3. Live, attenuated organisms: Cultured organisms are weakened in various ways so that they
132 ● Status of Biomedical Research and Related Technology for Tropical Diseases

will remain infectious and stimulate immunity but

will not cause disease. Polio, measles, rubella, and

BCG vaccines are of this type. Among the problems with this approach are the lack of assurance

that the attenuation procedures really eliminate

pathogenicity while maintaining appropriate antigenicity; the possibility of mutational reversion

to virulence; the introduction of pathogen genomic segments into host DNA; and a small but measurable incidence of severe side effects such as encephalitis. The antiparasite vaccines in use in

animals are live, attenuated organisms, the most

successful of which are the radiation-attenuated

bovine lung-worm larvae employed in some countries since 1959 (26). Field trials have also been

reported with irradiated larval schistosomes in

cattle (338).

4. Killed (inactivated) organisms: By heat, formalin, or other methods, the disease-producing

organism is inactivated without destroying its immunogenic potential. Typhoid, pertussis, plague,

and Rocky Mountain spotted fever vaccines are

examples of vaccines produced in this way.

S. Products of bacteria: When a specific biochemical product of bacteria produces the disease,

this toxin may be altered to a nonpathogenic toxoid, which can be used to immunize. Tetanus and

diphtheria vaccines are toxoids.

6. Parts of organisms (subunit vacanes): The

purified pneumococcal or meningococcal’ capsule

polysaccharide subunit vaccines are prepared by

isolating an immunogenic portion of the outer surface of the pneumococcal or meningococcal bacterium. Isolated polysaccharides are only weakly

immunogenic, especially in children. Efforts to increase immunogenicity by coupling polysaccharides to carrier proteins are in progress. The term

“subunit vaccine” also is used to describe vaccines

made from subunits of bacterial toxins (e.g., the

subunit-b vaccine for cholera).

Synthetic Subunit Vaccines.—Much current

work is directed toward the development of vaccines consisting of uniquely specific antigenic molecules of completely known structure. It is hoped

that such materials, of uniform high quality, can

be synthesized in production volumes, providing

inexpensive, safer, and more effective products.

1. Recombinant DNA products: Three strategies using recombinant DNA technology are useful in synthesizing vaccines:

a.

b.

c.

Antigenic substances that stimulate protective immunity are identified or extracted

from the pathogenic organism. Appropriate segments of pathogen DNA, coding for

production of those antigens, are spliced

into the DNA of another organism. In some

cases, the host organism is a yeast or bacterium which thereafter synthesizes the

desired molecule. The protective antigen is

then separated from other materials in the

culture and prepared as a vaccine (see fig.

7-l).

The host organism (from a above), if nonpathogenic, can be used to infect the vaccinee, who then produces an immune response against the harmless carrier organism

and the “piggy-backed” antigen. The nonpathogenic carrier organism can be either

a related strain of the disease-producing

DNA donor or an unrelated species.

A pathogenic organism can be “attenuated”

by deleting the specific gene causing disease,

thereby rendering the organism nonpathogenic but still antigenic.

2. Total synthetic antigens: If a specific, defined

protein or polypeptide (amino acid chain) is determined to induce protective immunity, scientists may be able to produce this antigen in pure

form by a total chemical synthesis (see fig. 7-1)

and use it to induce an immune response (314).

3. Anti-idiotype vaccines: In order to make

anti-idiotype vaccines, scientists produce a passively protective MAb against the disease organism, then produce a second MAb against the first

MAb. The second MAb (in effect, an “anti-antibody”) may then be used as an “antigen” to stimulate production of the original protective antibody in a host. This application of MAbs would

be most significant where an immunogenic protective antigen is in short supply and not amenable to synthesis by recombinant DNA or chemical techniques.
Ch. 7—Immunization Technologies: Selected Tropical Diseases ● 133

Figure 7-1 .—Methods Used To Prepare Subunit Vaccines Against Viral Diseases:

The Recombinant DNA Method and Chemical Synthesis Method

Chemical Synthesis Method

Viral nucleic

acid

! I Determine

\

Chemically

synthesize the

protein and

purify vaccine

r----- - *

Extract

protein /

Vaccine

In the chemical synthesis method, proteins that comprise the viral surface are isolated, often with the use of monoclinal antibodies (MAbs).

The protein sequence is then determined. Based on the sequencing information, large amounts of the protein or portions of the protein are

made chemically for use as the vaccine.

In the recombinant DNA method, the gene that encodes the viral surface protein is isolated and cloned into an appropriate vector (e.g., plasm id),

transformed into a host (e.g., a bacterium or yeast), and the host is grown in large quantities. Formation of the protein by the recombinant DNA

and isolation of the protein results in the subunit vaccine.

SOURCE: Adapted from Office of Technology Assessment, Commercial Biotechnology: An International Analysis, 1984
134 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Culture of Disease-Producing Organisms

Major steps toward producing vaccines against

tropical diseases have been made possible by recent advances in the culture of disease-producing

organisms. A method for in vitro cultivation of

P. falciparum (343), for example, removed a

major roadblock to further research in malaria,

namely, the requirement for maintaining this

organism in animal hosts. Other advances include

success in cultivating in vitro the complete life cycle of Trypanosoma brucei, which causes African sleeping sickness (156), and the discovery that

armadillos are an excellent culture medium for

Mycobacterium leprae, which causes leprosy.

These technical advances have increased the availability of experimental material for research use,

but further advances are still needed.

VACCINATION AND VACCINE RESEARCH:

CURRENT STATUS FOR SELECTED TROPICAL DISEASES

Malaria

The development of a vaccine against malaria

is one of the great challenges that is now being

addressed. Steady progress over the past decade

has encouraged the scientific community, and

hopes for an effective vaccine within the next decade have been voiced. A detailed account of malaria vaccine development to the present appears

in Case Study B: The Development of a Malaria

Vaccine.

Human malaria is caused by four species of protozoan parasites of the genus Plasmodium: P. falciparum, P. vivax, P. ovale, and P. malariae.

Other species of Plasmodiurn are infective to a

wide variety of vertebrates other than humans

(e.g., P. berghei, P. yoeli, and P. chabaudi to rats

and mice, and P. knowlesi to rhesus monkeys).

Malaria parasites (Plasmodium spp. ) have a

complex life cycle (see ch. 4) with several distinct

life-stages that could provide points of intervention for vaccines. One of the primary targets of

current malaria vaccine research efforts is the

sporozoite life-stage. Plasmodial sporozoites are

introduced into a vertebrate host’s bloodstream

during the bite of an infected Anopheles mosquito. Once in the host’s bloodstream, sporozoites

move within a short time to infect the liver cells,

initiating malaria infection. For a sporozoite vaccine to be effective in preventing malaria in humans, every sporozoite would have to be killed.

Another target of malaria vaccine research are

the erythrocytic or blood stages-either schizonts

within red blood cells or free merozoites—of the

malaria parasite. Merozoites, the stage most directly responsible for the symptoms of human malaria, develop from sporozoites in the infected

host’s liver cells (exoerythrocytic cycle), burst out

of the liver cells, enter the host’s circulatory system,

and invade the person’s red blood cells (erythrocytes). There they undergo asexual reproduction,

forming schizonts, which release more merozoites

to infect more red blood cells (erythrocytic cycle). In chronic infections, this cycle may be repeated many times. Merozoites are the main target of blood stage vaccine research.

A third target of malaria vaccine research is the

gamete, the sexual stage of the parasite. Gametes

mature in the mosquito from gametocytes that are

picked up when the mosquito bites an infected

person. If the person had been vaccinated against

the gamete stage, the mosquito would also pick

up antibodies circulating in the host’s blood

formed against the gamete. In the mosquito, the

human antibodies would prevent the gametes

from completing their life cycle, and thus prevent

the mosquito from spreading malaria. Since the

gamete vaccine would not protect the vaccinee

from acquiring malaria, this type of vaccine is

called an “altruistic” vaccine.

Two other potential vaccination targets are the

infected liver cells and the infected red blood cells,

whose surface membranes may be altered by the

presence of parasites in the cell.

Most evidence indicates immunity is stage-specific in malaria. A vaccine against merozoites re-
Ch. 7—immunization Technologies: Selected Tropical Diseases ● 135

leased from red blood cells, therefore, will not

control exoerythrocytic merozoites released from

liver cells (291). Successful immunization in human populations will probably require a combined vaccine against several stage-specific determinants, although a monovaccine against the

sporozoite stage may have value to short-term visitors to areas where malaria is endemic.

Early Attempts at Immunization

Experimental malaria immunization was achieved

using sporozoite, merozoite, and gametocyte vaccines during the late 1960s to mid-1970s.

Sporozoite Stage.—Immunization with radiation-attenuated sporozoites was shown to give excellent protection in rodents, birds, and human

volunteers (19,61,62,261,262). One problem with

this sporozoite vaccine is the difficulty of obtaining enough antigenic material in a sufficiently purified form to immunize even selected populations.

Erythrocyte Stage. —Vaccination with merozoites was achieved in rodents, birds, and nonhuman primate models using crude whole parasite antigens or partially purified antigens, but the

use of adjuvants (nonspecific stimulators that enhance the immune response) was required (65,67,

68,241,242). (Because of side effects, especially tissue death at the injection site, at least some adjuvants cannot be used in humans. ) Vaccination of

rhesus monkeys with merozoite preparations of

P. knowlesi in Freund’s complete adjuvant conferred complete protection against an otherwise

fatal infection. Immunity to blood stage parasites

was also induced in Aotus monkeys with P. falciparum and P. chabaudi in mice. Various antigen preparations were used in these experiments:

whole blood-stage P. falciparum parasites with

adjuvant; merozoites obtained by “natural release” methods; and glutaraldehyde-fixed and

freeze-thawed merozoites. Purified antigens of P.

yoeli merozoites induced protection in mice. Rhesus monkeys were also vaccinated with an immunogenic protein isolated from the membrane of

P. knowlesi-infected red blood cells.

Gamete Stage. —Gamete vaccination was demonstrated in chickens and rhesus monkeys. The

mechanism of action is transfer (when the mosquito feeds) of antigamete antibodies produced

in the animal host which act on the gametes in

the mosquito gut (46,142).

Cultivation of Plasmodium

Although the possibility of protective immunization against malaria had been established by

the mid-1970s, major progress towards producing vaccines required advances in the culture of

malaria parasites.

In 1976, Trager and Jensen (343,344) described

a method for in vitro cultivation of P. falciparum

blood stages that removed a major roadblock to

further research in malaria. Previously, malaria

research had depended on maintenance of the

parasite in animal hosts. This situation was unsatisfactory for several reasons: 1) it is expensive

to keep animals; 2) large-scale production and isolation of the parasite is usually difficult; 3) biological phenomena of the host affect the malaria

parasite and contaminate and increase the variability of any preparations.

In vitro cultivation of P. falciparum has become

standard laboratory procedure to provide researchers with sufficient parasite material to perform

other laboratory research on malaria. Because research on P. falciparum is most urgent (because

of fatal consequences in P. falciparum infection),

it was fortuitous that cultivation of P. falciparum

erythrocytic stages proved easier than other malaria species. Success in cultivating merozoites of

the other human malaria species is still elusive.

The next stage of the malaria parasite to be cultivated in vitro was the gametocyte, which when

fed to mosquitoes through artificial membranes,

led to the development of infective sporozoites

(42). This was important for subsequent work on

the sporozoite antigen because it ensured adequate

amounts of material for study.

Liver stages of the parasite are now also being

cultivated in vitro. The complete exoerythrocytic

cycle of P. berghei (a mouse malaria) has been

cultivated in the laboratory (158) (in a cell line

of human embryonic lung cells). Attempts to cultivate the exoerythrocytic stages of monkey and

human malaria are in progress. The tissue culture

system P. berghei provides an in vitro assay to

test for protective antibodies. It also provides a
136 ● Status of Biomedical Research and Related Technology for Tropical Diseases

model for study of possible chemotherapeutic

compounds.

Recent Research

Sporozoite Vaccine Research.—Using the P.

berghei mouse malaria model and the in vitro tissue culture system, scientists have made important progress in the study of protective sporozoite

antigens (63,422). The surface of the mature

sporozoite is covered by a major species- and

stage-specific membrane protein, the “circumsporozoite protein, ” which for P. berghez” is named

Pb44 for its molecular weight of 44,000. Synthesis of the circumsporozoite protein is a major

metabolic activity of the mature sporozoite (immature forms of the sporozoite isolated from the

mosquito midgut do not have this circumsporozoite protein, nor do later erythrocytic stages).

A very small quantity of purified MAb to Pb44

protects mice against sporozoite challenge (passive immunity). If sporozoites are incubated in

vitro with the complete MAb or the antibodyspecific portion of the MAb known as the “Fab”

fragment, they lose their ability to infect liver cells

(279). The antibody prevents attachment to, and

penetration of, the mammalian target cells by P.

berghei sporozoites in vitro (422). Thus, immunity to sporozoites in mice depends on binding

of antibody to the malaria surface protein, preventing interaction with target cells.

Sporozoites infect liver cells by first attaching

to a receptor on the cell membrane and then penetrating by means of a process involving movement

between parasite and host cell (3). Attachment requires a protein receptor, and penetration requires

movement of host cell components. Irradiated

sporozoites attach, enter, and transform to the

next liver stage (trophozoites), but do not develop

to the mature liver (schizont) stage.

MAbs, which permit the recognition and isolation of individual antigens in pure form, have

identified protective antigens from several species

of Plasmodium (P. knowlesi, P. chabaudi, P. falciparum, P. vivax), including polypeptides analogous to Pb44 (64,89,431).

Building from the availability of sufficient antigen and a suitable model, scientists have now applied recombinant DNA technology to the production of pure protective sporozoite antigens;

biosynthesis of the antigen by expression in Escherichia coli of the gene coding for protective

sporozoite surface antigen has been achieved

(109,432). The immunogenic region of the antigen has been identified and chemically synthesized

(134,434). This work has been reproduced with

the P. falciparum gene. Recently, the antigen

produced by this process has been shown to be

protective against P. falciparum challenge.

Erythrocyte Vaccine Research.—Erythrocyte

antigens are being studied in rodent and primate

malaria models and in P. falciparum. This research has been successful in producing scientific

knowledge, but the implications for a blood stage

vaccine are not yet clear.

Species- and stage-specific antigens have been

identified on the surface of malaria-infected red

blood cells (P. chabaudi, P. knowlesi, P. falciparum, and MAbs to at least one antigen showed

protective activity (P. chabaudi). Membrane antigens of P. knowlesi and P. falciparum have been

compared; clinical immunity to P. knowlesi correlated with the presence of antibody to a particular molecule (MW 74,000), which was then

isolated and used to immunize monkeys. Antibody to this protein inhibited in vitro multiplication. Thus, there is encouraging evidence that

a parasite-derived antigen(s) is at the erythrocyte

surface, and it stimulates protective immunity. Recombinant DNA technology is being applied in

several laboratories to produce protective erythrocyte stage antigens (16).

However, antigenic variation has been shown

in P. falciparum in Aotus monkeys (159,160).

Strain-specific determinants on the surface of red

blood cells infected with late-stage parasites were

recognized by immune serum. These determinants

varied according to host factors. Variant antigens

can be identified by surface immunofluorescence,

and protection is correlated with the presence of

antibody to surface determinants. MAbs against

geographic isolates of P. falciparum can distinguish between isolates, and some can block merozoite invasion of red blood cells.

These findings imply that functionally important antigens, which appear to protect by induc-
Ch. 7—immunization Technologies: Selected Tropical Diseases ● 137

ing antibodies, do exist in the erythrocytic stages,

but differ between P. falciparum isolates, and that

P. falciparum has the ability to vary surface antigens in response to the body’s immune response.

The possibility of developing a protective immunization against the merozoite stage appears to

be rather poor, if the antigen that stimulates protective antibodies varies not only over place, but

also over time. On the other hand, if there is one

or more determinant common to all or most strains

of the parasite, in spite of the variability, then a

vaccine may be feasible.

One fact that may be exploited for a merozoite

vaccine is that merozoites recognize receptor “gly -

coprotein” molecules on red blood cells. These

glycoproteins play an indirect role in the invasion

of red blood cells.

Other Immunologic Research. -Mechanisms of

immunity are being investigated, and greater understanding of cell-mediated immunity (T-cells,

macrophages, natural killer cells) has been

achieved. Generalized microphage activation, induced by substances unrelated to malaria, provides some protection to mice from P. berghei infection, perhaps by increasing phagocytosis of

parasitized red blood cells (334).

Researchers at the National Institutes of Health

(NIH) have demonstrated that P. falciparwninfected red blood cells attach to the cells that line

blood vessels (endothelial cells) and to cells of a

particular melanoma (skin cancer) cell line. This

is confirmation of the longstanding hypothesis

that P. falciparum somehow differs from the other

species by being “sticky” and thus adheres to and

clogs the blood vessels. This in vitro model allows further investigation of this phenomenon,

especially in relation to the pathogenesis of cerebral malaria. The implication for vaccine research

is that somehow the surface of malaria-infected

red blood cells are different from normal red

blood cells, a difference which perhaps can be exploited to selectively eliminate these cells.

Tropical splenomegaly syndrome (TSS), a complication of malaria infection, found especially in

children in endemic areas, is being studied, and

evidence obtained indicates that antibody responses in TSS are specific to malaria. Since TSS

may be some type of immune disease brought on

by the body’s response to infection, it must be

considered and evaluated in the development of

a vaccine.

The immunosuppressive effect of extracts of

malaria parasites is being investigated with the P.

berghei model. Certain components of malaria

parasite chemical extracts have been shown to

suppress T-cell-dependent immune responses to

other pathogens. This finding may help to explain

why malaria infection lowers the body’s immunity to other infections and reduces the immune

response to immunization for other diseases (e.g.,

measles vaccine). Further research in this area is

needed.

Schistosomiasis

Schistosomiasis affects about 200 million people in parts of Africa, the Caribbean and South

America, and the Orient. The major schistosomes

infecting humans (Schistosoma mansoni, S. japonicum, and S. haematobium) have life cycles

requiring certain freshwater snails as intermediate hosts.

Immunologic Research

Studies of schistosome infection using immunologic techniques have provided much new information of relevance to immunology itself, such

as the first knowledge about the biological function of eosinophils (a type of white blood cell) and

one of the immunoglobulins. In fact, it is only a

slight overstatement to say that schistosomiasis

has done more for immunology than vice versa.

The eosinophil is one of several types of white

blood cells involved in cell-mediated immunity.

The schistosome model is the most extensively

studied laboratory system with respect to eosinophil

involvement in immunity. There is mounting evidence that acquired resistance in schistosomiasis

involves an antibody-dependent (humoral) mechanism that somehow activates the eosinophil to

kill the foreign organism (19). The schistosomulae (the earliest life-stage of the schistosome in the

body of the human host) appears to be open to

attack by two mechanisms: 1) schistosomulae are

coated with IgG antibody and then are attacked

by eosinophils (39); and 2) IgE antibody-coated
138 ● Status of Biomedical Research and Related Technology for Tropical Diseases

schistosomulae are attacked by activated macrophages, specialized cells of the immune system

(44).

In addition to functioning at sites of skin penetration by schistosome larvae, eosinophils predominate in granulomatous lesions which develop

around schistosome eggs entrapped in host tissues.

The egg granuloma has been shown to be a principal factor in the pathogenesis of liver and spleen

involvement in schistosomiasis, and histologic

studies have indicated that the eosinophils destroy

schistosome eggs. If eosinophils are eliminated by

administration of anti-eosinophil serum, there is

marked reduction in size of egg granulomas and

concomitant decrease in pathology. This is an example of pathological damage to the human host

arising from the host’s own immune function.

Many laboratories have produced antischistosomal MAbs for identification of relevant antigens or for species identification (e.g., 83,84,99,

234,236,332,336). Dissous and colleagues (96)

have reported protection against S. Mansoni challenge in rats achieved by passive transfer of a rat

antischistosomal MAb. This MAb recognized an

antigen on the surface of immature worms and

a different antigen on adult worms, both antigens

sharing a common epitope. Other investigators

have also reported passive protection against S.

rnansoni challenge, using monoclonals in mice

(320,436).

Current Status

A live irradiated larval vaccine for schistosomiasis has been administered to cattle in the Sudan with encouraging results (338), but it is not

acceptable for use in humans (because the larvae

must infect the host, even though they do not mature to egg-producing adults).

Prospects for a subunit vaccine for schistosomiasis are not clear. Immunologic research has

demonstrated the complexity of the immune response to the parasite and the parasite’s protective responses. Developing schistosomes acquire

host antigen on their surface, which masks them

from the host immune response (130). The adult

worm is largely protected from immune attack by

acquisition of host antigen, shedding of antigen,

and/or antigenic variation. The adult schistosome

worm can survive for many years, apparently immune to host attack, yet stimulating immunity

against further infection by larval schistosomes,

thus becoming a chronic infection. Most evidence

points to the early schistosomulae (the first immature infective stage) as the most likely point

of attack for a vaccine.

Trypanosomiasis

Trypanosomiasis is a group of clinically differing diseases of the blood and tissues caused by

several species of the genus Trypanosozna. The

important human diseases are: African sleeping

sickness (African trypanosomiasis), an acute form

caused by T. brucei rhodesiense, and a chronic

form caused by T. b. gambiense; and Chagas’ disease (American trypanosomiasis) caused by T.

cruzi.

African Sleeping Sickness

(African Trypanosomiasis)

Current Status. —Currently, there are no vaccines against African sleeping sickness. One of the

larger research efforts in vaccination for African

trypanosomiasis is being carried out at the International Laboratory for Research on Animal Diseases (ILRAD) in Nairobi, Kenya, in an effort to

prevent a similar disease in cattle (nagana) that

has a devastating economic impact. The Special

Program for Research and Training in Tropical

Diseases (TDR) has a very modest effort that defers to ILRAD’s effort.

Recent Progress. —In the past few years, scientists have made significant advances in demonstrating genetic mechanisms of antigen switching

in African trypanosomes, with corresponding regulation of the expression of variant surface glycoprotein (VSG) genes. This work is important to

vaccine development because surface proteins are

the most likely antigens to stimulate the immune

system.

Research on VSGS is now being pursued intensively, and it appears that at least two distinct

mechanisms operate to cause shifts in VSGS: 1)

changes in the gene itself, including duplication

of a preexisting invariable basic copy of the VSG

gene, followed by transposition of the newly duplicated “expression-linked copy” to an area of
Ch. 7—immunization Technologies: Selected Tropical Diseases “ 139

the genome in which it can be expressed; and 2)

changes in gene activation related to a small segment spliced on to one end of the corresponding

messenger RNA. Several types of genomic rearrangements have been described (22). Complementary DNA (cDNA) has been prepared for a

variety of VSGS. Scientists have found that the

VSGS are encoded by a fairly large number, perhaps up to 1,000, separate genes. DNA sequencing has been done on several VSG genes, which

show considerable similarities over a large segment (290). The complete nucleotide sequence of

cDNA coding for a VSG of T. brucei has been

worked out by Boothroyd and colleagues (25),

and the manner in which the VSG is made from

more complex precursors is becoming known.

Current work on African trypanosomes by American investigators is centered largely on the

molecular biology of antigen switching of VSGS.

Roughly half a dozen laboratories are working intensively on mechanisms of transcription and expression, the nature of requisite messenger RNA

splicing, evolution of the VSG repertoire, and similar problems. Although some workers are attempting to characterize the complete antigenic

repertoire of African trypanosomes and are

searching for nonvariant antigens across strains,

there is little hope of developing a vaccine in the

near term. The hope is that this research will pay

off in the short term by identifying discriminatory proteins for use in immunodiagnosis and metabolic targets for chemotherapy.

Chagas’ Disease (American Trypanosomiasis)

Current Status.—The current status of vaccination for Chagas’ disease is generally considered

unpromising. Experimental vaccines have been

made by conventional methods (339) but have not

been widely adopted. T. cruzi appears to be a

good candidate for application of MAbs to develop vaccines. Immunization against T. cruzi is

complicated, however, by the possibility that the

severe immunopathologic events of Chagas’ disease may be exacerbated by a vaccine.

Wood and colleagues (410) have reported production of a MAb against rat nerve tissue that

cross-reacts with T. cruzi. Thus, a trypanosome

antigen, which is identical to, or close to, a protein that is in host tissue, may be important in

the pathology of this infection. It maybe that the

host immune system is stimulated to produce antibody against the parasite, that coincidentally

cross-reacts with host tissue in a type of autoimmune response. The implication is that a vaccine that stimulates a protective antibody may

stimulate immunity and pathology at the same

time.

Recent Progress.—MAbs to culture and intracellular stages of T. cruzi have been made by

several workers. Araujo and colleagues (10) reported production of 17 different secreting hybridomas, of which 5 recognized the antigens of

intracellular stages only, 2 recognized culture

forms only, and 10 reacted with both. Immunofluorescence procedures pinpointed the specific

antigenic sites on the T. cruzi organisms. Snary

and colleagues reported production of MAbs recognizing different stages, but not different species

of trypanosomes (323), demonstrating the presence of stage-specific determinants.

In a remarkably original experiment, Crane and

Dvorak (80) reported the fusion of T. cruzi organisms directly with vertebrate cells using a procedure analogous to hybridoma cell fusion. Three

hybrid clones continued to express T. cruzi antigens for at least 14 weeks of serial cultivation, suggesting the continued presence of functional parasite DNA. A great deal of research activity with

MAbs is under way. The research is aimed at identifying protective antigens which may be suitable

for use in a vaccine, but progress is still elusive.

An animal model for Chagas’ disease that mimics

the human disease is being pursued by several laboratories. The objective is to have an experimental

system for evaluating immunologic responses to

T. cruzi and the protective effect of any candidate vaccines.

Research Needs. —A vaccine against Chagas’

disease is badly needed, but researchers are not

highly optimistic that one can be developed (16).

In lieu of a vaccine, one investigator demonstrated

some protection in mice by passive immunization

with a MAb, suggesting that a different immunologic approach to parasite reduction maybe feasible (16). Improvements in drug therapy for
140 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Chagas’ disease, rather than vaccination, maybe

a more urgent and feasible priority.

Leishmaniasis

Leishmaniasis is caused by several species of the

protozoan genus Leishrnarzia. Differefit species

cause different clinical diseases:

1.

2.

3.

cutaneous Ieishmaniasis;

mucocutaneous leishmaniasis, which begins

as a skin lesion that progresses to destruction of tissue and cartilage of the face, nose,

and throat; and

a visceral form of leishmaniasis known as

“kala azar.”

Current Status

Leishmaniasis vaccination using living cultured

organisms has been attempted since the early

years of the 20th century, and field trials with a

frozen whole organism vaccine have recently been

undertaken in Israel (138). Experimental studies

in Australia using inactivated whole Leishmania

(frozen and thawed infected culture cells) to infect genetically resistant and susceptible strains

of mice are under way to devise standardized vaccination protocols. It has been demonstrated that

avirulent parasite strains can induce protection

in mice against related virulent strains (353). The

theoretical possibilities of immunization are being developed and evaluated.

Research Needs

In view of the large number of strains and types

of Leishnxmia and scientists’ limited ability to

identify organisms, there is doubt about whether

a leishmaniasis vaccine will ever be practical. Each

isolate is different. However, there is encouraging evidence from animal experiments and practical evidence of the potential for inducing active

immunity. Further research on the current leads

is needed.

Filariasis

Filariasis is the collective term for several distinct parasitic infections by insect-transmitted,

tissue-dwelling nematodes (roundworms). The

most important worldwide are Wuchereria bancrofti and Brugia malayi, the agents of filarial

elephantiasis; and Onchocerca volvulus, the agent

of onchocerciasis (river blindness) in west Africa,

also found in Central and South America.

Current Research

Until recently, filarial organisms were difficult

to maintain for laboratory study, with complicated life cycles requiring insect vectors at one

point. The lack of suitable animal models for

many filarial diseases also hindered research. Investigators have achieved successful in vitro cultivation of B. malayi and a related species B. pahangi from the larval stage to young adult stage.

They have also achieved some success with leaf

monkeys (Presbytis melalophos and P. cristata)

as experimental hosts (16).

Evidence of humoral immunity has been demonstrated by passive transfer of serum in mice and

cattle previously inoculated with crude parasite

preparations or live microfilariae.

Antifilarial MAbs are being produced for diagnostic purposes, but they can also be used for

identification and purification of protective antigen. Attempts to identify specific antigens of possible significance to protection against Wuchereria, Brugia, and Onchocerca are under way. At

least one U.S. Government laboratory, one industry laboratory, and one university laboratory

are conducting or planning recombinant DNA

work for expression of such filarial antigens (16).

Leprosy (Hansen% Disease)

Leprosy is the only bacterial disease among the

six tropical diseases targeted by TDR. It is mainly

a disease of skin and peripheral nerves, but is

characterized by a wide array of clinical presentations.

The severe form of leprosy, lepromatous

leprosy, occurs in patients with defective cellmediated immunity. These patients develop massive, contagious infections with extensive nerve

damage. The nerve damage in severe cases seems

to result from an overactive cell-mediated immune

response. Most patients, however, develop a high
Ch. 7—immunization Technologies: Selected Tropical Diseases ● 141

level of cell-mediated immunity which kills and

clears bacilli from the tissues. These patients, with

tuberculoid leprosy, have a good prognosis.

The role of humoral immunity in leprosy still

is not clear, but may be detrimental to the host.

In the process of isolating Mycobacteriwn leprae

bacilli from armadillo tissues, investigators have

obtained significant quantities of a phenolic gly -

colipid that is the only unique antigen present in

M. leprae and not other mycobacteria. In vitro

studies now indicate that immune reactions to this

antigen may actually increase disease, without

producing a protective effect (222). Characterization of the immune defect in lepromatous leprosy patients is being investigated using techniques

of biotechnology and also by analysis of the patients’ genetic makeup. Understanding the immune response to infection with M. leprae will

help in developing immunization strategies.

Current Research

The tuberculosis vaccine BCG (an attenuated

strain of a bovine mycobacterium) has been tested

for an immunoprophylactic effect in large-scale

prospective trials in Uganda, Burma, Papua New

Guinea, and India. Long-term followup in these

trials indicated a variable and not very effective

protection against leprosy.

An interesting development in leprosy vaccination is the work of Convit and collaborators

during the past decade in Venezuela (72). These

researchers conducted clinical trials in a variety

of individuals, including some with early disease,

using a vaccine made of heat-killed M. leprae

(grown in armadillos) combined with BCG vaccine. They found clinical, histopathologic, and immunologic changes in many patients with disease

of moderate severity. The investigators’ claim of

conversion to effective cell-mediated immunity in

lepromatous leprosy patients and more rapid

clearance of bacilli in patients treated needs confirmation. Vaccine field trials using killed M.

leprae only, BCG vaccine only, and M. leprae

with BCG vaccine are planned. Such trials will

require considerable amounts of M. leprae. Since

leprosy is a disease of low incidence and long incubation period, the evaluation of efficacy of a

vaccine will require 10 years or more of observation in large populations.

Carefully controlled human trials of leprosy

vaccine from TDR are now under way in Norway

in individuals never exposed to leprosy. It remains

to be seen whether this vaccination will simply

mimic natural exposure to leprosy (i.e., stimulating cell-mediated immunity in most people, without helping the smaller proportion of people who

cannot mount such a response), or if it will support the earlier results of Convit and colleagues

(72) of having some effect even in the lepromatous leprosy patient.

Extensive and careful planning of the production, purification, and testing of the leprosy vaccine, plus selection and epidemiological characterization of test populations has been under way

in the TDR project for several years (349,350,353).

At least two groups are planning recombinant

DNA work to try to make a leprosy vaccine, in

order to eliminate the need for recovering M.

leprae from armadillos (72). Some workers feel

that subunit vaccines are not likely to confer significant protection, since they induce a humoral

response but not cell-mediated immunity. One

group of workers has isolated and characterized

DNA from M. leprae and investigated its relationship to other mycobacteria (170). Further work

may identify important DNA segments that could

be cloned and used to produce quantities of a specific antigen.

Research Needs

Obtaining antigens from M. leprae grown in

armadillos is a slow process and not suited to producing large quantities. In vitro culture methods

are needed to facilitate leprosy research.

A concentrated effort is needed to use recombinant DNA methods and T-cell clones to sort out

M. Ieprae-specific epitopes pertinent to protection.

Better knowledge of cell-mediated immunity and

the cellular aspects of host reactions is needed.

Further evaluation of BCG vaccine and M.

leprae antigens through immunization trials is

continuing. The fact that the tuberculosis vaccine

(BCG vaccine) may prove to be useful against leprosy points out the importance of further study

of the relationship of M. leprae to other mycobacteria, especially M. tuberculosis.
142 • Status of Biomedical Research and Related Technology for Tropical Diseases

Tuberculosis

Tuberculosis remains a major threat to health

in many parts of the world, causing several million deaths annually. Even in the United States,

some 30,000 new cases of pulmonary tuberculosis were reported in 1980 (70). There is a tuberculosis vaccine, BCG vaccine, but its efficacy is

uncertain and controversial. (For a review of immunology and microbiology of tuberculosis, see

Collins (70) and Wayne (398).)

Current Status

BCG vaccine is an attenuated strain of a species of bovine mycobacterium. Since the early

1950s, BCG vaccination has been used extensively

in tuberculosis control programs around the

world. Even in the 1950s, however, it was known

that BCG vaccine did not offer complete protection against tuberculosis. Results of controlled

field trials were contradictory, with protection

varying from O to 80 percent.

All the BCG vaccines used throughout the world

today are derived from the original strain developed at the Pasteur Institute more than 50 years

ago. There is no reliable method for standardization of BCG vaccines. Most people agree that the

probable variation of different vaccine preparations, compounded by differences in immunologic

response of populations, is the main reason for

the enormous discrepancies in results between

otherwise well-conducted BCG trials.

Because of the lack of definitive evidence of

BCG’S efficacy, a controlled double-blind field

trial in southern India was started in 1968 (414).

Two highly ranked vaccines were used. After 7%

years of careful followup, there was no evidence

of a protective effect in BCG-vaccinated groups.

The Indian field trial was meticulously reviewed

(415,417) and found methodologically sound.

Quite possibly, the explanation for the results in

this trial is that the population had already developed some resistance to tuberculosis through

exposure to a widespread south Indian nonhuman

mycobacterium; and BCG vaccine could not add

to that resistance. The results from this trial may

not be applicable to other parts of the world, depending on local conditions. BCG vaccine is still

considered useful in tuberculosis control programs.

Immunologic research on tuberculosis has been

given relatively little attention over the years, and

the field trial results in southern India clearly revealed a large gap in knowledge about the immunology of tuberculosis (417). Application of recombinant DNA methods to M. tuberculosis is

in its infancy, but several investigators are planning projects. MAb work is under way in a handful of laboratories, including some at NIH and

several academic institutions. MAbs are being

reacted with M. tuberculosis to obtain purified

antigen probes. Attempts are being made to isolate specific antigens for diagnostic skin tests (see

ch. 8) in an attempt to make such tests less crossreactive to infections with other types of mycobacteria, but such antigens may also have value

for immunization.

Research Needs

A great deal of fundamental knowledge about

tuberculosis immunology is needed to resolve the

pending questions about immunization in general,

the efficacy of BCG vaccine, and the susceptibility of various related mycobacteria to vaccination and their importance in the disease process.

Better understanding of tuberculosis immunology and vaccination and clarification of the effectiveness of BCG vaccine are still needed.

Diarrheal and Enteric Diseases

Diarrheal diseases are caused by a variety of

viruses, bacteria, protozoa, and worms. Development of water and sanitation facilities where

they do not now exist is a long-term solution to

control these diseases. Nevertheless, some of the

agents of diarrheal disease may well be susceptible to immunologic attack, thereby permitting

prevention of disease with vaccines.

A few injectable vaccines inducing serum antibodies have been available for years (for cholera,

typhoid, paratyphoid, Shigella), but these have

limited effectiveness. Oral vaccines, which in most

cases appear to induce a more appropriate immune reaction in the gut, are currently the focus

of much research.

Although some research will have cross-over

potential for several agents, it is generally expected
Ch. 7—immunization Technologies: Selected Tropical Diseases ● 143

that each pathogen will need a tailored approach.

Immunologic research has made it clear that the

vaious etiologic agents evoke and are fought by

a variety of immune responses. A further challenge of diarrheal disease research is the continuing discovery of new important etiologic agents.

Viral Infections

Rotaviruses cause about one-third of all diarrheal disease in the world and up to 50 percent

of the hospitalized cases of diarrheal illness in children under 2 years (21,167,325,331). Animal rotaviruses are also important because of their economic significance in farm animals, and some

veterinary vaccines have been developed and are

in use.

Characterization of the rotavirus genome is being actively pursued with the aid of recombinant

DNA techniques. Rotavirus RNA has been synthesized in vitro, reverse transcribed, and inserted

into Escherichia coli; the genes from which most

of the clones derive have been identified (117);

bacterial clones containing copies of each rotaviral

gene have been identified; some of the genes have

been analyzed to determine the amino acid sequence of the proteins for which they code. This

work is a promising means of identifying critical

genes coding for the pathogenic characteristics or

antigenic determinants of the organism. If successful, production of a strain of rotavirus with deletions of the pathogenic genes (similar to the cholera research described below) or transfer of the

DNA segments that specify the antigenic determinant(s) and production of the antigen may be

possible, forming the basis of a vaccine.

A major obstacle to vaccine development has

been the lack, until recently, of a cell culture system for human rotaviruses (192,429). It may now

be possible to develop attenuated vaccines through

the various conventional methods (cell culture

passage, cold adaptation, chemical mutagenesis,

and reassortment) (371,385). In fact, volunteer

studies in adults are being conducted to provide

a test system for such candidate vaccines (181,182).

Animal rotaviruses have been successfully

grown in culture, and attenuated strains are available as veterinary vaccines. Studies in calves,

piglets, and lambs have demonstrated the importance of intestinal rotaviral antibody in preventing or attenuating illness. Because human and calf

rotavirus strains share a common group antigen

(179,711), calves inoculated in utero with bovine

rotavirus were protected against challenge with

human rotavirus. Furthermore, piglets infected

with bovine rotavirus and later challenged with

human rotavirus showed cross-protection (204,205).

On the basis of this evidence, an oral, live, attenuated bovine rotavirus vaccine was recently

tested and found protective in a test population

of human adults and children (385). The vaccine

has now been tested and found to be safe and effective in protecting infants from natural rotavirus

infection (386). This is a very promising development that is being tested further.

Bacterial Infections

Most of the vaccine research on the bacterial

enteric pathogens is being done on enterotoxigenic

E. coli, Salmonella typhi (the cause of typhoid fever), Vibrio cholerae (the cause of cholera), and

Shigella (the cause of bacillary dysentery). Most

research groups are studying several of these

agents. Fewer laboratories are studying Campylobacter, Yersinia, other species of Salmonella and

V.brie, and other pathogenic intestinal bacteria.

Approaches to vaccine development are generally

similar to those described below for E. coli.

Progress against several of these agents is described below.

Coliform Infection.—Various strains of E. coli

are now recognized as major causes of diarrheal

disease in older children and adults living in developing countries. They also appear to be a chief

cause of “traveler’s diarrhea.” The disease-causing

characteristics of E. coli are under genetic control, and investigators in several laboratories are

actively identifying and cloning the genes that

code for attachment and colonization, virulence,

toxin production, and antibiotic resistance. The

goal of this work is the development of strains

that stimulate specific immunity but are not themselves pathogenic. Successful development of vaccines against enterotoxigenic E. coli in animals has

stimulated work towards a vaccine for humans

(427).
144 . Status of Biomedical Research and Related Technology for Tropical Diseases

Typhoid Fever. —Injectable, killed S. typhi vaccines against typhoid fever have proved to be protective for adults and older children living in endemic areas. These vaccines are unsatisfactory,

however, because they frequently induce adverse

reactions, they are not entirely protective, and

they do not stimulate local intestinal immunity

(419).

While oral, killed typhoid vaccines have provided only minimal protection in volunteer and

field trials, several live attenuated S. typhi strains

are under study and good progress has been made.

A strain developed by the Swiss Serum and Vaccine Institute may well become available for widescale use very soon. This strain has been tested

for stability, safety, and efficacy in volunteers

(127), and in field trials in Egypt (391), all with

positive results. The results of a trial in Chile, in

collaboration with Walter Reed Army Institute

of Research, unfortunately do not match the

Egyptian results (24). The WHO Program for the

Control of Diarrheal Diseases is planning for the

use of this vaccine by collecting baseline information on the incidence of typhoid fever in different countries and reviewing other relevant data

(423,427).

Cholera.—Cholera is relatively rare compared

to the other diarrheal diseases, especially those

caused by rotavirus and E. coli. However, the epidemic and pandemic potential of cholera is great

enough to make cholera an important tropical disease. The severe diarrhea of cholera is caused by

a toxin produced by V. cholerae, acting on the

gut wall.

Although injectable, killed whole-cell and toxoid cholera vaccines are currently in use, these

vaccines do not stimulate effective, long-lasting

protection against cholera (47,86,200,201,260).

Scientists using traditional nonrecombinant

mutagenesis have produced a number of attenuated mutant strains of V. cholerae that do not

produce the cholera toxin (nontoxigenic strains).

These attenuated, nontoxigenic strains have been

used experimentally as oral vaccines with some

encouraging results, but genetic instability or poor

colonizing ability in the gut make them unsuitable.

Researchers have very recently succeeded in

using recombinant DNA techniques to produce

a live, attentuated V. cholerae strain by deleting

gene coding for the cholera toxin (178,223). A recombinant strain such as this could provide a vaccine that colonizes the gut and stimulates immunity without producing the toxin and without the

capability of mutating back to the pathogenic

type. Very early clinical studies are under way

to assess the safety and efficacy of this new vaccine. Unfortunately, the V. cholerae with the

genes for cholera toxin deleted cause diarrhea in

volunteers, presumably from a different toxin

(230).

Shigellosis.—Shigella dysenteriae is associated

with serious disease and fatality (bacillary dysentery), but S. sonnei and S. flexneri, which cause

less severe disease, account for a major portion

of all isolates in diarrhea patients. The available

injectable, killed vaccines are not effective, and

oral vaccines have not been developed until recently. Shigela will colonize primates only, making work with animal models difficult and expensive. One group of researchers has taken an

attenuated, live typhoid vaccine bacterium developed recently in Switzerland and has spliced into

it the genes coding for production of an important S. sonnei antigen, thereby inducing immunity to both typhoid fever and S. sonnei dysentery to vaccinees (126,123). Testing in humans is

getting under way. The same group has isolated

genes for attachment factors in Shigella and inserted these through plasmid vectors into noninvasive E. coli. These novel recombination as well

as a number of attenuated strains are under investigation as potential vaccines, but solid success

is still elusive.

Acute Respiratory Infections (ARIs)

ARIs area heterogeneous mixture of diseases,

caused by a variety of viruses, bacteria, and other

agents, some of which also cause infections out-
Ch. 7—immunization Technologies: Selected Tropical Diseases ● 145

side the respiratory system. As detailed below,

some vaccines for ARIs are available, and progress is being made in developing new vaccines.

Viral Infections

Many viruses cause acute respiratory illness in

humans. The following groups have been identified as most important (see Anderson, et al. (5)

for a recent extensive review):

Influenza. -Outbreaks of influenza may occur

annually, major epidemics every 2 to 3 years, and

pandemics at 10- to 15-year (or more) intervals.

The last pandemic (as of July 1985) was in 1968.

In pandemics, mortality rates are devastating, particularly in the elderly. Three main types of influenza viruses, with numerous subtypes and strains,

cause infection in humans. Infection sometimes

confers long-term strain-specific immunity or partial immunity. Because of antigenic drift and shift,

influenza vaccines must be reformulated every

year against the predominant virus. The annual

production of influenza vaccine targeted for the

strain predicted to be prevalent is now well implemented in the United States. However, the need

for annual renewal makes mass influenza vaccination in developing countries impractical at this

time.

Knowledge of the chemical and antigenic structure of influenza virus has increased greatly and,

combined with the application of current molecular biology, is leading to better influenza vaccines.

More rational approaches to vaccine development

have emerged, e.g., the discovery of chemicals

able to release surface glycoproteins from the viral particle without affecting their antigenicity,

thus producing an efficacious and safer (fewer side

effects) killed vaccine; the use of “high yield” influenza A viruses are being used to produce seed

strains for better vaccine production (264).

Inactivated influenza vaccines are in use in developed countries. Killed vaccines administered

by injection are made from virus particles disrupted by chemical treatment. This treatment

lowers the antigenicity but also reduces the adverse side effects. Purified vaccines that contain

only the immunogenic antigen, free of nonimmunogenic proteins, and are well tolerated by children can be produced (198,347). The vaccines are

usually reserved for selected high-risk populations, such as the elderly, chronic disease patients,

or economically important public service groups.

Although these vaccines do not provide complete

protection (225) and their effectiveness is dependent on annual renewal according to the prevalent strain (163), the Centers for Disease Control

has conducted field studies that show inactivated

influenza vaccine to be extremely useful in preventing or attenuating influenza infection (54,

374).

Live, attenuated influenza vaccines for intranasal administration are being developed. The attenuated vaccines can be rapidly and reliably produced by the transfer of genes from a cold-adapted

attenuated donor virus to any new wild-type influenza isolate. Live vaccines, produced by reassortant RNA, eliminate the pathogenic genes but

retain the immunogenic surface antigens. Recent

research has indicated that these vaccines may be

superior to the inactivated vaccines (60). In a

study in adult volunteers, the live, attenuated vaccine completely protected against illness, while the

licensed inactivated vaccine did not. Furthermore

the nasal administration (by nose drop) is more

convenient for patient and medical personnel.

Even with the live, attenuated vaccine, however,

there is a need for annual administration. Longer

term protection will be possible only if there are

common antigens among strains.

Much effort by 8 to 10 laboratories in the

United States is focused on the molecular biology of influenza. This reflects, to a large extent,

the importance of influenza as a public health

problem in the United States. With immunity

under active study (76) and the practical value of

immunization under review (271,304), hundreds

of publications in recent years have resulted. A

large number of genes have been identified and

sequenced. Synthetic peptides have been found

to be antigenic and are undergoing evaluation, but

preliminary studies have shown that such vaccines

have greatly diminished capacity to induce neutralizing antibodies. In an effort to understand

virulence factors, investigators have applied innovative methods, such as “tryptic peptide fingerprinting,” “RNA-RNA hybridization,” and “oligonucleotide mapping” (16).
146 ● Status of Biomedical Research and Related Technology for Tropical Diseases

One interesting development is the cloning of

influenza genes in vaccinia virus (the smallpox

vaccine) and the subsequent expression of both

vaccinia and influenza antigen resulting in simultaneous immunization of hamsters against both

agents (319). Because the vaccinia virus is well

characterized, and several extra genes can be

spliced into its genome, there is speculation that

this experimental work could be developed into

a method for delivering several vaccines in one

organism. Many safety questions need to be answered before the vaccinia vector approach can

be evaluated fully. The rate of adverse reactions

from smallpox vaccine is higher than those for

vaccines in current use, and may not be acceptable when smallpox is not the target.

Respiratory Syncytial Virus (RSV).–The several known strains of RSV are the most important causes of lower respiratory disease (pneumonia and bronchiolitis) in children under 2 years.

The disease tends to occur in sharp outbreaks.

Previous immunization efforts have not been successful, but new techniques show promise. Cloning of surface proteins or production of genetically engineered attenuated vaccines may meet

with success.

Early work using inactivated RSV showed that

vaccinated children developed antibodies, but

were not protected against infection (180,186). Various immunologic phenomena have been hypothesized and studied without clear resolution.

Vaccination with attenuated RSV administered

through the respiratory tract has not been successful. The degree of attenuation is variable, and

the vaccines are not very stable (49,286). Injecting wild-type RSV grown in cell culture induced

antibodies in young children without causing

symptoms of disease (0o). However, there is no

evidence that RSV given by injection will reproduce, nor that the vaccine will not interfere with

passively transferred maternal antibodies in infants. The lack of vaccine stability and the uncertainty of a viral replication after injection are

important, because a virus that fails to reproduce

may act as an inactivated antigen that can potentate disease due to natural infection, and because

the greatest need for effective RSV immunization

is in the first months of life when maternal antibodies are still circulating in the infant (passive

immunity). Another important finding that may

hamper vaccine development is the report of antigenic variation in a new strain of RSV, which

means that any single vaccine may be ineffective

against some RSV strains (154).

Parainfluenza. —There are four main types

(serotypes) and two subtypes of parainfluenza

viruses. These viruses are the second most common agents of ARI (after RSV), producing croup,

bronchitis, pneumonia, and bronchiolitis in infants and children.

Vaccine research thus far has shown that serum

antibodies circulating in the blood against type

1 virus do not protect against infection, but antibodies in nasal secretions directly correlate with

resistance to reinfection (317). Vaccines of formalin-inactivated virus administered by injection induce high titers of neutralizing antibody without

preventing infection, though severe illness is prevented. Results with experimental live, attenuated

vaccines administered intranasally have been encouraging. Whether killed or attenuated virus is

used, the usefulness of a parainfluenza vaccine

would have to be measured by the vaccine’s protective effect in children, especially infants, who

often become infected during the first months of

life and suffer the most severe symptoms. Establishing the safety of a vaccine will be difficult

(198,264).

Adenoviruses. —At least 41 different types of

adenoviruses have been identified as causes of ARI

(especially a problem in military recruits in the

United States), as well as of epidemic keratoconjunctivitis (an eye infection) and a venereal

disease.

Vaccines to prevent adenovirus infection have

been developed for use in military recruits. A live

vaccine cultured in human cell culture has replaced the earlier killed vaccine, which was highly

protective but was inconsistent in different

batches. The live vaccine, containing two different serotypes responsible for adenovirus infection

in military recruits, is given orally (264).

Unfortunately, the experience with the recruits

is not directly transferable to the general population, for two fundamental reasons. First, the
Ch. 7—immunization Technologies: Selected Tropical Diseases ● 147

strains of adenovirus that cause most disease in

recruits are different from those prevalent in the

population at large. Second, while oral administration works for recruits, it is unsuitable for children, because recruits, in general, do not pass the

vaccine virus on to each other, but children do,

and a good deal of disease can be caused in that

way.

Rhinoviruses.—Rhinoviruses, with more than

100 serotypes, are the most common human ARI

agents and the single most important cause of the

common cold.

Inactivated rhinovirus vaccines have shown

some protection when administered intranasally,

but not by injection. Live, attenuated vaccines

may provide more protection, but the large number of serotypes, the fact that rhinovirus strains

may not completely lose their infectivity (51), the

indications of antigenic variation (120), and the

possible development of genetic reassortant variants as a result of dual infection all hamper the

future development of rhinovirus vaccines (264).

In spite of the problems, some commercial firms

are working on the rhinoviruses.

Coronaviruses, Coxsackieviruses, Echoviruses.—

These viruses are important causes of common

cold-like illness in children and adults. Coronavirus disease is self-limiting (about 7 days) and will

not be a priority for vaccination. Coxsackieviruses

induce a resistance after infection that is long lasting. With many different types prevalent in different areas and the predominant type in any area

varying every few years as immunity in the population rises, there will be obstacles to vaccination. Echoviruses share epidemiologic features of

coxsackieviruses. Immunization does not seem to

be a practical method of preventing infection because of the large number of serotypes (129,264).

There currently is little, if any, vaccine-related research on this group of viruses (16).

Measles.—Measles is an ARI for which an excellent vaccine exists. The vaccine is widely used

in the United States, as part of a measles eradication program, and in other parts of the world,

in particular, as part of WHO’s Expanded Program on Immunization (see ch. 5). Measles vaccination has been controversial in developing

countries because of the variable seroconversion

rate (percentage of vaccinated individuals who

later have measurable antibodies) and protection

rates (percentage of vaccinated individuals who

do not get the disease). A problem that has impeded success of vaccination programs is that infants must be vaccinated in the narrow interval

between the waning of maternally derived antibody and the time of exposure to natural measles.

Recent clinical trials of live, aerosolized measles virus vaccines (298,299,300) have demonstrated that the intranasal route of administration

may be particularly useful in developing countries. In contrast to vaccines injected through the

skin, the aerosolized vaccine can induce seroconversion in the presence of maternal antibodies, in

infants as young as 4 months. The aerosolized

vaccine raises the respiratory tract antibody level,

offering better protection at the portal of entry

rather than in the serum. It also simplifies the logistics of vaccine delivery. A recent study indicates that not all measles vaccines stimulate an

equally high level of protection in humans, even

though they meet potency standards at manufacture (301). Better definition of potency standards

is required. Further clinical trials are needed to

assess the true efficacy of the aerosolized measles

vaccine (4). In addition, the practicality of delivering aerosol vaccines in developing countries requires investigation. Getting the dosage right with

aerosols, for instance, is much more difficult than

it is with other methods of vaccination.

Bacterial and Mycoplasmal Infections

Streptococcus pneumoniae.—S. pneumoniae

causes pneumococcal pneumonia, a disease that

can be fatal when complicated with bacteremia

(bacteria in the bloodstream). S. pneumonia accounts for approximately 80 percent of all bacterial pneumonias; it is an important cause of hospital admissions in developing countries and an

important cause of middle ear infections and bacterial meningitis (12,248,292).

Pneumococcal vaccine is available, and its protective effect is well characterized (356). Immunity is specific to each serotype (subgroup of a

species characterized by common antigens), so

vaccines must include the most common serotypes

found in the area of use, The polyvalent formu-
148 ● Status of Biomedical Research and Related Technology for Tropical Diseases

lation used for the vaccine contains capsular polysaccharides from a number of pneumococcal types.

The vaccine is ineffective in children, especially

under 2 years. It is usually targeted to older people whose risk of infection and case-fatality rates

are greatest. Some new techniques of conjugation

show promise in improving effectiveness in young

children, but a major problem is the lack of information about which strains are prevalent in developing countries.

Streptococcus pyogenes.—The approximately

70 serotypes of group A streptococci (S. pyogenes)

cause a variety of diseases, including streptococcal sore throat, scarlet fever, and rheumatic heart

disease. Attempts to produce a streptococcal vaccine have encountered two serious problems:

cross-reaction of streptococcal antigens with heart

tissue, and the large number of serotypes.

A synthetic peptide approach offers the possibility of eliminating cross-reacting antigens while

preserving those components that confer immunity. A synthetic streptococcal vaccine has been

successfully tested in animals, and development

of a vaccine against rheumatic fever is possible

within the next 5 years.

Bordetella pertussis. —Pertussis (whooping

cough) is endemic worldwide, and epidemics also

occur. Children under 5 years of age have the

highest incidence of morbidity, and about 70 percent of mortality is among children less than 1

year old (21). Immunization is available with a

killed pertussis vaccine that is highly effective.

This pertussis vaccine is usually given in a triple

vaccine known as DPT (diphtheria, pertussis,

tetanus). The use of DPT vaccine among children

is promoted through WHO’s Expanded Program

on Immunization (see ch. 5). Intensive research

to reduce or eliminate the adverse reactions seen

with the current vaccine is under way.

Hernophilus idhrenzae. -Immunization with

the polysaccharide antigen of H. influenzae results

in antibody response. Antibodies can also be induced by E. coli, which has a polysaccharide that

cross reacts with H. influenza. Attempts are being made to increase the immunogenicity of the

antigen by coupling it to proteins (264). Prospects

for licensing a new vaccine that is immunogenic

in infants and children look very good.

Mycoplasma pneumoniae. —M. pneumonia is

now recognized as a major cause of primary atypical pneumonia. Infection induces serum antibodies, but pneumonia can occur in individuals with

positive antibody titers. While vaccination studies

seem to demonstrate that serum antibodies correlate with protection, this and the role of respiratory tract antibodies still are not fully understood.

Although antibodies appear in young children (2

tos years old), peak incidence of disease is in older

children and young adults.

Field trials with inactivated M. pneumonia

vaccines have shown encouraging results, and attenuated vaccines have also been tested. However,

some vaccinees have developed disease and have

not produced detectable antibodies, and lung lesions may have resulted from an immune reaction.

Further cautious work is needed to resolve these

problems (50,264).

Arboviral and Related Viral Infections

Arboviruses include literally hundreds of distinct viruses that are widely distributed throughout the world. Arboviral infections such as yellow fever, dengue fever, and various types of

encephalitis occur worldwide in endemic and epidemic forms.

Current Status

Vaccines are available for only a handful of arboviral infections. A live, attenuated vaccine for

yellow fever (the 17D strain) was developed decades ago and is well established and successful in

preventing yellow fever. It has proved to be extremely safe and effective. The immunity stimulated persists for more than 10 years. Yellow fever vaccine is grown in embryonated chicken eggs

following classic procedures of viral vaccine production (373).

Formalin-inactivated vaccines are used against

Japanese B encephalitis and Russian springsummer encephalitis with good results. Experimental attenuated (Venezuelan equine encepha-
Ch. 7—Immunization Technologies: Selected Tropical Diseases ●

litis and western equine encephalitis) vaccines under research and development have been used in

horses.

Research Needs

The most critical need is for prevention of

dengue fever, not only because of dengue fever’s

debilitating clinical syndrome, but also because

of dengue hemorrhagic fever (DHF), a serious

complication of dengue infection that often leads

to a shock syndrome and death (with symptoms

similar to yellow fever).

Several laboratories, both military and civilian, are studying the biology of the four main

serotypes of dengue virus. MAbs for early serotype-specific identification have been developed

and are now generally available. These may also

help to identify protective antigens. Attenuated

vaccines are being developed using classical vaccine production techniques. Each of the four

known serotype strains of dengue virus requires

SUMMARY

Effective vaccines exist for some viral and bacterial diseases important in developing countries,

but there are currently no antiparasite vaccines.

The vaccines available in developing countries are

largely those that are also used in the developed

countries, mainly against childhood illnesses. Efforts to develop vaccines against rotavirus infection, which causes a large percentage of diarrheal

illness, and malaria, one of the world’s biggest

health problems, are among the first efforts whose

products will benefit mainly the developing countries, although there is some work in the direc149

a separate vaccine. A serious question of vaccine

efficacy relates to the origin of DHF. It is now believed that the sequential infection with two different dengue serotypes leads to DHF. It also appears

that type 2 is the critical precipitator. Therefore,

the hope is that vaccination against type 2 can

prevent DHF; however, careful evaluation will be

needed to ensure that vaccination will not lead

to increased incidence of DHF as a side effect

(143,144).

Very few vaccines exist against the arboviral

agents, but previous successes argue for a continued research effort. Because there are literally

hundreds of arboviruses that cause ill health, routine vaccination is still a long way off. Molecular characterization and increased taxonomic

studies are needed to lead the way to “generic vaccines” (polyvalent) for large groups of related

viruses, rather than serotype-specific vaccines. To

develop vaccination strategies, there is also a need

for better understanding of the epidemiology of

arboviruses.

tion of vaccine development for virtually every

disease.

The successful application of immunization

technologies, sadly, does not rest on exciting developments in research and development. The

vaccines that are already available are far from

universally applied because of financial and political constraints, and there is no reason to believe that new vaccines will fare any better unless

they are supported by international and bilateral

health programs.
Diagnostic Technologies:

Selected Tropical Diseases
Contents

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Conventional Diagnostic Techniques . . . . . . . . . . . . . . . . .

Direct Examination . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Serologic Diagnostic Techniques . . . . . . . . . . . . . . . . . . .

Genetic Tools for Improving Diagnostic Techniques . . . .

Monoclinal Antibodies . . . . . . . . . . .

Nucleic Acid Hybridization Probes.

Diagnosis: Current Status for Selected

Malaria . . . . . . . . . . . . . . . . . . . . . . . . .

Schistosomiasis . . . . . . . . . . . . . . . . . .

Trypanosomiasis . . . . . . . . . . . . . . . . .

Leishmaniasis. . . . . . . . . . . . . . . . . . . .

Filariasis . . . . . . . . . . . . . . . . . . . . . . . .

Leprosy . . . . . . . . . . . . . . . . . . . . . . . . .

Tuberculosis . . . . . . . . . . . . . . . . . . . . .

Diarrheal and Enteric Diseases . . . . .

Acute Respiratory Infections . . . . . . .

Arbovira

Summary .

. . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . .

Tropical Diseases

. . .

. . .

. . .

. . .

. . .

. . .

. . .

. . .

. . .

‘and Related Viral Infections .

. . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . .

...,, . . . . . . . . . . . . .,

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . .!

..•...................... . . . . . . . . . . . . . . .. .. ...,

.. .. .. .. ... . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . .,

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . .

LIST OF FIGURES

Figure No,

8-1.

8-2.

8-3.

Enzyme-Linked Immunosorbent Assay Technique for

Antigen or Antibody . . . . . . . . . . . . . . . . . . . . . . . . .

Thin-Layer Immunoassay Technique for Detecting Antigen or

. . .

Detecting

. . . . . . . . . . . . . . . . . . .

Antibody . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Reverse Passive Hemagglutination Technique for Detecting Viruses . . . . . . .

Page

153

155

155

155

161

161

162

163

163

164

165

167

168

169

170

172

173

176

177

Page

158

159

160
8

Diagnostic Technologies;

Selected Tropical Diseases

INTRODUCTION

Diagnosis has three important functions. One

is to determine the nature of the individual’s disease for the purpose of deciding on an appropriate course of treatment. A second function is to

determine the prevalence of specific disease-producing organisms or agents in populations, to allow assessments of the impact of public health interventions. The third is to find out about the

range of diseases affecting a population, the immune stages of populations, etc., for the purposes

of research.

Diagnosis of tropical diseases is a challenge for

the clinician or epidemiologist, because many of

the diseases mimic a wide variety of infectious

processes by presenting similar or ambiguous

symptoms. In countries where these diseases are

endemic, symptomatic ambiguity can also result

from the common occurrence of multiple infections in a single individual. The proper identification of the species of the organism that is causing disease can be critical in clinical management,

especially in serious illness.

Although each tropical disease can be diagnosed

by a number of different methods, diagnostic techniques vary in their usefulness under different conditions. There are several parameters to consider

in evaluating the usefulness of a diagnostic test:

sensitivity, specificity, predictive value, crossreactivity, and precision (388).

●

●

Sensitivity is the ability of a test procedure

to find a disease-producing agent or disease

when it is present. A very sensitive test correctly identifies all infected individuals. Negative tests for individuals who have the disease are “false negatives. ”

Specificity is the ability of a test procedure

to correctly determine that a disease-producing agent or disease is not present. A very

●

●

●

specific test will correctly identify all uninfected individuals. Positive tests for individuals who do not have the disease are “false

positives.”

Predictive value is an interaction of sensitivity, specificity, and the prevalence of the infection in the population being tested. The

sensitivity and specificity of a test will give

different numbers of false negative and false

positive results depending on the prevalence

of the infection in the population tested. If

a test is conducted in a group of patients

strongly suspected of having the disease, very

few false positives would be expected, simply because very few people tested will actually be negative. Thus, positive results will

be readily accepted as true. Conversely, if the

same test is conducted in a population in

which very few people are thought to have

the disease, the positives would have a higher

probability of being false positives, and in

fact, false positives may outnumber true positives. The negative test results will mostly be

true negatives.

Cross-reactivity is an aspect of immunodiagnostic tests related to, yet different from,

specificity. A cross-reaction occurs when the

test correctly identifies the appropriate chemical entity, but that chemical happens to exist in a different organism than the one tested

for. A false positive results.

Precision is reproducibility, the ability of a

test procedure to give consistent results in

repeated trials of the same sample.

Broadly generalizing, there are two categories

of conventional diagnostic technologies:

1. Direct examination of blood, stool, urine,

sputum, tissue biopsy, or cultured isolates

153
154 ● Status of Biomedical Research and Related Technology for Tropical Diseases

2.

using simple equipment (e.g., a light microscope) and reagents.

Serologic examination to detect antibodies

to the pathogen or to detect the pathogen or

its byproducts (antigen) in a sample of the

patient’s blood. Serologic methods have important advantages over direct examination,

but require in most cases specialized equipment and reagents and have never achieved

wide practical use.

Biotechnology is leading to the development of

new diagnostic methods based on the use of monoclinal antibodies (MAbs) and nucleic acid (DNA

or RNA) hybridization and promises to revolutionize diagnosis by offering quick, simple, accurate tests when and where they are needed.

The conventional methods of diagnosing infectious diseases and identifying disease-producing

organisms include the following:

●

●

●

●

●

the clinical impression of the physician or

other health worker (this is the most widely

used method of diagnosis);

direct examination of clinical specimens (e.g.,

blood, stool, urine, sputum, or tissue biopsy)

using light or electron microscopy to identify the disease-producing organism (e.g.,

malaria parasites, intestinal amebae and helminth eggs, leishmanial organisms, mycobacteria that cause tuberculosis and leprosy,

filaria);

X-ray or computed tomography (CT) scan

to image internal pathogens (e.g., amebic abscesses, echinococcal cysts); occasionally useful, but not routine;

xenodiagnosis, by permitting an insect vector to feed on a patient and then examining

the insect by light microscopy to look for the

disease-producing organism after it has multiplied to a detectable density (e.g., to diagnose Chagas’ disease);

culture of a specimen from a patient in a test

medium, tissue culture, or by inoculation

into an animal to allow it to multiply to a

detectable density, followed by direct exam-

●

●

●

ination for the disease-producing organism,

or use of serologic techniques (e.g., for many

bacteria and viruses);

skin tests, demonstrating hypersensitivity reactions to disease antigen (e.g., to diagnose

tuberculosis);

serologic methods to detect antibodies to the

pathogenic organism in the patient’s serum;

and

demonstration of a pathogen byproduct,

such as an antigen by chemical methods (e.g.,

schistosomes).

Photo credit: Office of Technology Assessment

Xenodiagnosis, by allowing laboratory-raised reduviid

bugs to feed on suspected Chagas’ disease victim.
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases ● 155

CONVENTIONAL DIAGNOSTIC TECHNIQUES

Direct Examination

Laboratory diagnostic methods that directly

identify a disease-producing organism by microscopy of clinical samples can provide a definitive

diagnosis. Constraints on this approach include

the following:

●

●

●

●

●

specially trained personnel are required;

the work quickly becomes boring and repetitious, while still demanding concentration

and attention to detail;

procedures are relatively time-consuming and

require equipment and materials that are

often inadequate in endemic countries;

in some cases, the organisms are not detectable either because:

—they are not present in routine clinical samples (e.g., the organisms that cause trichinosis, toxoplasmosis, hydatid disease,

hepatic capillariasis, toxocariasis);

—they are not detectable in routine clinical

samples until a later stage of infection (e.g.,

filaria); or

—their densities are very low, making sensitive diagnosis difficult (e. g., in chronic

malaria and chronic Chagas’ disease); and

identifying the disease-producing organism

is difficult because some species appear similar (e.g., intestinal amebae, malarial parasites, the African schistosomes with terminally spined eggs, leishmanial organisms).

Serologic Diagnostic Techniques

Serologic diagnosis depends on two types of immunologic methods, which are discussed further

below:

methods for detecting specific antibodies (the

immunoglobulins IgG, IgM, IgA, IgE, or IgD)

in a sample of the patient’s blood; and

methods involving the use of immunoglobulins for the detection of antigen (the diseaseproducing organism or its byproducts) in a

sample of the patient’s blood.

For the detection of antibodies, there are a variety of immunologic methods, all of which can

be adjusted by dilution of reagents to give a measure of intensity of reaction (a titer). Because the

various immunoglobulins have different roles and

develop at different times in an infection, different titers will be obtained at various stages in the

infection. (For instance, the immunoglobulin IgM

is usually an early but short-lived immune response to infection, whereas IgG production develops more slowly and then continues long after resolution of the infection. ) A positive test for

antibodies may indicate current infection or the

immunity that follows exposure to the infectious

agent. In some cases, the test may remain positive for extended periods after the infectious agent

has been eliminated by drug treatment.

For the detection of antigen, methods have not

been widely developed until recently, for technical reasons. The current advances in MAb technology make antigen detection more feasible.

Serologic Methods for Detection of Antibody

The principal serologic methods for detecting

antibodies in a patient’s blood are described

below,

1. Complement fixation (CF) test. This method

is well established and has been applied to all viral, bacterial, and parasitic diseases. Complement,

a substance normally present in the blood, fixes . -

or binds antigen to antibody. The CF test is performed by adding the known antigen and complement (prepared in the laboratory) to an individual’s serum in a test tube. If antibodies are present

in the patient’s serum, antigen-antibody complexes are formed. No reaction indicates a lack

of specific antibody. The CF test requires wellstandardized reagents, some of which have limited

shelf-life or limited availability in the tropics.

The CF test is widely used for viral diagnosis

and is most effective for parasite diagnosis of

American trypanosomiasis (Chagas’ disease) and
156 ● Status of Biomedical Research and Related Technology for Tropical Diseases

schistosomiasis. It is highly specific, but relatively

insensitive. It requires large amounts of antigen

per test. The CF testis performed adequately only

at specialized centers, and it is not considered a

test for general, practical use in the tropics (388).

2. Agglutination assays. Clumping of particles

due to the interaction of antibody and antigen is

the basis of all agglutination tests. In most variations, the disease-producing organism or its byproduct (antigen) is fixed to particles, and the particles are added to a test sample that may contain

the complementary antibody. If the complementary antibody is present, the particles agglutinate.

Each variation of this technique has acquired its

own name. The principal agglutination techniques

for detection of antibody are described below.

(The complementary techniques for detection of

antigen are noted in a later section. )

a. Direct agglutination test. In the direct agglutination test, whole organisms suspected of causing disease are added to the individual’s serum.

If antibody is present in the serum, the organisms

clump together. The test is simple, but it requires

pure, stable antigen preparations (from culture in

vivo or in vitro), the test result interpretation is

subjective, and there can be complicating autoagglutination reactions that result in false positives.

b. Indirect hemagglutination (IHA) test. In IHA

tests, the antigen is attached to a carrier, such as

specially prepared red blood cells or latex particles. When the individual’s serum is added, the

particles clump if specific antibody is present. II-1A

tests have been developed for malaria, Chagas’

disease, leishmaniasis, amebiasis, rotavirus, and

hydatid disease. IHA tests are simple to perform,

can be used to test minute volumes of serum, and

can be automated but the end-point reading is subjective. The preparation of test particles is not reproducible, the antigens have short shelf-lives,

and there are problems in standardizing antigens

from batch to batch.

c. Hemagglutination inhibition (HI) test. Various micro-organisms can clump or agglutinate red

blood cells from test animals under specified conditions. In the HI test, prepared red blood cells,

an appropriate micro-organism (e.g., a virus), and

a test serum are combined. If the individual’s

serum contains antibody specific to that virus, the

hemagglutination will be inhibited, because the

antibody combines with the organism and cannot

react with the red blood cells. The lack of agglutination indicates a positive test.

3. Neutralization test. Many viruses damage

cultured cells in certain detectable ways. When

test serum containing a specific antibody is added

to a test well containing the cell culture and the

virus (both known quantities prepared in the laboratory), the damage is inhibited, indicating a

positive test for the antibody. Neutralizing antibody is usually the first to appear, early in the

acute phase of the infection.

4. Precipitin (or immunodiffusion) test. With

this method, two separate wells are cut into a

semisolid substrate such as agar. Serum components are put in one well and antigen in the other.

As they migrate, an observable precipitation line

is formed where antibody and antigen combine.

No precipitation line forms in the absence of

antibody.

Counterimmunoelectrophoresis (CIE) and immunoelectrophoresis are adaptations of the precipitin test in which an electric current is passed

through the substrate, causing the reactive components to migrate more rapidly and with greater

resolution.

The relatively simple diffusion methods are

suited to the tropics, but these methods are of

limited practical value because of their insensitivity and the long period of time before results. The

electrophoretic methods require equipment and

a power supply, and the results are complex and

somewhat difficult to interpret (98).

5. Labeled immunodiagnostic reagent assays.

The various types of labeled immunodiagnostic

reagent assays all involve a similar procedure. The

test antigen is attached to a slide or test well (e.g.,

blood drops from malaria-parasitized monkeys

are dried on microscope slides; in vitro cultured

organisms are fixed to plastic test wells). The patients’ serum is incubated as a drop on the slide

or in the well. If antibody is in the serum, it binds

to the fixed antigen. The excess serum is washed

off, leaving the antibody sticking to the antigen

on the slide. Then a second antibody is added that

reacts with any antibody-antigen complexes that
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases ● 157

were formed. This second reaction depends on the

unusual structure of antibodies—one end of an

antibody is a totally unique fit for just one antigen, and the other end is a totally generic molecule. The second lab-prepared antibody has a

unique end which recognizes the generic end of

other antibodies. This second antibody is also

linked to a chemical marker that is detectable—a

fluorescent molecule, a radioactive molecule, or

an enzyme.

One type of labeled immunodiagnostic assay,

the indirect fluorescent antibody (IFA) test, uses

slides prepared with antigen attached (usually a

fixed, cultured organism). After serum antibody

binds to the antigen, the second lab-prepared antibody is added to react with any antibody-antigen complexes that were formed. This second

antibody is linked to a chemical marker that

fluoresces under ultraviolet light. A special microscope is used to detect fluorescence. If the slide

fluoresces, it indicates that the original human serum contained the antibodies in question. Microscope slides are easily prepared with antigens stable for long periods for a number of viruses,

bacteria, and parasites. Disadvantages of IFA tests

are that a well-maintained, carefully calibrated,

and expensive fluorescent microscope is needed,

the test is time-consuming and somewhat subjective, and with many helminths, there are crossreactions (388,421)

The enzyme-linked immunosorbent assay (ELISA)

is complex to describe, yet simple and elegant in

its determination. It is rapidly being adapted to

the diagnosis of a wide range of organisms. As

shown in figure 8-1, the ELISA for the detection

of antibody involves attaching specific antigen (a

known quantity of test organisms prepared in the

laboratory) to a test well or plate, then adding

the patient’s serum with suspected specific antibody, then adding a lab-prepared second antibody

which is linked to an enzyme. The final component added is a chemical substrate whose color

changes by the action of the enzyme. The degree

of color change is read qualitatively by eye or

quantitatively by photometric instrument to give

an indication of the amount of antibody present

in the serum specimen.

The ELISA uses small sample volumes. Large

numbers of specimens can be processed, so the

procedure is useful for epidemiologic studies. For

developing countries, the ELISA has clear advantages, since it can be done in simple laboratories,

and the reagents are stable if refrigerated. Widespread field testing of ELISA procedures is under

way for African sleeping sickness and Chagas’ disease, leishmaniasis, amebiasis, malaria, filariasis,

and schistosomiasis (82,353,389) One great advantage of the ELISA procedure is that the result

can often be judged positive or negative by the

naked eye. ELISA tests are rapidly being developed and improved.

The radioimmunoassay (RIA), a procedure similar to ELISA, can be carried out using a second

antibody labeled with a radioactive compound,

which is then read in a scintillation’ counter. RIA

is very sensitive and very quantitatively accurate,

but it requires a laboratory equipped to deal with

radioactive isotopes, which carry a risk and have

a short shelf-life. RIA has been a valuable research

tool, but not a practical diagnostic tool (335).

6. Other tests: circumoval precipitin tests

(COPT) are examples of the variety of serologic

tests used for detection of specific pathogens. The

COPT for schistosomiasis uses standardized prepared schistosome eggs obtained from animal infections. The eggs are incubated in patient sera,

which causes a characteristic precipitate to form

around the egg, if antischistosomal antibodies are

present.

The thin-layer immunoassay for the detection

of antibody is depicted in figure 8-2. This technique uses antigen in test plates to capture specific antibody from test serum. The antigen-antibody complexes form a thin layer that attracts

water. When the test plate is exposed to water

vapor, water visibly condenses on the immune

complexes in large droplets which form a pattern

distinct from the background. The thin-layer immunoassay has been used to detect the presence

of viruses, schistosomes, and amebae. It requires

relatively large amounts of antigen, however, and

in some cases has low sensitivity (421).
158 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Figure 8-1 .—Enzyme=Linked Immunosorbent Assay (ELISA) Technique for

Detecting Antigen or Antibody

1. Detection of antigen in a clinical sample:

● Antibody to the antigen being tested for is adsorbed to a plastic plate.

● The clinical sample is added. If the specific antigen is present, antigen-antibody complex forms on the

plastic plate.

● A second antibody to the antigen, with a specific enzyme attached, is added to the plate. The enzymeIabeled antibody adheres where antigen-antibody complexes were formed.

● An enzyme substrate is added. If it comes in contact with the enzyme, a reaction occurs that produces

a visible color change, If no complexes are formed, there is no reaction.

Il. Detection of antibody in a clinical sample:

● The process is the same as the one for detecting antigen, but includes one additional step: known antigen is added, resulting in one more layer in the final complex.

SOURCE: World Health Organization, Rapid Laboratory Techniques for the Diagnosis of Viral Infections, Technical Report Series

#661 (Geneva: WHO, 1961).

Serologic Methods for Detection of Antigen after clearance of an infection. Thus, false negaThe serologic techniques described above focus tive and false positive results often occur.

on antibody-detection; which has two inherent Direct immunologic detection of the antigen itdisadvantages (388): 1) there is always a delay be- self can be preferable. A serious drawback of this

tween infection and development of a detectable approach is that, compared to the presence of anlevel of antibody; and 2) antibody levels persist tibodies, antigen presence is a short-lived occur-
● 159
160 ● Status of Biomedical Research and Related Technology for Tropical Diseases

rence, especially for viral diseases. Current serologic methods for detecting antigen are described

below.

1. Labeled immunodiagnostic reagent assays.

The principal methods for detecting antigen are

labeled reagent immunoassay that are modifications of the ELISA, RIA, or IFA tests for antibodies (described above) and employ a known specific, lab-prepared antibody as the first reagent

to adhere to the plastic well. The ELISA technique

for the detection of antigen is shown in figure 8-1.

Labeled immunodiagnostic assays for detecting

antigens have been applied to viral infections,

amebiasis, toxoplasmosis, onchocerciasis (93),

schistosomiasis (84), malaria, Chagas’ disease,

other protozoan infection (8), acute respiratory

infectious (ARIs) caused by bacteria, and for detection of certain bacterial toxins causing diarrheal

diseases. These tests can identify minute quantities of antigen, but their sensitivity has still been

less than the sensitivity of direct examination techniques. As newer methods using MAbs are developed, sensitivity should improve greatly.

2. Agglutination assays. As indicated earlier,

clumping of particles due to the interaction of antibody and antigen is the basis of all agglutination tests. In the following variations, the diseaseproducing organism or its byproduct (an antigen)

is detected by complementary antibody fixed to

particles which are added to a test sample.

In the coagglutination (COA) test, a bacterium

(Staphylococcus aureus) is used as the earner particle for a specific antibody. When the antibody

is mixed with a serum sample, it will agglutinate

if the bacteria of interest are present. In a similar

test, the latex agglutination (LA) test, latex particles are the earners. Technical considerations determine which method is superior for the diagnosis of any particular agent.

The reverse passive hemagglutination (RPHA)

test, shown in figure 8-3, is used for rapid detection of viruses (smallpox, arboviruses, and hepatitis B). Here again, specific antibody is fixed to

particles that agglutinate if the appropriate antigen is present in the test serum.

Figure 8-3.—Reverse Passive Hemagglutination

(RPHA) Technique for Detecting Viruses

Detection of viruses in a clinical sample:

● Known antibody is added to and adheres to the surface of treated

erythrocytes (red blood cells).

● A clinical sample, suspected of containing the specific virus, is

added. If the virus is present, complexes are formed linking

antibody-coated erythrocytes together, visible in the test tube

as an agglutination pattern.

SOURCE: World Health Organization, Rapid Laboratory Techniques for the

Diagnosis of Viral Infections, Technical Report #661 (Geneva: WHO,

1981).
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases ● 161

Constraints on Serologic Diagnosis

Serologic diagnostic techniques provide some

attractive technical options. Such methods can be

very effective in epidemiologic surveys (203).

Some of these methods have been adapted for field

use in tropical areas (e.g., by absorbing a few

drops of blood onto a filter paper strip for later

examination in the laboratory). Serologic methods

also are more easily automated than techniques

of direct examination and thus have increased precision. Despite these features, however, very few

such immunologic methods have proved to be of

practical value in routine diagnostic laboratory

practice. Most have remained in the research laboratory (388).

Several constraints limit the usefulness of serologic techniques in medical practice or for public

health measures:

● Serologic procedures require sophisticated

laboratory instruments and equipment. In

many developing countries, basic services

such as transport, electricity, and water are

inadequate or unreliable, and trained personnel are lacking.

● In industrialized countries, where the expensive, sophisticated tests are feasible, few

●

●

●

●

centers specializing in “exotic” diseases have

the demand to justify doing the procedures.

The result is that most medical facilities rely

on the conventional techniques of direct examination.

Parasite antigens are highly cross-reactive.

Cross-reactivity, coupled with the presence

commonly of more than one organism in an

individual, makes the interpretation of serologic tests difficult.

Test reagents vary widely, and different laboratories can report grossly divergent results

using the same batch of antiserum. The need

for standardization and for high-quality antigens is acute (177,388).

“Paired sera” (from patients in the acute and

convalescent stages of illness) may be required if changing antibody titer is a criterion

for diagnosis.

Many individuals are reluctant to have blood

drawn.

Many of the antigens needed for serodiagnostic tests can be cultured in vitro. In vitro cultivation techniques are improving rapidly for many

of the agents of tropical diseases (296). The other

major source of precisely defined antigens for

diagnosis will be recombinant DNA technology.

GENETIC TOOLS FOR IMPROVING DIAGNOSTIC TECHNIQUES

It is one thing for a university medical center

to undertake the diagnosis of a single patient recently returned from the tropics and quite another

for a survey team to apply diagnostic methods

under field conditions to large numbers of local

people in isolated, endemic, tropical areas. For

use in the field, there is a great need for simple,

reproducible, and inexpensive diagnostic methods that can provide a clear-cut indication of the

status of an infection, and a specific identification of the causative agent. This need is especially

acute because of the current work on developing

vaccines for a variety of tropical diseases (see ch.

7). Without a means for accurate determination

(or at least reliable estimation) of the prior exposure and immune status of vaccinees and controls, and their post-inoculation followup, valid

field testing of these vaccines will not be possible.

Biotechnology (see ch. 5) is leading to the development of new diagnostic methods based on

the use of MAbs and recombinant DNA (or recombinant RNA) techniques. These techniques,

which are advancing rapidly, hold out a promise

of accuracy and simplicity. In addition, they generally require smaller amounts of clinical samples

than have heretofore been necessary. DNA and

RNA hybridization tests hold great promise, but

further development and testing are necessary before many of the tests can achieve widespread use

in the field (137).

Monoclinal Antibodies (MAbs)

Immunodiagnosis using MAbs is building on

the past 20 years of research in serologic diagnostics. The revolution is in making these diagnostic
162 . Status of Biomedical Research and Related Technology for Tropical Diseases

test procedures much more sensitive and specific,

more reproducible, faster, and more economical.

Three diagnostic uses of MAbs are being developed (84):

●

●

●

immunoassay of high specificity, i.e., making existing serologic diagnostic tests better

by production of purer reagents;

“two-site” immunoassay of antigen (the pathogenic organism itself or its byproducts) from

appropriate body fluids, using two monoclonals directed against different binding sites

of one antigen—one MAb to catch the antigen and another to label it for detection; and

the use of cross-reactive MAbs to absorb certain common antigens out of crude mixtures,

making possible the sensitive and specific detection of an antigen of interest.

MAbs have been used to distinguish between

organisms that cross-react in conventional serologic tests, e.g., Trypanosoma cruzi and Leishmania braziliensis (71); between closely related

species of New World Leishmania (283); between

strains within single species of Theeileria (277) and

Leishmania (145); between genetic variants (43)

or life cycle stages (111) of Trypanosoma rhodesiense; and between larval and adult Schistosoma mansoni (332).

Assays of antigen in which MAbs are affixed

to cellulose fibers or other solid substrates (e.g.,

plastic test wells) have been referred to as “dipstick technology” (136). Antigen present in blood,

urine, stool, etc., binds to the MAbs, and the combination is detected by a second antibody through

an ELISA-like color reaction. Immunodiagnosis

using MAbs as probes for parasite antigen has

been applied to such diverse conditions as tuberculosis (153), hydatid and other larval tapeworm

diseases (78,79), Chagas’ disease (9), onchocerciasis (93), and schistosomiasis (83,84,239). ELISA

procedures using MAbs have been described for

rotavirus infection (424), toxoplasmosis (8), and

schistosomiasis (240). MAbs have been used to

identify malarial organisms within mosquitoes

without the need for exhaustive dissection and

microscopy.

Nucleic Acid Hybridization Probes

The use of a nucleic acid (DNA or RNA) hybridization probe to identify the actual DNA or

RNA of a disease-producing organism (112) is a

promising diagnostic technique that has some important advantages, perhaps most important of

which is high specificity. Scientists are now able

to isolate and then reproduce (“clone”) DNA segments from known organisms, label the segments

radioactively, separate them from the normal

double-stranded state to a single-strand state, and

then test them against unknown specimens of

DNA (e.g., in a stool sample) for the ability to

hybridize (re-form double-stranded DNA). If the

suspected organism is present, then the labeled

DNA and the sample DNA, being the same, will

hybridize. The radioactivity or fluorescence or enzymatic color change can then be detected.

The nucleic acid hybridization techniques for

different organisms vary, but such techniques can

be developed to be simple, practical, and relatively inexpensive. Using these techniques, investigators can screen large numbers of samples at

one time. Samples can be collected and stored for

several weeks before the test is completed. Nucleic acid probes should be useful for large-scale

epidemiologic and surveillance studies (117,246).

The major disadvantages of the tests that use

radioactive labels are the radiation hazard and the

short shelf-life of the radiolabeled reagents. These

problems can probably be circumvented in most

cases by substituting tests that use color change

methods in place of radiolabels (196,202).

The uses of nucleic acid hybridization techniques to detect malaria parasites (122), Leishmania spp. (409), Escherichia coli (246), and rotavirus, and to detect Salmonella bacteria in food

products (116) are described below. The utility

of nucleic acid hybridization probes in the field

in developing countries must be further evaluated,

but good results have been reported in detection

of rotavirus and E. coli in stool sample dots. Although nucleic acid hybridization is still in its early

development phase, it may well progress to a predominant role in diagnosis.
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases ● 163

DIAGNOSIS: CURRENT STATUS FOR SELECTED TROPICAL DISEASES

Malaria

Since the emergence of drug-resistant malaria

parasites in the late 1950s, the importance of

specific diagnosis, including a determination of

whether the malaria parasites are drug resistant,

has been greatly elevated. Previously cheap, safe,

effective, and completely standardized antimalarial treatment regimens were available, and, presumptive treatment, without definitive diagnosis,

was freely administered. Now, however, appropriate treatment rests on accurate diagnosis.

Conventional Diagnosis

Diagnosis of malaria based on physical examination alone can be difficult, because malaria’s

symptoms are protean. For that reason, laboratory diagnosis of malaria is important. The standard method of diagnosis is by microscope examination of a stained blood smear made from a

finger-prick. The presence of malaria parasites is

definitive. Under field conditions, there are generally quite a few false negatives, because people

with malaria do not always have large numbers

of malaria parasites circulating in their blood. In

the absence of microscopically confirmed infection, a presumptive diagnosis of malaria may still

be made, based on clinical symptoms. Treatment

is given to reduce the parasite load of the population and, in effect, to prevent mosquitoes from

acquiring malaria.

Although methods for the serologic diagnosis

of malaria have long been available (CF, IHA, and

IFA tests), they are not widely used for two reasons. First, the need for equipment and materials

to perform the tests cannot always be met. Second, the tests demonstrate the presence of antibodies, which persist after cure and may indicate

previous infection. In an endemic country where

infection is always possible, such tests do not

greatly help the decision to treat. Nevertheless,

these tests have been useful for epidemiologic surveys (especially IHA and IFA tests (168)), and for

special purposes such as establishing the presence

of antibody in a particular patient in whom infection is suspected but cannot be demonstrated

directly (e.g., in a blood donor who is suspected

of having transmitted malaria to a blood recipient).

Recent Progress

Serologic Diagnosis. —Improved serologic

methods for the detection of blood stage malarial antigens are being developed: RIA and ELISA

tests for the detection of parasitized red blood cells

have been developed. Sensitivity of parasite detection with these tests is encouraging, and reproducibility should improve as standard reagents become available (353).

In the hope of establishing standard reagents

for malaria serology, the World Health Organization’s (WHO) Immunology Research and Training Center, Geneva, has established a registry of

malarial MAbs collected from other laboratories

and evaluated for potential value as serodiagnostic

reagents.

Sporozoite Diagnosis.—Immunologic work on

plasmodial sporozoites has led to the development

of two methods to test for sporozoites in mosquitoes. Such testing has importance for epidemiologic studies in determining the degree to

which various mosquitoes function as vectors of

species of malaria parasites.

In one method, MAbs are used to identify

sporozoites in mosquito squashes by a direct binding assay, which is species-specific. This method

was developed as an RIA, but an ELISA procedure is being developed as well. Although the test

procedure must be carried out in a centralized

laboratory, the samples are stable without refrigeration and easily handled, and the results are

available within an acceptable time interval for

epidemiologic purposes.

A second method, another new immunoradiometric test (inhibition of idiotype-anti-idiotype interaction, or “4 i-assay”), has been developed to

detect circumsporozoite protein (280) and is based

on inhibition of binding of two MAbs. The first

MAb is against malarial antigen, and the second

MAb is against the first MAb (thus called an idiotype, which resembles the original antigen). When

both MAbs are mixed in a test well, they bind to-
164 . Status of Biomedical Research and Re/ated Technology for Tropical Diseases

gether in a detectable way, unless the test sample

contains antigen. If the sample contains antigen,

the antigen will bind specific antibody, thus inhibiting the two MAbs from interacting (inhibition indicates the presence of true parasite antigen). This test is sensitive enough to distinguish

between different species of malaria parasites. The

method has general applicability. Furthermore,

because the two immunoglobulin reagents are

MAbs, this method does not require cultivation

and purification of antigen from parasite, and the

two immunoglobulin reagents are completely pure

(422).

DNA Hybridization. —Work on development

of a rapid diagnostic test using specific DNA

hybridization is proceeding. A recent publication

describes experimental success in identifying Plasmodium fa]ciparum parasites in samples of blood

from in vitro culture and from malaria patients

(122). This method is still at a very preliminary

stage in relation to any practical use, because the

sensitivity is no better than microscopic examination of a stained blood film, and the procedure

involves a number of laboratory steps that take

about 24 hours to complete.

In Vitro Cultivation of Malaria Parasites and

Microtest of Drug Sensitivity .-With the current

situation of widespread resistance to drugs by malaria parasites, diagnosis of drug susceptibility in

parasite isolates is an important epidemiologic

task. The method for in vitro cultivation of P.

falciparum, the malaria species with widespread

drug resistance, has been adapted to several techniques for testing drug susceptibility against the

primary antimalarial. The tests are available in

kit form from WHO.

Research Needs

Greater standardization of serologic tests for

the detection of malaria antigens and antibodies

is needed. These tests need to be adapted for field

applications, both as quick and easy diagnostics

at remote or poorly equipped treatment posts and

to assess any vaccination trials that may be attempted in the future. Assessment of vaccination

trials will rely on detecting antibodies in individuals who did not have antibodies before vaccination as well as determining infection rates

post-vaccination.

A quick and easy field method is needed to

establish the existence of infection (to conserve

drug for true cases) and to differentiate species

of Plasmodium (for appropriate drug type). Similarly, a field method to detect infection in mosquito vectors is needed to assess the impact of vaccination on the overall risk of transmission.

A less urgent but important need is a method

of screening blood bank donations to prevent

transfusion malaria.

Schistosomiasis

The major schistosomes infecting humans are

Schistosoma mansoni, S. japonicum, and S. haematobium. Current control for schistosomiasis

calls for the identification and treatment of all infected persons at regular intervals (usually 6

months) (353). Quick and reliable diagnosis is essential to identify people infected and for assessing the effectiveness of the treatment. To determine optimal treatment, it is necessary not only

to diagnose the presence of infection but also to

determine how heavy the parasite load is.

Conventional Diagnosis

Direct examination of feces or urine for characteristic eggs is the classical method of diagnosis of schistosomiasis. There are a number of

methods for concentrating the sample to maximize

the chance of detection and a number of methods for accurately estimating the number of eggs

in order to estimate the parasite burden in the

host. Cytoscopy or sigmoidoscopy (viewing through

instruments inserted into the body) is occasionally used to detect lesions, and rectal biopsy is

sometimes used. There is a CF test, an intradermal skin test, and the COPT to help confirm

diagnosis.

Recent Progress

Two new techniques have been developed for

S. haematobium diagnosis. One is the filtration

of urine using a reusable plastic woven filter which

isolates excreted eggs. The second is a prototype

image processing and pattern recognition apparatus for automated S. haematobium egg counts

that has been tested in the laboratory and is undergoing field trials in endemic areas (353).
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases ● 165

A new diagnostic kit has recently been made

available by the Program for Appropriate Technology in Health, a nonprofit, nongovernmental

organization. The kit is for the diagnosis of S. haematobium, and it is designed for quick, practical, and reliable field use.

In a special collaborative study sponsored jointly

by WHO and the Edna McConnell Clark Foundation, eight research laboratories evaluated a

number of procedures for immunodiagnosis of

schistosomiasis (247). Included were the COPT,

ELISA, IHA, IFA, RIA, and other procedures,

testing a pool of banked sera against a variety of

schistosomal antigens. A study of this type is of

great value in developing and standardizing materials and methods for immunodiagnosis.

Several candidate antigens for immunodiagnosis have been identified (59). There are several

modifications of the ELISA procedure, including

an inhibition-ELISA used to detect and characterize schistosomal antigens (l). Many laboratories have developed MAbs for identification of

relevant antigens or for species diagnosis (e.g.,

1,83,84,99,237,238,239,240) .

Many workers are attempting to identify and

isolate relevant protective antigens, and at least

half of those are actively engaged in gene cloning experiments. Several laboratories are devising improved diagnostic methods, primarily with

ELISA-based tests. Most schistosomiasis experts

believe that effective MAb-based diagnostic tools

are very near or within 5 years of introduction

(16).

Research Needs

Field methods for quick and easy diagnosis of

schistosomiasis are being introduced but still require evaluation and standardization.

Trypanosomiasis

African Sleeping Sickness

(African Trypanosomiasis)

There are two forms of African sleeping sickness in humans. Trypanosoma brucei gambiense

causes the chronic form found in west Africa. T.

b. rhodesiense causes the acute form in east Africa

and also infects livestock over large areas of the

continent.

Conventional Diagnosis.—Direct examination

for African trypanosomes is done from stained

blood preparations, but parasites are extremely

difficult to find. In the chronic form (T. b. gambiense), fluid drawn by needle from lymph nodes

in the neck is examined. The number of parasites

detected varies daily. In the acute form (T. b.

rhodesiense) too, parasites may vary in density,

making diagnosis difficult. Cerebrospinal fluid

may also be examined. Inoculation of laboratory

animals or culture on appropriate media to allow

the parasites to multiply is sometimes useful.

Immunodiagnostic techniques have been available for many years (IFA and CF tests), but these

must be performed in central laboratories. The

ELISA has also been found effective in the laboratory but not for the field (353). All of these

tests are valuable for epidemiologic studies, but

the delay in reaching a diagnosis limits their usefulness for patient care.

Recent Progress. —Tests for antibodies against

T. b. gambiense, which can be read within minutes and carried out with blood obtained from a

finger-prick, are under development and evaluation in the field. They are the card agglutination

test for trypanosomiasis (CATT), the Cellognost

test (a commercial technique based on indirect

haemaggIutination), and the Tryptest. CATT is

being evaluated on a large scale in west Africa.

This test has been found to be as specific as the

IFA test. It is easily transported to the field, requires little technical skill, and gives results within

minutes. The lack of stability of the antigen and

storage under field conditions are problems to be

worked on. Antigens to diagnose T. b. rhodesiense are being sought (353).

Two other techniques are also being evaluated:

the miniature anion exchange column technique

and the microhaematocrit buffy coat centrifugation method. Both have shown greater sensitivity than blood film examination (208). Development of the double centrifugation technique for

the detection of trypanosomes in cerebrospinal

fluid has improved diagnosis of central nervous

system involvement (353).
166 ● Status of Biomedical Research and Related Technology for Tropical Diseases

The three subspecies of the Trypanosoma brucei species complex are morphologically indistinguishable. Two species (T. b. rhodesiense and T.

b. gambiense) are infective to humans, causing

sleeping sickness; the third (T. b. brucei) is infective to wild and domestic animals but not to humans. Scientists’ inability to distinguish between

human and animal forms has epidemiologic importance for determining risk to humans where

animals are found infected. Culturing of trypanosomes in human serum is widely used for determination of infectivity to humans. Isoenzyme

electrophoresis, DNA hybridization, and comparative IFA with standardized sera have also been

used recently.

Research Needs. —Several avenues of research

may be nearing fruition for the diagnosis of African trypanosomiasis. There is a need to develop

sensitive and specific techniques suitable for field

use.

Chagas’ Disease (American Trypanosomiasis)

Chagas’ disease, caused by the protozoan parasite Trypanosoma cruzi, affects more than 12 million people in Latin America (229). Acute and

chronic phases of the disease vary somewhat from

one region to another (166). Most damage is done

by tiny, nonflagellated forms within the cells of

heart muscle and certain nerve ganglia.

Conventional Diagnosis.—The clinical picture

of Chagas’ disease is variable. Examination of

blood during the acute phase of the disease may

reveal parasites. Because of commonly low parasite density in blood, several culture techniques

are used to allow the parasite to multiply to a detectable level: inoculation into laboratory animals

followed by periodic examination over 60 days;

in vitro culture of blood; xenodiagnosis (allowing clean, uninfected reduviid bugs to feed on the

suspected patient and then examining the hindgut of the bug for trypanosomes after 2 weeks).

All these methods often require repeated attempts.

A CF test (the Machado-Guerreiro test) using

T. cruzi antigen is available, as is other serologic

diagnosis for the direct detection of antibody.

Conventional serologic tests for Chagas’ disease

cross-react with leishmaniasis, leprosy, and

syphilis antigens and are not sufficiently sensitive

to detect Chagas’ disease with assurance. A number of groups are working on development of an

immunodiagnostic reagent, and optimism is generally high. Immunodiagnosis of specific antibody

is available as a procedure in centralized laboratories, but standardization between laboratories

is a problem.

Recent Progress. —Standardization of serodiagnostic techniques has been promoted by the Special Program for Research and Training in Tropical Diseases (TDR). A serum reference bank is

now providing standardized lyophilized (freezedried) serum samples to laboratories throughout

Latin America, and a network of collaborative

laboratories has developed protocols for the

standardization of reagents, techniques, and procedures (353).

Many workers have developed MAbs that may

be useful for the detection of various trypanosome

antigens (9,259). Development of ELISA diagnosis is under way in a number of laboratories. Investigators have shown that circulating antigens

in acute infections can be detected (7).

A specific diagnostic test is being evaluated

using purified cell membrane antigens of T. cruzi

which are fixed on polyamide strips. After exposure to suspected serum, the strips are treated

as in ELISA-type tests to detect any antigenantibody complexes that would form if the serum

were from a person with Chagas’ disease. Preliminary results indicate that the test greatly reduces,

but does not eliminate, nonspecific reactions.

Blood-transfusion-transmitted Chagas’ disease

in Brazilian hospitals is a serious problem, and

better screening methods for donated blood are

needed. An agglutination test for rapid screening

of donated blood for T. cruzi infection is undergoing evaluation in Brazil, where it was developed, and in a network of collaborating laboratories (353).

A DNA-DNA hybridization probe shows

promise of detecting the presence of T. cruzi organisms or DNA fragments present in the blood. T.

cruzi, has a unique, highly variable type of DNA

(kinetoplast DNA) that can be used to distinguish

species and strains within species. A technique for

isolating DNA, cutting it into pieces, and then sep-
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases ● 167

arating the DNA into recognizable patterns (restriction endonuclease finger-printing of kinetoplast DNA) promises to provide a valuable new

tool for epidemiologic and clinical purposes. New

subdivisions of T. cruzi strains can be demonstrated using this method. These refinements in

the taxonomy of the parasites may help to explain

the variability of the disease in different localities, including the variation in clinical symptoms

and the variable susceptibility or resistance in

hosts (353).

Research Needs.—A quick, easy, and reliable

method is needed for diagnosis of acute Chagas’

disease (when treatment might be prophylactic,

and because the treatment is toxic). A means to

predict prognosis in different geographic areas

also is desirable.

A sensitive test for screening donated blood is

also particularly important in Chagas’ disease, as

infection by transfusion is a serious problem in

endemic areas.

.“

Leishmaniasis

Leishmaniasis is a disease with three clinical

presentations depending on the leishmanial

parasite species. Cutaneous leishmaniasis, caused

by either L. tropica, L. mexicana, or L. brazilienis

(depending on geographical location), is a self-limiting and usually self-resolving sore at the point

of infection. Mucocutaneous leishmaniasis, caused

by L. brazilienis, begins as a sore but commonly

metastasizes and proliferates in the nasal and

pharyngeal mucous membranes. Visceral leishmaniasis, or “kala-azar,” is caused by L. donovani

and affects the spleen, liver, bone marrow, and

lymph glands.

Conventional Diagnosis

Diagnosis of leishmanial organisms is complicated by the rather uniform appearance of different species under the light microscope, and crossreactivity of different species with conventional

serologic diagnosis. Until recently, microscopic

identification and conventional serologic techniques were the only techniques available for diagnosing leishmaniasis.

●

Photo credit: Office of Technology Assessment

Leishmanial organisms as seen through a

light microscope.

The inability to distinguish correctly between

species of Leishmania can have serious consequences for patients. For example, the lesions of

L. mexicana and L. braziliensis are very similar

at first appearance, and both species overlap in

many parts of South America. Without treatment,

L. mexicana is self-resolving, but L. braziliensis

progresses to gross destruction of the nose and

throat. Thus, treatment and followup for L.

braziliensis infection are critical. But the treatment

is itself highly toxic and clearly not to be used for

those not requiring it.

Recent Progress

Investigators have developed a DNA hybridization probe to distinguish between L. mexicana
168 ŽStatus of Biomedical Research and Related Technology for Tropical Diseases

and L. brazdiensis (409) using kinetoplast DNA,

a unique form of DNA in Leishmania and Trypanosoma species. Kinetoplast DNA is extracted

from growing cultures of various species of Leishmania organisms, processed and labeled radioactively. Test material is collected from the patient

as “touch preparations” on nitrocellulose filter paper from suspected lesions. Hybridization and

analysis are then carried out. Kinetoplast DNA

hybridization is highly species-specific and provides a relatively rapid means of diagnosis direct

from infected tissue. This has now been tested for

the diagnosis of human patients and shows promise.

Another test, called the “DOT-ELISA” test, has

been developed, and represents a major advance

in the rapid field diagnosis of visceral leishmaniasis. It also is useful for field surveys for identifying infected vectors (267).

In recent years, scientists have prepared MAbs

against a variety of antigenic determinants in

Leishmania species (71,91,139,145,171) and used

them to probe for specific morphologic and taxonomic differences. Among the many monoclonals

produced, some recognize antigens common to

all kinetoplastid (Leishmania and Trypanosoma)

species tested; others bind only to certain strains

within a single species. The process of sorting out

these specificities and defining the precise nature

of the reactive leishmanial antigens should produce advances in knowledge and diagnosis of

these organisms.

Research Needs

The diagnosis of Leishmania species is very important, because different species produce similar lesions but have very different long-term consequences. DNA hybridization looks promising

but needs development for practical use.

In the clinical-epidemiologic area, there are

many strains and types of Leishmania, but a lack

of a good classification system. More field surveys

to determine prevalence of disease are needed, and

those surveys will require practical field tests.

Filariasis

Filariasis is a collective term for several distinct

parasitic infections by insect-transmitted, tissuedwelling nematodes. The principal species are

Wuchereria bancrofti and Brugia malayi, which

cause filarial elephantiasis; and Onchocerca volvulus, the cause of onchercerciasis (river blindness) in west Africa, also found in Central and

South America.

Conventional Diagnosis

Conventional diagnosis of filarial infections

has severe limitations. For lymphatic filariasis

(Wuchereria and Brugia), diagnosis is made by

microscope examination of stained blood films to

find microfilariae. Density of microfilariae is usually very low, especially during the early stages

of infection; and for several species, it is cyclical

according to a circadian rhythm, so at times there

may be no microfilariae present in the blood.

Diagnosis of onchocerciasis (in which the subcutaneous tissues are infected) is made by using

a special surgical punch to take tiny snips of skin

and then examining this skin under the microscope. Because of cross-reactions with other organisms, no dependable serologic diagnostics for

filarial infections are available.

Microfilariae of Onchocerca volvulus were found in

skin snips from these nodules.
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases •169

Recent Progress

A number of MAbs to filarial antigens have

been produced, but immunodiagnosis has been

hindered by a high level of cross-reactivity with

other helminth antigens. Excretory-secretory antigens, which are released by the parasite and circulate in the host’s blood, and surface antigens

on the parasite itself are being assessed for use in

immunodiagnosis of filaria. The demonstration

of circulating antigens in Onchocerca infections

has made a diagnostic MAb feasible. A good

correlation between circulating antigen and presence of parasites was found in one study, using

a MAb in an RIA (93). However, the false positive rate was high, suggesting that further refinement of this test is needed. At a workshop held

in 1983, 26 researchers from around the world

brought about 50 antifilarial MAbs representing

the total successful effort to that date (16).

Tests for detection of specific antibodies against

filarial worms, for circulating parasite antigen,

and for parasite surface antigen are being developed. There is optimism about the possibility of

a breakthrough in filarial detection within the next

few years. At least one Federal Government laboratory, one in industry, and one in a university

are conducting or planning recombinant DNA

work for expression of such filarial antigens.

A problem in diagnosing filariasis is the identification of parasites found in wild-caught vector insects. The problem of confusing intermediate life stages of disease-producing organisms with

other organisms of little public health significance

is common to vector-borne diseases, and represents a special problem in diagnosis which is

amenable to solution by modern methods. In

areas of west Africa with active vector control

programs against onchocerciasis, new insects with

potential vectorial capacity are entering controlled

areas from adjoining regions. Some of these insects are naturally infected with filarial larvae,

possibly parasites of wildlife, that cannot be distinguished by direct examination from larval Onchocerca. The development of probes, either by

MAbs or possibly DNA hybridization, would permit identification of these nematode larvae and

determination of whether they pose a threat to

humans.

Research Needs

Very generally, improvements in all aspects of

the diagnosis of filarial infections are needed. Because much remains to be learned about the epidemiology and pathology of filariasis, useful diagnostic tests are needed. It is important both for

patient care and for research purposes to be able

to distinguish animal and human filariae, a task

which at present is not always possible.

Leprosy (Hansen’s Disease)

Leprosy, caused by the bacillus Mycobactetiuzn

leprae, is the only bacterial infection among the

six diseases targeted by TDR. Since the 1950s, the

WHO-recommended strategy of leprosy control

through early case finding, followup of contacts,

and chemotherapy of patients has proved to be

difficult to implement and sustain in many countries (353).

Conventional Diagnosis

The initial diagnosis of leprosy is through recognition of areas of skin that lack feeling (anesthesia) and may be discolored or slightly raised.

Definitive diagnosis is made by acid-fast staining

and microscope examination of skin biopsy smears

from these suspected lesions. This technique is useful in all forms of leprosy, but there maybe few

bacteria in milder cases, making detection uncertain.

The lepromin or Mitsuda test, a skin test similar to the tuberculin skin test (see below), is used

as a prognostic test after leprosy is diagnosed. It

tells where the patient is along the immunopathologic scale of disease. A crude suspension of killed

bacilli (derived from human leprosy patients) is

injected under the skin of the patient. About 3

weeks later, the skin reaction is assessed, allowing a prognosis to guide treatment.

A lymphocyte transformation test has been

available for a decade as an indicator of M. leprae

infection and the potential course of the disease

(133). This test is an indicator of cell-mediated immune response. Despite its technical difficulties

and subjective interpretation, the lymphocyte

transformation test has given useful results in experimental field studies of exposure to leprosy an-
170 ● Status of Biomedical Research and Related Technology for Tropical Diseases

tigen, though it has not become a practical method

for routine use. Another test of the cell-mediated

immune response, microphage migration inhibition, is also not used routinely (168). It also possible to isolate and diagnose leprosy by inoculation of a clinical specimen from a suspected case

into the footpad of a mouse.

Recent Progress

A phenolic glycolipid molecule has recently

been isolated and identified as a unique and specific antigen that is abundant on M. leprae and

in the skin lesions of leprosy patients. This antigen has been used in an ELISA and has proved

to be specific for antileprosy antibody (56). It has

been tested experimentally to detect antibody in

the blood of leprosy patients, with excellent results (433). This antigen has the potential to be

used in a specific diagnostic test for leprosy. Investigators have found that contacts of known

cases are more likely than other people to develop

antibodies to this antigen. The phenolic glycolipid

test may therefore be useful in screening persons

at increased risk of developing clinical disease.

A number of apparently M. leprae-specific antigens (other than the phenolic glycolipid antigen)

have been reported (30,41,149), and MAbs have

been produced, which may also prove useful as

diagnostics. Serologic tests using IFA, ELISA, or

RIA are being developed for epidemiologic studies.

The fluorescent leprosy antibody absorption test

has shown high specificity and is being evaluated

for predictive value in long-term epidemiologic

studies (2). Skin tests have been developed for

monitoring delayed-type hypersensitivity reactions to M. leprae after immunization, though the

current test antigens are crude extracts that lack

specificity. Other skin test preparations using

soluble antigens are being evaluated in the field

for predictive value (224,232,346).

Research Needs

A method of culturing M. leprae in vitro is an

urgent need. Such a method could lead to improved methods for early diagnosis, which in turn

would lead to earlier treatment and favorable

prognosis. Practical techniques to diagnose leprosy for epidemiologic studies need to be developed to allow better understanding of transmission and susceptibility.

Progress in development of serologic tests that

are sensitive and specific has raised optimism

about prospects for achieving practical diagnostic techniques for leprosy. The phenolic glycolipid

antigen is most promising at present and may be

developed into a useful diagnostic test. At present,

the main source of this antigen and others is from

M. leprae grown in armadillos. Obtaining antigens from M. leprae grown in armadillos is a slow

process. It may be necessary or useful for diagnosis to make important molecules from M. leprae

using recombinant DNA.

A method for the differentiation of patients

with lepromatous leprosy (the severe form with

poor prognosis) and tuberculoid leprosy (the mild

form with good prognosis) is needed to improve

understanding of the epidemiology of the disease

and for the early recognition of individuals at high

risk of developing disease. The evidence for a

genetic basis to resistance needs further study.

Tuberculosis

Tuberculosis remains a major threat to health

in many parts of the world, causing several million deaths annually. Tuberculosis control programs are built around vaccination coupled with

early case detection, treatment, and followup of

active cases. Diagnostic methods and early diagnosis before symptoms are overt are critical to the

control of this disease.

Conventional Diagnosis

The consideration of clinical symptoms and examination of direct sputum smears for the presence of the causative tubercle bacilli are the conventional means of diagnosing tuberculosis. If no

bacilli are found in a direct smear but tuberculosis still is suspected, culture isolation of bacteria

from a clinical specimen (usually sputum, though

urine, spinal fluid, or tissue biopsy may be appropriate) is attempted. Culturing also is used to confirm a diagnosis made by a direct smear. Isolation of bacilli from culture is the preferred

method, but several serious problems arise: the

need to decontaminate the specimen to prevent
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases ● 171

overgrowth by contaminating bacteria from the

oral cavity; the need to collect and culture multiple samples from a suspected case; and the slow

multiplication of the bacilli, which means that it

may take 3 to 6 weeks for growth to appear.

Chest X-rays continue to be of great value in

the diagnosis of tuberculosis, particularly in areas

where other diseases (like histoplasmosis) with

similar X-ray appearance are absent. X-rays are

particularly useful for determining (by noting differences between earlier and later films) that a dormant case has been reactivated.

Microscope examination of stained sputum

smears can be used to quickly identify tuberculosis-like bacteria, but does not distinguish virulent tubercle bacilli from look-alikes. In areas of

the world where active pulmonary tuberculosis

is common, a presumptive diagnosis can be made

on the basis of numerous bacteria with particular staining qualities (acid-fast) in the sputum. It

is clear, however, that many different species of

Mycobacterium, in addition to M. tuberculosis,

can infect humans and cause tuberculosis-like disease. Such environmental mycobacteria (called

“atypical mycobacteria”) are diagnosed by their

characteristics in culture, and cannot be distinguished on direct sputum examination. Some of

these (the M. avium-intracellulare complex) are

associated with acquired immunodeficiency syndrome (commonly known as “AIDS”).

The tuberculin skin test is used to identify people infected with tubercle bacilli, by means of an

allergic reaction to tuberculosis antigens (delayedtype hypersensitivity). A small amount of tuberculoprotein is introduced into the skin, and the

person is observed for an inflammatory reaction

2 to 3 days later. Old tuberculin (a crude concentrate of tubercle bacilli in culture medium) has

been replaced by purified protein derivative

(PPD), a purer, more standard material.

Three techniques can be used to introduce the

tuberculin: 1) the Mantoux test, in which PPD is

injected intradermally; 2) the Vollmer patch test,

in which tuberculin is applied to the skin on a

gauze adhesive strip; and 3) the Tine test, in which

the tuberculin is dried on the points of a standard puncture device that is pressed into the skin.

All of the tests have advantages and disadvantages. None of these tests can be used to test anyone already vaccinated against tuberculosis (a

large percentage of the population in many developing countries and in a number of developed

countries), because vaccinated individuals should

react to the challenge. The Mantoux is the most

sensitive and reliable test, because a standard volume and amount of tuberculin is injected directly

into the skin; however, the procedure of injection

by hypodermic is a practical disadvantage. The

Tine test is a rapid and easy test for use in large

population groups, but it gives a relatively high

number of “false positives” (people who test positive but who actually are not infected); positives

must be followed up by a Mantoux test. The

Vollmer patch testis useful for skin-testing infants

and children, but it is less sensitive than the Mantoux test.

A positive tuberculin skin test may be caused

by infection with other species of mycobacteria

or as a result of previous vaccination. A negative tuberculin test is strong evidence against the

tuberculosis diagnosis, but can also result from

loss of potency in stored PPD, and is occasionally observed in far advanced cases.

Recent Progress

ELISAS have recently been applied to the diagnosis of active tuberculosis pulmonary infection

(183). Application of recombinant DNA methods

to M. tuberculosis is in its infancy, but several

investigators are planning projects. In a handful

of laboratories including the National Institutes

of Health and several academic institutions, MAbs

are being reacted with M. tuberculosis to isolate

and purify antigens that may be useful as diagnostic targets and may be the basis for a new tuberculosis vaccine.

Attempts are being made to isolate specific antigens for skin tests in an attempt to make tuberculin skin tests less cross-reactive to infections

with other types of mycobacteria. Several tuberculosis researchers also work with M. leprae, the

related bacterium that causes leprosy.
172 . Status of Biomedical Research and Related Technology for Tropical Diseases

Research Needs

Tuberculosis may be the most serious infectious

disease problem in many developing countries,

as much a social as a scientific dilemma. There

is an obvious need for better vaccination and diagnosis. Diagnostic methods are needed to differentiate the “atypical mycobacteria, ” such as M. intracellulare and M. smegmatis, which hinder

diagnosis and seem to interfere with the effectiveness of BCG (Bacillus Calmette-Guerin) immunizations.

Diarrheal and Enteric Diseases

Diarrhea] and enteric diseases are caused by a

variety of viruses, bacteria, protozoa, and worms,

and a single individual often is infected with several at one time. Full understanding of the various etiologic agents will come only when simpler

techniques of diagnosis are available. Clinical

diagnosis of diarrheal disease calls for immediate

institution of therapy. Fortunately, dehydration

therapy is appropriate for all diarrheal disease (see

ch. 9). Nonetheless, identification of specific pathogens is essential for understanding the distribution of different agents and to establish priorities

for specific actions.

Viral Infections

Viruses are now recognized as important agents

of gastroenteritis. The principal agents are rotaviruses and Norwalk agents, though adenoviruses,

astroviruses are enteroviruses, coronaviruses, and

calciviruses are also found in fecal specimens (15).

Conventional Diagnosis.—Most of the viruses

that cause diarrheal and enteric diseases do not

grow under ordinary cell culture conditions, so

direct diagnosis of viral antigen in stool specimens

or detection of a serologic response is necessary.

Electron microscopy is a sensitive and simple

method for detecting the presence of virus (if an

electron microscope is available), but only limited

numbers of samples can be processed.

A variety of immunologic methods have been

developed for detection of viral antigen in stool

samples. The ELISA, IFA test, RIA, and CIE produce good results, though ELISA and RIA have

the greatest sensitivity.

Recent Progress.—Progress has been achieved

in isolating and cloning rotavirus DNA and then

developing DNA hybridization assays (117). Cloned

DNA hybridization probes have been used on

stool specimens for rotavirus diagnosis. The utility

of DNA hybridization probes in the field in developing countries must be further evaluated, but

good results have been reported in detection of

rotavirus in stool samples taken in remote areas

of Venezuela (146).

A test “kit” for rotavirus based on an ELISA

has been developed and evaluated and is now in

use by more than 50 investigators in the field. A

second generation ELISA test based on the use of

MAbs is being developed (427).

Bacterial Infections

Conventional Diagnosis.—Definitive identification of bacteria causing diarrhea is by the isolation of the agent through culture of clinical samples (e.g., stool samples for Shigella and Vibrio

cholerae; blood or stool samples for Salmonella.

Culture of V. cholerae is relatively simple and

gives a result in 18 hours.

Serologic diagnosis of Salmonella typhi, the

cause of typhoid fever, is possible because specific agglutinins appear in the blood at 7 to 10 days

of illness (the Widal reaction). For cholera, rise

in titer of specific agglutinins or antibodies confirms the diagnosis.

Two tests for the isolation and identification

of enterotoxigenic E. coli have been available for

a number of years: one uses a miniculture of adrenal

cells (302), and the other uses suckling mice (90).

Recent Progress. —Although E. coli is one of

the most intensely studied of all organisms, with

many thousands of research publications on all

aspects of its biology and biochemistry, much remains to be learned about its relation to diarrheal

disease. The several strains of E. coli that produce

diarrheal and intestinal disease in humans are recognized and described on the basis of clinical

pathology (“enterotoxigenic,” “enteropathogenic,”

“enteroinvasive,” see ch. 4). All of these characteristics are under genetic control, and investigators in several laboratories are identifying and

cloning the genes that code for attachment and
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases ● 173

colonization, virulence, toxin production, and antibiotic resistance. With the genes in hand, diagnostic procedures can be developed.

DNA hybridization probes for field identification and typing of E. coli are under development

in several laboratories. A DNA hybridization test

for identification of enterotoxigenic E. coli has

been available for several years (41,42) and now

has been tested in the field (103,312). The DNA

hybridization test has been shown to be very specific, reliable, stable, and sensitive (1,000 times

more sensitive than the standard assays). It appears to be a valuable tool for epidemiologic

studies.

The WHO Control of Diarrheal Diseases (CDD)

program has evaluated the “Biken” gel diffusion

test for detection of certain strains of E. coli. The

test is simple to perform, accurate, reproducible,

and has potential for use in developing countries.

Commercial production and marketing of the materials and reagents used in the test is being pursued (427).

The CDD program is also evaluating ELISAS

for diagnosis of strains of E. coli. Results indicate

a potential for widespread application, though

further development is needed to make it suitable

for routine diagnostic use (427).

A DNA-DNA hybridization probe was used by

one team of researchers to detect Salmonella bacteria not in stool samples but in food products

(116). This type of application may provide far

more serotype-specific identification of contaminating organisms than conventional culture methods, without the need for incubators, sterile media, and glassware.

Cholera is being studied by four or five researchers in the United States. Recombinant DNA

libraries are being constructed to collect gene sequences from wild-type organisms. Studies are under way on the transmissible genetic elements isolated from V. cholerae from endemic areas such

as Bangladesh, and differences between toxigenic

and nontoxigenic organisms are being defined (16).

Campylobacter jejuni is now recognized as an

important diarrheal agent, but epidemiologic

study is hampered by lack of a serotyping technique. Antigenic studies are under way to develop

a serotyping system. A simple slide agglutination

technique to identify antigens is under evaluation

in a number of developing countries (49).

Acute Respiratory Infections (ARIs)

ARIs are caused by a range of etiologic agents—

viruses, bacteria, rickettsia, and parasites—presenting great diagnostic complexity. Although

most of these agents can be specifically diagnosed,

the process normally requires multiple serologic

tests of various kinds.

Many diagnostic procedures based on isolation

and culture of the organism or based on comparison of an initial and a later serum sample (“paired

sera”) do not provide timely enough information

to be relevant to individual patient care. Also, to

a large degree, the diagnosis of ARIs can be and

is made on the basis of clinical symptoms, because

no matter what the diagnosis, specific therapeutics are lacking, especially for the many viral

diseases.

Because much of the treatment of ARIs consists

of providing symptomatic relief, the identification of specific disease-causing agents is not so important. In lieu of specific diagnosis flowcharts

or decision trees, using a patient’s symptoms as

criteria may be more appropriate for use by medical staff at all levels of the health systems of developing countries.

Still, in many cases, the etiologic agent needs

to be identified. For the treatment of the individual patient, diagnosis permits rational use of available therapy. The nonspecific prophylactic or

placebo use of antibiotics for any undiagnosed

ARI is greatly abused. Bacterial infections can be

treated with common antibiotics, but for viral

agents, only symptomatic support and relief are

proper (with the exception of viral influenza for

which an antiviral drug is available).

For public health needs, specific diagnosis is

necessary to assess, plan, implement, and evaluate interventions against epidemic outbreaks and

endemic infections.

Conventional Diagnosis

Faced with an individual patient with respiratory infection, the clinician or epidemiologist must
174 ŽStatus of Biomedical Research and Related Technology for Tropical Diseases

make simplifying judgments to decide which diagnostic procedures are in order. Simple microscope

examination of nose or throat secretions can be

useful, though limited, in ruling out possible etiologic agents.

Culture and isolation of the pathogen leads to

a definitive diagnosis, but the growth takes time

(usually at least 2 days), delaying the diagnosis,

For viral agents and even bacterial agents, this

procedure requires careful collection; special handling; storage and transport; tissue culture facilities (including appropriate culture media, and cell

types for virus growth); and equipment for serologic diagnosis (e.g., immunofluorescent microscope, scintillation counter, electrophoretic equipment). Prior antibiotic treatment or contamination

of the sample at any of several points from collection to inoculation, in the mature medium can

easily lead to incorrect results.

Serologic diagnosis by most of the standard

tests can demonstrate specific antibody in the individual’s serum, but this is not definitive, since

antibody from previous infections can persist after cure. Active infection is demonstrated when

the serum titer of antibody in a later convalescent sample (after 1 to 3 weeks) is higher than an

early acute sample (“paired sera”). This means the

diagnosis is often confirmed after the infection has

been resolved. This is acceptable for epidemiologic use, but not so useful for individual patient

care.

Recent Progress

Viral Infections.—With the IFA test and ELISA,

the diagnosis of respiratory viruses can now be

made in a few hours after specimen collection

(52,123,125,418). This is a great improvement

over isolation by tissue culture which takes several days. With appropriate antisera, the following viral antigens can be identified by the IFA test:

influenza (types A and B), respiratory syncytial

virus (RSV), measles, adenovirus, parainfluenza

1,2,3,4 (125,264). The economy of time permits

large numbers of specimens to be processed.

ELISA appears to be very useful for detecting several viruses, but additional experience is needed

to evaluate it (264).

Even with a rapid serologic diagnosis, the evidence is only indicative because of possible contamination (i.e., if the clinical sample is negative

but the culture is positive, contamination of the

culture is suggested rather than a positive diagnosis).

influenza.-Because influenza infection may be

fatal, and outbreaks occur annually, with major epidemics and pandemics occurring sporadically, the

three main types of influenza, with numerous subtypes and strains, are monitored epidemiologically.

Precise strain typing is carried out in order to prepare effective vaccines. Virus isolation and identification from tissue culture is available in 48 hours,

but the definitive result may take a week or more.

HI and CF tests produce a result in 24 hours, but

tests of acute and convalescent serum (samples

“paired sera”) separated by an interval of 2 to 3

weeks provide definitive diagnosis. The neutralization test is useful but expensive and time-consuming.

The direct or indirect immunofluorescent technique

can be used with cells obtained from the respiratory tract for a rapid diagnosis even when the patient has no symptoms.

Respiratory Syncytial Virus (RSV).–Precise diagnosis of RSV requires culture and isolation of the

virus from tissue culture, then identification with

an IFA test. CF or neutralization tests can be used

for serologic diagnosis in the patient. Completing

the positive identification may take up to 2 weeks,

though preliminary results can be obtained by early

examination of the cell culture.

Fluorescent antibody tests have been developed

to permit rapid diagnosis of RSV infections. If the

blood sample is transported promptly to the laboratory and processed immediately, the diagnosis

can be made in 4 to 6 hours (264).

Parainfluenza. —Precise diagnosis of parainfluenza viruses requires culture and isolation. Hemagglutination tests identify the virus after 5 to 10 days,

while immunofluorescence tests can make the diagnosis in 24 to 72 hours. Detection of a rise in serum

antibodies in the individual can be made with the

HI test or CF titration, but nonspecific responses

make serology an unreliable diagnostic tool. The

IFA test has been used for rapid diagnosis, but it

has not achieved widespread use.
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases ● 175

Adenoviruses. -Definitive diagnosis is by culture,

isolation, and use of CF, IHA, or neutralization

tests. These three tests can be used to establish the

diagnosis using paired serum samples from the

patient.

Rhinoviruses. —For the most common agent of

the common cold, with over 100 serotypes, routine serologic diagnosis is not very practical, and

is not routinely available even in developed countries, though the neutralization test can be used.

Even tissue culture isolation is often unsuccessful in detecting the infection, because the conditions for successful isolation require special handling (246).

Corona viruses. —Diagnosis of coronavirus by

culture isolation is not a routine procedure and

needs specific types of culture cells. Serologic diagnosis can be made using the CF test.

Bacterial and Mycoplasmal Infections.—The

diagnosis of bacterial agents of respiratory infections has been greatly improved over the last decade. Rapid and accurate diagnosis is now available in hours rather than days through the use

of a variety of diagnostic methods to detect intact bacteria, or in some cases soluble antigen, by

the COA and LA tests, ELISA, RIA, CIE, and the

IFA test (264).

The COA test can be used for detection of bacteria in specimens such as sputum, serum, urine,

and cerebrospinal fluid. It can also be used for

strain typing of culture isolates. The test is simple, rapid, sensitive, and specific when the antiserum used is of good quality. Each bacterium to

be identified needs a specific antiserum reagent

with appropriate antibodies.

The LA test has been used to detect Streptococcus pneumonia and Hemophilus influenza

in body fluids such as serum, cerebrospinal fluid,

and urine. False positives result from nonspecific

autoagglutinations and from reaction to antigens

common to pathogenic and nonpathogenic organisms.

ELISA can be used to detect bacteria in body

fluids. It is highly sensitive for detecting H. influenzae type B, as well as pneumococcal antigen.

There are two variations: the direct assay uses a

specific antibody (against the bacterial antigen)

that is labeled with the enzyme. The indirect assay uses unlabeled specific antibody. The antigenantibody complex is then identified with labeled

antibody that binds to it. The indirect method is

very sensitive and more useful, because it limits

the need to just one labeled antibody.

RIA is extremely sensitive for detecting S. pneumoniae and H. influenza. To evaluate the practical use of RIA, however, more experience is

needed. CIE has been successfully used to detect

pneumococci, streptococci, and H. influenza in

respiratory secretions and body fluids.

The IFA test can be used to identify bacteria

but not soluble antigens. It is very sensitive for

detecting H. influenzae, S. pneumonia, and Bordetella pertussis. The IFA test can also be used

to identify various organisms in culture.

Streptococcus pneumoniae. —Direct microscopic

examination of sputum can indicate pneumococci,

but the predictive value of this test is variable,

because several organisms resemble pneumococci,

healthy individuals can carry pneumococci, and

a sputum negative for pneumococci does not rule

out infection. Sputum culture is the standard procedure. Serologic diagnosis is not practical, because antibodies persist for long periods of time.

Efforts to detect antigen in respiratory tract secretions, blood, and urine by CIE (97,345), as well

as by the COA and LA tests (342), have been successful.

Streptococcus pyogenes. —Diagnosis is made

by isolation of the streptococci from culture of

throat samples. Group determination is made with

specific antiserum—the Lancefield precipitation

test is considered the standard, though direct fluorescent antibody test is also useful. IFA of throat

swab isolates can be obtained after 2 to 24 hours

of culture. CIE can be used 6 hours after culture.

The COA and LA tests have also been used. There

are three convenient tests for detection of serum

antibodies.

Bordetella pertussis.—Definitive diagnosis is

made 2 to 3 days after culturing a throat sample

on specific media, by agglutination with specific

antiserum or the IFA test. A rapid diagnosis is

available with the direct fluorescent antibody test.
176 . Status of Biomedical Research and Related Technology for Tropical Diseases

Hemophilus influenzae. -After isolation and

culture of a clinical sample, diagnosis is made by

microscopically detecting a surface change in the

bacterium (the Quellung test). Rapid diagnosis of

antigen in secretions and body fluids is available

by CIE, the LA test, and ELISA (264).

Mycoplasma pneumonia. —Definitive diagnosis is made by culturing a throat sample on appropriate media, with subcultures made weekly for

8 weeks. When colonies appear, they can be identified visually, though IFA staining confirms the

diagnosis.

Rapid diagnosis of sputum by CIE has been

demonstrated, but more experience is needed to

validate this technique (404). Serologic diagnosis of paired sera is done with the CF test. Detection of antibody in one sample is useless for diagnosis, because high titers persist long after initial

infection.

Research Needs

There are many procedures for the diagnosis

of ARIs, but various constraints reduce their practical use. Direct demonstration of causative agents

is only now becoming an option. Serologic diagnosis using paired sera and serologic diagnosis of

culture isolates do not provide timely information for the treatment of the individual patient.

Requirements for well-equipped and well-supplied

laboratory facilities are another constraint that

limit the use of diagnostic procedures in tropical

countries.

Continued development of the available and

promising technologies for the diagnosis of ARIs

is needed. With rapid, simple, reliable diagnostic tests, both the needs of the patient and the community could be better met. Ideally, such tests

would not require expensive equipment, reagents,

or highly trained operators. In some cases, however, if the tests have certain sophisticated requirements, it may be possible to use them in centralized laboratories.

Arboviral and Related Viral Infections

Because of the general lack of effective treatment for arboviral infections, diagnosis is of less

importance to individual patients than to the community, where it is critical for recognizing outbreaks and initiating vector control measures.

Conventional Diagnosis

Clinical diagnosis of the diseases produced by

the arboviruses recognizes three syndromes: 1)

fevers of an undifferentiated type, frequently

called “dengue-like,” with or without rash and

usually relatively benign; 2) encephalitis, often

with a high case fatality rate; and 3) hemorrhagic

fevers, also frequently severe and often fatal (418).

Conventional diagnosis of arboviral disease involves isolation of the virus in newborn mice or

cell culture, followed by serologic diagnosis using

HI, CF, or neutralization tests. The serologic diagnosis itself is quick and straightforward, but successful culture requires appropriate laboratory

equipment, tissue culture materials, and time (usually at least 2 days). More sensitive cell cultures

have become available using mosquito cell lines

from which early detection can be made using the

IFA test (340). It is also possible to inoculate clinical samples into live mosquitoes and then identify the virus using an IFA test of the salivary

glands. This technique is very sensitive, but slow

(10 to 14 days) (190). Other new methods, such

as the RPHA test, are being developed to detect

the virus earlier in the culture cycle.

Serologic diagnosis with paired sera still uses

conventional methods (HI, CF, and neutralization

tests). Single radial hemolysis has been introduced

with some important advantages (124). It is as sensitive as the HI test for dengue fever, tick-borne

encephalitis, yellow fever, West Nile fever, and

Venezuelan equine encephalitis, yet simpler to perform. The method looks promising but needs further evaluation with different viruses.

Direct detection of virus from the patient is dependent on sufficient virus in specimens. The IFA.

test has been used with some success for a few

diseases (Japanese B encephalitis, Colorado tick

fever, Rift Valley fever). ELISA and RPHA methods are being evaluated for detection of viral antigen in body fluids and respiratory secretions.

CIE has been used to detect dengue virus in sera

from patients with acute diseases but the test has

low sensitivity. In all cases of individual direct

diagnosis, a negative result is not conclusive.
Ch. 8—Diagnostic Technologies: Selected Tropical Diseases “ 177

Detection of specific IgM antibody (which is an

early immune response in the acute phase of infection) is carried out by the HI test, ELISA, and

the IFA test. This is used for the diagnosis in convalescent patients (when virus has disappeared)

and for primary dengue virus.

Recent Progress

MAbs have been developed for certain of the

Togaviruses, Bunyaviruses, and Arenaviruses,

and nucleic acid hybridization techniques are in

various stages of development for some of these

viruses. ELISAS are also being developed for members of each group and field tests for some have

begun. MAbs to antigens common to groups of

viruses have been developed which allow “generic” diagnosis of disease, frequently sufficient

to initiate medical therapy and epidemic prevention measures (197).

MAbs for early type-specific identification of

the four main serotypes of dengue viruses have

SUMMARY

There is great variability in the availability of

diagnostic technologies for diseases of importance

in developing countries. In general, however, lack

of effective diagnosis is a major obstacle to health

care only when health care systems are adequate

to act on diagnoses. This situation is not the norm

today. While diagnosis is an integral part of medbeen developed by the U.S. Army at the Walter

Reed Army Institute of Research (197) and are

now generally available. However, dengue virus

isolation is still difficult as most patients have low

virus concentrations, and the viruses grow poorly

in cell cultures. The U.S. Public Health Service

laboratory in Fort Collins, CO, also is producing dengue monoclonals in collaboration with

WHO. MAbs have also been made to specific surface antigens of several viruses, and ELISA tests

based on these reagents are being evaluated (16).

Research Needs

Development of synthetic peptides following sequence analyses of the alphaviruses and flaviviruses could be very important for developing

diagnostic reagents to detect serum antibodies (as

well as for vaccine development). With regard to

the needs of individual patients, rapid diagnostic

methods will become increasingly important as

drug treatments for arboviruses are developed.

ical care, in some cases, diagnostic technologies

have even greater value in providing information

about the incidence, prevalence, and natural history of diseases of importance to developing countries. Development and use of diagnostic technologies in research could lead to effective integrated

disease control strategies.
9 ■

Therapeutic Technologies:

Selected Tropical Diseases
Contents

.

Introduction . . . . . .

Therapies:

Malaria

Current

. . . . . . . .

Schistosomiasis .

Trypanosomiasis

Leishmaniasis

Filariasis . . . .

Leprosy . . . . .

Tuberculosis.

. . .

. ..

. . .

. . .

status

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

for

. . .

. . .

. . .

,..

. . 

. . .

. . .

“Selected

. . . . . . . .

. . . . . . . .

. . . . . . . .

. . . . . . . .

. . . . . . . .

.$, . . . . .

. . . . . . . .

Diarrheal and Enteric Diseases . . . . .

Acute Respiratory Infections . . . . . . .

Tropical

. .

. .

. .

. .

. .

. .

. .

. .

. .

Arboviral and Related Viral Infections

Summary . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

.,,.. .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

Diseases . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . ,

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . . . . . . . . . . . . . . . . . .

Page

181

182

182

185

186

187

188

188

189

190

193

196

197
9.

Therapeutic Technologies:

Selected Tropical Diseases

INTRODUCTION

The purpose of therapy is to alter the course

of disease so that its consequences are less severe,

or to make being ill more tolerable for the patient.

Therapy for infections may treat symptoms,

which themselves can be dangerous, or may

directly attack the responsible organism.

Symptomatic treatment is quite common. Thus,

for instance, it is usual to take measures against

high fevers, regardless of the cause, because very

high body temperatures can result in brain damage. There may or may not be treatment to eliminate the organism causing the infection, and the

organism may not even be identified; nevertheless, controlling fever is important. Another symptomatic treatment that has quickly become one

of the most potent tools in the tropics is oral dehydration therapy (ORT) for the dehydration that

accompanies diarrheal diseases (see Case Study

A: Oral Dehydration Therapy for Diarrheal

Diseases).

Symptomatic treatment is adequate for some

infections that are self-limited. For infections that

the body cannot eliminate, however, the goal of

therapy is the eradication of the disease-producing

organisms, not simply alleviation of symptoms,

although this goal is difficult if not impossible to

achieve for many tropical diseases.

In general, therapy against bacterial infections

is safe, effective, and usually lasts about 1 week.

Some bacterial infections (e.g., urinary tract infections) can be treated with one dose; others

(e.g., those causing enlargement of the heart, endocarditis) may require 6 weeks of therapy.

Prolonged treatment, usually 6 months or more,

is the norm for tuberculosis, and lifetime treatment is necessary in the case of lepromatous leprosy. Many tropical diseases caused by helminths

can now be adequately treated with 1 to 6 days

of therapy, while for others, there is no adequate

therapy. The treatment for diseases caused by protozoa is similar. The development of antiviral

drugs is still in its infancy.

The mere existence of adequate therapy does

not guarantee that it will be used. The availability and quality of health care varies from country to country and from one region of a country

to another. Some areas in developing countries,

principally the cities, have quite modern health

facilities; other areas have only dispensaries or

often nothing at all. This variability results in a

marked inconsistency in the ability to make specific diagnoses and administer pathogen-directed

treatments. In most developing countries, antimicrobial are readily available over-the-counter.

The result is two common abuses of these drugs:

use for conditions in which they are ineffective

and use in inadequate doses. Both extensive use

and underdosing promote drug resistance in the

pathogens. The prevalence of organisms resistant

to the antimicrobial most commonly available

is high and thus poses significant therapeutic

problems.

The antimicrobial that are generally available

in developing countries are penicillin, chloramphenicol, various sulfonamides, tetracycline,

streptomycin, isoniazid (INH), chloroquine, pyrimethamine-sulfadoxine (P/S), and some antihelminthic drugs. These are relatively inexpensive

compared to other, newer agents, but have significant drawbacks. It is to these agents that

resistance in some areas is widespread and growing. Alternatives are usually marketed, but frequently not available where they are needed, often

because they are too expensive. Many of the older

drugs, particularly the antihelminthics, are toxic

to the patient. Drugs for chronic infections often

require months or years of treatment, so their ef181
182 • Status of Biomedical Research and Related Technology for Tropical Diseases

fective use in developing countries, particularly

in rural areas, is unlikely.

Some progress is being made. Older chemotherapeutic agents are being reexamined; newer ones

are being screened in the laboratory and in

animals. The process of drug development is slow,

however, and a decade can easily elapse before

a promising chemical is marketed as an approved

drug. There is a great need for safe, effective, inexpensive, oral, single dose therapies for tropical diseases. Alternatives to the present toxic

agents and alternatives for use against drugresistant organisms are particularly pressing

needs. A major problem with new drug development for important pathogens in developing countries is that some diseases at-e relatively rare and

others are prevalent only in areas with limited economical resources. Thus, financial incentives for

the pharmaceutical companies best equipped to

develop new agents are lacking.

THERAPIES: CURRENT STATUS FOR

SELECTED TROPICAL DISEASES

Malaria

Human malaria is caused by four species of the

genus Plasm odium: P, falciparum, P. malariae,

P. vivax, and P. ovale. There are many groups

of drugs available for the prevention and treatment of this disease. Their use is determined by

the stage of disease, the immunologic status of the

patient, and the probability that the parasite is

susceptible to a particular drug (usually geographically determined). Some drugs are effective

against sporozoites (the invasive stage of Plasmodium), some against hypnozoites (the latent stage

of P, vivax and P. ovale in the liver), some against

the merozoites (the erythrocytic or red blood cell

stage), and still others against gametocytes (the

sexual form which is picked up by the mosquito

during feeding and perpetuates malaria transmission after further development in the mosquito).

Some of the drugs affect more than one stage of

the parasite.

Resistance of P. falciparum to most agents is

growing. Resistance of P. falciparum to chloroquine has been present for some years in Panama,

parts of some South American countries, India,

Southeast Asia, Indonesia, China, the Republic

of the Philippines, and other Pacific islands. It has

spread from some parts of South America to include most parts of Bolivia, Venezuela, French

Guyana, and northern Peru. There is growing

prevalence of resistance in east Africa, which includes Kenya, Tanzania, eastern Zaire, Burundi,

Uganda, Rwanda,

clan, Madagascar,

In some areas,

resistant to drugs

as P/S, quinine,

Malawi, Zambia, northern Suand the Comoro Islands (377).

P. falciparum strains are also

other than chloroquine, such

and mefloquine (see below).

Strains resistant to P/S were thought originally

to be present only in Southeast Asia. Now there

appear to be some resistance in Kenya, Tanzania, and the Amazon basin. Most of these are

resistant to both chloroquine and P/S. Some

strains in Southeast Asia are resistant to chloroquine, P/S, and mefloquine, and relatively resistant to quinine. These multiply resistant strains

are mostly limited to the Thailand-Kampuchean

border.

Agents for Treatment of Acute Malaria

Specific chemotherapy for acute malaria is at

least 400 years old. During the 1600s, it was

known that the bark of a Peruvian tree, the cinchona tree, was effective in the treatment of intermittent fever. The active ingredient, quinine,

was isolated in 1820 by two French chemists,

Pelletier and Caventou. Since then, new compounds have been discovered that are useful in

the treatment of malaria; however, the ideal drug

is yet to be discovered. The treatment of malaria

becomes less satisfactory almost every year,

mainly because more areas of the world report

P. falciparum resistant to currently available

drugs and drugs in clinical trials.
Ch. 9—Therapeutic Technologies: Selected Tropical Diseases ● 183

Quinine. -Quinine was the first medication for

the treatment of malaria, and it continues to play

a key role. Quinine is effective mainly against the

merozoite stage of Plasmodium and therefore is

effective in the treatment of acute malaria of all

species. It was the sole specific chemotherapeutic agent for the treatment of acute malaria until

World War I. As new drugs such as quinacrine

(1930) and chloroquine (1934) were discovered,

the importance and use of quinine diminished,

With the emergence of chloroquine-resistant,

quinine-sensitive strains of malaria parasites in

1959 in Venezuela (274), however, quinine has

again become invaluable, and recent research has

helped clarify its proper use.

Quinine is used parenterally (by injection) or

orally as the drug of first choice in patients with

falciparum malaria in areas of the world where

chloroquine-resistant, P/S-resistant P. falciparum

is prevalent. Unfortunately, it is the most toxic

of all antimalarials used regularly to treat acute

attacks, and this toxicity limits its use. Adverse

effects from quinine are relatively common, ranging from sudden death (which is rare, but may

occur from rapid intravenous infusion or hypersensitivity), to more common ringing in the ears,

nausea, visual disturbance, and headache. Quinine is also associated with hypoglycemia (low

blood sugar), hemolytic anemia (breakdown of

red blood cells), and a decrease in clotting factors. Furthermore, increasing resistance of P. falciparum to quinine has recently been reported in

Thailand and has been associated with treatment

failures when quinine has been administered in

standard doses (48).

4-Aminoquinolines. —Since quinine (and quinacrine) was in limited supply during World War

11 and was quite toxic, efforts were made to find

alternatives. Through a cooperative research program in the United States during World War II,

investigation of 4-aminoquinolines was undertaken after one of these compounds was reported

by the French to be well tolerated and highly

active.

Chloroquine. —Chloroquine is the most valuable and most extensively used of the 4-aminoquinolines. It has activity mainly against the

merozoite. It continues to be the drug of first

choice for treatment of acute malaria in “nonallergic” persons with chloroquine-susceptible strains

of the malaria parasite and for prophylaxis for

travelers in areas without chloroquine-resistant

strains. Chloroquine can be given orally, intramuscularly, or intravenously and is generally

available in the geographical areas where it is

needed. It is well tolerated in the usual treatment

and prophylactic dosages. Occasional side effects

include mild transient headaches, nausea, diarrhea, visual disturbances, and pruritus (itching).

Pruritis may occur in anyone, but more commonly in blacks. It probably does not cause birth

defects, and is therefore presumed to be safe during pregnancy.

Amodiaquine, —Amodiaquine, another 4-

aminoquinoline, was first found to be effective

against nonhuman malaria in 1946. It, like chloroquine, is effective in preventing acute malaria

and in treating patients with acute malaria due

to all four species of Plasmodium, except chloroquine-resistant strains of P, falciparurn. Amodiaquine is slightly more effective than chloroquine

against these resistant strains; however, this fact

probably has little clinical significance (328). The

side effects of amodiaquine are similar to, but possibly slightly less severe than, those of chloroquine. Amodiaquine is available only in an oral

form, and since it does not have a bitter taste like

chloroquine, it is more acceptable to children,

Pyrimethamine-Sulfadoxine .—It appeared for

a short period of time that the introduction of

chloroquine (1934) had brought to an end the

search for malaria treatment. Then resistance to

chloroquine was reported. Again an extensive

search began for drugs effective against chloroquine-resistant strains of P. falciparurn. Many previously available drugs were screened for activity against malaria parasites. In course, it was

discovered that by combining pyrimethamine

with a sulfa derivative, pyrimethamine resistance

could be overcome. Thus, P/S (Fansidar) was

marketed as another effective regimen in the treatment and prevention of acute malaria due to all

four species, excluding P/S-resistant P. falciparum strains.

P/S activity is against the merozoite. This combination was, and still is, one of the most widely
184 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

used regimens for the treatment of uncomplicated

chloroquine-resistant P. falciparum infections.

P/S is also frequently used in Africa for the treatment of chloroquine-sensitive acute malaria.

Blacks frequently have itching from chloroquine

and therefore use P/S as a substitute. Unfortunately, others without chloroquine-induced itching continue to use P/S when chloroquine would

be adequate. Many strains of chloroquine-resistant P. falciparum are now resistant to P/S. This

situation is thought to result partially from the

indiscriminate use of P/S for prophylaxis and partially from treatment with inadequate doses.

Adverse effects are those related to pyrimethamine and sulfadoxine individually. Pyrimethamine has few side effects. Sulfadoxine has the

usual adverse effects associated with any sulfa,

including gastrointestinal and skin reactions which

may be severe, and can be life threatening in persons with sulfa allergy. P/S has not been available in all countries, and it has only recently become available in a parenteral form.

Pyrimethamine has also been combined with

other sulfas (sulfalene, dapsone) both for treatment and prophylaxis.

Trimethoprim.—Sulfas have also been combined with trimethoprim for use mainly in bacterial infections, and these combinations have

been used with quinine in the treatment of uncomplicated chloroquine-resistant (known or

suspected) P. falciparum. Trimethoprim’s main

activity is against the red blood cell stage of Plasmedium. It has relatively few side effects at doses

usually given for malaria.

Tetracycline. —Tetracyclines are antibacterial

agents available in various forms since 1948. They

do have activity against the red blood cell form

of Plasmodium, but this effect is extremely slow.

They are not used alone, but mainly serve as adjuncts to quinine in the treatment of chloroquineresistant, P/S-resistant P. falciparurn infections.

Alteration of the intestinal flora usually occurs

within 48 hours following daily administration of

the usual therapeutic dosages. This alteration of

the normal intestinal flora increases one’s susceptibility to enteric (gut) pathogenic bacteria such

as Salmonella. Tetracycline also sensitize the skin

to sunburn, besides causing nausea, vomiting, diarrhea, and discoloration of the teeth during development, the latter limiting their usefulness in

children and pregnant women.

Quinacrine (Mepacrine).-Quinacrine is essentially an obsolete antimalarial because of its side

effects. It is important historically because of its

widespread use during World War II.

Mefloquine. —Mefloquine is one of a number

of 4-quinolinemethanols developed during the

1970s by the U.S. Army Research and Development Command. This group of drugs has been

studied since World War II. Mefloquine, the most

active of this group, has been found effective in

both prophylaxis and treatment of acute malaria

due to all species, including chloroquine-resistant,

P/S-resistant P, falciparum infections. Unfortunately, there already have been reports of resistance of P, falciparum strains to mefloquine both

in vivo and in vitro (321). Thus, when used

prophylactically, mefloquine is combined with

P/s.

Agents for the Treatment of

Persistent Malaria (Radical Cure)

So far, this discussion has focused on the drugs

used for treating acute clinical malaria. Of the four

species of Plasmodium that cause malaria, only

two, P, vivax and P. ovale, have persistent liver

stages. Plasmodial forms in the liver are called

hypnozoites because they can remain quiescent

for long periods, up to 3 years in cases of P. vivax. Following subsequent development, hypnozoites can rupture out of the liver and invade

red blood cells (erythrocytes), producing clinical

malaria. While hypnozoites are in the liver, patients are asymptomatic.

Quinine, chloroquine, mefloquine, and most of

the other antimalarial effective in treating acute

malaria have no effect on the liver stages. Since

susceptible P. falciparum and P. malariae do not

have liver stages, these infections can be cured by

standard treatment regimens effective against the

merozoites. However, since P. vivax and P. ovale

do have liver stages, these infections can recur following treatment with agents effective only

against the erythrocytic form.
Ch. 9—Therapeutic Technologies: Selected Tropical Diseases ● 185

In 1924, an 8-aminoquinoline, pamaquine, was

found to be effective in the treatment of malaria.

Too toxic for general use, it has been supplanted

by a drug in the same family, primaquine (1950).

Primaquine has good activity against the liver

stages of P. vivax and P. ovale and currently is

the only drug besides quinocide, another of the

same family, which is effective in eliminating the

hypnozoites. Although both pamaquine and

primaquine also have activity against the erythrocytic stages, the necessary doses are toxic. Unfortunately, some strains require increased doses

and duration of therapy for complete elimination

of the hypnozoites (115).

Primaquine’s use is limited by gastrointestinal

symptoms and hemolytic anemia. Hemolytic anemia occurs in people with a deficiency of a certain enzyme, G6PD, most frequently occurring

in blacks and people of Asian or Middle Eastern

descent. Hemolysis in blacks is usually selflimited; however, in Asians, hemolysis may not

resolve even after primaquine is withdrawn.

Agents for Malaria Prophylaxis

Drugs that prevent acute malaria are said to be

used as prophylaxis. Some of these, chloroquine

and P/S, have already been mentioned. Chloroquine is effective in preventing multiplication of

merozoites. P/S is effective against the sporozoites

and merozoites. Unfortunately, again, both chloroquine and P/S are limited in geographical use

because of drug resistance.

Another drug, proguanil (chloroguanide) has

been available since 1945 and has been effective

in the suppression and treatment of both P. vivax and P. falciparum infections. Like pyrimethamine, proguanil now has limited use because of

the emergence of resistant strains of P. fulciparum. It is not useful for the treatment of acute attacks, but may be used as prophylaxis in areas

where P. falciparum is susceptible to it. Not

infrequently, resistance to pyrimethamine and

proguanil exist together.

Recent Advances in the Supportive

Treatment of Malaria

Advances in the treatment of severe malaria

apart from antimalarials include the discovery of

the deleterious effect of steroids, used previously

in the treatment of cerebral malaria (395), hypoglycemia associated with severe malaria and quinine therapy (148,228,403), quinine pharrnacokinetics (absorption, distribution, and excretion of

the drug) (401,402), and the effectiveness of exchange transfusion for high levels of parasitemia

(191).

Summary of Current Malaria Therapy and

Outlook for the Future

The armamentarium of chemotherapeutic agents

to be used against malaria is barely adequate in

areas of the world where there are resistant strains

of P. falciparum. Strains of P. falciparum resistant to chloroquine and P/S are spreading (327).

Quinine and mefloquine resistance is emerging.

In areas where there are both chloroquine- and

P/S-resistant strains of malaria parasites, no effective prophylaxis is available except for quinine

and mefloquine. As these drugs are used for prophylaxis, and used in inadequate doses, as so frequently happens in developing countries, resistance to them will probably increase. Primaquine

is an effective cure for the hypnozoite stage; however, the increased dosage required for some

strains suggest that its effectiveness may be diminishing. (Strains such as the “Chesson strain” are

thought to have intrinsic resistance, in contrast

to resistance acquired from drug exposure. ) It is

clear that new effective antimalarial agents are

needed as alternatives for most of the ones currently available.

Schistosomiasis

Recently, the treatment for all forms of schistosomiasis has changed significantly. Antimony

potassium tartrate was initially found to be effective in 1918. It was subsequently replaced by

trivalent antimonials such as stibophen, and these

were largely replaced by hycanthone (1960s) and

niridazole (1966), which were the drugs of choice

until recently. Three agents which have been studied since about the 1970s—metrifonate, oxamniquine, and praziquantel —are effective given

orally and have few adverse effects compared to

hycanthone and niridazole, but are expensive.

Were it not for the expense, these agents would
186 ● Status of Biomedical Research and Related Technology for Tropical Diseases

probably replace hycanthone and niridazole in the

treatment of schistosomiasis.

Hycanthone and Niridazole.—Hycanthone is

effective against Schistosoma mansoni and S. haematobium, but must be given intramuscularly in

a single dose and commonly causes nausea and

diarrhea. Niridazole can be given orally, is effective against S. mansoni, S. haematobium and

somewhat against S. japonicum, but may cause

necrologic side effects necessitating observation

during therapy.

Metrifonate.—Metrifonate is effective only

against S. haematobium. It can be administered

orally, but must be given three times separated

by 2-week intervals and its effectiveness varies

greatly.

Oxamniquine.-Oxamniquine, effective against

S. mansoni, maybe given orally. Side effects are

usually mild. Strains of S. mansoni vary in susceptibility, from highly susceptible strains in the

Western Hemisphere, requiring only one dose, to

less susceptible strains in Africa, requiring about

four times the dose given over a 2- to 3-day

period.

Praziquantel.—Praziquantel is a welcome addition to the antischistosomal armamentarium. Effective against all known human schistosomes and

many other trematodes, it can also be given in

one oral dose and is well tolerated. Cure rates

range from 70 to 95 percent with S. mansoni and

S. haematobium.

Oltipraz.—Oltipraz, now undergoing clinical

trials, appears to have promise for the future. It

is well tolerated and has produced cure rates of

greater than 80 percent with both S. hematobium

and S. mansoni.

Amosconate.—Amosconate, a new antischistosomal drug, has demonstrated efficacy against

S. mansoni and S. japonicum and is effective

against S. haematobium infections of primates besides other worms. It also has some serious side

effects.

Trypanosomiasis

African Sleeping Sickness

(African Trypanosomiasis)

Therapy for African sleeping sickness, whether

due to Trypanosoma brucei gambiense or to T.b.

rhodesiense, differs depending on the stage of the

disease.

The early stage of African sleeping sickness can

be treated with suramin, which has been available since the 1920s. It is given on days 1, 3, 7,

14, and 21 slowly intravenously, and then weekly

twice after that. Suramin has serious side effects,

the immediate ones consisting of nausea, vomiting, seizures, loss of consciousness, and shock.

The most important delayed effect is kidney damage. Other side effects have been reported. The

drug is not effective once the pathogenic organisms have invaded the central nervous system.

Another drug that is effective in the early stage

is pentamidine, which has been available since the

mid-1930s. Pentamidine’s use is limited by side

effects which frequently result in an abbreviated

course of therapy. The drug must be administered

intramuscularly, resulting in pain at the injection

sites. It can also cause abscesses, hypoglycemia

(low blood sugar) or hyperglycemia (high blood

sugar), pancreatitis, and hypotension (low blood

pressure) if inadvertently given intravenously.

Pentamidine is generally not as effective as

suramin.

The late stage of African trypanosomiasis can

be treated with melarsoprol, an arsenical first

shown to have activity against trypanosomes in

1940. Melarsoprol is effective against both subspecies and both stages of disease, but its use is

limited by its toxicity. This agent is used only after therapy with other agents has failed. It must

be given intravenously daily, or every other day,

for three doses, and then this sequence must be

repeated twice more after some time has elapsed.

Side effects include intensely irritating local reactions upon leakage into tissue and a potentially

fatal reaction of the brain.
Ch. 9—Therapeutic Technologies: Selected Tropical Diseases ● 187

In summary, therapy for African sleeping sickness requires the use of toxic and frequently only

partially effective drugs over an extended period

of time. Hospitalization is usually required for

administration and observation of intravenous

therapy. Effective and appropriate treatment will

require new drugs that require short courses, and

are easy to administer. Such treatment is not now

foreseeable.

Chagas’ Disease (American Trypanosomiasis)

Chagas’ disease has two clinical stages: acute

and chronic. The acute stage can be suppressed

by the drug nifurtimox, which has been available

since the 1970s. Following treatment with nifurtimox for 120 days, one study has claimed an

80-percent cure rate at 2 years (258). However,

strains of T. cruzi vary in susceptibility depending on the area—strains from Argentina and Chile

are more susceptible than some Brazilian strains.

Nifurtimox is fairly well tolerated, although side

effects such as gastrointestinal upset and abnormalities of nerves occur in 40 to 70 percent of patients. Another drug reported to be effective in

the acute stage in a large percentage of patients

is benzonidazole (258). Both are given orally.

Leishmaniasis

Leishmaniasis, caused by protozoans of the genus Leishmania, has three major forms, depending on the species of parasite: 1) cutaneous leishmaniasis, a localized ulcer caused by one of three

species, L. tropica, L. mexicani, or L. braziliensis (determined by the geographical location); 2)

mucocutaneous leishmaniasis, an invasive destructive lesion caused by L. braziliensis; and 3)

visceral leishmaniasis, a systemic disease caused

by L. donouani. Visceral, mucocutaneous, and

complicated cutaneous leishmaniasis require

therapy.

Antimonials.—Antimony-containing compounds

have been the mainstays of therapy for all types

of leishmaniasis requiring treatment since tartar

emetic was shown to be effective in 1912 (114).

Sodium stibogluconate was found effective as

early as 1937 and continues to be the treatment

of choice along with meglumine antimonate. Both

of these drugs must be given parenterally for 10

to 30 days depending on the susceptibility of the

Leishmania parasites.

Appropriate regimens of therapy for each strain

from each geographic area have not been precisely

determined. Ten-day periods of treatment are effective in the majority of patients in China, India,

and the Sudan, but persons infected with Kenyan

strains usually receive a 30-day course. Therapy

is frequently interrupted by a rest period of 10

to 14 days.

Recently, therapy in India has been increased

from 10 to 20 days, decreasing the relapse rate

from 13 to 0.5 percent. Also, the pharmacokinetics of sodium stibogluconate have recently been

studied, suggesting that daily doses need to be increased.

Side effects of nausea, anorexia, and malaise

are generally tolerable. Antimonials do cause

changes in the electrocardiogram, generally in

doses higher than those used against leishmaniasis.

Amphotericin.-Amphotencin BIDD, an agent

used in the United States mainly for its antifungal activity, has also been found to be effective

against leishmaniasis. It must be given intravenously, usually every other day or daily, until

2.5 grams total dose is achieved. This dose takes

at least 1 month to administer. Because amphotericin is extremely toxic to the kidneys, renal

function must be monitored. The facilities to do

this are frequently not available where the patients

are in developing countries.

Pentamidine and Stilbamidine.-Another agent

useful in leishmaniasis is pentamidine. As mentioned previously in relation to trypanosomiasis,

its use is limited by its side effects. A similar compound, stilbamidine, maybe more effective, but

its use is limited by its effects on the nervous system. Hydroxystilbamidine is also effective, but

not generally available in developing countries.

Allopurinol Riboside. -Allopurinol riboside is

a drug most often used in the United States to decrease uric acid excretion by the kidney in the

treatment of gout. A derivative has shown some

in vitro activity against L. donovani and is almost

ready to undergo clinical trials in cutaneous leishmaniasis at the Gorgas Memorial Laboratory.
188 . Status of Biomedical Research and Related Technology for Tropical Diseases

Other Agents.—The role, if any, of other

agents such as cycloguanil, pyrimethamine,

metronidazole, benzimidazole, rifampin, and

nitrofurtimox remains to be clarified.

Summary

At present, management of leishmaniasis is usually hospital-based, since most therapy is parenteral, and side effects common and frequently serious. There is, therefore, a need for effective, safe,

oral agents in the treatment of these diseases.

Filariasis

There are at least eight types of human filarial

infections. The three most important are infections

by Wuchereria bancrofti, Brugia malayi, and Onchocerca volvulus. The therapy for all filarial infections is similar. There is effective therapy

against microfilariae (the immature worms that

cause the symptoms of filariasis), but no nontoxic

therapy is available against all the species of adult

worms.

Diethylcarbamazine. —Diethylcarbamazine kills

microfilariae and is therefore effective as prophylaxis or in treatment of symptomatic infections.

It also has an effect on the adults of W. bancrofti

and Loa loa (the agent of African eyeworm disease). Diethylcarbamazine can be given orally.

Side effects include adverse diethylcarbamazine

reactions to itself and reactions to dying worms.

The latter can be severe and exacerbate eye

damage.

Suramin.—Suramin is also effective against

microfilariae and adults of O. volvulus. Following an initial test dose, there must be 1 week of

observation. If no adverse reactions occur, doses

may be increasea at weekly intervals. Therapy

usually requires 6 weeks. Suramin must be given

intravenously. Side effects include adverse reactions to the drug itself and reactions to the dying

worms.

New Drugs.—New drugs undergoing evaluation are ivermectin, mebendazole, and flubendazole. The early results with ivermectin are very

encouraging. A report of the first study of ivermectin in human beings with onchocerciasis (river

blindness) was published in 1982 (14). The side

effects that occur with the existing drugs were absent, and ivermectin cleared or greatly reduced

the microfilarial load in all 32 subjects after one

dose. Those results have been duplicated in at least

one subsequent study (14,77). Both mebendazole

and flubendazole have activity against adult

worms.

Summary

In summary, there is a great need for inexpensive, well-tolerated, oral agents effective against

adult worms. Of the new drugs under consideration, ivermectin may ultimately have the potential for controlling filariasis in endemic areas, but

that prospect is not near at hand.

Leprosy (Hansen’s Disease)

Treatment for leprosy varies depending on the

severity of disease, which ranges from tuberculoid

leprosy (localized lesions with few Mycobacterium

leprae organisms) to lepromatous leprosy (which

is widespread disease and infection with large

numbers of M. leprae bacteria). The course of

treatment ranges from about 3 years for some patients at the tuberculoid end of the spectrum who

have relatively good immunity to lifetime treatment for patients with lepromatous leprosy who

have little or no immunity.

Dapsone.—Specific chemotherapy for leprosy

was not available until 1940, when dapsone, a

sulfa derivative, was shown to be effective when

administered by injection to patients at the U.S.

Public Health Service leprosarium at Carville, LA.

Dapsone was shown to be effective when given

orally in 1947 and continues to be an effective,

safe, inexpensive agent. Acedapsone is an injected

drug which releases dapsone slowly. Both have

the usual side effects of any sulfa-containing drug.

Dapsone is bacteriostatic, able to inhibit, but

not kill, M. /eprae. Viable dapsone-susceptible

bacilli can be isolated from patients after 10 years

of treatment even though they have no evidence

of active disease.

Resistance to dapsone, first reported in 1964,

is now an important problem. Up to 62.5 percent

of newly diagnosed and 9.3 percent of previously

treated patients have resistant strains of M. leprae.
Ch. 9—Therapeutic Technologies: Selected Tropical Diseases ● 189

Resistance to dapsone develops, like resistance to

many antimicrobial, when patients do not follow the prescribed treatment regimens, for whatever reasons. Irregular compliance with treatment

frequently occurs as the result of unsupervised

administration. It is hoped that combination

chemotherapy (see below) will minimize the development of resistance. Development of widespread resistance could lead to loss of control of

leprosy in a community.

Clofazimine. —Clofazimine is another bacteriostatic agent with a unique additional property

of being anti-inflammatory. Its use is limited by

its cost and side effects, consisting mainly of red

skin pigmentation, nausea, and diarrhea.

Rifampin.—The only bactericidal (able to kill

bacteria) agent available for the treatment of

leprosy is rifampin. It is rapidly effective in reducing the number of bacteria. However, even after 5 years of treatment, viable bacteria can be

recovered from patients. Its use again is limited

by its cost and side effects.

Combination Chemotherapy .—Since viable

bacteria can be recovered from patients treated

with all of the agents mentioned above, and since

dapsone resistance is quite prevalent, combinations of the above drugs are now recommended

as standard treatment.

Summary

The difficulties in treating people with leprosy

are readily apparent. Treatment with the agents

that are currently available necessitates many

years of supervised chemotherapy. Cure is virtually impossible in many patients who require

lifetime treatment. When dapsone resistance develops, there are few effective alternatives. There

is great need for new bactericidal agents. Since

recovery from leprosy will probably not occur until the body’s natural defenses against the organism are adequate, immunopotentiating agents,

drugs able to stimulate the natural defenses of the

body, may be the solution in the future.

Tuberculosis

It has been 40 years since the discovery that

streptomycin was effective in arresting the growth

of Mycobacterium tuberculosis, the bacterium

that causes tuberculosis. Two years later, paraaminosalicylic acid (PAS) was also found to be

an effective “tuberculostatic” agent, arresting the

growth of the bacteria, but not killing them. With

the development of M, tuberculosis bacteria resistant to streptomycin in 1947, PAS became the

companion drug of streptomycin to prevent the

growth of resistant organisms. This combination

was only partially effective in eliminating resistant strains.

It was not until 1952, with the introduction of

the tuberculocidal drug INH (isoniazid) that the

era of effective chemotherapy for tuberculosis began. INH actually kills the organisms, rather than

holding them at bay. INH is not adequate as

monotherapy, however, because of the development of resistant organisms. For that reason, INH

was combined with PAS or streptomycin. In 1968,

with the introduction of ethambutol, another

tuberculostatic agent, antituberculosis therapy

seemed adequate and safe. Because the duration

of therapy was long, 12 to 24 months, however,

patient compliance with the treatment regimen remained a problem.

Rifampin was introduced in 1971, but its special advantage, rapid tuberculocidal activity, was

not realized until the late 1970s. Rifampin helps

cure the patient, and it reduces the period of time

during which the patient can transmit disease to

others. M. tuberculosis bacteria are transmitted

in sputum, particularly when the patient coughs.

Combining rifampin with INH rapidly clears the

sputum of bacteria, decreasing the infectivity of

the patient and thereby decreasing transmission

of disease. The duration of therapy has been reduced from the previously recommended 18

months with INH and ethambutol to 9 months

with INH and rifampin. INH is prohibitively expensive for use in many less developed countries,

however, and for that reason, simply is not available where it is most needed.

Short-Course Chemotherapy .—Abbreviated

courses of therapy for tuberculosis are an important goal because such courses increase patient

compliance with the treatment regimen. Inadequate treatment promotes the growth of drug-
190 • Status of Biomedical Research and Related Technology for Tropical Diseases

resistant organisms, and failure to achieve cure

promotes spread of the disease.

In an effort to reduce the duration of therapy,

various regimens of short-course combination

chemotherapy have been tested in developing

countries, many sponsored by the British Medical Research Council. These courses primarily involve combinations of INH, rifampin, pyrazinamide, and streptomycin. The results indicate that

duration of therapy maybe reduced to 6 months

or possibly even less when these drugs are used

in certain combinations.

Although compliance problems are less with

shorter courses of therapy, they have not been

eliminated. Liver toxicity continues to occur with

most drug combinations, but usually does not require discontinuing of therapy.

Problem of Resistance.—It is fortunate that

agents described above are so effective in treating most patients with tuberculosis. For tuberculosis patients with resistant organisms, however,

therapy is only barely adequate. For these patients, more drugs are required, resulting in more

side effects, problems of compliance, and higher

cost. Isolation of INH-resistant organisms from

newly diagnosed cases of tuberculosis is common

in the developing world, and there are some bacilli

resistant to both INH and streptomycin (101).

The main limitation in these areas, however,

is not drug resistance but cost. Rifampin is very

expensive for most developing countries with high

prevalence of tuberculosis. Rifampin treatment

for an adult costs $0.44 to $1.60/day, INH $0.001

to $0.004/day, pyrazinamide $0.09 to $0.72/day

(53). In a 9-month course (2 months of ethambutol, INH, and rifampin followed by 7 months

of INH and rifampin), rifampin accounted for 96

percent of the cost of the regimen (121). If this

regimen were used, for example, in the Philippines

in 1976, it would have cost about $200/patient

(121). With an estimated 141,040 persons needing therapy, the total cost of treatment would be

approximately $28 million. These countries, therefore, continue to use relatively long courses of less

expensive chemotherapeutic agents, with predictable compliance problems and a predictable lack

of control of tuberculosis.

Summary

In summary, there is adequate treatment available now for most tuberculosis. Although effective agents are available, many countries with

tropical environments are unable to benefit from

them because of financial restrictions and compliance problems.

A need for new drugs, particularly to be used

in combination with INH, however, remains. The

current companion drugs have significant toxicities or are associated with compliance problems.

There are drug-resistant strains of M. tuberculosis. Because some strains are resistant to more than

one agent, there is a need for new effective tuberculocidal agents with rapid onset of action, especially as an alternative to INH and rifampin.

Diarrheal and Enteric Diseases

General Treatment

Until the late 1960s, treatment of moderate diarrhea required hospitalization and intravenous

hydration therapy. In 1964, it was shown that patients with cholera responded well to oral dehydration with glucose, bicarbonate, potassium, and

sodium chloride (276). Three years later, it was

shown that glucose facilitated the absorption of

these electrolytes and decreased the total stool volume (337). In 1968, oral administration of a solution containing glucose and the electrolytes mentioned above was demonstrated to be effective

therapy for moderate dehydration secondary to

cholera (250). Since then, much has been written

concerning the role of ORT (oral dehydration therapy). WHO now has made recommendations for

the composition of the oral dehydration solution.

In 1983, 29 million packs containing the ingredients to be mixed with water for ORT were supplied to various countries.

ORT has made a tremendous impact on the resources needed to manage diarrheal illnesses, particularly in infants and children (see Case Study

A: Oral Dehydration Therapy for Diarrheal Diseases). The oral dehydration solutions can be given

by mothers at home and mothers can be trained

by paramedical personnel, thus saving both the

expense of hospitalization and time of the medi-
Ch. 9—Therapeutic Technologies: Selected Tropical Diseases •191

Photo credit: Dr. Robert .Edelrnan, National Institutes of Health

Intravenous dehydration for most cholera patients, shown

here, is being replaced by the simpler, more cost-effective

alternative, oral dehydration therapy (ORT).

cal personnel. About 95 percent of all patients

with diarrhea given ORT respond. Those who do

not can be treated with intravenous solutions. Severely dehydrated patients, however, require initial intravenous therapy. ORT is effective in diarrheas from many causes. The success of the

therapy depends significantly on the cooperation

of the patient and/or mother. In some areas, tradition still impedes behavioral changes necessary

for successful ORT. In many areas, sanitary water

is unavailable. Water must be boiled prior to making the oral dehydration solution, and mothers frequently may be reluctant to do this.

Treatment for Specific Organisms

Viruses. -Viruses cause a significant amount

of diarrheal illnesses in the tropics. Viral agents

of diarrheal diseases include rotavirus and Norwalk agent, possibly the two most important

agents, adenoviruses, enteroviruses, etc., as well

as other agents which have not been specifically

identified.

There is no effective specific therapy for any

of the viral agents of diarrhea] diseases. None of

the generally available antiviral agents are useful in treating diarrheal illnesses. Interferon, a

molecule which can protect host cells from viruses, and interferon inducers are being studied

in some viral illnesses. In general, prevention of

viral illnesses through vaccination is the hope for

the future.

Bacteria.—Many bacteria have been identified

as the cause of diarrheal illness, and the list is

growing as laboratory methods for identification

improve. Other bacteria have been isolated from

patients with and without diarrhea, and some of

these are clearly pathogenic. In many areas, the

facilities to identify etiologic agents of diarrhea

are unavailable.

Most episodes of bacterial diarrhea are selflimited and specific. There are effective antibacterial agents for most of the pathogenic organisms,

but these agents are frequently unavailable or too

expensive. Alternatives in special situations (e.g.,

allergy to an agent or resistance) often are not

readily available. Resistance to drugs by many enteric bacterial pathogens is growing.

Vibriocholerae. —Tetracycline has been the

drug of choice for treating cholera, but it has several drawbacks. It causes staining of teeth in children, the group in which most diarrheal illness

occurs. Tetracycline also sensitizes the skin to

sunburn and is not recommended for pregnant

women during the stages of bone and teeth development. Alternatives to tetracycline are furazolidone, chloramphenicol, and trimethoprim/sulfamethoxazole (T/S).

Cholera strains had remained almost universally susceptible to tetracycline until 1977, when

76 percent were found resistant after 5 months of

an epidemic Tanzania (226). In December 1980,

18 percent of strains of Vibriocholerae isolated in

Bangladesh were multiply resistant to tetracycline,

ampicillin, kanamycin, streptomycin, and T/S

(131). Some of these strains are able to transfer

their resistance factors to Escherichia coli (131).

Shigella.—Many species of Shigella cause diarrhea. Shigella usually cause endemic diarrhea,

but may cause epidemics. In general, effective

agents such as ampicillin, chloramphenicol, and

T/S are available to treat shigellosis, but there is

a growing problem with resistance. Sulfonamide

resistance was described in Japan in the 1950s and

was described subsequently elsewhere (185). Areas

which had previously had uniformly sensitive

strains began to have consistently resistant strains.
192 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Subsequently, strains were isolated that were

resistant to more than one antimicrobial. Investigators discovered that these Shigel/a were probably acquiring resistance factors from other

organisms present in the gastrointestinal tract,

specifically E, coli. The transfer of resistance factors between bacilli has become an important

complication of the treatment of many bacterial

infections; some of these are mentioned later in

this chapter.

The exact reason for the increase in the incidence of resistance factors in Shigella is not entirely clear, but is, in general, thought to result

from the widespread use of therapeutic agents in

a population, especially when they are used indiscriminately or in inadequate dosages. Such use

frequently occurs when the agents are available

over-the-counter, as they are in many developing countries. Shigella resistance not only is a

problem in the developing world, but also has

been reported in Washington, DC (294). Up to

93 percent of strains in some areas have been resistant to more than one drug (294).

Salmonella, —For many years, chloramphenicol

has been the most reliable drug in the treatment

of typhoid and typhoid-like enteric fever (infection due to nontyphi species of Salmonella). With

the development of new antimicrobial, alternatives such as ampicillin, amoxicillin, amdinocillin, and T/S have become available.

Drug resistance in Salmonella was not much of

a problem before 1972. Since then, there have

been outbreaks of disease with resistant organisms

in more than one country (128). Strains of both

typhi and nontyphi species have been found to

be resistant (30). More importantly, multiply

resistant strains of S. typhi have been isolated.

Occasionally, these strains are resistant to both

cloramphenicol and ampicillin (38). More recently, strains resistant to chloramphenicol, ampicillin, and T/S have been isolated (188). Again,

some of these resistant strains are able to transfer resistance factors to other bacteria (187).

Escherichia coli. —Antimicrobials are ineffective in decreasing the duration or volume of diarrhea associated with E. coli infection, but are

sometimes recommended if the patient’s diarrhea

continues beyond 3 days or if the patient has bacteremia (bacteria in the bloodstream). Various

strains of E. coli vary in their susceptibility to

drugs, but they may be susceptible to ampicillin,

chloramphenicol, and T/S.

Enterotoxigenic strains of E. coli (those that

cause disease by a toxin produced by the bacterium) frequently cause diarrhea in travelers. A

number of antimicrobial have been shown to be

effective prophylactically including doxycycline,

minocycline, trimethoprim, and T/S. Doxycycline-resistant strains have been isolated from

Peace Corps volunteers in Thailand (102). Besides

nausea, the possibility of skin photosensitivity

from doxycycline and minocycline exist, although

reactions to these agents are less frequent than

those due to tetracycline.

Campylobacter. —The diarrhea resulting from

Campylobacter infections frequently lasts only a

few days, and therapy usually is not necessary.

Campylobacter are generally susceptible to

erythromycin, tetracycline, furazolidone, chloramphenicol, and clindamycin; erythromycin

continues to be the drug of choice. In some areas,

strains are resistant to tetracycline (113).

Other Causes. —Other bacterial causes of diarrhea are species of Aeromonas, Pleisiomonas,

Yersinia, Vibrio (noncholera), Staphylococcus,

Clostridium difficile, and C. perfringens. Aeromonas produces a toxin that causes diarrhea that

is usually self-limited; it can usually be treated

with erythromycin, tetracycline, and chloramphenicol, but is resistant to ampicillin. V. parahemolyticus is one of the noncholera vibrios

which cause diarrhea. Treatment is usually unnecessary. S. aureus and C. perfringens are common causes of food-borne outbreaks of gastroenteritis which are self-limited, and no treatment

is advocated. C. difficile is an important cause of

antibiotic-induced colitis in developed countries,

although the incidence of this disease is not

known, since detection methods are not available.

The preferred drug is vancomycin, but the cost

of one course of therapy is $200 to percent 500,

more than most citizens of developing countries

can afford. Metronidazole, bacitracin, and cholestyramine are cheaper alternative agents.

Protozoa.—Giardia lamblia and Entameba

histolytica are common causes of diarrhea world-
Ch. 9—Therapeutic Technologies: Selected Tropical Diseases •193

wide, but are particularly prevalent in countries

with tropical environments. Giardiasis usually

responds to either quinacrine, metronidazole, or

tinidazole, and these agents are fairly well tolerated in the doses required. Therapy is not always

effective, however, and retreatment may be necessary.

Symptomatic and asymptomatic intestinal

ameba infections (amebiasis) respond to a number of drugs that are given orally and are well

tolerated. Amebic liver abscess responds to a

group of drugs known as “imidazoles,” which includes metronidazole and tinidazole, and to more

toxic drugs such as emetine, dihydroemetine, and

chloroquine. Metronidazole in high oral doses

causes abdominal cramps, nausea, and bloating

and therefore is not tolerated by some people.

Tinidazole is better tolerated and as effective. In

patients who may not tolerate metronidazole and

tinidazole, emetine and dihydroemetine are alternatives. Emetine and tinidazole may cause many

adverse effects, including abnormalities of the

heart rhythm, however, and it is recommended

that patients receiving either of these drugs remain

hospitalized.

Four other protozoa should be mentioned as the

cause of diarrhea: Balantidium coli, lsospora belli,

Cryptosporidium spp., and Sarcocystis spp.

Balantidiasis responds to tetracycline and paromomycin. There is little information available on the

proportion of diarrhea in tropical populations

caused by the other three, and data on the effectiveness of treatment are only now becoming

available. Furazolidine, T/S, and pyrimethaminesulfa have been reported to be effective.

Helminths.—There area number of nematodes

(roundworms) implicated as causes of diarrhea,

including Ancylostoma duodenale and Necatur

americanus (hookworms), Capillaria phillippinensis, Strongyloides stercoralis (threadworms), and

Trichuris trichura (whipworms). For many of

these, treatment may not be recommended the

worm burden is not heavy and the risk of reinfection, is high, as is often the case in developing

countries. For most roundworms, effective, safe,

short-duration therapy is available.

Hookworms can usually be treated with bephenium, pyrantel pamoate, levamisole, or mebendazole in one or two doses for 3 consecutive

days. Frequently, however, only the more toxic

drug tetrachloroethylene is available.

S. stercoralis usually responds to thiabendazole,

but this is one drug which is toxic and frequently

causes anorexia, nausea, vomiting, diarrhea, and

vertigo. Furthermore, thiabendazole is not always

effective. Mebendazole is better tolerated, but still

less effective.

T, trichura infections were a problem until the

advent of mebendazole in 1971; now over 80 percent of these infections can be cured by a twice

daily course for 6 days. Other alternatives effective against trichuriasis are oxantel, flubendazole,

and albendazole.

Infection with C. philippinensis is a continuing problem. Mebendazole and thiabendazole are

sometimes effective after 20 and 30 days respectively.

Praziquantel has revolutionized the treatment

of trematodes (parasitic flatworms), many of

which are associated with diarrhea: Fasciolopsis

buski, Echinostoma itocanum, heterophyids,

clonorchids, opisthorchids, and schistosomes. Virtually all of these respond to praziquantel.

Summary

Most cases of diarrhea can be treated successfully by treating the patient’s symptoms with

ORT. ORT is of particular benefit to infants and

children, who may otherwise die from the dehydration that accompanies bouts of diarrhea.

There are also safe, effective drug treatments for

most of the bacterial infections that cause diarrhea, but not for viral infections. Protozoa and

helminths that cause diarrhea can usually be eliminated with drugs, but the drugs are often toxic

and not entirely effective. In general, however,

the worldwide impact of these specific treatments,

even if applied universally and successfully, would

not approach the worldwide benefit of ORT if

ORT were universally applied.

Acute Respiratory Infections (ARIs)

ARIs due to bacteria or viruses are usually either the major cause of death or second only to

diarrheal disease in tropical environments. They
194 . Status of Biomedical Research and Related Technology for Tropical Diseases

are a major cause of morbidity everywhere. From

a clinical point of view, infections are divided between the upper respiratory tract infections

(URTIS) and lower respiratory tract infections

(LRTIs), though the causative agents overlap to

some degree. URTIS are infections that occur in

and around the teeth, gums, sinuses, throat, tonsils, epiglottis, middle ear, larynx, and trachea.

The most important LRTIs result in pneumonias.

In many tropical countries, the methods necessary to make the cause-specific diagnosis of respiratory infections are not available. Many areas

in developing countries do not have bacteriology

or serology, not to mention the more sophisticated

facilities necessary for culturing and identifying

viruses.

Viruses

Most of the URTIS in tropical countries appear

to be viral in origin, usually caused by the same

viruses caused by disease in temperate environments—rhinoviruses, coronaviruses, adenoviruses, Ebstein-Barr virus, cytomegalovirus, influenza, parainfluenza viruses, etc. Most of these

infections are self-limited, and although there is

some morbidity resulting in loss of workdays,

there is little mortality in adults. In children, there

can be acute mortality associated with the development of a cycle of infection and malnutrition.

Many of the viruses that cause URTI also cause

LRTI. The viruses that may cause severe pneumonia and death are measles, respiratory syncytial virus, and influenza. These generally cause

death from overwhelming pulmonary involvement and respiratory failure. In many localities,

the oxygen therapy that frequently is necessary

is unavailable.

Antiviral chemotherapy is still in its infancy.

Amantidine and rimantadine are effective in reducing the incidence and severity of symptoms

due to infection with influenza A if given prophylactically or within the first 48 hours after the

onset of symptoms. These agents are fairly well

tolerated, but they are generally unavailable in

the developing countries and relatively expensive.

There is no treatment available for the other

common viral causes of URTI. Enviroxime is an

antiviral agent available in Europe but not in the

United States, and evidently effective against

rhinovirus, one of the causes of the common cold.

Bacteria

Upper Respiratory Tract Infections (URTIs).—

Four bacterial causes of URTI deserve specific

mention. One, Corynebacterium diphtheria,

causes diphtheria, an acute, life-threatening pharyngitis. Another, Bordetella pertussis, is the cause

of pertussis (whooping cough). The treatment of

diphtheria is with antitoxin and penicillin or

erythromycin. Antimicrobial are not effective in

changing the course of pertussis. Both of these diseases can be prevented by immunization.

Streptococcus pyogenes, a Group A streptococcus known best as the organism precipitating rheumatic fever, is a common cause of pharyngitis. The disease is responsive to penicillin and

erythromycin, among other antimicrobial.

Haemophilus influenzae causes ear infections,

bronchitis, sinusitis, infection of the epiglottis, and

meningitis, primarily in children. Ampicillin has

generally been the drug of choice, although T/S

and tetracycline also may be effective. Many

strains of H. influenza are now resistant to ampicillin. The preferred drug for serious infections

involving ampicillin-resistant strains is chloramphenicol.

Lower Respiratory Tract Infections (LRTIs).—

Effective antimicrobial are available for most of

the pathogens causing pneumonia acquired in the

community, although the variety of drugs is usually limited. Hospital-acquired pneumonias (nosocomial infections) in developing countries, just as

in the developed countries, are often caused by

drug-resistant bacilli or Staphylococcus aureus.

Streptococcus pneumonia, —S. pneumonia,

the cause of pneumococcal pneumonia, is one of

the most common causes of bacterial pneumonia,

affecting almost all ages in both tropical and temperate climates. Pneumococcal pneumonia continues to be a frequent cause of mortality in some

groups of patients worldwide even though effective antimicrobial are usually available and even

though these patients are treated (13). S. pneu-
Ch. 9—Therapeutic Technologies: Selected Tropical Diseases . 195

moniae is generally responsive to oral penicillin,

cephalosporins, tetracycline, erythromycin, or

chloramphenicol, as well as other antimicrobial.

Patients who are very ill must receive these drugs

by injection.

In 1967, a drug-resistant strain of S. pneumonia was isolated from a patient in Australia (147).

Subsequently, relatively resistant strains have

been isolated from many parts of the world, especially South Africa and New Guinea, and some

of these strains are multiply resistant with relative resistance to penicillin, tetracycline, and

chloramphenicol. Patients with these strains have

not responded to the usual doses of penicillin

(394).

Streptococcus pyogenes, —S. pyogenes is a

cause of pneumonia as well as URTI and is frequently associated with complications such as empyema (infection between the inside of the chest

and the outside of the lung causing a collection

of pus in that site). It responds to penicillin, but

hospitalization and parenteral therapy are often

required, at least initially.

Staphylococcus aureus, —Staphylococcal pneumonia not infrequently complicates recovery from

viral infections, especially in children. Hospitalization is recommended with parenteral administration of semisynthetic enzyme-resistant penicillins

or cephalosporins for about 2 weeks. Antistaphylococcal antimicrobial are effective and generally available.

Haemophilus influenza. —Besides causing ear

infections and meningitis, H. influenza may cause

pneumonia, most frequently in children. Most

strains are sensitive to ampicillin, chlorampheniCOl, T/S, and the newer cephalosporins. However, drug resistance is prevalent in some tropical environments. Some strains are resistant to

ampicillin, but occasionally there is resistance to

both ampicillin and chloramphenicol, the antimicrobial most commonly recommended for

treatment (40).

Klebsiella pneumonia and Escherichia col.—

K. pneumonia and E. coli are less common than

S. aureus or H. influenza as causes of community-acquired pneumonia. These organisms are

usually susceptible to aminoglycosides such as

gentamicin. Aminoglycosides must be given

parenterally and are toxic to the kidney. Frequently, aminoglycosides are not available in developing countries.

Pseudomonas pseudomallei. —This organism

causes melioidosis, a spectrum of disease ranging

from acute to chronic pneumonitis, soft tissue infection, and other manifestations. Most cases have

been reported from Southeast Asia. Treatment

consists of tetracycline, chloramphenicol, and/or

T/S.

Yersinia pestis.—Y. pestis is the etiologic agent

of plague. It may cause fulminant pneumonia,

which is highly contagious. Streptomycin is the

drug of choice. Alternatives may be chloramphenicol, tetracycline, or T/S.

Others. —Other less common kinds of bacterial

pneumonias are associated with brucellosis, leptospirosis, anthrax, glanders, salmonellosis, typhus, and Q fever. These pneumonias are usually only one manifestation of a more generalized

illness. If the diagnosis can be made, treatment

is usually available.

Mycoplasma

Mycoplasma are bacteria-like organisms that

usually cause URTIS and atypical pneumonia in

adolescents and young adults. Tetracycline and

erythromycin are effective therapy and usually

do not have significant adverse effects.

Protozoa

Pneumocystis carinii is a protozoan which

causes pneumonitis in immunosuppressed patients

in developing countries, mainly in malnourished

children. In the United States, it is common

among patients with acquired immunodeficiency

syndrome (more commonly known as “AIDS”).

The drug of first choice, a combination of trimethoprim and sulfamethoxazole, or a sulfa with

pyrimethamine is effective in about 75 percent of

cases. The alternative, pentamidine, is not readily available and is not entirely effective either.

It must be given parenterally and has some serious adverse effects.
196 . Status of Biomedical Research and Related Technology for Tropical Diseases

Amebiasis can sometimes cause acute and

chronic pulmonary disease, but usually responds

well to oral metronidazole.

Fungi

There are a number of fungi that cause acute

and/or chronic pneumonitis and pleural disease

in tropical environments. Most are responsive to

amphotericin B. Amphotericin is toxic to the kidney, must be given parenterally, and is frequently

unavailable in developing countries. Amphotericin-induced kidney failure is difficult to manage without the aid of dialysis, which also is frequently unavailable.

Helminths

Most adult helminths in developing countries

do not initially infect the respiratory tract, but

cause disease if the larval forms migrate through

the lungs. Most of the time, this stage of the infection is self-limited and does not need treatment.

Echinococcus granulosus and other related species are “tapeworms. ” The larval worms form

cysts in various organs, including the lungs. There

is no effective medical treatment for tapeworm

infection of the lung. Recently, a new drug, albendazole, has been demonstrated effective in a

small number of patients. Paragonirnus westermani, a fluke that causes small pulmonary cysts,

has recently been shown to respond to praziquantel. The eggs of S. mansoni and S. japonicum can

end up in the lung, causing a buildup of fibrous

tissue around them. There is no available treatment for this stage of the disease.

Summary

Drug therapy is available for most bacteria, but

not viral ARIs. Even with treatment, however,

mortality rates from ARIs are high, especially

among children. The general state of good health

of the population that keeps mortality from ARIs

low in the United States and other developed

countries, is probably the most important factor

lacking in developing countries.

Arboviral and Related Viral Infections

The 80 or so arboviruses known to infect humans cause four basic types of clinical conditions.

Two are generally benign and self-limited, consisting of: 1) fevers of short duration, with or

without a rash; and 2) painful joints and rash of

short duration. Complications can develop from

either of these two conditions, but they are the

exception. The two more serious clinical syndromes caused by arboviruses are: 1) acute central nervous system disease usually with inflammation of the brain (encephalitis), ranging in

severity from mild aseptic meningitis to coma,

paralysis, and death; and 2) hemorrhagic fevers,

with extensive hemorrhaging, and associated with

shock and high case fatality rates (liver damage

and jaundice accompany these symptoms in yellow fever).

There have been no specific treatments available for any of these viral illnesses. Management

of the more serious conditions is exclusively supportive, directed toward alleviating major symptoms and preventing spread. Correction of fluid

and electrolyte imbalance, oxygen, and blood

transfusions are the most common measures.

Immune therapy with monoclinal antibodies

has been effective in protecting mice against

Sindbis virus and arboviruses (311). Plasma containing antibodies to one hemorrhagic fever virus, given to patients before the eighth day of illness, reduced mortality from 16.5 to 1.1 percent.

However, these patients had more frequent

relapses (211). Similar results were obtained with

immune therapy in monkeys experimentally infected with Machupo virus (211). Immune therapy and ribavirin, a broad spectrum antiviral, is

effective in decreasing the mortality from Lassa

fever in monkeys (172). Both are undergoing clinical trials in Sierra Leone (219).
Ch. 9—Therapeutic Technologies: Selected Tropical Diseases • 197

SUMMARY

This chapter has briefly outlined the present malaria to leprosy and other bacterial infections.

therapy available for the selected tropical diseases For other diseases, therapy is available, but quite

covered in this report. For some diseases, there toxic. For still others, such as most viral infechas been, and continues to be, effective therapy, tions, no therapy is available at all. Although

although there is a growing problem of drug re- tremendous progress has been made, the deficiensistance in diseases ranging from P. falciparurn cies are great, and much work remains to be done.
Case Studies
Page

Case Study A: Oral Dehydration Therapy for Diarrheal Diseases . . . . . . . . . . . . 201

Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202

Definitions of Infant Diarrhea and Dehydration. . . . . . . . . . . . . . . . . . . . . . . . . . 202

Etiology of Infant Diarrhea . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203

Magnitude of the Problem of Infant Diarrhea . . . . . . . . . . . . . . . . . . . . . . . . . . . 203

Nutritional Consequences of Infant Diarrhea . . . . . . . . . . . . . . . . . . . . . . . . . . . . 204

The Interrelationship of Infant Diarrhea With Other Major

Public Health Problems of the Less Developed World . . . . . . . . . . . . . . . . . . 204

Points of Intervention . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 205

History of the Development of ORT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 205

Progression of Clinical Research Studies Leading to

ORT as It Is Practiced Today . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 209

Clinical Indications, Regimens, and Limitations of ORT . . . . . . . . . . . . . . . . . . 211

Composition of Oral Rehydration Solutions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 213

ORT To Prevent Dehydration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 215

Effect of ORT on Nutritional Status . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 216

Effect of ORT on Infant Mortality . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 216

Dissemination of ORT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 216

Attitudes of U.S. Pediatricians Toward ORT . . . . . . . . . . . . . . . . . . . . . . . . . . . 217

The Economy of ORT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 218

Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 219

Case Study Preferences. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 219

Case Study B: The Development of a Malaria Vaccine . . . . . . . . . . . . . . . . . . . . . 225

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 225

The Malaria Parasite . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 226

Malaria and Immunity. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 228

A Malaria Vaccine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 230

Funding Sources . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 236

The Future of Malaria Vaccine Research . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 238

Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 240

Case Study P references . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 240

TABLES

Table No. Page

A-1. Infectious Agents That Cause Infant Diarrhea . . . . . . . . . . . . . . . . . . . . . . . . . 203

A-2. WHO Glucose/Electrolytes Solution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 211

FIGURES

Figure No, Page

A-1. Incidence of the Main Groups of Infectious Diseases Among Children

During the First 3 Years of Life in Less Developed Countries . . . . . . . . . . . . 204

A-2. Interrelationship of Factors Perpetuating Diarrheal Disease . . . . . . . . . . . . . 204

A-3. infant Mortality by Prominent Causes in New York City, 1898-1930 . . . 206
Case Stud A.- Oral Dehydration Therapy

for Diarrheal Diseases

Myron M. Levine, M. D., D.T.P.H.

Professor of Medicine and Pediatrics

University of Maryland School of Medicine

Summary

In the living conditions prevalent in the less developed world, characterized by a lack of potable water,

sanitation, and refrigeration, the bacteria and other

pathogens that cause diarrheal diseases are easily transmitted to young children by contaminated water, hands,

and food. As a result, infants in less developed countries suffer on the average six to eight separate episodes

of diarrheal disease per child.

The two major consequences of infant diarrhea are

dehydration and malnutrition. Diarrhea causes a loss

of body water and salts (electrolytes), which, if sufficiently copious and not adequately replaced, may result in dehydration. Approximately 1 of every 150 to

200 episodes of infant diarrhea results in severe, lifethreatening dehydration. In the last decade, diarrheal

dehydration was one of the major causes of infant

mortality in virtually all less developed countries,

All episodes of infant diarrhea have nutritional consequences, in part because ill infants eat less and in

part because the pathogens that cause diarrhea often

diminish the ability of the small intestine to absorb nutrients. Repeated diarrheal infections within a short

period of time can lead to clinically overt malnutrition.

Irrespective of the specific infectious agent causing

diarrhea, the treatment of diarrheal dehydration is the

same and involves the replacement of body water and

electrolytes (salts in solution). In the 1950s and 1960s,

this treatment was accomplished by intravenous rehydration—i.e., infusion of sterile water and electrolytes into the vein of an infant. Although intravenous

therapy is very effective, it was poorly suited to use

in the less developed world where it was most needed.

The drawbacks of intravenous dehydration include its

expense; the need for sterility of the fluids, tubing, and

needles; the requirement for relatively sophisticated

health workers to administer the infusion; and the

difficulties of distribution and supply of all the

materials.

In recent years, thanks to important contributions

by a number of laboratory scientists, clinicians,

epidemiologists, and health delivery experts, a simple,

efficacious, technologically appropriate alternative to

intravenous dehydration has become available—namely,

oral dehydration therapy (ORT) using sugar/electrolyte solutions. An application of ORT other than for

treating dehydration involves its use early in the course

of infant diarrhea to prevent dehydration. The ingredients used in oral dehydration solutions are widely

available, easily transported, and need not be sterile.

Furthermore, sophisticated health workers are not required to administer ORT.

The efficacy of an oral glucose/electrolyte solution

to promote sodium and water absorption was demonstrated in clinical intestinal balance studies by U.S.

investigators in the Philippines in the early 1960s. By

the end of that decade, clinical studies by U.S. physicians and their Bangladeshi and Indian collaborators

had established that ORT could diminish by 80 percent the intravenous fluid requirements of adults with

severe cholera. Similar studies were carried out in children with cholera and adults and older children with

severe diarrhea not due to cholera.

Since 1977, investigators have carried out a number of clinical studies perfecting the use of ORT in noncholera infant diarrhea in many less developed areas

of the world. These studies have examined modifications in oral dehydration formulas including the sugar,

base, sodium, and potassium content; they have resulted in practical regimens for health workers to follow in orally dehydrating infants; they have demonstrated the efficacy of oral dehydration, irrespective

of the etiology of the diarrhea; and they have defined

the limitations of ORT.

Several methods have been devised for the preparation of simple sugar/salt solutions that are safe and

can be prepared in the home. These simple solutions

are just as effective as more complex balanced glucose/

electrolyte solutions in stimulating sodium and water

absorption by the intestine, though infants treated with

such solutions have inadequate potassium replacement

and can become potassium depleted, a state with adverse clinical consequences.

ORT has been perfected in recent years to the point

where it represents one of the most powerful weapons in the armamentarium against disease in the less

developed world. Increasingly, less developed countries are undertaking national diarrheal disease control programs in which ORT constitutes the keystone.

201
202 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Many U.S. pediatricians were initially somewhat

resistant to ORT. Through a series of recent seminars,

reviews, and publications in American pediatric journals, however, information on the efficacy and advantages of oral dehydration has been widely disseminated,

The development of ORT illustrates how basic physiological and biochemical research can lead to a highly

effective therapeutic modality and practical public

health tool. Specifically, the observation that during

diarrhea, the intestine maintains its ability to absorb

glucose and that sodium and water absorption accompany glucose absorption provided the physiologic basis for ORT as a clinical tool.

Introduction

More than one-half of the world’s population lives

in less developed areas that are characterized by a lack

of potable water, inadequate means for disposal of human fecal waste, and intense crowding in rudimentary

housing. Under such conditions of poverty, underdevelopment and lack of education, the pathogenic

bacteria, viruses, and protozoa capable of causing diarrheal disease in humans are readily transmitted to

young children. Indeed, under such conditions, the

hands that touch infants, the weaning foods that are

fed to infants, and the water that infants drink are

heavily contaminated with fecal material, and therefore with diarrhea-causing pathogens. As a consequence, in the first 2 years of life, young children in

the less developed countries bear an enormous burden of diarrheal diseases.

This case study examines the causes of infant diarrhea, its consequences, the magnitude of the problem,

and the interrelationship between infant diarrhea and

other major public health problems of the less developed world. The primary focus of the case study is

on ORT. The discussion that follows reviews basic

scientific contributions that provide a physiologic basis for this therapy, obstacles that had to be overcome

to give it widespread acceptance, and hurdles that are

now having to be passed in order to realize implementation of ORT on a global scale.

Definitions of Infant Diarrhea

and Dehydration

The term diarrhea stems from the Greek word meaning “to flow through” and denotes an increased number of stools per day which are in an unformed or liquid state. Since the daily stool patterns of infants vary

greatly owing to diet, feeding frequency, etc., the most

useful operational definition of infant diarrhea is when

the mother (who best knows her baby’s stool pattern)

states that her child has diarrhea (71). Typically, a state

of diarrhea is characterized by at least three loose

stools within a 24-hour period. Since infancy in American pediatric terms refers to the first 12 months of life,

strictly speaking, infant diarrhea would refer to illness

within that period only. However, since diarrheal diseases remain an important cause of morbidity and

mortality for at least the first 24 months of life in less

developed areas, the term “infant diarrhea” in common usage has come to refer to the illness in children

up to 2 years of age (71),

Seventy percent of the human body, by weight, consists of water. Approximately 55 percent of the body

water is located inside tissue cells and is associated with

potassium ion (K ‘); the remaining 45 percent is extracellular and is associated with sodium ion (Na + ).

Irrespective of the specific infectious cause of an episode of diarrhea, the common denominator from the

pathophysiologic point of view is the loss of body

water and salts (electrolytes) in the diarrheal stool. Diarrhea represents mainly loss of extracellular water and

N a

+

, but K+ is also lost in significant quantities.

In the older child or adult, diarrhea is usually a nuisance rather than a mortal danger (although the copious purging that occurs with cholera can dehydrate

even adults). In children under 2 years of age, however, diarrhea of any etiology is potentially lifethreatening if stool losses are copious. The reason is

that the losses of body water and electrolytes resulting from loose stools can lead to significant dehydration, which, in the face of continuing unreplenished

losses, can result in acidosis, shock, and death. In virtually all the less developed countries of the world,

diarrheal dehydration is either the first, second, or

third most common cause of infant deaths (22,48,71,

121,159).

Infants are at enhanced risk of development of severe dehydration for several reasons:

●

●

●

Per kilogram of body weight, an infant each day

requires 2% times more water than an older child

or adult (67). This requirement is mainly due to

the greater surface area of the infant per kilogram

of body weight, which results in proportionally

more water loss by transpiration through the infant’s skin.

An infant cannot talk to specifically communicate

thirst and request fluids.

An infant cannot walk to a source of water or

fluids to help himself or herself quench thirst. The

infant is thus totally dependent on its mother or

caretaker. In many cultures across the world,

mothers actually decrease the amount of fluids

(and breast milk) offered to infants with diarrhea
Case Study A—Oral Dehydration Therapy for Diarrhea/ Diseases ● 203

in the incorrect belief that this will benefit the infant by “resting the gut.”

 • Certain homeostatic mechanisms of the kidney

may be less efficient in the young infant, thereby

diminishing the body’s capacity to adjust successfully to physiologic derangements (3,6).

Etiology of Infant Diarrhea

Prior to the early 1970s, the etiology of most episodes of infant diarrhea was largely unknown, either

in industrialized societies or in less developed areas

(59,62,88,111,124,125,163,164,165). Since the early

1970s, however, there has been a veritable explosion

of knowledge on the causes of infant diarrhea with

many new agents being discovered.

The most important infectious agents that cause infant diarrhea are listed in table A-1. Currently, an etiologic agent can be identified in approximately 65 to

75 percent of the cases of infant diarrhea that occur

in less developed countries if comprehensive microbiologic techniques are used (11,12,83,148).

Black and colleagues, working in Bangladesh, noted

that two etiologic agents, rotavirus and enterotoxigenic

Escherichia coli, were found in approximately 60 percent of the cases of diarrhea] dehydration severe

enough to be seen in a dehydration center (11). Black

and colleagues also followed a cohort of infants prospectively in a rural Bangladeshi village (9). This more

intensive surveillance detected mild as well as more

severe cases of diarrhea, including cases that would

not usually be seen in a treatment center because of

their mildness. Enterotoxigenic E. coli was found to

be the most common cause of diarrhea, accounting for

three episodes per child per year. One-fourth of the

episodes of E. coli diarrhea lasted longer than 10 days

(9).

Magnitude of the Problem

of Infant Diarrhea

In the United States, the infant mortality rate is 11

per 1,000 live births (i.e., 11 of every 1,000 babies born

live fail to survive to their first birthday) (161), and

diarrheal dehydration is an uncommon cause of death.

In contrast, in less developed countries, infant mortality rates typically range between 100 to almost 250

deaths per 1,000 live births, and diarrheal diseases are

almost everywhere 1 of the three major causes of infant deaths. In general, when the infant mortality rate

exceeds 100 per 1,000 live births, at least one-third of

all infant deaths are attributable to diarrheal diseases

(32,101).

In the United States and other industrialized countries, infants experience one to two separate episodes

of diarrhea per year (58,68). These episodes are usually mild, and because of easy access to health care,

adverse consequences of the diarrheal illness rarely occur. In the less developed world, however, infants may

experience six to eight separate episodes of diarrhea

per year in the first 2 years of life (7,81,82). Results

of a study of infectious disease incidence among children in less developed countries are summarized in figure A-1, showing the incidence of diarrhea] disease to

be much higher than rates for the other major causes

of morbidity. Since each separate episode of clinical

diarrhea lasts for several days, one calculates that infants in less developed countries spend approximately

15 percent of their entire life experience in the first 2

years, when rates are highest, suffering with diarrhea.

The World Health Organization (WHO) undertook

to quantitate the magnitude of the problem of infant

diarrhea in the less developed world by reviewing the

published literature to determine the incidence rates

of infant diarrhea and applying the calculated rates to

the 1980 estimates for the population of children under 5 years of age (145). In an extremely conservative

estimate, the investigators calculated that there are at

least 750 million to 1 billion episodes of diarrhea and

4.6 million deaths each year due to diarrhea in children less than 5 years of age in Africa, Asia (excluding the People’s Republic of China), and Latin America. They also estimated that approximately 1 in every

167 episodes of diarrhea in young children results in

death.

Table A-1 .—Infectious Agents That Cause Infant Diarrhea

Type of agent Major importance Lesser importance

Bacteria. . . . . . . . . . . . . Enterotoxigenic Escherichia coli Salmonella

Carnpylobacter jejuni Aeromonas hydrophila

Enteropathogenic E. co/i Vibrio cholerae

Shigella

Viruses . . . . . . . . . . . . . Rotavirus 27NM Gastroenteritis viruses

Atypical adenoviruses Astroviruses

Calciviruses

Protozoa . . . . . . . . . . . . Giardia Iamblia Cryptosporidium

Entameba histolytica
204 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Figure A.1 .—Incidence of the Main Groups of

Infectious Diseases Among Children During the

First 3 Years of Life in Less Developed Countries

(data from a cohort of 45 children observed from

birth to 3 years of age)

-

0-5 6-11 12-17 18-23 24-29 30-35

Age in months

SOURCE: Adapted from L. J. Mata, R. A. Kronmal, and H. Villegas, “Diarrheal Diseases: A Leading World Health Problem, ” in Cholera and Related Diarrheas, O. Ouchtertony and J. Holmgren (eds.) (Basel: S. Karger, 1980).

Nutritional Consequences

of Infant Diarrhea

Besides dehydration, the other major consequence

of infant diarrhea is malnutrition. Virtually every infectious agent that causes acute diarrhea results in a

temporary state of intestinal malabsorption that may

last up to several weeks, during which time ingested

nutrients are not properly digested and absorbed

(14,63,74,76,129). Furthermore, each episode of diarrhea has other nutritional consequences. The diarrheal

infection itself, particularly if associated with fever,

results in increased catabolism (i.e., increased breakdown of body mass to provide energy sources). Studies

in Guatemala by Mata and colleagues (81,82) showed

that infants and toddlers readily consumed the recommended daily caloric intake during periods when they

were not ill. However, young children ill with diarrheal or respiratory infections simply did not consume

the recommended intake of calories during the episode

of illness. When episodes of illness occurred one after

the other, children would fall off the normal growth

curve and malnutrition would become evident. Another

problem faced in many parts of the world is that

mothers in many cultures will decrease offerings of

food to infants with diarrhea.

The Interrelationship of Infant Diarrhea

With Other Major Public Health Problems

of the Less Developed World

Certain health problems are common to all less developed countries. These include high infant mortality, diarrheal disease as a major cause of infant morbidity and mortality, high fertility (manifested as high

population growth), and malnutrition (see fig. A-2).

Because of interrelationships among these various

health problems, control of some of them can have

an important ameliorating effect on others. The interrelationships and the critical points of intervention are

briefly discussed below.

The fertility rate of a country refers to the average

annual number of live births per 1,000 women of childbearing age (15 to 49 years). High fertility leads to

rapid population growth. One school of thought contends that high fertility is in great part the consequence

of high infant mortality, since the latter creates compelling pressures for parents to produce more children

in order to ensure survival to adulthood of some children (16,82,84). For example, in a particular culture,

if two sons are believed to be required for the wellbeing and perpetuation of an agrarian family, the parents aim to produce a sufficient number of “insurance”

children to ensure the survival of the desired number.

Multiple infant deaths and multiple pregnancies lead

to the maternal deprivation syndrome—the poorly

nourished, anemic, exhausted, women ubiquitously

Figure A.2.—lnterrelationship of Factors

Perpetuating Diarrheal Disease

SOURCE: Office of Technology Assessment, 1985.
Case Study A—Oral Dehydration Therapy for Diarrheal Diseases ● 205

encountered throughout the less developed world. The

last infants of such women have even less chance of

survival because of their mothers’ diminished lactational capacity.

High fertility can lead to extraordinary population

growth and pressure on land resources and limited

food supplies. Some less developed countries (e.g.,

Kenya) have annual population increases of more than

3 percent. Consequently, the population is young

(about 40 percent are less than 15 years of age) and

doubles in size every 20 to 25 years!

There is an intimate relation between infant diarrhea and malnutrition (9,10,29,71,80,81,102, 121,130,

143,152,154). Simply stated, multiple bouts of diarrhea lead to malnutrition. Malnutrition, in turn, predisposes to increased incidence, severity, and case fatality of diarrheal disease. Puffer and Serrano’s (121)

classic study of childhood mortality in the Americas

documented the intimate correlation between malnutrition and death due to diarrheal disease.

There is evidence that the malnourished child has

more frequent bouts and increased severity of diarrhea. Protein malnutrition leads to decreased stomach

acid (49); stomach acid is perhaps the most important

nonspecific defense mechanism against bacteria that

cause diarrhea. Diminished stomach acid allows inordinately large numbers of pathogenic bacteria to reach

the small intestine.

Points of Intervention

How is it possible to intervene effectively to diminish or break the vicious cycle portrayed in figure A-2

and described above? In the past, attempts were made

via nutritional intervention and family planning programs. Nutritional programs by themselves have largely

failed, and family planning is least successful where

infant mortality rates are high.

Some countries have broken this cycle in the course

of just 25 to 30 years as a consequence of rapid capital development, industrialization and striking improvement in the general socioeconomic level. Just

how rapidly these changes can occur is demonstrated

by the New York City’s experience between 1900 and

1930. Figure A-3 shows that the infant mortality rate

in New York City in 1900 was 140 deaths per 1,000

live births, a typical rate for a developing country. The

single most important cause of infant mortaIity was

infant diarrhea (referred to as “cholera infantum”). By

1930, New York’s infant mortality rate had plummeted

to approximately 50 per 1,000 live births. This extraordinary fall in the rate was almost entirely attributable

to the decrease in infant deaths from diarrhea] disease.

It is important to note that this decrease in New York’s

infant mortality from diarrheal disease occurred in the

absence of modern treatments such as dehydration

therapy or antibiotics and without the benefit of specific vaccines. Rather, the rapid improvement in living conditions (provision of potable water and sanitation systems) and in the educational level of the

population resulted in a striking decrease in the incidence of diarrheal infections. As the population of

New York City went from living conditions of underdevelopment to those of modern industrialized societies, the transmission of the micro-organisms that cause

diarrhea in young children was sharply curtailed.

Thus, both the incidence of diarrhea and mortality due

to diarrhea] diseases sharply decreased. The epidemiologic changes described in New York City were also

experienced in European countries (including England,

France, and Germany) at approximately the same time.

Although fundamental changes in quality of living

conditions represent the optimal mode of intervention,

such changes are unlikely to occur in many less developed countries in the near future because the means

for rapid capital development and industrialization

simply are not available. Other more practical, simplified interventions must be employed if they are to have

an expectation of success in the short term.

Up to the last decade, there was no simple and effective intervention to attack the vicious cycle in figure A-2 at the point involving infant diarrhea. However, such an intervention now exists: ORT. This is

an inexpensive, highly effective means of replacing the

deficits of body water and electrolytes in dehydrated

infants, ) that is technologically appropriate for use in

less developed countries. ORT constitutes a revolutionary innovation in the treatment of diarrheal disease that has the potential to greatly diminish infant

mortality throughout the world.

History of the Development of ORT

Intravenous Dehydration

Dehydration is the most important complication of

diarrhea; untreated it can lead to death. An important therapeutic advance in pediatrics was the demonstration in the first half of the 20th century that

deaths from diarrheal dehydration could be prevented

if the dehydrated infant’s deficits in body water and

electrolytes were replaced by infusions into a vein of

sterile electrolyte-containing solutions (54).
206 ● Status of Biomedical Research and Related Technology for Tropical Diseases

140

120

100

80

60

40

20

0

Figure A.3.—lnfant Mortality by Prominent Causes in New York City, 1898-1930 (rates per 1,000 births)

Respiratory sy stem

Mostly the pneu monias

 , injury at birth, ateotatasis ,

1898 ’99 ’01 ’02 ’03 ’04 I ’06 ’07 ’08 ’09 I ’11 ’12 ’13 ’14 I ’16 ’17 ’18 ’19 I ’21 ’22 ’23 ’24 I ’26 ’27 ’28 ’29

-

’31

1900 1905 1910 1915 1920 1925 1930

SOURCE: W. McDermott, “Modern Medicine and the Demographic-Disease of Overly Traditional Societies: A Technologic Misfit,” J. Med. Educat. 41 (Sept. Suppl.):

137-162, 1966, used by permission from the Journal of Medical Education

This life-saving intravenous therapy gained widespread use in the United States in the 1940s, 1950s, and

1960s for the treatment of diarrheal dehydration in infants (54). Intravenous dehydration used by U.S. Navy

physicians during large outbreaks of cholera in Egypt

in 1947, in Thailand in 1958-60, and in the Philippines

in 1962 dropped the case fatality rate for severe cholera from 20 to 50 percent down to 1 percent (108,

158,160).

Intravenous dehydration for the treatment of dehydrated infants also became increasingly popular in

less developed countries in the 1950s and 1960s. However, its impact was limited because availability was

generally restricted to hospitals in cities. Even in those

facilities where it was used, it represented a significant

drain on financial and personnel resources.

The disadvantages of intravenous dehydration include the following:

● Intravenous dehydration is expensive.

● The intravenous fluids and needles and plastic

tubing required to administer them must be sterile; often these materials must be imported with

●

●

●

●

hard currency.

In general, a specially trained person is required

to start the intravenous infusion, select the appropriate fluid, and supervise the volume and rate

of administration.

Use of the same needles and tubing in more than

one patient without proper sterilization (a common practice in less developed areas) in an effort

to decrease costs encourages the transmission of

infections such as hepatitis from one patient to

another.

The mother is not directly involved in care of the

infant.

There are logistical problems in ensuring the delivery of supplies of intravenous fluids, tubing,

needles, etc., to rural and less accessible areas.

Oral Rehydration—An Alternative

During the past 20 years, there has evolved an alternative form of therapy, ORT, that has multiple advantages over intravenous dehydration and is well-
Case Study A—Oral Dehydration Therapy for Diarrhea/ Diseases ● 207

suited for use in less developed countries. This new

therapy involves rehydration by the oral route using

solutions containing glucose or sucrose (sugars) and

electrolytes (salts in solution). The advantages of ORT

include the following:

●

●

●

●

●

●

ORT is inexpensive.

The basic ingredients are found within most less

developed countries.

Sterile materials are not necessary.

Highly specialized personnel are not directly required to administer the therapy.

While therapy is in progress, the patient does not

require the same degree of vigilance as a patient

receiving intravenous rehydration.

The mother can be directlv involved in treatment

by administering the oral rehydration solution to

the infant.

Physiologic Basis of Oral Dehydration

The normal intestine has tremendous absorptive capacity. In the normal state in the healthy adult, 7 to

8 liters of endogenous fluids (including saliva, bile,

stomach fluid, intestinal juice, pancreatic juice) are

secreted into the intestine. In addition, approximately

2 to 3 liters of exogenous fluids are consumed each day.

Yet at most, only 200 ml of water are lost each day

in normal feces. The normal intestine absorbs 98 percent of the fluid that enters it in the normal state. In

contrast, during diarrhea, the intestine exhibits net

secretion.

The physiologic rationale for the efficacy of glucose/

electrolyte oral rehydration solution is the fact that

even during diarrhea, the simple sugar glucose continues to be absorbed by the upper small intestine (the

jejunum) via an active transport mechanism (56,107,109).

As molecules of glucose are actively absorbed, molecules of water and sodium (Na + ) are also absorbed;

bicarbonate (HCO, – ) and potassium (K+) are absorbed during diarrhea even without glucose.

The first clinical balance studies documenting the

importance of glucose as an actively transported substrate that promotes water and Na + absorption during diarrhea were carried out in 1962 in the San Lazaro

Hospital in Manila, Philippines by Phillips and coworkers (108) of the U.S. Naval Medical Research

Unit—2 (NAMRU-2), Taipei.

Prior to that time, several clinical investigators had

published reports describing the use of glucose/electrolyte mixtures in oral rehydration of patients with diarrhea, but none of these investigators was aware of

the critical importance of glucose in the absorption of

Na + and water during diarrhea. Rather these workers had included glucose merely as a source of calories

for the child with diarrhea. Darrow (34) and Harrison (44,52) used such solutions to treat infants with

mild dehydration. Chatterjee (21) treated mild cholera patients with oral dehydration. The most extensive and impressive experience was reported by

Menenghello and colleagues (86) from Santiago, Chile.

Each summer during the 1950s, epidemics of infant

summer diarrhea in Santiago overwhelmed the available health services. These Chilean workers set up oral

rehydration units and rehydrated infants with a glucose/saline solution administered into the stomach

through a nasogastric tube. In a comparative study,

these Chilean workers noted that oral rehydration with

their glucose/saline solution was as effective as intravenous fluids in rehydrating moderately dehydrated

infants and was much more practical, simple, and economical. Menenghello and colleagues’ experience was

reported in the American pediatric literature in 1960

in the annual publication Advances in Pediatrics, but

little notice was given to it.

Prior to 1962, there also had been several publications in the scientific literature establishing that in animal intestine glucose is actively absorbed and promotes the absorption of Na +

. Fisher and Parsons (45)

are credited with being the first to show that glucose

is actively absorbed by rat intestine and that its absorption promotes water absorption. Riklis and Quastel

(127) in 1958, and Curran (31) in 1960, demonstrated

that glucose also promoted the transport of Na + and

Cl– ions. Finally, Schedl and Clifton (140) as early

as 1963, reported that glucose exhibited a stimulator

effect on the absorption of Na + and water by normal human intestine.

It is not clear whether Phillips and associates were

aware of the above reports when they commenced

their studies in Manila in 1962. Van Heynigan and Seal

discusses this question in their book Cholera—The

American Scientific Experience 1947-1980 (155). They

quote one associate of Phillips, Craig Wallace (p. 229),

as believing that Phillips was indeed aware of these

early physiologic studies reporting the effect of glucose on Na+ and water absorption. In contrast, they

point out that another friend and collaborator of Phillips, Sir Graham Bull (p. 229-230), was convinced that

Phillips was unaware of the importance of glucose in

stimulating absorption of Na + and that his inclusion

of glucose in solutions given to certain patients in

Manila in 1962 was “purely to make the solution

isosmolar, ”

Whether Phillips did or did not know of the earlier

studies is probably irrelevant in the historical context.

What is important is that he carried out elegant and

precise clinical balance studies on adult patients with

cholera who ingested various solutions. In the course
208 ● Status of Biomedical Research and Related Technology for Tropical Diseases

of these studies, he demonstrated for the first time the

salutary effect of glucose on absorption of both Na +

and water in patients with severe diarrhea.

Phillips first published his observations in 1964

(107). In these balance studies, Phillips measured diarrheal stool output (in ml/kg/hr) and quantitated the

electrolyte content of the cholera stools. He also carefully measured the volume and rate of administration

of the various oral solutions that he tested. By subtracting the hourly oral intake from the hourly stool

output, Phillips was able to gauge the effects of various oral solutions on stool output.

In this way, Phillips clearly documented that pure

distilled water, devoid of any electrolytes or other solutes, was readily absorbed by an adult cholera patient and was able to replace the water losses in moderate cholera. Phillips reported that the patient with

cholera drank two glasses of water (500 ml) each hour

without difficulty. However, choleraic stools of adults

contain considerable Na + (approximately 13s mMol/1)

as well as other electrolytes. Since plain water does

not replace the Na + losses, patients with moderate

cholera rapidly become deficient in Na +

. The Na +

deficits must be repaired or the patient’s life would

soon be endangered.

Phillips had cholera patients drink a balanced oral

solution that contained concentrations of electrolytes

closely resembling those found in the stools of an adult

with cholera. Phillips noted that in contrast to plain

water, the electrolyte solution resulted in no absorption whatsoever of water or Na+ even with the patient drinking 700 ml each hour. Only K+ was readily absorbed. Phillips next demonstrated that when a

healthy adult without diarrhea drank the same electrolyte solution at the same rate, no diarrhea occurred.

This showed that the healthy intestine could easily absorb the water and electrolytes in the large volumes

of the solution ingested.

In the next step, Phillips gave an electrolyte solution containing a much higher concentration of Na +

(230 mMol/1) than cholera stool (135 mMol/1) or serum (140 mMol/1). Under these circumstances, as well,

water and Na + losses increased in the stool.

Phillips next decided to “investigate the effect of oral

solutions supplemented with nonelectrolytes such as

D-glucose.“ The solution he used included 95 mMol/1

of Na + and 100 mM/1 of glucose. The patient drank

the solution at the rate of 500 ml/hr. Phillips found

that glucose was absorbed during cholera, and along

with it water, Na +

, Cl –

, HCO3 

–

, and K+. Phillips exact words in the 1964 report are of interest (107):

Furthermore, it was found that the glucose was absorbed, indicating that the glucose transport mechanism was not inactivated in cholera.

The use of the term “glucose transport mechanism”

strongly implies that at least at the time of preparation of his 1964 manuscript, Phillips was aware of

studies showing intestinal absorption of glucose by an

active transport mechanism.

The last clinical experiment described by Phillips involved the evaluation of increasing concentrations of

glucose from so to 400 mMol/1 with the electrolyte

concentrations remaining the same. As the glucose

concentration increased, Na + losses in the stool decreased (i.e., Na + absorption increased).

Phillips’ closing statement is prescient and worth

noting in its entirety (107):

Oral therapy in cholera. From the above studies it

is evident that oral solutions can replace the HZO, K+,

and some of the HC03– losses in cholera. This means

that the only intravenous requirement is the replacement of the Na + and Cl — losses. We have further

evidence which suggests that by incorporation of glucose in an oral solution that one may be able to develop an oral treatment regimen which in the average

case might completely eliminate the requirement for

intravenous fluids. I would like to urge caution on this

particular point. Such a regimen can only be validated

by careful balance studies of the type reported here.

The literature on cholera abounds with treatment regimens which had been enthusiastically urged by their

proponents in whose hands there has been great success. When the same treatment was tried by others,

it appeared to be useless, I am sure this audience will

readily understand why we believe it is necessary to

carefully document any new therapeutic regimens suggested for the treatment of cholera.

Further Studies of Intestinal Absorption

In the 5 years following Phillips’ 1964 report on the

effect of glucose on intestinal absorption of Na + and

water in patients with cholera, important research

proceeded rapidly in two independent but mutually

related areas. First, physiologists and gastroenterologists began to study intensively the precise mechanisms

by which glucose enhanced Na + and water absorption in the intestine of healthy humans and animals

(5,40,46,47,128,141,144). Second, concomitantly, Americans and Bangladeshis in Dhaka, East Pakistan (now

Dacca, Bangladesh), and American and Indian investigators in Calcutta, India, carried out clinical intestinal flux studies to expand on and confirm Phillips’ observations in the Philippines, again establishing the

feasibility of using oral dehydration in the treatment

of cholera (56,109).

Schultz and Zalusky (141) suggested that the enhancement of Na+ transport by glucose is due to an

interaction with the sugar transport per se, i.e., there

occurs a glucose-coupled Na + transport. Sladen and
Case Study A—Oral Dehydration Therapy for Diarrheal DiseasesŽ 209

Dawson (144) came to the same conclusion from studies of intestinal absorption in humans. Fordtran and

coworkers (47), who also studied glucose and Na +

absorption in humans, concluded in 1968 that while

a small fraction of Na + absorption occurs by active

transport coupled to glucose, most Na + absorption

occurs in the osmotic flow of water molecules created

by the active absorption of glucose. (This means that

as glucose molecules are actively absorbed, osmosis

results in absorption of water molecules as well. And

as the water molecules are absorbed, Na + is dragged

along. ) In a 1975 publication, Fordtran (46) reiterated

that both active and passive sodium transport occurs

in the jejunum and that the relative importance of each

depends on the glucose and Na+ concentrations in the

jejunal lumen. Fordtran again concluded that solvent

drag, rather than active cotransport, is the most important mechanism by which glucose stimulates net

Na + absorption in the human jejunum.

Other investigators showed that substrates other

than glucose, including certain amino acids, also promote the absorption of Na + and water as they are

actively transported (50,128,142,147,151). This finding raises the possibility of including more than one

actively absorbed substrate in an oral dehydration solution to stimulate absorption of water and Na +

.

Hirschhorn and colleagues (56), working at the Pakistan Southeast Asia Treaty Organization Cholera Research Laboratory (CRL) in Dacca, and Pierce and

colleagues (109), working at the Johns Hopkins International Center for Medical Research and Training

(ICMRT) at the Infectious Disease Hospital in Calcutta, carried out elegant clinical balance studies to expand the preliminary work initiated by Phillips (107)

in patients with cholera. These clinical studies provided

a sound physiologic basis in humans for proceeding

with clinical trials of oral glucose/electrolyte therapy

in cholera and other diarrheal diseases.

Hirschhorn and colleagues (56) infused various

sugar/electrolyte solutions into the stomach or small

intestine of eight patients with cholera. The electrolyte concentration of the solution was fixed (and included 134 mMol/1 of Na + ), but varying concentrations of glucose, galactose, or fructose were added.

Hirschhorn’s group confirmed that glucose caused enhanced Na + and water absorption. Fructose, in contrast, was shown to have no enhancing effect.

Pierce and colleagues (109) carried out balance studies in 14 adults with cholera who were divided into

three groups to receive one of three solutions by nasogastric tube. All three solutions contained approximately the same concentrations of K+ and HCO3 

–

,

while the Na + and glucose concentrations varied.

These investigators confirmed that the intestine’s ability to absorb glucose remained intact during cholera

and that glucose stimulated absorption of Na + and

water. They reported that raising the glucose concentration of the solution infused into the stomach from

40 to 160 mMol/1 doubled the absorption of water and

Na +

. However, further raising the glucose concentration from 160 to 220 mMol/1 did not further enhance water and Na + absorption.

Progression of Clinical Research Studies

Leading to ORT as It Is Practiced Today

Early Studies in Adults With Cholera

The first publication investigating the therapeutic

efficacy of an oral glucose/electrolyte solution was reported by Nalin and coworkers (95) from the CRL in

Dacca and involved adults with moderately severe

cholera. After receiving intravenous fluids to treat

shock, 29 patients were randomly assigned to one of

three groups. The first group of 10 patients continued

to receive intravenous fluids, the standard therapy for

cholera; the second group of 10 patients received a glucose/electrolyte solution by stomach tube after they

were treated for shock with intravenous fluids; and

the third group of 9 patients drank the glucose/electrolyte solution after they were successfully treated for

shock.

Nalin’s group (95) demonstrated for the first time

that the intravenous fluid requirements of patients with

severe cholera could be diminished by 80 percent if

they either drank a glucose/electrolyte solution or had

it infused through a stomach tube. The composition

of the oral glucose/electrolyte solution used in this

study contained 120 mMol/1 of Na + and 110 mMol/1

of glucose. Cholera patients drank 400 to 1,050 ml each

hour without difficulty, and vomiting was not a

problem.

In 1969, Pierce and coworkers (110) in Calcutta reported a similar favorable experience using an oral glucose/electrolyte solution in 20 adult patients with cholera. The solution contained 120 mMol/1 of glucose

but had a somewhat lower Na+ concentration (100

mMol/1) than the solution used by the CRL group (95).

Only one patient in the oral therapy group required

some additional intravenous fluids to recover. The results of the Calcutta group corroborated the report

from CRL,

In 1979, Cash and coworkers (17) reported from the

CRL in Dacca that adults with moderate cholera and

dehydration short of overt shock could be entirely

treated by oral dehydration using a solution that contained 120 mMol/1 Na+ and 110 mMol/1 glucose,

completely obviating the need for intravenous fluids.
210 ● Status of Biomedical Research and Related Technology for Tropical Diseases

In other studies at this time, Nalin and Cash (93) compared glucose/electrolyte solutions containing 2 percent glucose (110 mMol/1) and 4 percent glucose (220

mMol/1), and found that net water balance was equally

good with the lower concentration. Since Pierce and

colleagues (109) had earlier shown that a solution containing 5 percent glucose offered no advantage over

a 3 percent glucose solution, this combined experience

led to the general acceptance of 2 percent glucose as

the concentration in all future clinical studies with oral

dehydration solutions.

Studies in Children With Cholera

Nalin’s group at the CRL in Dacca next examined

oral dehydration in children with cholera (94,96). In

the early 1970s, the CRL investigators were routinely

using an oral glucose/electrolyte solution containing

120 mMol/1 of Na+ and 110 mMol/1 of glucose. The

results in children were as good as those obtained in

adults with cholera. The intravenous fluid requirements of children with severe cholera were reduced by

80 percent, and children with mild or moderate cholera could be dehydrated entirely by the oral route. Evidence was rapidly accumulating showing the efficacy

of oral glucose/electrolyte solutions in treatment of patients of any age with cholera.

Studies in Patients With Noncholera Diarrhea

The next major contribution of clinical investigators

working in the Indian subcontinent was the demonstration that oral glucose/electrolyte solution was

effective in treating adults and older children with

diarrheal dehydration of etiology other than cholera

(92,136). Most of these adult patients were later found

to be infected with enterotoxigenic E. coli (137).

Preliminary Use of ORT

in Field Dehydration Centers

Although experience with ORT in Dacca and Calcutta was rapidly increasing and the scope of its use

was being expanded, ORT was at first used only in

clinical research studies under carefully controlled conditions. Beginning in 1970, however, reports of evaluations of ORT under field conditions in rural treatment centers began to appear (18,78). Cash and

coworkers (18) showed that ORT could be used successfully under field conditions in rural East Pakistan

during an epidemic of cholera. As in the controlled

environment, the requirement for intravenous fluids

was decreased by 70 percent.

In 1975, a notable report by Bangladeshi researchers showed the promise of ORT (78) by describing its

use in a crowded camp for Bangladesh refugees during a cholera epidemic. Polyethylene packets of glucose and salts were prepared by hand. When mixed

in 1 liter of water, the solution yielded 90 mMol/1 of

Na + and 121 mMol/1 of glucose. No potassium was

included since K + salts were unavailable. Under extremely adverse, primitive conditions, 3,703 patients

with clinical cholera were treated with oral glucose/

electrolyte solution and only a small supply of intravenous fluids. The case fatality rate was only 3.6 percent in this epidemic, largely because of the availability of ORT.

Early Reports of ORT in Infants With

Diarrheal Dehydration

Despite the expanding use of ORT in Dacca and in

Calcutta in the mid-1970s, there were few reports of

its use in infants and young children outside of the Indian subcontinent. This represented a problem since

the greatest need for oral dehydration worldwide was

in the treatment and prevention of dehydration in infants and young children with noncholera diarrhea

rather than in the treatment of adults with cholera.

Up through 1977, almost all of the reports describing oral dehydration of pediatric patients originated

from Asia (almost all from the Indian subcontinent)

and often included children with cholera. Thus, a false

perception was perpetuated elsewhere in the world that

oral therapy was mainly for cholera.

The earliest reports of the use of oral glucose/electrolyte solution outside Asia were those of Hirschhorn

and coworkers (55,57), who treated moderately dehydrated Apache infants with oral dehydration in Arizona. In these pioneer efforts, the investigators clearly

demonstrated ORT’S efficacy by means of monitoring

body weights, stool volume, and plasma protein concentration. The method of dehydration Hirschorn and

colleagues described was referred to as “ad Iibitum”

treatment, implying that when glucose/electrolyte solution is offered to an infant, the homeostatic mechanisms of the infant would lead the infant to drink until

dehydration was established.

The “ad libitum” treatment regimen was not readily adopted by pediatricians or other health workers

for three reasons:

1.

2.

The “ad libitum” treatment does not make use of

the concept of estimation of fluid deficits and

replacement of the calculated deficit. This is the

fundamental physiologic concept on which all rational dehydration therapy, oral or parenteral,

should be-based (79). -

-

The treatment is not easy to teach to unsophisticated health workers.
Case Study A—Oral Dehydration Therapy for Diarrheal Diseases • 211

3. A small percent of overtly dehydrated infants do

not readily drink oral dehydration solutions and

must be actively coaxed. The fluid needs of such

infants would not be recognized if a pure “ad libitum” method were followed.

Clinical Studies of ORT in

Infant Diarrhea Since 1977

Since 1977, there has been a veritable explosion of

information on the use of ORT in infants in many

areas of the world including Central and South America, Africa, Southeast Asia, and the Caribbean basin.

In careful clinical research studies, the remaining voids

of knowledge were filled and obstacles impeding widespread acceptance of ORT were overcome. Much of

this research was carried out by five groups: from the

University of Maryland School of Medicine; the National Children’s Hospital, San Jose, Costa Rica; the

International Center for Diarrheal Diseases Research,

Bangladesh (ICDDR, B); the Baltimore City Hospital;

and the Kothari Pediatric Gastroenterology Center,

Calcutta. Most of these studies in infants and children

used the glucose/electrolyte solution recommended by

WHO (shown in table A-2) or a very similar solution.

These clinical studies established the efficacy of ORT

in infants, identified its limitations, examined efficacy

in relation to etiology, and evaluated modifications in

the sugar and base content, and Na + and K+ concentration in oral dehydration solutions. These studies

are summarized below.

Clinical Indications, Regimens,

and Limitations of ORT

Development of Practical Methods

To Rehydrate Dehydrated Infants

Several groups of researchers have described regimens of ORT that were highly successful in dehydrating dehydrated infants (except those in overt shock)

(24,28,100,114,139). These methods used the concepts

of percent dehydration and fluid deficit estimations.

Three methods (28,100,114) included offerings of plain

water to infants, in addition to sugar/electrolyte solutions, to prevent the development of hypernatremia

(a condition in which the serum Na + concentration

exceeds 1.50 mMol/1).

Two groups (24,139) have demonstrated that dehydrated infants can be safely rehydrated with sugar/

electrolyte solution alone without hypernatremia developing. Other investigators (113,132) have corroborated these findings.

Table A.2.—WHO Glucose/Electrolytes Solution

Ingredients:

NaCl. . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.5 gm

NaHCOs. . . . . . . . . . . . . . . . . . . . . . . . . 2.5 gm

KCI . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.5 gm

Glucose. . . . . . . . . . . . . . . . . . . . . . . . .20 gm

HzO . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.0 liters

Electrolytes in resulting solution (mMol/liter):

Na+cl- HC0 3

- Glucose

90 20 80 30 111

Acceptability of Oral Solutions

and Intestinal Water Absorption

Many researchers have documented that the vast

majority of overtly dehydrated infants, despite fever,

malaise, and lethargy, avidly drink sugar/electrolyte

solutions (19,20,24,26,28,73,100,113,114,118,132,139),

even though the taste is described as unpleasant by

healthy adults. These overtly dehydrated infants gain

weight from absorption of water and electrolytes (19,

20,24,26,28,73,100,113,114,118,132,139). Approximately 95 percent of dehydrated infants, even those

10 to 12 percent dehydrated (but not in shock) can be

successfully rehydrated with oral sugar/electrolyte solutions. (Dehydration with loss of more than 10 to 12

percent of body weight is almost always accompanied

by clinical shock. Mild dehydration refers to up to 5

percent loss of body weight; moderate dehydration

means loss of 6 to 9 percent; 10 percent or greater loss

of body weight represents severe dehydration. ) The

demonstration that infants with 8 to 12 percent dehydration could be orally rehydrated was important,

because it dispelled the contention of some pediatricians (101) that oral dehydration alone was incapable

of dehydrating infants with moderate and severe dehydration short of shock.

ORT’S Efficacy in Relation

to the Etiology of Diarrhea

Studies from Costa Rica (100) and those of investigators in other parts of the world, including Dacca

(13,134,150), have shown that ORT is effective in 95

percent of cases of diarrheal dehydration, irrespective

of etiology. ORT is equally efficacious whether diarrhea is due enterotoxigenic bacteria (e. g., enterotoxigenic E. coli. Vibrio cholerae), invasive bacteria (e.g.,

Shigella spp., Campylobacter jejuni), enteropathogenic

E. coli, or viruses.

Of particular interest is ORT’S efficacy in rotaviral

infection. Davidson and coworkers (35) showed that

in isolated loops of piglet small intestine heavily in-
212 • Status of Biomedical Research and Related Technology for Tropical Diseases

fected with human rotavirus, there was no evidence

for glucose-coupled Na+ absorption. On the basis of

these observations, they inferred that oral glucose/

electrolyte solutions might be ineffective in infants with

rotavirus infection. In fact, however, ORT is as effective in infants with rotavirus infection as it is in nonrotavirus infection (13,100,134,150).

The pathophysiologic explanation is that rotavirus

causes patchy involvement of the small bowel with islands of denuded intestinal surface surrounded by normal or minimally affected areas. While the severely

affected areas undoubtedly are defective in active glucose absorption, the nonaffected surface of the bowel

absorbs sufficient glucose to permit excellent clinical

results. When Nalin and coworkers (100) measured

stool glucose concentration during oral therapy in infants with rotaviral diarrhea and infants with nonrotaviral diarrhea, the concentration was significantly

higher in the former, demonstrating that indeed glucose absorption was somewhat impaired. Nevertheless, sufficient active transport of glucose occurred to

allow 92 percent of rotavirus-infected infants to be successfully treated with ORT. In Costa Rica, rotavirus

is by far the major single cause of diarrheal dehydration in infants (83), and ORT is the mainstay of therapy in the Emergency Room Service of the National

Children’s Hospital. Similar success in the treatment

of dehydration due to rotavirus has been reported in

Bangladeshi infants (13,150).

Oral Dehydration and Vomiting

Infants with diarrhea commonly manifest vomiting.

Pediatricians and nurses unfamiliar with ORT often

conclude that ORT cannot be instituted or that it will

not be effective in those cases. This is a fallacy. In carefully monitored clinical studies in Costa Rica and Honduras (28,100,114), most of the infants had a history

of vomiting on admission and most also experienced

one or more episodes during ORT. However, careful

balance studies in which the volume of vomitus was

carefully recorded showed that with rare exceptions,

the volume vomited was only a small fraction of the

total volume of glucose/electrolyte solution ingested

(28,73,100,114). From the perspective of the fluid and

electrolyte status of the patient, it is not the frequency

of vomiting that is of importance but the net balance

of intake versus emesis (vomitus) and stool volume.

Studies in Costa Rica (114) have shown that vomiting tends to be more frequent during the early hours

of ORT and diminishes thereafter. This observation

suggests that emesis in some infants may be the consequence of electrolyte imbalance and acidosis.

The rate of emesis is also related to the sugar in the

solution and to the method of administration. In comparative trials between glucose/electrolyte and sucrose/electrolyte solutions, children who received the

sucrose/electrolyte solution had higher rates of vomiting (13,28). In summary, careful balance studies have

provided the evidence to demonstrate that vomiting

is rarely an obstacle to successful ORT.

ORT of Hypernatremic and

Hyponatremic Dehydration

In approximately 70 percent of cases of diarrheal

dehydration in infants, the concentration of Na+ in

serum remains normal (131 to 149 mMol/1). In a minority of cases, the serum Na + may be elevated (hypernatremia, –>150 mMol/1) or abnormally low (hyponatremia, < 130 mMol/1).

Hypernatremic diarrheal dehydration represents a

serious complication. It usually occurs in young infants and can be accompanied by seizures, cerebral

hemorrhages, and death. Hypernatremia is notoriously

difficult to treat because of the propensity of infants

to convulse during therapy. In the 1950s and 1960,

hypernatremia was encountered in approximately 20

percent of infants with diarrheal dehydration seen in

North America and Western Europe. Older American

and European pediatricians and family practitioners,

therefore, have great respect for this complication of

infant diarrhea, consequent to their experiences several decades ago. In general, hypernatremia has been

uncommon in infants in the tropics.

Some pediatricians have expressed concern that oral

glucose/electrolyte solutions of the type recommended

by WHO contain a Na+ concentration (90 mMol/1)

that is too high for infants, particularly those in industrialized countries (6,9,64,153,156,157). They suggest that in these well-nourished, predominantly formula-fed infants, who suffer mainly from rotavirus

diarrhea, glucose/electrolyte solutions with Na+ concentrations of 90 mMol/1 are potentially unsafe and

could lead to or exacerbate hypernatremia.

Considerable confusion has resulted from discussions on this point because of the failure to differentiate clearly between the use of oral glucose/electrolyte

solution to replace fluid deficits in overtly dehydrated 

infants versus administration of glucose/electrolyte solution early in the course of diarrhea to prevent dehydration. The use of solutions with Na + concentrations of 90 mMol/1 in nondehydrated infants with

diarrhea must be accompanied by equal volumes of

low solute fluids or plain water to allow the kidneys

to handle Na+ loads which are greater than the stool

Na+ losses. However, in the overtly dehydrated infant, even if hypernatremia is already present, there

exists a total body deficit of Na + as well as water.

In overtly dehydrated infants, a glucose/electrolyte so-
Case Study A—Oral Dehydration Therapy for Diarrheal Diseases ● 213

lution containing 90 mMol/I is physiologically sound

and clinically safe; indeed, it is an ideal dehydration

solution to replace deficits (26,113,115,119).

Pizarro and coworkers have carried out several

studies (113,115,119) in Costa Rica clearly establishing the safety of ORT of infants with hypernatremic

dehydration. The problem with treatment of hypernatremic dehydration, intravenous or oral, is the

occurrence of seizures (15,42,65,75,119). The last report by Pizarro and coworkers (113) described the use

of slow oral dehydration in 35 infants with no episodes

of seizures during therapy.

ORT also has been shown successful in treating infants with hyponatremia (serum Na + concentration

<130 mMol/1) (113,119,139).

Experience With ORT in Neonates

At the National Children’s Hospital in Costa Rica,

the use of ORT in neonates has been exhaustively studied by Pizarro and colleagues (116,117). An extraordinary collective experience involving several hundred

newborns documents that ORT is as safe and effective in this age group as in older infants. Furthermore,

neonates with hypernatremic and hyponatremic dehydration can be as successfully rehydrated as older children (116,117).

Rapidity of Oral Dehydration

Complete replacement of the water and electrolyte

deficit, accompanied by clinical signs of normal hydration and weight gain, is complete by 6 hours after beginning ORT in approximately 60 percent of cases; by

12 hours in 90 percent; and in 95 to 98 percent of instances by 18 hours (24,28,73,100,114,118). At the

point of successful dehydration, infants are switched

to the maintenance phase of dehydration, which includes the introduction of soft foods. Depending on

social factors, the infant may be sent home at this point

after the mother has been instructed in how to provide maintenance fluids at home and has been given

packets to prepare the solution (118).

Limitations of ORT

Even in highly experienced units such as those in

Costa Rica, Honduras, Bangladesh, and India, 2 to 5

percent of overtly dehydrated infants will fail on ORT

alone. The most frequent causes of failure include the

following:

● Glucose or sucrose intolerance. Overtly dehydrated infants often are severely malnourished

and have intestinal bacterial infections. When

given sugar/electrolyte solutions, these infants

greatly increase their purge rates. This situation

is seen in perhaps 1 percent of cases in less developed areas.

• Intractable vomiting. Occasionally, a dehydrated

infant will simply be unable to drink without

vomiting virtually the entire fluid volume that

was just ingested. Such instances are rare.

Ž Continuing high purge rates. Most infants with

diarrheal dehydration have diminished purge

rates following replacement of fluid deficits or

purge at rates that can be adequately replaced

orally. Some infants, however, particularly those

with cholera, continue to purge copiously at rates

that cannot be easily replaced with ORT.

● Abdominal distention. Paralytic ileus or abdominal distention occasionally occurs, precluding

continued ORT.

ORT is labor-intensive. Someone must continue to

administer the fluids to the infant. If the mother or

another guardian is not available, this requirement becomes a limitation. ORT units are dependent on the

full cooperation of the infant’s mother or a surrogate.

Composition of Oral

Dehydration Solutions

Comparison of Glucose and Sucrose

Because of the lesser expense and greater availability of sucrose (table sugar), trials have been carried

out to compare the efficacy of sucrose/electrolyte

versus glucose/electrolyte solutions (13,99,103,134,

135). Sucrose molecules are “double sugars” twice the

size of glucose molecules. For sucrose to be effective,

it must be broken down by intestinal enzymes to its

single sugar constituents, glucose and fructose, whereupon the actively absorbed glucose molecules promote

water and glucose-coupled Na + transport. Fructose,

in contrast, is not avidly absorbed and thus exerts a

“solute drag” osmotic effect (56). In summary, the

comparisons show that glucose/electrolyte solutions

are slightly superior to sucrose/electrolyte solutions.

However, the differences in overall efficacy are minimal so that if economic or logistic considerations are

paramount, sucrose-based solutions can be routinely

used with expectation of excellent clinical results.

Comparison of “Low” and “High”

Sodium Dehydration Solutions

Several controlled studies have been carried out in

which dehydrated children were rehydrated orally

with either “high” Na + (90 mMol/1) or “low” Na

(50 to 60 mMol/1) glucose/electrolyte solutions (4,20,
214 ● Status of Biomedical Research and Related Technology for Tropical Diseases

98,133,138,139). High- and low-sodium rehydration

solutions appeared equally efficacious in replacing

fluid deficits.

In a study in Jamaica (98), Nalin and coworkers detected transient hypernatremia in 4 of 25 minimally

dehydrated infants who were given the WHO glucose/electrolyte solution without free water. The

hypernatremia was mild (150 to 156 mMol/1) and was

not associated with any adverse clinical signs or symptoms. Chatterjee and colleagues (20) in India reported

similar results. Two of their dehydrated infants who

had normal serum Na + concentrations on admission

developed asymptomatic mild hypernatremia during

ORT with a glucose/electrolyte solution containing 90

mMol/1. In contrast, Santosham and colleagues working in Panama and in the United States did not encounter hypernatremia (138,139) in their orally rehydrated

infants who also received the solution without free

water.

In the study in Jamaica (98), net Na+ absorption

was significantly less in the infants who received the

low Na+ (60 mMol/1) solution and hyponatremia developed in 3 of 31 infants. Furthermore, several infants

who were hyponatremic on admission remained so

during oral rehydration with the low Na+ concentration solution. A comparison of “high” and “low” Na +

sucrose/electrolyte solutions by Saberi and Assaee

(133) in Iran gave comparable results as seen in the

glucose/electrolyte comparisons.

In summary, while in “low” and “high” Na + oral

glucose/electrolyte solutions give equivalent results in

most dehydrated infants, the high Na + solution is

safer for hyponatremic and hypernatremic infants.

Optimal Potassium Concentrations

Losses of potassium ions (K + ) in diarrheal stool can

be significant, particularly in infants in less developed

countries who have repeated episodes of diarrhea. In

such infants, insufficient replacement of K + losses can

lead to total body K+ depletion, accompanied clinically by muscle weakness, ileus, cardiac arrhythmias,

and hypokalemic kidney disease. Nalin and colleagues

(98) compared K+ balance in mildly dehydrated

infants treated with the WHO oral dehydration solution (which contains 20 mMol/1 K+) versus a modified solution containing 35 mMol/1 K+. Net K+ absorption at 24 hours was more than twice as high in

the infants that received the solution containing the

higher K + concentration (35 mMol/1), and none of

these infants had abnormally low or high potassium

blood levels at 6 or 24 hours after beginning ORT. In

contrast, low potassium levels were detected in about

one-fifth of the infants treated with the low K + solution. Based on these studies in Jamaica, the formula

of the oral rehydration solution used in Costa Rica was

modified to provide more K+.

Acetate or Citrate in Place of Bicarbonate

The shelf life of optimally prepared glucose/electrolyte packets containing HCO3

– is 3 years. However,

the shelf life of poorly made porous packets is limited

because of the discoloration and caramelization that

occurs when moist glucose and bicarbonate mix. Sodium acetate and sodium citrate are being studied as

substitutes for sodium bicarbonate to see whether they

prolong the shelf-life. In the body, acetate and citrate

are converted to bicarbonate. Pizarro and colleagues

(112) and Patra and colleagues (105) recently have carried out double-blind randomized comparisons of glucose/electrolyte solutions containing citrate or acetate,

respectively, versus the standard solution containing

bicarbonate. The solutions with citrate or acetate were

equally efficacious.

“Super Solutions”

In 1970, Nalin and colleagues (97) reported results

of a clinical study in which a glucose/electrolyte solution was compared with an electrolyte solution that

contained the amino acid glycine as well as glucose.

Both glucose and glycine molecules are actively transported, but by different mechanisms. The objective of

this study was to determine if a solution containing

two distinct actively transported substances would result in faster and increased water and electrolyte absorption than a solution containing only glucose and

electrolytes. Nalin and coworkers observed that the

glucose/glycine/electrolyte solution was superior to

the glucose/electrolyte solution in promoting water

and Na + absorption in patients with severe diarrhea.

In fact, the glucose/glycine/electrolyte solution was

so much better that the duration of diarrhea and total

stool volume were significantly diminished in patients

who received the solution as compared to the group

who received glucose/electrolyte solution.

The implications of this important observation were

not widely appreciated at the time. Many public health

authorities believed that the lesser availability of glycine and the increased cost of solutions having both

glucose and glycine ruled against widespread use of

such solutions. Since 1982, however, the concept of

“super solutions” with more than one substance actively transported by the gut has reappeared with increased popularity (2,77,144). Patra and coworkers

(144) in Calcutta compared glucose/electrolyte and

glucose/glycine /electrolyte solutions in oral dehydration of infants with diarrheal dehydration. Their results corroborated those in adults with cholera 15 years
Case Study A—Oral Dehydration Therapy for Diarrhea/ Diseases • 215

earlier. The duration of diarrhea and the total stool

volume significantly diminished in infants given the

solution containing both glucose and glycine.

Work is under way to carry these investigations further. The goal is to develop a “super solution” containing up to three actively transported substrates that

not only replaces water and electrolytes but actually

diminishes diarrheal stool volume to the point where

losses are no longer clinically relevant (2).

Some groups of investigators have been examining

naturally occurring complex substrates as opposed to

pharmaceutically prepared formulas in making of “super solutions” (90,106,162). They hope in this way to

accomplish the same goals at lesser cost and with

greater availability. Most work in this area uses rice

powder (30 to 50 gm per liter) with the same electrolyte concentrations as in the WHO soIution. Rice contains polymers of glucose (glucose molecules strung together in a chain) which can be broken down in the

small intestine to single glucose molecules (90). Rice

also contains glycine (30 to 36 mg/100 gm of rice).

Clinical studies have shown rice powder/electrolyte

solutions to be at least as effective as glucose/electrolyte solutions in oral dehydration of infants with diarrheal dehydration (90,106). Some studies in infants

(106) demonstrate a clear superiority of a 5-percent rice

powder/electrolyte solution over the glucose/electrolyte solution, with the former significantly lowering

stool output, duration of diarrhea, and intake of oral

dehydration fluid.

ORT To Prevent Dehydration

Up to this point, ORT has been discussed in this case

study as a substitute for intravenous dehydration to

treat 95 percent of infants with overt clinical dehydration. Used this way, ORT is practiced by health care

providers in health care facilities.

The second major use of ORT is to initiate the use

of oral sugar/electrolyte solutions early in the course

of diarrhea in an attempt to prevent dehydration.

Ideally, this intervention is carried out with a balanced,

physiologically sound solution containing appropriate concentrations of K+ and HCO3 

– (or other base)

in addition to Na+ and Cl—

. In infants, stool Na +

concentrations in noncholera diarrhea usually range

from 25 to 70 mMol/1, with a mean of approximately

45 to 50 mMol/1 (89,100,101,149). Thus, a glucose (or

sucrose) /electrolyte solution containing 40 to 50

mMol/1 of Na + is ideal to replace ongoing diarrheal

losses early in the course of diarrhea in young children (43,44). The WHO solution can also be utilized

to replenish ongoing diarrheal losses in infants but in

this instance, an equal volume of low solute fluids (or

plain water) is given following each volume of glucose/electrolyte solution to provide free water to handle Na+ loads.

Simple Sugar/Salt Solutions

It is not possible economically or logistically to provide a packet of balanced sugar/electrolyte powder to

treat every episode of diarrhea in all young children

in developing countries, For that reason, some observers have advocated the use of simple dehydration solutions of table salt and sugar, prepared and administered in the home (15,53,66,70,85,91,120). Depending

on the purity of the sugar and salt, simple solutions

may contain only Na , Cl + —

, and sucrose.

Two major obstacles had to be overcome before the

use of home-made simple sugar/salt solutions could

be advocated with confidence. First, it was necessary

to identify simple, technologically appropriate methods for producin solutions containing safe and effective levels of Na + and sucrose. This was regarded as

critical by many authorities who feared that improper

mixing could result in solutions with unacceptably high

Na + and sucrose concentrations, which could conceivably induce hypernatremic dehydration, accompanied by convulsions, intracerebral hemorrhage, and

high case fatality. Second, it had to be shown to what

degree simple sugar/salt solutions promoted water and

Na + absorption, how well they combatted acidosis

(since they lacked base), and what clinical and biochemical problems might accrue from the lack of K +

in the fluids.

Many methods for measuring sugar and salt in preparation of simple sugar/salt solutions have been described. Some methods, such as the “finger pinch of

salt and fistful scoop of sugar” technique, use the human hand for measurement (25,91). Other methods

make use of a teaspoon or bottle cap, and a glass or

a 750 ml or 1.0 liter bottle (30,38,69,70,72). Some authorities favor the use of a special double-ended plastic spoon with one trough for measuring salt and

another for sugar (79,53). There is great controversy

over the reliability of some of these methods (8,23,25,

30,32,33,38,51,53,66,69, 70,72,85,91,120,126, 146).

Many health workers consider certain methods such

as the “pinch and scoop” inherently so variable that

they are unacceptable (32,69,72). Others disagree and

believe that the simplicity of the pinch and scoop

method justifies its use (25,91,146). A large rural ORT

program in Bangladesh (Bangladesh Rural Advancement Committee) makes use of the pinch and scoop

method (33).

Despite controversy over particular methods of

preparation of simple sugar/salt solutions, all involved

parties appear to agree on two fundamental points.
216 • Status of Biomedical Research and Related Technology for Tropical Diseases

The first is that one must adapt methods of preparation of simple sugar/salt solutions to local conditions

using local utensils. The second is that the more supervision and teaching involved, the greater the reliability of the method and the greater the safety of the

resultant solutions.

Simple Sugar/Salt vs. Glucose/Electrolyte

Oral Dehydration Solutions

Although many health authorities avidly supported

the use of simple sugar/salt solutions and many national diarrheal disease control programs relied heavily on them, prior to clinical studies in Honduras (28),

there were no physiological data to support their efficacy or show their limitations. Clements and coworkers (28) treated 61 Honduran infants 3 to 18 months

of age with diarrheal dehydration with either a simple sugar/salt solution (60 mMol/1 Na +

, 3.0 gm percent sucrose, no K+ or HCO3

–

) or with the WHO

glucose/electrolyte solution. The simple sugar/salt solution was found to be equal to the glucose/electrolyte solution in stimulating Na+ and water absorption and almost as good in combatting acidosis.

However, because of the lack of K+ in the simple sugar/salt solution, significantly more infants treated

with this solution developed abnormally low serum

K+ concentrations. Small amounts of banana puree

were unable to replace sufficient K+. It would require

at least 320 ml of banana puree (280 ml of mashed

banana, 2 to 3 whole bananas) to provide as much K+

as contained in the amount of WHO glucose/electrolyte solution given over 24 hours to an infant with diarrhea (27).

In summary, properly prepared simple sugar/salt

solutions are highly effective in promoting water and

Na+ absorption, thereby restoring a normal blood

volume and blood flow through the kidneys which

combat metabolic acidosis. However, the lack of K +

in most simple sugar/salt solutions represents a critical drawback which can lead to abnormally low serum K + concentrations. Until the problem of adequate K + replacement can be resolved, simple

sugar/salt solutions must be regarded as suboptimal

alternatives for use only where a balanced sugar/electrolyte solution is unavailable.

Effect of ORT on Nutritional Status

A study in the Philippines showed that children who

received ORT with glucose/electrolyte solution during bouts of diarrhea gained significantly more weight

in ensuing months than matched control children who

did not receive ORT (60,61). Studies in two other

countries (Turkey and Iran) had similar results (7,39).

Field workers, in the Gambia, however, have not been

able to corroborate the results (131). One possibility

is that ORT stimulates the appetite of children with

diarrhea, and this accounts for the increased weight

gain. Since early reintroduction of feeding is part of

the ORT intervention, it is difficult to determine what

role ORT per se plays with respect to food consumption. More information about ORT’S effect on food

consumption and nutritional states is needed.

Effect of ORT on Infant Mortality

Implementation of ORT programs in less developed

countries would be expected to diminish diarrhearelated infant mortality. Unfortunately, however, few

epidemiologic data are available to demonstrate this

effect. Reports from India, Bangladesh, and Egypt provide some insights, but none is completely convincing because of methodologic problems (1,66,123).

More data on this important question need to be

gathered,

Dissemination of ORT

Many international, national, religious, and private

agencies are involved in disseminating the use of ORT

throughout the world. The most important agencies

are the following:

1. World Health Organization (WHO),

2. Pan American Health Organization (PAHO),

3. U.N, International Children’s Emergency Fund

(UNICEF), and

4. U.S. Agency for International Development

(AID).

World Health Organization

ORT is the keystone in WHO models for national

diarrheal disease control programs.

WHO’s Diarrheal Disease Control Program has

played a pivotal role worldwide in stimulating the use

of ORT. This has involved:

●

●

●

●

Pan

provision of treatment guides, manuals, etc., in

several languages;

provision of technical consultants to countries;

assistance in organization of national and regional

workshops on ORT and diarrheal disease control;

and

provision of research grants to support basic, clinical, and operational research related to ORT.

American Health Organization

PAHO is both the WHO regional office for the

Americas and the Pan American Sanitary Bureau.

Both independently and in conjunction with WHO,
Case Study A—Oral Dehydration Therapy for Diarrheal Diseases ● 217

PAHO has pursued dissemination of ORT. This has

involved:

● provision of technical consultants;

. support of national and regional meetings. PAHO

has been active in promoting workshops on diarrheal diseases and ORT; and

● provision of research grants, equipment, and materials to clinical and public health investigators.

In the late 1970s, the first generation of PAHO ORT

consultants were mainly North Americans. These consultants trained a second generation of PAHO ORT

consultants from Latin America and the Caribbean

who have since become internationally recognized authorities.

U.N. International Children’s Emergency Fund

UNICEF has been a major supplier of packets of glucose/electrolyte powders to allow countries to start national pilot programs. UNICEF also has supplied some

countries with machines to package their own ORT

powders. ORT is an integral part of “GOBI/FF” program (growth chart, oral therapy, breast feeding, immunizations/family planning, female literacy).

U.S. Agency for International Development

AID has made major contributions to the dissemination of ORT. Some of the most important contributions include:

● provision of technical consultant support.

● support for meetings and workshops, and

● special projects.

The AID-supported international meeting on ORT

in Washington, DC, in June 1982 was an important instrument to disseminate ORT. One of AID’s special

projects is the Mass Media Diarrhea Project, which has

pilot programs in Honduras and the Gambia. Mass

communications techniques (mainly radio) are being

used to spread health information about diarrhea, dehydration, and ORT. The effectiveness of this program

in Honduras is very impressive. Another special AID

project has been direct financial support for the International Center for Diarrheal Diseases Research, Bangladesh. This is the offspring of the CRL (Cholera Research Laboratory), which has played an important

role carrying out basic, clinical, and operational research on ORT.

Attitudes of U.S. Pediatricians

Toward ORT

The vast majority of pediatricians practicing in the

United States were trained in programs where the following attitudes were fostered:

●

●

●

●

ORT helps to prevent dehydration and is useful

in treatment of mild dehydration.

ORT solutions for U.S. children should not contain more than 60 mMol/1 of sodium in order to

preclude the development of hypematremia (a

much feared complication).

Children with moderate or more severe dehydration must be given intravenous dehydration.

If a child is sufficiently dehydrated to receive intravenous fluids, nothing further should be given

by mouth for 24 hours. The bowel should be

“rested.”

These have been the fundamentals of diarrhea therapy taught to two generations of American pediatricians.

Because of the large number of reports on ORT that

have been published in recent years in the most prestigious pediatric journals in the United States (Journal of Pediatrics, Pediatrics, and the American Journal of Diseases of Children) and Britain (Archives of

Disease in childhood), most U.S. pediatricians are

aware of the expanded use of ORT. Most are also cognizant of the composition of the WHO oral dehydration solution and accept its use as an important advance for the treatment of diarrhea in the less

developed world. Some pediatric training programs

in the United States have adopted ORT and use the

WHO (as well as other) dehydration solutions. Throughout the United States, however, there remain many

pediatricians who are skeptical of ORT and who are

particularly resistant to use of a formula like the WHO

glucose/electrolyte solution which contains 90 mMol/1

of sodium.

The American Academy of Pediatrics has played an

important role in fostering information on ORT and

in attempting to forge agreement on the role of solutions containing 90 mMol/1 of sodium in treatment of

diarrhea in the United States. At its autumn national

meeting in New York City in October 1982, the Academy convened a workshop on ORT. Other presentations on oral dehydration were made at American

Academy of Pediatric meetings in May 1983, in Philadelphia, and in October 1983, in San Francisco.

In February 1983, the Academy convened a task

force on ORT which met in New York. The assignment of the task force was to prepare recommendations on the following issue:

the advantages and disadvantages of Oralyte, a

high electrolyte solution (distributed worldwide by

UNICEF/WHO) in the treatment of infants with diarrhea and dehydration.

Initially, the task force was not able to come to an

agreement. Most of the task force members wanted

to make Oralyte (the WHO solution) a prescription

drug in the United States for fear that some infants
218 ● Status of Biomedical Research and Related Technology for Tropical Diseases

would develop hypematremia (abnormally high serum

Na + concentrations) if it were used without medical

supervision. One dissenter argued that Oralyte had

been proved safe worldwide and was safer than aspirin (which is sold without prescription). Some task

force members believed that adopting a prescription

requirement for Oralyte in the United States could

have serious repercussions internationally, perhaps

impeding the dissemination of in less developed

countries.

A compromise was accepted for the task force to

prepare recommendations, which were sent to the

Academy Committee on Nutrition and then transmitted to the Food and Drug Administration to assist that

agency in developing policy and regulations. The major points of the compromise include the following

(28a):

There is a role for two different sugar/electrolyte

solutions in the treatment of diarrhea in the

United States. The solutions would differ in sodium concentration. A solution for deficit replacement should contain 61 to 90 mMol/1 of Na+

.

A solution for maintenance should contain 40 to

60 mMol/1 of Na +.

The solution for deficit replacement should be

used preferably in a fixed health care facility (physician’s office, emergency room, etc. ) in overtly

dehydrated children.

The higher sodium solution also may be used in

prevention of dehydration or maintenance of

hydration but must be accompanied by provision

of ample additional low-solute fluids to provide

free water.

A preferred maintenance solution is the lower (40

to 60 mMol/1) sodium solution.

All the dehydration solutions should be considered “medical foods. ” (Distribution of medical

foods does not require a prescription, but such

foods are accompanied by explicit instructions. )

The Economy of ORT

ORT makes the treatment of diarrheal disease more

economical in two ways: 1) by diminishing the cost

of the dehydration fluids, and 2) by shortening the hospital stay of the pediatric patient admitted for the treatment of dehydration.

The costs of treatment for two hypothetical patients

treated by ORT versus intravenous dehydration are

compared below.

The first patient is a 10 kg American infant admitted

to a large Eastern municipal hospital with 10 percent

dehydration, In typical pediatric care in the 1970s, this

infant would have a hospital stay of approximately

3 days and would receive intravenous fluids. Typically, he or she would be given nothing by mouth for

the first 24 hours and then would have slow introduction of clear fluids and dilute formula over the subsequent 2 days. Some of the costs of treating this infant

intravenously would include: sterile intravenous fluids

at $1.50/liter (approximately 3 liters might be used in

this patient over 2 days); sterile plastic intravenous

tubing ($2.62); and at least one sterile butterfly needle ($0.40). The relevant cost of intravenous therapy

for the infant would be $4.50 for intravenous fluids,

plus $3.02 for sterile tubing and needle and $600 for

3 days of hospitalization.

In the 1980s, this same infant could be treated with

ORT and early reintroduction of feeding with a probable hospital stay of only 24 hours. The cost of ORT

would be $0.25 per liter for WHO oral dehydration

solution. The infant might require 3 liters ($0.75) of

the solution, but no sterile tubing or needles would

be needed. So the relevant cost of ORT for the infant

would be $0.75 for oral dehydration solution, plus $200

for 1 day of hospitalization.

The second hypothetical patient is a 16-year-old, 70

kg east African male who presents with severe cholera during a cholera outbreak. He arrives at the hospital in a severely dehydrated state and in shock.

Treated with intravenous fluids, this patient might

typically require 20 liters for treatment. This would

cost $30 in fluids alone ($1.50 per liter) plus at least

$2.00 for sterile tubing and needles. Thus, the total

cost for intravenous therapy for this patient would be

$32.00.

With ORT, this patient’s intravenous fluid requirement could be reduced to 3 liters ($4.50). The patient

would also require approximately 26 liters of oral dehydration solution (at $0.15 per liter in east Africa).

So the total cost of treating this patient with ORT

would be $3.90 for 26 liters of oral glucose/electrolyte solution, $6.50 for 3 liters of intravenous solution,

infusion tubing, and a sterile needle to treat shock; the

relevant costs total $10.40.

The comparative costs for purely intravenous treatment of a patient with severe cholera in shock, $32.00,

versus the use of ORT, $10.40, are notable. Many patients with somewhat less severe cholera can be treated

entirely with ORT alone, further reducing the costs.

Savings of this magnitude in therapy of diarrheal diseases are of critical importance to less developed countries where financial resources are severely limited in

comparison with health care needs.
Case Study A—Oral Dehydration Therapy for Diarrheal Diseases • 219

Conclusion

Diarrheal disease is one of the most important causes

of morbidity and mortality in infants and young children in the less developed world. Dehydration is the

most common precipitating cause of death in infant

diarrhea, and malnutrition is the most important

chronic consequence of such diarrhea.

Even during diarrheal infection, the intestine maintains its ability to absorb glucose and many amino

acids; as glucose is absorbed, sodium and water also

are absorbed. This observation has allowed the development of ORT—a highly efficacious form of therapy

for diarrheal dehydration that is technologically appropriate for use in less developed countries.

ORT, which involves oral dehydration with solutions containing glucose or sucrose and electrolytes,

now is in use worldwide, in programs sponsored by

international agencies and national governments. Although the basic breakthrough has been made, efforts

to improve the solutions and to increase the accessibility to ORT by those in greatest need are continuing.

Case Study A References

1.

2.

3.

4.

5.

6.

7.

8.

9.

Anonymous, “Oral Therapy for Acute Diarrhea,” Lancet 2:615-616, 1981.

Anonymous, “Management of Acute Diarrhea,”

Lancet 1:623-625, 1983.

Aperia, A., Broberger, O., Thodenius, K., et al.,

“Development of Renal Control of Salt and Fluid

Homeostasis During the First Year of Life, ” Acta.

Paediat. Scand. 64:393, 1975.

Aperia, A., Marin, L., Zetterstrom, R., et al.,

“Salt and Water Homeostasis During Oral Dehydration Therapy, ” J. Pediat. 103:364-369,

1983.

Barry, R. J. C., Smyth, D. H., and Wright, E. M.,

“Shortcircuit Current and Solute Transfer by Rat

Jejunum,” j. F%ysio]. 181:410-431, 1965.

Bart, K. J., and Finberg, L., “Single Solution for

Oral Therapy of Diarrhea,” Lancet 2:633-634,

1976.

Barzgar, M. A., Ourshano, S., and Nasser Amini,

J., “The Evaluation of the Effectiveness of Oral

Dehydration in Acute Diarrhoea of Children

Under Three Years of Age in West Azerbaijan,

Iran,” J. Trop. Pediat. 26:217-222, 1980.

Biehusen, F. C., and Schlegel, R. J., “Hypernatremia and the Use of Homemade Oral Electrolyte

Solutions,” A4iht. Med. 133:287-290, 1968.

Black, R. E., Brown, K. H., Becker, S., et al.,

“Longitudinal Studies of Infectious Diseases and

Physical Growth of Children in Rural Bangla10.

11.

12.

13.

14.

15<

16(

17.

18.

19,

20,

21,

22,

desh. 11. Incidence and Etiology of Diarrhea, ”

Am. J. Epidemiol. 115:315-324, 1982.

Black, R. E., Brown, K. H., and Becker, S., “Influence of Acute Diarrhea on the Growth Parameters of Children, ” Acute Diarrhea: Its Nutn”tional

Consequences, J. Bellanti (cd.) (New York: Raven

Press, 1983).

Black, R. E., Merson, M. H., Huq, I., et al., “Incidence and Severity of Rotavirus and Escherichia

coli Diarrhoea in Rural Bangladesh: Implications

for Vaccine Development, ” Lancet 1:141-143,

1981.

Black, R. E., Merson, M. H., Rahman, A. S. M. M.,

et al., “A Two Year Study of Bacterial, Viral and

Parasitic Agents Associated With Diarrhea in Rural Bangladesh,” -1. Infect. Dis. 142:660-664, 1970.

Black, R. E., Merson, M. H., Taylor, P. R., et al.,

“Glucose vs. Sucrose in Oral Dehydration Solutions for Infants and Young Children With Rotavirus-Associated Diarrhea, ” Pediatrics 67:79-83,

1981.

Blacklow, N. S., Dolin, R., Fedson, D. S., et al.,

“Acute Infectious Nonbacterial Gastroenteritis:

Etiology and Pathogenesis, ” Ann. Int. Med.

76:993-1008, 1972.

Bruck, E., Abal, G., and Aceto, T., “Therapy of

Infants With Hypertonic Dehydration Due to Diarrhea, ” Am. ]. Dis. Child. 115:281-301, 1968.

Bryant, J., Health and the Developing World

(Ithaca, NY: Cornell University Press, 1969).

Cash, R. A., Nalin D. R., Forrest, J. N., et al.,

“Rapid Correction of Acidosis and Dehydration

of Cholera With Oral Electrolyte and Glucose Solution,” Lancet 2:549-550, 1979.

Cash, R. A., Nalin, D. R., Rochat, R., et al., “A

Clinical Trial of Oral Therapy in a Rural CholeraTreatment Center, ” Am. J. Trop. Med. Hyg.

19:653-656, 1970.

Chatterjee, A., Mahalanabis, D., Jalan, K. N., et

al., “Evaluation of a Sucrose/Electrolyte Solution

for Oral Rehydration in Acute Infantile Diarrhea,” Lancet 1:1333-1335, 1977.

Chatterjee, A., Mahalanabis, D., Jalan, K. N., et

al., “Oral Dehydration in Infantile Diarrhea:

Controlled Trial of a Low Sodium Glucose/Electrolyte Solution, “ Arch. Dis. Child. 53:284-289,

1978.

Chatterjee, H. N., “Control of Vomiting in Cholera and Oral Replacement of Fluid, ” Lancet

2:1063, 1953.

Chen, L. C., Rahman, M., and Sarder, A. M.,

“Epidemiology and Causes of Death Among Children in a Rural Area of Bangladesh, ” Int. J.

Epidemiol. 9:25-33, 1980.
220 • Status of Biomedical Research and Related Technology for Tropical Diseases

23,

24,

25.

26.

27,

28,

28a.

29.

30.

31.

32.

33.

34.

35.

36.

Child-to-Child Programme, Newsletter 1, Institute of Child Health, 1979.

Chiriboga, E,, Tejada, L., Calderon, E., et al.,

“Rehidration con Sales en Nines Menores de Un

Anode Edad Deshidratados por Diarrea Aguda.

Estudio Doble Ciego con y sin Administration de

Agua Libref ” submitted for publication.

Church, M. A., “Fluids for the Sick Child,” Trop.

Doct. 2:119-121, 1972.

Cleary, T. G., Cleary, K. R., DuPont, H. L., et al.,

“The Relationship of Oral Dehydration Solution

to Hypernatremia in Infantile Diarrhea, ” J.

Pediat. 99:739-741, 1981.

Clements, M. L., Levine, M, M., Black, R. E., et

al., “Potassium Supplements for Oral Diarrhoea

Regimens, “ Lancet 2:854, 1980.

Clements, M. L., Levine, M. M., Cleaves, F., et

al., “Comparison of Simple Sugar/Salt Versus

Glucose/Electrolyte Oral Rehydration Solutions

in Infant Diarrhoea, ”J. Trop. Med. Hyg. 84:189-

194, 1981.

Committee on Nutrition, American Academy of

Pediatrics, “Oral Hydrating Solutions for Pediatric Use in the United States, ” report to the Food

and Drug Administration, U.S. Department of

Health and Human Services, typescript, June

1983.

Condon-Paoloni, D., Cravioto, J., Johnson, F. E.,

et al., “Morbidity and Growth of Infants and

Young Children in a Rural Mexican Village,” Am.

J. Publ. Hlth. 67:651-656, 1977.

Conteh, S., McRobbie, I., and Tomkins, A., “A

Comparison of Bottle Tops, Teaspoons and WHO

Glucose-Electrolyte Packets for Home Made Oral

Rehydration Solutions in the Gambia, ” Trans.

Roy, Soc. Trop. Med. Hyg. 76:783-785, 1982.

Curran, P. F., “Na, Cl, and Water Transport by

Rat Ileum in Vitro,” J. Gen. Physiol. 43:1137-

1148, 1960.

Cutting, W. A., “Rehydration Solutions and Domestic Measurements, ” Lancet 2:663-664, 1977.

Cutting, W. A.M., and Ellerbrock, T.V., “Homemade Oral Solutions for Diarrhea,” Lancet

1:198, 1981.

Darrow, D. C., Pratt, E. L., Flett, J., Jr., et al.,

“Disturbances of Water and Electrolytes in Infantile Diarrhea, ” Pediatrics 3:129-156, 1949.

Davidson, G. P., Gall, D. G., Petric, M., et al.,

“Human Rotavirus Enteritis Induced in Conventional Piglets, ” J. C]in. Invest. 60:1402-1409,

1977.

De, S., Chaudhuri, A., Dutta, P., et al., “Oral

Fluid Therapy for Cholera and Non-Choleraic Diarrhoeas in Children, ” J. c’om. Dis. 7:124-128,

1975.

37.

38.

39.

40.

41.

42.

43.

44.

45.

46.

47<

48,

49

50.

51.

52

53

Delvoye, P., Delogne-Desnoeck, J., Robyn, C,,

“Serum Prolactin in Long-Lasting Lactational

Amenorrhea,” L.ancet 2:288-289, 1976.

DeZoyza, 1., Kirkwood, B., Feachem, R., et al.,

“Preparation of Sugar-Salt Solutions, ” Trans.

Roy. Soc. Trop. Med. Hyg. 78:260-262, 1984.

Egemen, A., and Bertan, M., “A Study of Oral

Rehydration Therapy by Midwives in a Rural

Area Near Ankara,” Bull. W.H.O. 58:333-338,

1980.

Faust, R. G., Leadbetter, M. G., Plenge, R. K., et

al., “Active Sugar Transport by the Small Intestine, ” J. Gen. Physiol. 52:482-494, 1968.

Finberg, L., ‘The Management of the Critically

111 Child With Dehydration Secondary to Diarrhea,” Pediat. 45:1029-1036, 1970.

Finberg, L., “Hypematremic (Hypertonic) Dehydration in Infants, ’’l’V. Eng. J. Med. 289:197-198,

1973.

Finberg, L., “The Role of Oral Electrolyte-Glucose Solutions in Hydration for Children: International and Domestic Aspects, ”J. Pediat. 96:51-

54, 1980.

Finberg, L., Harper, P. A., Harrison, H. E., et al.,

“Oral Rehydration for Diarrhea,”J. Pediat. 101:

497-499, 1982.

Fisher, R. B., and Parsons, D. S., “Glucose Absorption From the Surviving Rat Small Intestine, ”

J. F’hysiol. 110:281-293, 1950.

Fordtran, J. S., “Stimulation of Active and Passive Sodium Absorption by Sugars in the Human

Jejunum,” J. Clin. Invest. 55:728-737, 1975.

Fordtran, J. S., Rector, F. C., Jr., and Carter,

N. W., “The Mechanisms of Sodium Absorption

in the Human Small Intestine, ” J. C’in. Invest.

47:884-900, 1968.

Gordon, J. E., Chitkara, I. D., and Wyon, J.B.

“Weanling Diarrhea,” Am. J. Med. Sci. 245:345-

377, 1963.

Gracey, M., Cullity, G. J., Suharjono, et al,, “Th~

Stomach in Malnutrition, ” Arch. Dis. Child.

52:325-327, 1977.

Hagihira, H., Lin, E. C. C., Samily, A. H., et al.,

“Active Transport of Lysine, Ornithine, Arginine

and Cystine by the Intestine, ” Biochem. Biophys.

Res. Comm. 4;478-481, 1961.

Harland, P. S. E. G,, Cox, D, L., Lyeu, M., et al.,

“Composition of Oral Solutions Prepared by

Jamaican Mothers for Treatment of Diarrhea,”

Lancet 1:600-601, 1981.

Harrison, H. E., “Symposium on Clinical Advances; Treatment of Diarrhea in Infancy, ”

Pediat. Clin. N. Amer. 1:335-348, 1954.

Hendrata, L., “Spoons for Making Glucose-Salt

Solutions,” Lancet 1:612, 1978,
Case Study A—Oral Dehydration Therapy for Diarrheal Diseases ● 221

54,

55

56.

57.

58.

59.

60.

61.

62.

63.

64.

65.

66.

67.

Hirschhorn, N., “The Treatment of Acute Diarrhea in Children: A Historical and Physiological

Perspective, ” Am. J. C]in. Nutr. 33:637-663,

1980.

Hirschhorn, N., Cash, R. A., Woodward, W. E.,

et al., “Oral Fluid Therapy of Apache Children

With Acute Infectious Diarrhea,” Lancet 2:15-

18, 1972.

Hirschhorn, N., Kinzie, J. L., Sachar, D. B., et al.,

“Decrease in Net Stool Output in Cholera During Intestinal Perfusion With Glucose-Containing

Solutions,” N. E’ng. J. Med. 279:176-181, 1968.

Hirschhorn, N., McCarthy, B.J., Ranney, B., et

al., “Ad Libitum Oral Glucose-Electrolyte Therapy for Acute Diarrhea in Apache Children, ” J.

Pediat. 83:562-571, 1973.

Hodges, R. G., McCorkle, L. P., Badger, G. F., et

al., “A Study of Illness in a Group of Cleveland

Families. XI. The Occurrence of Gastrointestinal

Symptoms,” Am. J. Hyg. 64:349-356, 1956.

Ingram, V. G., Rights, F. L., Khan, H. A., et al.,

“Diarrhea in Children of West Pakistan: Occurrence of Bacterial and Parasitic Agents, ” Am. J.

Trop. Med. Hyg. 15:743-750, 1966.

International Study Group, “A Positive Effect on

the Nutrition of Philippine Children of an Oral

Glucose-Electrolyte Solution Given at Home for

Treatment of Diarrhea. ”

International Study Group, “Beneficial Effects of

Oral Electrolyte-Sugar Solutions in the Treatment

of Children’s Diarrhoea, 1. Studies in Two Ambulatory Care Clinics,”J. Trop. Pediat. 27:62-67,

1981.

Jelliffe, D. B., “The Etiology of Diarrhea in Early

Childhood, ” J. Pediat. 68:792-793, 1966.

Jones, T. C., Dean, A. G., and Parker, G. W,,

“Seasonal Gastroenteritis and Malabsorption at

an American Military Base in the Philippines. II.

Malabsorption Following the Acute Illness, ” Am.

J. Epidemiol. 95:111-127, 1972.

Kahn, A., and Blum, D., “Hyperkalemia and

UNICEF Type Dehydration Solutions, ” Lancet

1:1082, 1980.

Kahn, A., Brachet, E., and Blum, D., “Controlled

Fall in Natremia and Risk of Seizures in Hypertonic Dehydration, ” Intensive Care Med. 5:27-

31, 1979.

Kielmann, A, A., and McCord, C., “Home Treatment of Childhood Diarrhoea in Punjab Villages, ” Environ. Child. Health Trop, Paediatr.

23:197-201, 1977.

Kooh, S. W., and Metcoff, J., “Physiologic Considerations in Fluid and Electrolyte Therapy With

Particular Reference to Diarrheal Dehydration in

Children, ” J. Pediat. 62:107-131, 1963.

68.

69.

70.

71

72

73.

74.

75<

76.

77.

78.

79,

80.

Koopman, J. S., Turkish, V. J., Monto, A. S., et

al., “Patterns and Etiology of Diarrhea in Three

Clinical Settings,” Am. J. Epidemiol. 119:114-123,

1984.

Levine, M. M., Clements, M. L., Black, R. E., et

al., “A Practical, Reliable Method for Preparing

Simple Sugar/Salt Oral Dehydration Solution, ”

J. Trop. Med. Hyg. 84:73-76, 1981.

Levine, M. M., Clements, M. L., Black, R. E., et

al., “Oral Dehydration With Simple Sugar/Salt

Solutions as an Alternative in Rural Areas When

Glucose/Electrolyte Solutions Are Unavailable,”

Acute Enteric Infections in Children, New Prospects for Treatment and Prevention. T. Holme,

J. Holmgren, M.H. Merson, and R. Mollby(eds. )

(Amsterdam: North Holland Biomedical Press,

1981).

Levine, M. M., and Edelman, R., “Acute Diarrheal Infections in Infants. 1. Epidemiology,

Treatment and Prospects for Immunoprophylaxis,” Hospital Practice 14:89-100, 1979.

Levine, M. M., Hughes, T. P., Black, R. E., et al.,

“Variability of Sodium and Sucrose Levels of

Simple Sugar/Salt Oral Dehydration Solutions

Prepared Under Optimal and Field Conditions, ”

J. Pediat. 97:324-327, 1980.

Levine, M. M., and Pizarro, D., “Advances in

Therapy of Diarrheal Dehydration: Oral Dehydration, ” Advances in Pediatrics 31, 1984.

Lindenbaum, J., “Malabsorption During and After Recovery From Acute Intestinal Infection, ” Br.

Med. J. 2:326-329, 1965.

Macauley, D., and Blackhall, M. I., “Hypernatremic Dehydration in Infantile Gastroenteritis, ” Arch. Dis. Child. 36:543-550, 1961.

MacLean, W. C., Jr., Klein, G. L., Lopez de

Romana, G., et al., “Transient Steatorrhea Following Episodes of Mild Diarrhea in Early Infancy,” J. Pediat. 92:562-565, 1978.

Mahalanabis, D., “In Search of a Super Oral Dehydration Solution: Can Optimum Use of Organic Solute Mediated Sodium Absorption Lead

to the Development of an Absorption Promoting Drug?” J. Diar. Dis. Res. 1:76-81, 1983.

Mahalanabis, D., Choudhuri, A, B,, Bagchi, N. G.,

et al., “Oral Fluid Therapy of Cholera Among

Bangladesh Refugees, ” Johns Hopkins Med. J.

132:197-205, 1975.

Mahalanabis, D., Sack, R. B., Jacobs, B., et al.,

“Use of an Oral Glucose-Electrolyte Solution in

the Treatment of Pediatric Cholera, ” J. Trop.

Pediat. Env. Child. Hlth. 20:82-87, 1974.

Martorell, R., Habicht, J-P., Yarbrough, C., et

al., “Acute Morbidity and Physical Growth in

Rural Guatemala Children,” Am. J. Dis, Child.
222 ● Status of Biomedical Research and Related Technology for Tropical Diseases

81

82.

83.

84.

85.

86.

87.

88.

89.

90.

91.

92.

93.

94.

129:1296-1301, 1975.

Mata, L. J., The Children of Santa Mm”a Cauque;

A Prospective Field Study of Health and Growth

(Cambridge, MA: MIT Press, 1978).

Mata, L. J., Kronmal, R. A., and Villegas, H., “Diarrhea Diseases: A Leading World Health Problem,” Cholera andl?elated Diarrheas, O. Ouchterlony and J. Holmgren (eds. ) (Basel: S. Karger,

1980).

Mata, L. J., Simhon, A., Padilla, R., et al., “Diarrhea Associated With Rotaviruses, Enterotoxigenic Escherichia coli, Campylobacter, an d

Other Agents in Costa Rican Children, 1976-

1981,” Am. J. Trop. Med. Hyg. 32:146-153, 1983.

McDermott, W., “Modern Medicine and the

Demographic-Disease Pattern of Overly Traditional Societies: A Technologic Misfit, ” J. Med.

Educat. 41 (Sept. suppl.): 137-162, 1966.

Melamed, A,, and Segall, M., “Spoons for Making Glucose/Salt Solutions, ” Lancet 1:1317-1318,

1978.

Menenghello, J., Rosselot, J., Aguilo, C., et al.,

“Infantile Diarrhea and Dehydration: Ambulatory Treatment in a Hydration Center, ” Advances in Pediatrics 11:183-208, 1960.

Moenginah, P. A., Supraptof Soenarto, J., et al.,

“Oral Sucrose Therapy for Diarrhea,” L.ancet

2:323, 1975,

Moffet, H. L., Schulenberger, H. K., and Burkholder, E. R., “Epidemiology and Etiology of Severe Infantile Diarrhea, ” J. Ped”at. 72:1-14, 1968.

Mona, A. M., Rahman, M., Sarker, S. A., et al.,

“Stool Electrolyte Content and Purging Rates in

Diarrhea Caused by Rotavirus, Enterotoxigenic

E. coli and V. cholerae in Children, ” J. Pediat.

98:835-838, 1981.

Mona, A. M., Sarker, S. A., Hossain, M., et al.,

“Rice-powder Electrolyte Solution as Oral Therapy in Diarrhoea Due to Vibrio cholerae and Escherichia coli, ” Lancet 1:1317-1319, 1982.

Morley, D., Pediatric Priorities in the Developing World (London: Butterworth, 1973).

Nalin, D. R., and Cash, R. A., “Oral or Nasogastric Maintenance Therapy for Diarrhoea of Unknown Aetiology Resembling Cholera,” Trans.

Roy. Soc. Trop. Med. Hyg. 64:769-771, 1970.

Nalin, D. R., and Cash, R. A., ‘The Optimal Oral

Therapy Formula for Cholera and Cholera-Like

Diarrheas, ” Proceedings of the Sixth Annual International Epidemiologic Association Scientific

Meeting (Belgrade: Savremena Administracya,

1971).

Nalin, D. R., and Cash, R. A., “Oral or Nasogastric Maintenance Therapy in Pediatric Cholera

Patients,” J. Pediat. 78:355-358, 1975.

95.

96.

97.

98.

99.

100,

101,

102.

103.

104.

105.

106

107,

108,

109,

Nalin, D. R., Cash, R. A., Islam, R,, et al., “Oral

Maintenance Therapy for Cholera in Adults, ”

Lancet 2:370-373, 1968.

Nalin, D. R., Cash, R. A., and Rahman, M., “Oral

(or Nasogastric) Maintenance Therapy for Cholera Patients in All Age-Groups, ” Bull. W.H.0.

43:361-363, 1970.

Nalin, D. R., Cash, R. A., Rahman, M., et al.,

“Effect of Glycine and Glucose on Sodium and

Water Absorption in Patients With Cholera,” Gut

11:768-772, 1970.

Nalin, D. R., Harland, E., Ramlal, E., et al.,

“Comparison of Low and High Sodium and Potassium Content in Oral Dehydration Solutions, ”

J. Pediat. 97:848-853, 1980.

Nalin, D. R., Levine, M. M., Mataf L., et al.,

“Comparison of Sucrose With Glucose in Oral

Therapy of Infant Diarrhea,” Lancet 2:277-279,

1978.

Nalin, D. R., Levine, M. M., Mata, M., et al.,

“Oral Dehydration and Maintenance of Children

With Rotavirus and Bacterial Diarrhoeas,” BuIZ.

W.H.O. 57:453-459, 1979.

Nichols, B. L., and Soriano, H. A., “A Critique

of Oral Therapy of Dehydration Due to Diarrhea]

Syndrome,” Am. J. C’h. IVutr. 30:1457-1472,

1977.

Palmer, D. L., Koster, F. T., Alan, A. K. M. J., et

al., “Nutritional Status: A Determinant of Severity of Diarrhea in Patients With Cholera, ” J. Infect. Dis. 134:8-14, 1976.

Palmer, D. L., Koster, F. T., Islam, A. F. M. R., et

al., “Comparison of Sucrose and Glucose in the

Oral Electrolyte Therapy of Cholera and Other

Severe Diarrheas, ” N. Eng. J. Med. 297:1107-

1110, 1977.

Patra, F. C., Mahalanabis, D., Jalan, K. L., et al.,

“In Search of a Super Solution: Controlled Trial

of Glycine-Glucose Oral Dehydration Solution in

Infantile Diarrhoea,’’Acta Paediat. Scand. 73:18-

21, 1984.

Patra, F. C., Mahalanabis, D., Jalan, K. N., et al.,

“Can Acetate Replace Bicarbonate in Oral Dehydration Solution for Infantile Diarrhea,” Arch.

Dis. Child. 57:625-637, 1982.

Patra, F. C., Mahalanabis, D., Jalan, K. N., et al.,

“Is Oral Rice Electrolyte Solution Superior to

Glucose Electrolyte Solution in Infantile Diarrhea,” Arch. Dis. Child. 57:910-912, 1982.

Phillips, R. A., “Water and Electrolyte Losses in

Cholera,” Fed. Proc. 23:705-712, 1964.

Phillips, R. A., “Cholera in the Perspective of

1966,” Ann. Int. Med. 65:922-930, 1966.

Pierce, N. F., Banwell, J. G., Mitra, R. C., et al.,

“Effect of Intragastric Glucose-Electrolyte Infu-
Case Study A—Oral Dehydration Therapy for Diarrhea/ Diseases ● 223

110,

111.

112.

113.

114.

115.

116.

117.

118.

119

120.

121.

122

sion Upon Water and Electrolyte Balance in

Asiatic Cholera,” Gastroenterology 55:333-343,

1968.

Pierce, N. F., Sack, R. B., Mitra, R. C., et al.,

“Replacement of Water and Electrolyte Losses in

Cholera by an Oral Glucose-Electrolyte Solution, ” Arm. ht. Med. 70:1173-1181, 1969.

Pierce, V., Ascoli, W., DeLeon, R., et al.,

“Studies of Diarrheal Disease in Central America. III. Specific Etiology of Endemic Diarrhea

and Dysentery in Guatemala Children, ” Am. J.

Trop. Med. Hyg. 11:395-400, 1962.

Pizarro, D., Chief, Emergency Room Service,

National Children’s Hospital, San Jose, Costa

Rica, personal communication, Nov. 15, 1983.

Pizarro, D., Posada, G., and Levine, M. M.,

“Hypernatremic Diarrheal Dehydration Treated

With ‘Slow’ (12 Hour) Oral Dehydration Therapy: A Preliminary Report, ” J. F’ediat. 104:316-

319, 1984.

Pizarro, D., Posada, G., Levine, M. M., et al.,

“Oral Dehydration of Infants With Acute Diarrhoeal Dehydration: A Practical Method,” J.

Trop. Med. Hyg. 83:241-245, 1980.

Pizarro, D. T., Posada, G., Levine, M. M., et al.,

“Tratamiento Oral de la Deshidratacion Hipernatremica, ” Acta. Med. Costa Rica 24:341-346,

1981.

Pizarro, D., Posada, G., and Mata, L., “Treatment of 242 Neonates With Dehydrating Diarrhea With an Oral Glucose-Electrolyte Solution, ”

J. Pediat. 102:153-156, 1983,

Pizarro, D., Posada, G., Mata, L., et al., “Oral

Dehydration of Neonates With Dehydrating Diarrheas,” Lancet 2:1209-1210, 1979.

Pizarro, D., Posada, G., Mohs, E., et al., “Evaluation of Oral Therapy for Infant Diarrhoea in an

Emergency Room Setting: The Acute Episode as

an Opportunity for Instructing Mothers in Home

Treatment,” BuIZ. W.H. O. 57:983-986, 1979.

Pizarro, D., Posada, G., Villavicencio, N., et al.,

“Oral Dehydration in Hypernatremic and Hyponatremic Diarrheal Dehydration, ” Am. J. Dis.

Child. 137:730-734, 1983.

Population Information Program, Oral Dehydration Therapy for Childhood Diarrhea, Population Reports, vol. 8, series L, No. 2, 1980.

Puffer, R, R., and Serrano, C. V., Patterns of

Mortality in Childhood, PAHO Scientific Publication No. 262 (Washington, DC: Pan American Health Organization, 1973).

Raghu, M. B., Deshpande, A., and Chintu, C.,

“Oral Dehydration for Diarrhoeal Diseases in

Children, ” Trans. Roy. Soc. Trop. Med. Hyg.

75:552-556, 1981.

123.

124.

125.

126.

127.

128.

129.

130.

131.

132,

133,

134,

1358

136

Rahaman, M. M., Aziz, K. M. S., Patwari, K., et

al., “Diarrhoeal Mortality in Two Bangladeshi

Villages With and Without Community-Based

Oral Dehydration Therapy,” Lancet 2:809-812,

1979.

Ramos-Alvarez, M., and Olarte, J., “Diarrheal

Diseases of Children, ” Am. J. Dis. Child.

107:218-231, 1964.

Ramos-Alvarez, M., and Sabin, A. B., “Enteropathogenic Viruses and Bacteria. Role in Summer

Diarrheal Diseases of Infancy and Early Childhood,” J. A.M.A. 167:147-156, 1958.

Ransome-Kuti, O., and Bamisaiye, A., “Oral

Therapy of Infant Diarrhea,” Lancet 2:471, 1978,

Riklis, E., and Quastel, J. H., “Effect of Cations

on Sugar Absorption by Isolated Surviving Guinea

Pig Intestine, ” Can. J. Biochem. Physiol. 36:347-

362, 1958,

Rohde, J. E., and Cash, R. A., “Transport of Glucose and Amino Acids in Human Jejunum During Asiatic Cholera, ”~. Infect. Dis, 127:190-192,

1973.

Rosenberg, I. H., Solomons, N., and Schneider,

R. E., “Malabsorption Associated With Diarrhea

and Intestinal Infections, ” Am. J. C/in. Nutr.

30:1248-1253, 1977.

Rowland, M. G. M., Cole, T. J., and Whitehead,

R. G., “A Quantitative Study Into the Role of Infection in Determining Nutritional Status in Gambian Village Children, ” Br. J. Nutr. 37:441-450,

1977.

Rowland, M. G., and Cole, T. J,, “The Effect of

Early Glucose-Electrolyte Therapy on Diarrhoea

and Growth in Rural Gambian Village Children, ”

J. Trop. Pediat. 26:54-57, 1980.

Roy, S. K., Rabbani, G. H., and Black, R. E.,

“Oral Dehydration Solution Safely Used in

Breast-Fed Children Without Additional Water, ”

J. Trop. Med. Hyg. 65, 1984.

Saberi, M. S., and Assaee, M., “Oral Dehydration of Diarrhoeal Dehydration: Comparison of

High and Low Sodium Concentration in Dehydration Solutions,” Acta Paediat. Scand. 72:167-

170, 1983.

Sack, D. A., Chowdhury, A. M. A. K., Eusof, A.,

et al., “Oral Hydration in Rotavirus Diarrhoea:

A Double Blind Comparison of Sucrose With

Glucose Electrolyte Solution, ” L.ancet 2:280-283,

1978.

Sack, D. A., Islam, S., Brown, K. H., et al., “Oral

Therapy in Children With Cholera: A Comparison of Sucrose and Glucose Electrolyte Solutions,” J. Pediat. 96:20-25, 1980.

Sack, R. B., Casells, T., Mitra, R., et al., “The

Use of Oral Replacement Solutions in the Treat-
224 ● Status of Biomedical Research and Related Technology for Tropical Diseases

ment of Cholera and Other Severe Diarrhoeal

Disorders,” Bull. W.H.O. 43:351-360, 1970.

137. Sack, R. B., Gorbach, S. L., and Banwell, J. G.,

“Enterotoxigenic Escherichia coli Isolated From

Patients With Severe Cholera-Like Disease,”J. infect. B“s. 123:378-385, 1971.

138. Santosham, M., Carrera, E., and Sack, R. B.,

“Oral Rehydration in Well-Nourished Ambulatory Children,“ Am. J. Trop. Med. Hyg. 32:804-

808, 1983.

139. Santosham, M., Daum, R. S., Dillman, L., et al.,

“Oral Rehydration Therapy of Infantile Diarrhea:

A Controlled Study of Well-Nourished Children

Hospitalized in the United States and Panama, ”

N. Eng. J, Med. 306:1070-1076, 1982.

140, Schedl, H. P., and Clifton, J. A., “Solute and

Water Absorption by the Human Intestine,” Nature 199:1264-1267, 1963.

141. Schultz, S. G., and Zalusky, R,, “Ion Transport

in Isolated Rabbit Ileum. II. The Interaction Between Active Sodium and Active Sugar Transport, ” J. Gen. Physiol. 47:1043-1059, 1964.

142. Schultz, S. G., and Zalusky, R., “Interactions Between Active Sodium Transport and Active

Amino-Acid Transport in Isolated Rabbit Ileum, ”

Nature 205:292-294, 1965.

143. Scrimshaw, M. S., Taylor, C. E., and Gordon,

J. E., Interactions of Nutrition and Infection,

WHO Monograph Series No. S7 (Geneva: World

Health Organization, 1968).

144. Sladen, G. E., and Dawson, A. M., “Interrelationships Between the Absorption of Glucose, Sodium and Water by the Normal Human Jejunum,” C]in. sci. 36:119-132, 1969.

145. Snyder, J. D., and Merson, M. H., “The Magnitude of the Global Problem of Acute Diarrhoeal

Disease: A Review of Active Surveillance Data, ”

Bull. W.H. O. 60:605-613, 1982.

146. Snyder, J. D., Yunus, M., Wahed, M. A., et al.,

“Home-Administered Oral Therapy for Diarrhoea: A Laboratory Study of Safety and Efficacy,” Trans. Roy. Soc. Trop. Med. Hyg.

76:329-333, 1982.

147. Spencer, R. P., “Intestinal Absorption of Amino

Acids,” Am. J. Clin. Nutr. 22:292-299, 1969.

148. Stoll, B. J., Glass, R. I., Huq, M. I., et al., “Surveillance of Patients Attending a Diarrhoeal Disease

Hospital in Bangladesh, ” Br. Med. J. 285:1185-

1188, 1982.

149, Tallet, S,, and MacKenzie, C,, “Clinical Laboratory and Epidemiologic Features of a Viral Gastroenteritis in Infants and Children, ” Pediatrics

60:217, 1977.

150, Taylor, P. R., Merson, M. H., Black, R. E., et al.,

“Oral Rehydration Therapy for Treatment of

Rotavirus Diarrhea in a Rural Treatment Centre

in Bangladesh, ” Arch. Dis. Child. 55:376-379,

1980.

1s1. Thier, S.0., Segal, S., Fox, M., et al., “Cystinuria: Defective Intestinal Transport of Dibasic

Amino Acids and Cystine, ” J. C’in. Invest.

44:442-448, 1965.

152. Tomkins, A., “Nutritional Status and Severity of

Diarrhoea Among Pre-School Children in Rural

Nigeria, “ Lancet 1:860-862, 1981.

153. Tripp, J. H., and Harries, J. T., “Oral Dehydration of Infants With Gastroenteritis, ” Adv. Biosci.

47:23-32, 1980.

154. Trowbridge, F. L., and Newton, L. H., “Seasonal

Changes in Malnutrition and Diarrheal Disease

Among Preschool Children in El Salvador, ” Am.

J. Trop. Med. Hyg. 28:135-141, 1979.

155. Van Heynigan, W. E., and Seal, J. R., Cholera—

The American Scientific Experience, 1947-1980

(Boulder, CO: Westview Press, 1983),

156. Walker, S. H., “Hypernatremia From Oral Electrolyte Solutions in Infantile Diarrhea, ” N. Eng.

J. Med. 304:1238, 1981.

157. Walker, S. H., Gahol, V. P., and Quintero, B. A.,

“Sodium and Water Content of Feedings for Use

in Infants With Diarrhea, ” Clin. Pediat. 20:199-

204, 1981.

158. Wallace, C. K., Cox, J. W., and Fabie, A., “The

Treatment of Cholera,”J. Phil. Med. Ass. 38:297,

1962.

159. Walshf J. A., and Warren, K. S., “Selective Primary Health Care: An Interim Strategy for Disease Control in Developing Countries, ” N. Eng.

J. Med. 301:967-974, 1979.

160. Watten, R. H., Morgan, F. M., Songkhla, V. N.,

et al., “Water and Electrolyte Studies in Cholera, ”

J. C’lin. Invest. 38:1879-1889, 1959.

161. Wegman, M. E., “Annual Summary of Vital

Statistics–1982, ” Pediatrics 72:755-765, 1983.

162. Wong, H. B., “Rice Water in Treatment of Infantile Gastroenteritis, ” Lancet 2:102-103, 1981.

163. Young, V. M., Lindberg, R. B., Ortiz, A., et al.,

“Studies of Infectious Agents in Infant Diarrhea.

III. Bacteria, Viral, and Parasitic Agents in Feces

of Puerto Rican Children, ” Am. J. Trop. Med.

Hyg. 2:380-388, 1962.

164. Yew, M. D., Melnick, J.L. Blattner, R. J., et al.,

“The Association of Viruses and Bacteria With

Infantile Diarrhea,” Am. J. Epidemiol. 92:33-39,

1970.

165. Yew, M. D., Melnick, J. L., Phillips, C. A., et al.,

“An Etiologic Investigation of Infantile Diarrhea

in Houston During 1962 and 1963, ” Am. J.

Epidemiol. 83:255-261, 1966.
Case Study B.—The Development of a Malaria Vaccine

Lydia Woods Schindler

Darnestown, MD

Introduction

The recognition that malaria stimulates natural immunity gave rise to the hope that a protective immune

response could be reproduced artificially. Experiments

in animals and humans have shown that this is indeed

feasible. Malaria vaccine research today is directed at

identifying the immunity-stimulating portions of the

parasite or its products, producing them in quantity,

and introducing them into the human body in such

a way that they stimulate immunity without causing

disease.

The undertaking is an ambitious one. A parasite

vaccine is difficult to engineer simply because a

parasite is so much larger and more complicated than

a virus or bacterium (targets of all familiar vaccines),

and carries a multiplicity of antigens. The problem is

complicated by the fact that antigens on the malaria

parasite vary according to the species of Plasmodium

involved and the stage of the parasite’s development.

Vaccines are now being developed against the various

types of malaria in all stages of the parasite’s life cycle, The ultimate vaccine will probably combine antigens to various stages. The main target species is P.

falciparum because it is so often lethal and the stakes

in preventing it are highest, though eventually a vaccine might protect against several species.

Although natural immunity to malaria develops

slowly, over a long period of time, and requires

repeated contacts with the parasite, vaccine researchers are encouraged by the prospect that an artificial

vaccine may be able to improve upon nature. By using

only the immunity-producing antigens—and not the

many proteins and contaminants carried by a whole

parasite, or in less pure vaccine preparations—they expect to be able to sidestep the multiplicity of immune

reactions that are triggered by the intact parasite, many

of which may actually favor the parasite’s survival.

Each of the three main life stages of the malaria parasite—the sporozoite, the merozoite, and the gamete—

has now been shown, under certain conditions, to

produce immunity in birds, rodents, and monkeys. Irradiated sporozoites, in an ingenious experimental system (mosquitoes do the inoculating) have also succeeded in immunizing a few human volunteers.

The farthest advanced work centers on the sporozoite, the infectious form transmitted to people by

mosquito bite. The immunogenic sporozoite antigen

has been isolated and characterized for several parasite

species. The gene that codes for the sporozoite antigen

of a monkey malaria has been identified, and the

antigen reproduced through genetic engineering. Moreover, because this antigen has proved to have a relatively simple structure, it has been possible to synthesize it, and the synthetic antigen has been used to

immunize rodents and monkeys. A dramatic announcement in the summer of 1984 revealed success in cloning and elucidating the structure of the antigenic sporozoite protein of the first human malaria, P. falciparum.

Although sporozoite research has captured the lead,

most malaria vaccine programs are concentrating on

other forms of the parasite. Progress has been impeded

by the fact that blood stage parasites carry many antigens, most of which do not elicit a protective immune

response. Moreover, blood stage parasite antigens

vary not only between stages and between species, but

also from one geographic strain to another. Once the

antigens are identified, the strategy for producing a

vaccine is similar to the strategy for sporozoites. One

set of blood stage antigens from a human malaria has

been cloned, and these antigens are being studied to

see if some of them are candidates for a vaccine.

A vaccine against gametes, sexual forms that occur

in the mosquito, would not prevent disease symptoms

but wouId block disease spread. Gamete antigens that

can elicit antibodies capable of preventing parasite fertilization have recently been identified and are under

study.

Several government and international organizations,

whose support has financed vaccine research, have

already met to make preliminary plans for field trials

of a sporozoite preparation; such trials are expected

to get under way by 1986 or 1987, although the details

have not yet been worked out.

The eventual large-scale production of a malaria

vaccine or vaccines, complicated by patent issues, and

the difficulties of delivering a vaccine to large numbers of people, many of whom inhabit remote and impoverished areas, will require thoughtful attention and

cooperation from the international community.

A first-generation sporozoite malaria vaccine is on

the threshold of becoming a reality. Scientists are convinced that they have at their fingertips the capability

of identifying and producing those elements of the ma225
226 ● Status of Biomedical Research and Related Technology for Tropical Diseases

laria parasite that evoke an immune response, and

public health planners anticipate the start of clinical

trials in 1986 or 1987.

This is not to say that work is complete. This first

“vaccine,” even if it proves safe and effective, would

probably constitute no more than one component of

the ultimate vaccine. Many formidable hurdles—

biological, immunologic, and chemical—are yet to be

met, and the logistical problems of field testing, mass

producing, and delivering a vaccine to target populations are enormous. Nevertheless, the prevailing mood

is one of great optimism.

The development of a malaria vaccine has been propelled by two distinct currents. One is the failure of

an international effort to eradicate malaria, due in

large part to the emergence of mosquitoes resistant to

pesticides and malaria parasites resistant to antimalarial drugs. The other is the recent explosion in biotechnology, which has provided scientists with tools

that have swept away obstacles that seemed insurmountable less than a decade ago.

A program to rid the world of malaria, which at the

time was estimated to affect 250 million persons annually, with 2.5 million deaths each year (5), was

launched by the World Health Organization (WHO)

in 1957. Through a combination of large-scale spraying of insecticides and medical treatment, the program

succeeded in eliminating malaria or greatly reducing

it in about 80 percent of the target areas by the mid1960s. Just a few years later, however, the picture had

begun to change dramatically. In Sri Lanka, where the

number of cases had shrunk from 3 million to only

18 reported cases, an epidemic of malaria broke out,

and more than 1 million people were affected (128).

Although malaria remained vanquished in temperate

regions of the world, the disease was making a strong

comeback in many tropical areas.

To some extent, the resurgence reflected the difficulties of carrying out such an ambitious and complex program in countries that lack a strong, central,

public health program (63,95,96). More fundamental,

however, was the fact that more and more malariacarrying Anophels mosquitoes were becoming resistant to DDT (dichloro-diphenyl-trichloroethane) and

other insecticides. By 1981, insecticide resistance had

appeared in 51 species of Anophels mosquitoes (96).

Because alternative insecticides, most of them petroleum products, are often too expensive for Third

World countries to use, spraying efforts have been curtailed (128).

The first reports that the malaria parasite could resist the effects of chloroquine appeared almost simultaneously in several countries of Southeast Asia and

South America around 1960. Resistant strains of P.

falciparum, the species that causes the most severe disease, have now surfaced in more than two dozen countries in Latin America, Asia, and Africa (128). New

drugs are being developed, but early reports indicate

that the parasite can become resistant to them, too.

In short, despite extensive research efforts, there is

little affordable on the shelf to fend off a disease for

which one-third of the world’s population is at risk,

which strikes an estimated 150 million persons a year,

and which causes at least 1 million deaths each year.

While efforts to conquer malaria through drugs and

insecticides have been faltering, research into the immunology of malaria, with a view to developing a vaccine that could prevent its symptoms and its spread,

has been surging ahead. One major breakthrough

came in 1976, when it first became possible to grow

P. falciparum in continuous culture, in the laboratory.

This ready source of raw material opened the way for

a stream of studies on the parasite and its ability to

stimulate immunity. The pace accelerated again with

the advent of the new biotechnologies. Since 1980, malaria researchers have been using monoclinal antibodies (MAbs) to identify those precise parts of the

parasite that produce an immune reaction. They then

produce this protective antigen in quantity through recombinant DNA technology, cloning those parasite

genes that code for the protective antigens. Alternatively, once the antigen structure has been fully

spelled out, scientists can synthesize the antigens chemically.

The Malaria Parasite

Malaria is caused by single-celled protozoan parasites of the genus Plasmodium. More than 100 different

species are known to cause disease in a variety of animals (24), Four species naturally infect humans: P.

falciparum, P. vivax, P. malariae, and P. ovale. Each

of the four has a distinctive appearance and life cycle;

each produces a somewhat different clinical effect (72).

The most dangerous is P. falciparum, which can cause

severe anemia, kidney failure, and brain damage; it

is often fatal, especially among children. In P. vivax

infection, the typical symptoms—cycles of chills, high

fever, and sweating with headache, muscle aches, and

nausea—are less severe. Although the disease is not

often fatal, relapses can occur periodically for up to

3 years. P. malariae infections can persist in the blood,

without producing symptoms, for life; chronic infections in children can lead to kidney damage.

Other species of Plasmodium cause malaria in a variety of vertebrates—reptiles, birds, rodents, rabbits

and monkeys. Several of these have been used as experimental models in vaccine research: P. gallinaceum
Case Study B—The Development of a Malaria Vaccine . 227

and P. lophurae in chickens and ducks; P. berghei, P.

yoelli, P. vinckei, and P. chabaudi in rats and mice;

and P. knowlesi in the rhesus monkey. In addition,

human malarias can infect certain higher apes and New

World monkeys. An important experimental model is

P. falciparum in the South American owl monkey.

All malaria parasites have a complex life cycle, alternating between vertebrate host and mosquitoes. In

vertebrates they reproduce asexually, first in the liver

and then, repeatedly, in the red blood cells (erythrocytes). In the mosquito, they reproduce sexually.

Human infection begins with the bite of an infected

female Anopheles mosquito. As she ingests a blood

meal to nourish her eggs, she simultaneously injects

a stream of saliva that can contain plasmodial sporozoites, which have been clustered in her salivary

glands. The motile, threadlike sporozoites quickly

leave the bloodstream and lodge in the cells of the liver

(hepatocytes). Within about an hour, all sporozoites

have disappeared from blood circulation.

Over the next week (P. falciparum and P. vivax)

or 2 weeks (P. malariae), each sporozoite that has invaded a liver cell becomes a schizont, a developmental structure that contains thousands of merozoites.

Liver stage parasites are known as “exoerythrocytic”

forms, to distinguish them from blood stage, or “erythrocytic, ” parasites. When the schizont is mature, it ruptures out of the infected liver cell and discharges

thousands of merozoites into the bloodstream. In P.

vivax and P. ovale malaria, some sporozoites become

hypnozoites, forms that remain dormant in the liver

for months or years before they start to proliferate

(62).

Merozoites, released into the bloodstream, invade

erythrocytes. The invasion process, which can be

observed by microscopy, takes about 20 seconds. P.

vivax and P. ovale parasites invade young erythrocytes, while P. malariae preferentially infect mature

erythrocytes; P. falciparum invades old and ‘young

cells alike—one reason why the concentration of parasites in the blood reaches dangerously high levels in

P. falciparum (82).

Most of the parasites that enter erythrocytes

undergo a second round of asexual reproduction, similar to but quicker and less prolific than that in the liver

cells. In 2 or 3 days, depending on the species, the intraerythrocytic parasite has developed from young

ring forms to trophozoites to the dividing form, again

known as a schizont. Red blood cells infected with P.

falciparum develop “knobs,” small, sticky, protrusions

of parasite origin that allow the infected erythrocyte

to adhere to the lining of small blood vessels while the

parasite matures (70,117). The infected cell is thereby

prevented from circulating through the spleen, where

it could be destroyed.

Depending on the species, each schizont contains 10

to 20 erythrocytic merozoites. When the schizont is

mature, these merozoites burst out of the erythrocytes

and invade yet other red blood cells, thus perpetuating

the cycle of infection. It is at this point, when the red

blood cells rupture, that clinical symptoms appear; because the cycle can repeat every 48 hours (P. vivax,

P. falciparum, and P. ovale) or 72 hours (P. malariae),

attacks of fever can occur every 2 or 3 days.

Plasmodial parasites thus continue to recycle untiI

they are brought under control through drug therapy

or through the host’s immune defenses, or until the

host dies. If reinfection does not occur, P. falciparum

infections will generally clear in 1 to 2 years; P. vivax

and P. mahriae may last 3 years. P. malariae, if untreated, can persist as an asymptomatic infection for

decades.

Some of the merozoites that invade red blood cells,

instead of developing asexually, differentiate into sexual forms, male and female gametocytes. Mature

gametocytes, enclosed within the erythrocyte membrane, circulate in the blood, available to feeding

Anopheles mosquitoes.

Blood ingested by the female Anopheles carries the

gametocytes into the mosquito’s stomach. There, perhaps triggered by changes in temperature and pH, they

shed the red blood cell envelope. Male gametocytes

rapidly transform into motile, spermlike structures and

fertilize the larger, egglike female gametes, forming

zygotes.

Forms that develop from zygotes in about a day,

called ookinetes, burrow into the mosquito’s stomach

wall, where they form oocysts; 9 to 14 days later the

oocysts rupture and release the motile, threadlike

sporozoites. The sporozoites infect the mosquito’s salivary gland. One cycle is complete.

The malaria parasite’s many life stages present a variety of possibilities for interrupting the infectious cycle by immunization. Potential targets include:

● sporozoites before they enter the liver cells;

. infected liver cells;

● merozoites before they enter the red blood cells;

• red blood cells carrying infectious schizonts; and

● gametes before fertilization occurs.

Each system has its advantages and disadvantages.

The ultimate vaccine is likely to combine antigens

against several stages of the parasite. A sporozoite

component would provide high immunogenicity; a

merozoite component would act as a backup, preventing disease should even one sporozoite escape the antisporozoite defenses. A gamete component, even

though it offers no direct benefit to the individual being vaccinated, would help to prevent disease spread

by interrupting the transmission of parasites from the

mosquito to a new vertebrate host.
228 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Malaria and Immunity

It is clear from the course of natural infections that

the Plasmodium parasite, in its various manifestations,

can stimulate immunity. Persons who live in areas

where malaria is endemic, and who are frequently exposed to infected mosquitoes, gradually develop some

immunity. But the development of immunity usually

takes repeated infections, over a period of years (82).

Vaccines are designed to mimic the natural process,

stimulating protection without producing any of the

adverse effects that accompany natural infection.

The Immune System

The immune system has evolved to protect an individual from invasion by “foreign” substances, including micro-organisms such as viruses, bacteria, and

parasites. Components of the immune system, including white blood cells called lymphocytes, recognize substances as being foreign by features of their

chemical makeup that are unlike “self. ” The chemical

entities recognized as such are called “antigens. ”

Once the immune system recognizes an antigen, it

can set in motion a variety of responses designed to

rid the body of the invader. One is the production of

antibody by a family of white blood cells known as

B-lymphocytes, or B-cells. Antibody is a substance

that “matches” the invading antigen and can inactivate

it or speed its uptake by scavenger cells. Another set

of responses involves T-lymphocytes, or T-cells. Some

subsets of T-lymphocytes work in collaboration with

B-lymphocytes, helping either to induce or suppress

the production of antibodies. Other T-lymphocytes

produce potent chemicals (one example is interferon)

that call into play yet other cells, and other responses.

Other types of immune cells, including “macrophages”

and “monocytes,” are scavengers equipped to take up

and digest foreign molecules and micro-organisms;

natural killer cells attack tumor cells and perhaps aid

in the elimination of parasites.

Some cells of the immune system become “memory”

cells. After the host’s initial encounter with a specific

antigen, the body’s defenses are primed to attack it

quickly: the individual acquires immunity.

Natural Immunity

P. falciparum typically takes its greatest toll among

children. Infants are protected temporarily by virtue

of antibodies they receive from their mothers in breast

milk (92). But by the second year of life, children who

live in highly endemic areas become victims of severe

and recurrent attacks. Those who survive gradually

acquire immunity; by 5 to 10 years of age they show

few or no further symptoms of disease. Immune adults

rarely experience acute attacks. However, such immunity is generally not complete. Even though the individual develops no symptoms, small numbers of parasites may continue to cycle through the red blood

cells. The combination of clinical immunity and continued low-grade infection, a condition known as

“premunition,” reflects a balance between parasite survival and host resistance. Importantly, the person with

asymptomatic low-grade parasitemia remains a reservoir for transmitting the disease.

If a person, once recovered, is not re-exposed to infection, immunity gradually wanes. Symptoms may

also flare up in otherwise immune individuals when

the immune system is disturbed by events such as surgery or pregnancy. Pregnant women in endemic areas,

especially those pregnant for the first time, are much

more likely than their nonpregnant counterparts to

contract acute P. falciparum malaria.

Innate Resistance

Some people inherit traits that make them naturally resistant to malaria. In general, these involve some

peculiarity of the red blood cell that makes it inhospitable to Plasmodium. Because they favor human

survival where malaria is endemic, these traits have

become prevalent in such areas.

One example is sickle-cell hemoglobin, which is

common in areas of west Africa where malaria is widespread. Under conditions of low oxygen tension—

which prevail in the small blood vessels where parasiteinfected red blood cells sequester—parasites within

cells containing sickle-cell hemoglobin die (33). Although children with sickle-cell hemoglobin develop

P. falciparum malaria, the disease is much less likely

to be fatal than in children with normal hemoglobin.

Another trait that confers resistance to malaria involves a receptor(s) on the surface of red blood cells;

the receptor is a prerequisite for parasite invasion. It

has long been known that most west Africans and

many American blacks are completely resistant to infection with P. vivax. These persons are also known

to be “Duffy blood-group negative”: their red blood

cells lack genetically determined surface markers

known as Duffy antigens A and B. Studies with parallel infections using P. knowlesi, a monkey infection

similar to P. vivax, indicate that these Duffy antigens

are closely associated with, if not identical to, the specific receptors that merozoites recognize. Cells lacking these receptors are unable to form a junction with

the parasite (74).
Case Study B—The Development of a Malaria Vaccine ● 229

Malaria Antigens

Malaria parasite antigens are remarkable for their

diversity. Each species and stage of the parasite carries its own characteristic surface structures that mark

it as immunologically distinct. Antigens of blood stage

parasites, but apparently not sporozoites (132), also

differ from one strain to another (67).

To offer protection, the immune system must tailor a response to fit each variation. A person who is

immune to P. falciparurn, for instance, can be susceptible to infection with P. vivax, and vice versa. Persons immune to P. falciparum in one country—or in

one part of a country—may become infected with a

different strain of falciparum when they travel. Even

within a given area, several strains may coexist; what

were once thought to be “relapses” may possibly represent a series of infections with different strains. This

phenomenon may also explain the slow development

of natural immunity, over the course of many infections (19,53).

In addition, some malaria parasites exhibit antigenic

variation. In response to changes in their environment

(for instance, the host’s deployment of effective immune defenses), the parasite changes surface antigens

(53,54).

Immune Responses

A malaria infection stimulates a spectrum of immune responses, not all of them beneficial. These include the production of antibody by B-cells and the

participation of various sets of T-cells, as well as the

activation of a variety of nonspecific responses, including macrophages, monocytes, and natural killer

cells.

Antibodies are clearly important to immunity

against malaria. Persons living in endemic areas develop increasing levels of serum antibodies to sporozoites as well as to blood stage parasites as they build

up immunity (80). However, much of the antibody

produced in response to malaria infection is nonspecific. Other antibodies, though specifically matched

to certain parasite antigens, are not protective.

Some antibodies, however, are both specific and

protective, and their role has been established in several ways. For one thing, immunity can be transferred

passively, by taking serum from immune individuals

or experimental animals and injecting it into nonimmune individuals. In a dramatic clinical demonstration, serum from immune adults living in west Africa

was given to 12 infants with severe malaria; it cured

their symptoms and sharply reduced the levels of asexual blood stage parasites, though the protection lasted

only a short time (17). However, it did not affect levels

of circulating gametocytes—an early indication that

the asexual and sexual stages carry different antigens.

It is also possible to induce protective antibodies by

vaccinating animals with antigens from defined stages

of the malaria parasite. Like immune serum, MAbs to

sporozoites, merozoites, and gametocytes can be used

to transfer passive immunity (97) and to inhibit the

growth of parasites in vitro (129).

Antibody appears to prevent sporozoites and

merozoites from entering their target cells; it can cause

them to agglutinate; it may coat them so they become

attractive targets for macrophages; it can prevent

gametes from forming zygotes.

T-cells are essential for the development of immunity in malaria (56,110). In addition to their major role,

which is assisting B-cells to produce antibody, they appear to secrete mediators that recruit and activate other

immune cells, including macrophages and monocytes.

They may possibly also exert a direct toxic effect on

parasites (36).

Several types of nonspecific mechanisms (that is,

those which do not depend on the recognition of a particular malaria antigen) can affect the malaria parasite.

For one, general potentiators of the immune system,

which carry no malaria antigens, can trigger general

cell-killing activity. Malaria-infected animals also have

increased numbers of macrophages in the spleen, liver,

and bone marrow. These activated macrophages are

probably responsible for the high levels of interferon,

a natural immunopotentiator, seen in such animals.

The macrophages can also secrete other soluble substances, monokines, that can trigger immune activity

(lo).

Natural killer cells are another nonspecific immune

defense. Although their known main target is tumor

cells, they can also attack virus-infected cells. Levels

of natural killer cells increase in people recovering from

P. falciparum malaria. Also, strains of mice that have

high levels of natural killer cells are more resistant to

plasmodial infection than mice with low levels, When

mice are immunized with irradiated sporozoites, levels

of both natural killer cells and interferon rise (91).

Adverse Immune Responses

Although the overall effect of the immune system’s

activity is to curb the parasite’s growth and gradually

eliminate it, some of the immune responses elicited by

plasmodial infections work to the host’s detriment. To

begin with, malaria leads to a general suppression of

the immune system, impairing the host’s ability to cope

with other, nonmalarial antigens, as well as the malaria infection itself. Children with malaria, as well as

animals infected experimentally, may be unable to

mount an effective response when they encounter an
230 . Status of Biomedical Research and Related Technology for Tropical Diseases

antigen for the first time. They may, for instance, respond poorly to vaccination against other diseases

such as typhus, meningitis, or measles. Children with

malaria are also prone to more, and more severe, viral diseases, including measles, respiratory tract infections, and gastroenteritis.

The immune response to malaria has been described

as “hyperactive and at the same time highly inefficient”

(126). While some aspects are suppressed, others are

overactive. Among the excess of antibodies produced

in response to malaria infection are autoantibodies

directed against a variety of the body’s own tissues.

Other antibodies may form potentially damaging immune complexes by combining with soluble parasite

antigens. (Soluble antigens are generally considered

detrimental to the host, a sort of decoy for the parasite. ) Immune complexes are believed to initiate the

kidney damage that occurs in malaria; the damage is

then thought to be sustained by autoantibodies.

A Malaria Vaccine

Early Vaccine Studies

The earliest reported attempt to vaccinate against

malaria dates back to 1910. Two Algerian brothers,

Etienne and Edmond Sergent, during the course of “a

lifetime of experimental work on malaria control by

every means” (41), inoculated birds with killed sporozoites (103).

These experiments came just three decades after

Alphonse Laveran, looking through his microscope at

blood smears from malaria patients, discovered “elements that seemed to me to be parasites” and identified

them as the principal cause of malaria. Only in 1898

did Ronald Ross put an end to theories incriminating

bad air (the Italians called it mal’ - aria) (41), decay,

or filth, by demonstrating that the source of malarial

infection was the mosquito.

Vaccination was not attempted again for several

decades. By that time a variety of studies had established that antiparasite antibodies existed, could be

detected, and could be used to transfer immunity

passively in monkeys (14).

During the 1930s and 1940s, it became clear that

vaccination was feasible. Birds, rodents, and eventually primates were immunized against malaria, using one

of two relatively accessible forms of the parasite, either sporozoites inoculated by feeding mosquitoes, or

the blood stages contained within infected erythrocytes. The usual strategy was to inactivate or kill the

parasites, inject them into an experimental animal,

wait for immunity to develop, and then challenge the

animals by exposing them to infection with virulent

parasites.

The first successful vaccine used P. gallinaceum

sporozoites to immunize fowls. Fifty percent of the

birds were able to survive a subsequent challenge (79).

In a series of experiments in the 1940s, Freund and

his colleagues studied killed blood stage parasites.

They used mature schizontsf derived from red blood

cells and inactivated by formalin, with and without

adjuvants, to immunize ducks (114). With adjuvants,

they succeeded in protecting rhesus monkeys from an

invariably fatal infection with P. knowlesi (32).

In 1946, Heidelberger attempted to vaccinate human

beings. Using formalin-killed blood stage P. vivax parasites, he inoculated a series of patients and volunteers.

The effort was not successful (44). According to current opinion, “perhaps the most remarkable result of

this trial was that eight injections of antigen given subcutaneously, intracutaneously, and intravenously over

a period of 2 weeks did not cause marked reactions

in the volunteers” (25).

During the 1950S, while dreams of eradicating malaria through vector control and drug therapy flourished, research on malaria vaccines was relatively

quiescent. Lacking any ready source of malaria parasites, and thus malaria antigens, researchers focused

on transferring immunity passively, by means of antibody-containing serum from immune individuals, or

to newborn animals through maternal milk. These

studies led, in the early 1960s, to the demonstration

that passive immunity could be effective in humans—

not preventing malaria infection but sharply reducing

the severity of the disease (17).

The pace did not pick up until the latter half of the

1960s. By that time, the success of the WHO malaria

eradication program was in doubt, and several laboratories began to make some headway in malaria immunology.

The U.S. Agency for International Development

(AID) began funding research toward a vaccine, an

effort that has been sustained and has been at least partially responsible for much progress in this area. The

first all-encompassing AID contract—to grow both

sporozoite and blood forms in culture, and to isolate

their protective antigens—was awarded to researchers at the University of Illinois in 1966; by 1972, the

program had expanded into a seven-site network and

was still growing (29).

Vaccination With Sporozoites

Most of the immune response to a natural infection

with malaria is elicited by blood stage parasite antigens. Sporozoites, however, are also highly immunogenic. Despite the facts that only a relatively small

number of sporozoites are inoculated by a mosquito

bite and that they spend a very brief time in the host’s
Case Study B—The Development of a Malaria Vaccine ● 231

bloodstream, sporozoites trigger antibody production

(81). Furthermore, experimental sporozoite vaccination—without the use of adjuvants—completely blocks

infection, but only for a few months.

For a sporozoite vaccine to be effective, it must kill

all sporozoites. If a single sporozoite escapes, it can

infect a liver cell and eventually give rise to up to

40,000 merozoites (in P. falciparum) that can then infect red blood cells, creating a fullblown attack of

malaria.

A drawback to sporozoite research has been supply.

Sporozoites live in the salivary gland of an infected

female mosquito. To produce sporozoites, mosquitoes

must be raised, then infected by feeding them

gametocytes. After the appropriate interval, laboratory workers must dissect the mosquitoes’ salivary

glands. Even though a single salivary gland teems with

sporozoites, dissection is a painstaking, labor-intensive

process that is obviously unsuited to large-scale production. Furthermore, the sporozoites must then be

purified of mosquito saliva and other contaminants.

The good news is that recombinant DNA technology promises to significantly ease sporozoite research.

The protective sporozoite antigen, which is distributed

uniformly over the surface of the sporozoite, consists

largely of a single antigen and short marker or epitope,

repeated several times. This antigen has now been

identified, using MAbs, and produced by recombinant

DNA technology. Moreover, the repeating epitope of

the P. knowlesi sporozoite has been synthesized, and

used to immunize monkeys (34).

Research in the 1960s and 1970s.—In the late 1960s,

Richards revived Mulligan’s studies of a quarter century earlier by showing that killed P. gallinaceum

sporozoites protected birds from challenge with

sporozoites, but not blood forms, of the malaria

parasite (98). Richards subsequently used the same

technique to protect mice against infection with P.

berghei and P. chabaudi (99).

Meanwhile, Nussenzweig and her coworkers at New

York University (NYU) had begun what would become

a major contribution to the development of a sporozoite

vaccine. NYU parasitologists had just succeeded in

growing the rodent parasite, P. berghei, through the

mosquito cycle. Taking advantage of this steady

supply of sporozoites, Nussenzweig showed that vaccination was effective. Repeated intravenous injections

of X-irradiated P. berghei sporozoites protected more

than 90 percent of the mice against an otherwise lethal challenge; immunity lasted about 2 months (87).

Subsequent studies elucidated other features of the

immune response to vaccination with intact sporozoites:

●

●

●

Sporozoites are more immunogenic if they are injected intravenously than if they are administered

intramuscularly, subcutaneously, or orally (109).

Not all sporozoites are immunologically equal:

only “mature” sporozoites obtained from mosquito salivary glands 17 to 18 days after the mosquito becomes infected are protective; younger

sporozoites collected from the mosquito stomach

wall are not effective because they lack an immunogenic surface antigen (121).

Antisporozoite immunity is species and stage spe- - .

cific. Mice immunized with P. berghei sporozoites

could be infected with erythrocytes (87) or schizonts (123). They were also susceptible to avian

and simian malarias (86). However, mice protected with one species of rodent malaria were

protected against all other rodent species tested

(89).

To get around the problem of injecting material that

was contaminated with a relatively large proportion

of mosquito salivary gland tissue, the NYU researchers turned to the mosquito for help. Infected female

Anopheles were exposed to enough X-irradiation to

render the parasites they were carrying noninfectious,

then they were allowed to bite experimental animals.

Mosquito inoculation met with most of the known criteria: mature sporozoites, inactivated by X-irradiation,

delivered intravenously, via multiple inoculations.

Mice repeatedly bitten by the infected, irradiated mosquitoes developed both circumsporozoite protein antibodies and protection against sporozoite challenge

(122).

The technique was soon applied to humans. For 84

days, three volunteers were exposed to a total of 397

mosquitoes that had been infected with P. falciparum

and then irradiated. On day 98, the volunteers were

challenged by exposure to heavily infected but nonirradiated mosquitoes. Two of the three men became

infected, but the third resisted infection. The third man

subsequently resisted challenge with other strains of

P. falciparum, but not P. vivax (12). It is thought that

the two volunteers who were not protected had probably been inoculated with too few sporozoites during

the 84-day immunization period.

The same technique was used to immunize another

volunteer in 1974 (100). The following year, a volunteer (the researcher himself) was protected sequentially against P. falciparum and P. vivax; the former

immunity lasted 3 months, the latter, 6 months (11).

Early attempts to immunize rhesus monkeys using

P. cynomolgi were not very successful (18,125). Results were better using multiple injections of large num-
232 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

bers of P. knowlesi sporozoites over a period of several months (39).

Recent Sporozoite Vaccine Research.—The search

for purified malaria antigens surged ahead when biotechnology made it possible to produce MAbs —that

is, antibodies that are secreted from clones of a single,

hybrid parent cell and which are thus identical, all

equipped to recognize and link to one single, specific

antigen. In the case of the malaria sporozoite, MAbs

were derived by fusing nonsecreting but long-lived cells

from a plasmacytoma (a plasma cell tumor) with antibody-secreting spleen cells from mice that had been

immunized with P. berghei sporozoites.

The resultant hybridoma secreted a MAb that singled out the immunogenic sporozoite antigen. This antigen, which has a molecular weight of 44,000 daltons

(a measure of mass), and is known as “Pb44” (129),

is distributed uniformly over the surface of mature

sporozoites, but it is not found on the other stages of

P. berghei (except for the very early stages within the

liver). Additional studies identified two more antigens,

Pb54 and Pb52 (130), which are recognized by the

same MAb; these have proved to be intracellular

precursors of Pb44.

The importance of this MAb in immune protection

has been demonstrated in several types of laboratory

tests (81,97,129). The antibody also prevents sporozoites from invading target liver cells in vitro (51). In

addition, the passive transfer of very small amounts

(10 Kg) completely protects mice against sporozoite

challenge. Similar MAbs were produced against P.

falciparum and P. vivax sporozoites (81) as well as

against P. knowlesi (13).

Work using these MAbs has identified one surface

antigen and one intracellular antigen on each of two

human malaria sporozoites, Pf58 and Pf67 in P.

falciparum, and PV45 and Pv5l in P. vivax. One surface antigen, Pk42, and two intracellular precursors,

Pk52 and Pk50, have been identified for P. knowlesi.

With the availability of genetic engineering, it became only a matter of time until a potentially protective sporozoite antigen was cloned. The first success

was reported in 1983 (27). Researchers at NYU extracted messenger RNA from mosquitoes infected with

P. knowlesi, converted the messenger RNA into complementary DNA, and inserted the complementary

DNA into a plasmid. The plasmid containing the genetic material from the P. knowlesi was then introduced into an Escherichia coli bacterium. Using MAbs

to identify the many proteins being produced by the

various E. coli colonies, they isolated three clones that

produced the sporozoite surface antigen.

Analysis of the DNA sequences in these clones

showed that they code for an epitope of 12 amino

acids, which is repeated 12 times in tandem (35). Because the shortest of the three DNA sequences cloned

contains nothing but the repeating unit, it was possible to deduce the amino acid sequence that the DNA

codes for. Then they corroborated their deduction by

chemically synthesizing it, and showing that it and the

native P. knowlesi circumsporozoite protein behave

identically (35), This peptide was then used to immunize rabbits and monkeys (34).

The announcement that the circumsporozoite protein gene of P. falciparum had been cloned, inevitable but still a cause for excitement, came in August

1984 (21). As predicted, a repeating sequence of nucleotides makes up a large portion of the molecule. The

surprise is that the repeat is shorter than the corresponding section in P. knowlesi, consisting of only

four amino acids, with some slight variation. The simplicity of this antigenic protein should be an advantage in the vaccine development work that now is proceeding.

Other avenues of research are yielding ingenious approaches to vaccine delivery and testing. In a twist of

genetic engineering, the gene encoding for the circumsporozoite protein of P. knowlesi has been incorporated into the genetic material of the vaccinia virus,

the agent of smallpox vaccine. When this recombinant

vaccinia virus is introduced into a cell, the infected cell

produces not only protective vaccinia proteins but also

the protective circumsporozoite protein.

The recombinant vaccinia virus system has already

been used to vaccinate animals against hepatitis, influenza, and herpes (108). Now rabbits have been vaccinated with a recombinant Plasmodium-vaccinia

virus, and have responded by developing antisporozoite antibodies (107). Because the vaccinia virus has

a large capacity for foreign DNA, it might eventually

be possible to incorporate genetic material for antigens

to several of the malaria parasite’s life stages (107),

A test to measure a sporozoite vaccine’s effectiveness—a pivotal concern of field testing—has evolved

from studies of the parasite’s cycle within the liver.

The new test, called the inhibition of sporozoite invasion assay, measures the ability of P. falciparum and

P. vivax sporozoites to invade cultured human

hepatocytes. Such invasion is blocked not only by

monoclinal antibodies to the circumsporozoite protein, but also by serum from immunized volunteers

and serum from persons living in endemic areas. Once

the test has been adapted for use in the field, it should

be possible to evaluate levels of preexisting immunity
Case Study B—The Development of a Malaria Vaccine ● 233

in a broad population, and to monitor the effects of

a sporozoite vaccine in clinical trials (51).

Vaccination With Blood Stage Parasites

As the form that induces disease as well as natural

immunity, blood stage parasites—either mature schizonts in red blood cells or free merozoites—are good

candidates for a vaccine, and blood stage antigens are

the object of most of the malaria vaccine research in

the world today. Although a blood stage vaccine

would not block either sporozoite or liver stage infection, those stages produce no symptoms. An effective

blood stage vaccine would prevent disease and interrupt further transmission.

Vaccines to blood stage parasites have succeeded in

producing immunity in several animal models, including P. knowlesi and P. fragile in the rhesus

monkey and the human malaria P. falciparum in the

Aotus monkey. However, the degree of protection depends on the nature of the antigen used and the way

it is administered. Moreover, it is typically necessary

to combine the parasite component with an adjuvant,

which independently and nonspecifically boosts the

immune response; most adjuvants used in animals are

unsuitable for use in humans because of adverse side

effects.

Another problem is that blood stage parasites are

antigenically very complex. In contrast to the sporozoite, which has a single immunodominant antigen

with a short repeating epitope, blood stage parasites

carry a mosaic of antigens, many of which elicit responses that are not protective. These antigens may

vary not only between stages and between species, but

also from strain to strain and even in the course of one

infection.

Evaluating the effectiveness of blood stage vaccines

is complicated by the use of experimental systems that

yield many different patterns of clinical immunity. The

same preparation—for instance, blood parasites attenuated by irradiation—will protect the rat, but give less

protection against more virulent infections in mice or

monkeys (15). Additionally, different programs have

used various doses and immunizing schedules, and different ways of measuring the outcome.

Malaria vaccine research took a tremendous leap

forward, when, in 1976, it became possible to grow

P. falciparum blood stage parasites in continuous

culture in the laboratory. Nevertheless, with current

techniques in vitro cultivation is not suitable for Iargescale production (24), nor has it eliminated problems

of contamination by cells in which the parasites are

grown.

Again, biotechnology holds out the promise of solutions. The major challenge now is to identify the protective antigens and produce them through cloning,

or possibly, chemical synthesis. Hybridomas are being used to produce MAbs, which are, in turn, being

used to isolate antigens that are protective.

Research in the 1960s and 1970s.—Attempts to induce immunity against the erythrocytic forms of malaria parasites have used a variety of approaches. In

some cases the animal (mouse, rhesus monkey, Aotus

monkey) was exposed to a severe or lethal infection

(transmitted by parasitized blood), then treated with

drugs or dietary manipulation; once cured, the animal was immune. Monkeys given subcurative drug

therapy, however, develop chronic recrudescent infections, and each recrudescence is associated with an antigenically different population of parasites (3). Such

antigenic variation may contribute to chronic infections and explain the slow development of natural immunity over the course of repeated infections (19,53).

In other immunization experiments, animals were

inoculated with parasitized erythrocytes that had in

some way been altered: attenuated (weakened by

growth in culture), heat-inactivated, killed, or combined with other adjuvants. Sometimes parasite fractions have been used. Such immunization can convert

lethal infections to chronic disease, with most animals

eventually recovering (P. berghei in mice and P.

falciparum in the Aotus monkey), or at least prevent

a portion of the animals from dying (P. knowlesi in

rhesus monkey) (4).

Many blood stage immunization studies have concentrated on the free-living merozoite. Immunity to

malaria is at least partly mediated by antibody, and

in vitro studies have shown that merozoites are the

target of this protective antibody; it prevents their entry into red blood cells (16).

Merozoites have proved to be an effective form of

vaccination. When P. knowlesi merozoites, combined

with Freund’s complete adjuvant were used to vaccinate rhesus monkeys, 100 percent of the animals survived this usually fatal infection (77, 78). Merozoite

vaccines, with and without adjuvants, have also succeeded in immunizing Aotus monkeys against P.

fa]ciparum infection, with 100 percent survival (78,

104). Merozoite vaccines have shown varying degrees

of effectiveness in birds, chickens, and rodents.

In Vitro Cultivation. —Until the mid-197&, merozoites for vaccine studies had to be separated out of

suspensions of schizont-infected red blood cells. Scientists had, for more than 60 years, been attempting to

develop a steady supply of blood stage parasites by

growing them in continuous culture. In 1912, two

malariologists, having cultivated the parasite through

three cycles (6 days), predicted that continuous culture

would be achieved within the year (71).
234 ● Status of Biomedical Research and Related Technology for Tropical Diseases

The approach that finally succeeded in 1976, capping

more than 30 years of research by Rockefeller University parasitologist William Trager. Trager, working

with James Jensen, devised an apparently simple system that resembles the normal physiological environment. P. falciparum blood stage parasites are grown

in human red blood cells in a culture medium that is

supplemented with normal human serum, in an atmosphere reduced in oxygen and enriched with carbon dioxide. As schizonts mature and rupture, extracellular

merozoites accumulate in the culture medium (115).

A similar system was devised independently by scientists at the Walter Reed Army Research Institute,

though the parasites recycled for only 21 days (43).

Because the system is easy to reproduce, it was soon

in widespread use and was improved upon. In addition to providing material for immunization experiments, the in vitro culture technique proved useful for

screening antimalarial drugs, for studying parasite-red

blood cell interactions, and for exploring the cellular

abnormalities underlying sickle cell anemia (116).

It also benefited research on vaccines against other

stages of P. falciparum, because with further manipulation, asexual blood forms could undergo development into gametocytes. These are proving useful not

only for gamete vaccine research, but also—when fed

to mosquitoes—as a means of producing sporozoites.

Recent Blood Stage Vaccine Research.-The search

for protective blood stage antigens—on the surface of

either the merozoite or the infected erythrocyte—has

been hindered by their tremendous complexity: P.

falciparum appears to carry about 40 antigens (47,93).

Blood stage antigens also change as the parasite develops and matures.

Monoclinal Antibodies.–MAbs made with hybridoma technology have been raised against the blood

stages of several species of malaria—P. yoelli (81), P.

knowlesi (23,28), P. falciparum (94), and P. berghei

(93,94)–but not against parasite antigens on the erythrocyte membrane (73).

Because most MAbs are produced by immunizing

mice with the entire schizont-infected erythrocyte, or

through mosquito-borne infection, and not by immunization with a pure parasite antigen, the clutch of

MAbs that results needs to be sorted out by laboratory screening. Then each MAb is tested for effectiveness: When mixed with merozoites in vitro, will it

cause the parasites to agglutinate, or otherwise prevent them from invading red blood cells? When injected into test animals, will it confer passive immunity? If a MAb passes these tests, its corresponding

antigen becomes a target for vaccine studies.

Purified Blood Stage Antigens. -Both immune

serum and MAbs have been used to identify potentially protective blood stage antigens. In general, such

antigens have proved to have relatively high molecular

weights, to be synthesized at a late stage in the schizont cycle, and to be processed into smaller, discrete

fragments (46).

One type of antibody target is antigens on the surface of the merozoite. Antibody to these antigens prevents merozoites from invading red blood cells by agglutinating the merozoites as they rupture from the

infected erythrocyte and also, perhaps, by blocking

specific receptors on the merozoites that allow them

to recognize and forma junction with the erythrocyte.

One merozoite surface antigen of the rodent malaria

parasite, P. yoelli, which has a molecular weight of

230,000 daltons, has been used to immunize mice; (47),

and a related MAb inhibited P. yoelli proliferation in

vivo (66). Comparable merozoite surface antigens have

been identified in P. knowlesi and P. falciparum

(22,28).

Numerous studies, using human serum and/or MAbs,

are examing the structure of these antigens and their

role in the immune response (2,93).

Second possible point of attack are antigens on the

surface of the erythrocyte. Antibody directed against

these antigens could destroy intraerythrocytic parasites in a number of ways. By coating the infected

erythrocyte, for example, antibody may make it attractive to macrophages. Alternatively, antibody

might imperil parasite survival by locking onto

“knobs,” thus preventing the infected erythrocyte from

sequestering in small blood vessels.

One erythrocyte-associated antigen is known as the

S-antigen. Although it stimulates antibody formation,

the S-antigen varies from one strain of P. falciparum

to another; many distinct types of S-antigen occur even

within a restricted geographical area. The S-antigen

is thus more likely to serve as a mechanism to help

the parasite evade host immune defenses than to help

the host destroy the parasite (19).

Evidence for another type of erythrocyte membrane

antigen comes from studies that show that antibody

(or other factors) can damage the P. falciparum parasite within infected erythrocytes, causing schizonts to

degenerate (94), or inhibiting intracellular growth

(57,111). Thus, there may be two (or more) groups

of antigens expressed on the parasitized erythrocyte

surface which elicit antibodies that can block normal

parasite metabolism. A purified erythrocyte membrane

antigen from P. know]esi infected cells has been successfully used to vaccinate rhesus monkeys (101).

Genetic Engineering. –In a novel attempt to study

blood stage proteins, Australian investigators manipulated the usual gene-cloning procedure. They extracted messenger RNA from the various blood stage
Case Study B–The Development of a Malaria Vaccine . 235

forms of P. falciparum, copied all of it into complementary DNA, and inserted the complementary

DNA into E’. coli. Then, instead of using a MAb to

isolate a single piece of DNA that codes for a particular protein, they used human immune serum containing many anti-P. falciparum antibodies to identify the

many clones manufacturing immunogenic proteins.

The result is a library of several hundred clones that

express a wide range of so-called “monoclinal antigens, ” each of which can be studied in detail. In their

ensemble, these clones probably represent a large portion of the antigens carried by P. falciparum (59).

However, it is not yet clear just what antigens they

include. The first of these clones to be examined

proved to code for a protein with a molecular weight

of 220,000 daltons. Although it contains repeated

epitopes, like the sporozoite surface protein, it is an

S-antigen that varies from strain to strain (19).

Several laboratories are working to streamline recombinant DNA production of malaria antigens. In

one approach, developed at the National Institutes of

Health (NIH), the genomic DNA itself (as distinct from

copies generated through messenger RNA and complementary DNA) is cut into fragments. These genecarrying fragments are cloned, and the clones screened

with MAbs to find which ones are producing the desired antigens (68).

Vaccination With Gametes

Like sporozoites and merozoites, male and female

gametes carry antigens that are capable of provoking

an immune response. Gamete immunization has prevented parasite fertilization in chicken, mice, and

monkey malarias, and blocked subsequent transmission of the disease.

The strategy for gamete vaccination follows a circuitous route. An individual is inoculated with gamete

antigens and makes antibodies to them. When a female

mosquito feeds on this person, she ingests not only

gametocytes, but also antibodies. In the stomach of

the mosquito, after the gametes emerge from the erythrocyte casings, they are exposed to these antibodies,

which quickly immobilize the male gametes and prevent fertilization.

Studies have shown that chickens immunized against

P. gallinaceum gametes produce antibodies that block

infectivity in mosquitoes (7,37). Although the chickens

were still susceptible to malaria infections, the mosquitoes that fed on those chickens developed no or few

oocysts. Subsequently, gamete vaccination against P.

yoelii infection in mice (69), as well as P. knowlesi in

the rhesus monkey, totally suppressed gamete infectivity in mosquitoes.

Gamete vaccine research moved ahead when, in

1981, it became possible to culture P. falciparum

gametocytes with regularity. Previously, blood stage

forms would only sometimes develop into gametocytes

in culture. When hypoxanthine, a substance found in

many body tissues, was added to the culture, gametocytes were found to predictably develop into mature

infectious parasites (55).

Two sets of target antigens have been identified on

P. falciparum gametes. The first is a set of three proteins with molecular weights of 250,000, 60,000, and

55,000 daltons. These antigens occur on both male and

female gametes, as well as newly formed zygotes, and

are shed shortly after fertilization. Antibodies to these

antigens block gamete fertilization and, in the presence of complement, destroy both gametes and zygotes. Unfortunately, these antigens, like some merozoite antigens, may vary within a species (8).

The second target of antigamete antibodies is a single

26,000-dalton protein which is synthesized by the zygote and expressed on the zygote surface. Antibody

to this antigen prevents zygotes from developing (8).

The gamete vaccine is known as an “altruistic” vaccine because it does not prevent infection or cure disease, or otherwise directly benefit the person being

vaccinated. Rather, it benefits the community by drying up the supply of infected parasites, preventing further malaria spread. As a result, it probably will be

used only if it is combined with a sporozoite and/or

merozoite vaccine. Moreover, its value in the field may

be difficult to prove. The effectiveness of a gamete,

or transmission-blocking, vaccine would depend on

how long immunity lasts, the proportion of the population that is immunized, and the intensity of transmission. In some parts of Africa, the rate of transmission is so high that a single infected individual can lead

to the infection of more than 500 others; in such an

area a gamete vaccine would hardly make a difference.

In other areas, such as India or Sri Lanka, where the

transmission is less intense, such a vaccine, possibly

combined with other control measures, would have

a chance of eliminating malaria (73).

Vaccination With Liver Stage Parasites

Any possibility of developing a liver stage vaccine

was impeded until recently by researchers’ inability to

grow intrahepatic parasites in culture. Although the

liver or exoerythrocytic stages of avian malarial parasites had been grown in continuous culture since 1966,

it was not until 1981 that mammalian malarial parasites were induced to grow and develop through a complete cycle, beginning with the entry of a sporozoite

into the target cell, through the parasite’s development
236 . Status of Biomedical Research and Related Technology for Tropical Diseases

into a liver schizont, complete with the release of

merozoites (49). Subsequently, the parasitic infection

was carried full cycle. The liver merozoites, injected

into mice, caused red blood cell infection, and some

of the blood stage parasites in the infected mice developed into gametocytes. Mosquitoes allowed to feed

on these mice developed sporozoites, which, when inoculated into the cell culture system, invaded the target cells and became liver schizonts.

The initial work was performed using P. berghei in

cultured human embryonic lung cells. More recently,

scientists have managed to grow both P. falciparum

and P. vivax (as well as P. berghei) in a cultured cell

line derived from a human liver cancer (48). This advance is particularly significant for P. vivax research,

since it has not yet been possible to grow P. vivax

blood stage parasites in continuous cultures.

Using the P. berghei system, it is possible to watch

as sporozoites encounter target cells, enter them, and

develop into trophozoites. The process of attachment

and entry, which seems to parallel in many ways the

invasion of merozoites into red blood cells, appears

to be effected by the circumsporozoite protein (Pb44);

conversely, MAbs to the circumsporozoite protein will

block sporozoite entry into liver cells (50).

The hepatoma culture system is also being used to

discover how some P. vivax liver stage parasites, instead of promptly developing into schizonts, lay dormant for long periods of time. When these dormant

forms, hypnozoites, later reactivate and develop into

schizonts that release merozoites, they produce a relapse. Electron microscopy is being used to document

the differentiation of liver parasites into hypnozoites

during the first few hours of development (48).

Scientists are currently working to isolate, purify,

and characterize the cell receptor that permits sporozoites to enter the liver cell. Once the nature of this

receptor is better understood, it may be possible to prevent infection by suppressing these receptors with

drugs. Alternatively, if parasite antigens can be detected on the surface of infected liver cells, it may be

possible to attack infected cells by linking antimalarial

drugs to antibodies that will recognize and join up with

these antigens.

The culture of liver stage plasmodia has yielded two

major spinoffs. One, the inhibition of sporozoite invasion assay described above, can be used to evaluate

the effectiveness of a sporozoite vaccine in the field.

The other is the adaptation of the liver parasitehepatoma culture system to test new antimalarial

drugs. As a fast and inexpensive alternative to testing

candidate antimalarial in costly and scarce primates,

the tissue culture system promises to revolutionize drug

development. Both the U.S. Department of Defense

(DOD) and WHO are exploring its use.

Funding Sources

U.S. Government, international, and philanthropic

institutions spent about $20 million in 1984 for malaria vaccine research. The biggest contributors are

AID and the National Institute of Allergy and Infectious Diseases (NIAID) of NIH. The next biggest are

the U,N. Development Program/World Bank/WHO

Special Program for Research and Training in Tropical Disease (TDR), DOD, the Centers for Disease Control (CDC), and the Rockefeller Foundation. In addition, a few pharmaceutical companies are conducting

malaria vaccine research.

In 1984, AID spent close to $8 million on the development of a malaria vaccine. The $8 million figure represents nearly a doubling of the Agency’s original commitment. In late 1983, sensing that the goal was within

reach, AID requested an increase in funds that would

bring its outlay for fiscal years 1983 to 1985 from $11.9

to $22.7 million.

Since launching the malaria vaccine program in

1966, AID has spent roughly $35 million, and expects

the program effort to cost an additional $15 to $25 million before a P. falciparum vaccine is ready for general use. By way of comparison, AID contributed more

than $1 billion (and other countries, an additional $4

to $5 billion) to the WHO malaria eradication campaign since its inception in the 1950s (29). AID funded

14 projects throughout the United States in 1984.

AID also provides a variety of support services. One

contractor, in addition to research, is charged with

testing and characterizing all materials injected into test

monkeys; another produces and supplies selected

strains of P. falciparum, as well as MAbs, to network

laboratories. Looking to the future, AID has hired experts to help contractor structure their research so that

it will answer Food and Drug Administration (FDA)

requirements, and other experts to counsel on matters

of patent rights (30). AID is currently taking the lead

in laying the groundwork for clinical trials.

NIAID sponsors both intramural and extramural research on malaria, with a total annual expenditure of

close to $4 million (120). Including basic research on

topics such as recombinant DNA technology or cell

receptors, which NIAID itself does not usually classify

as “vaccine research” (90), NIAID spends roughly $2

million for intramural research related to malaria immunology, either in the Laboratory of Parasitic Diseases or the Laboratory of Microbial Immunity. The

Institute awards an additional $2 million in grants (not

contracts) to about 20 institutions, primarily universities, throughout the United States.

Over the past 15 years, the focus of NIAID’s malaria research has shifted heavily in favor of immunology and vaccines, and funding commitments have
Case Study B—The Development of a Malaria Vaccine • 237

shown a steady growth. During the first 4 years of this

decade, when the tempo of research was rapidly accelerating, NIAID’s outlay doubled. However, it represents only a minute fraction of the NIH budget. In

1981, when the NIH was spending a total of $3.6 billion, NIAID’s share was $232 million or 6 percent; of

this, $27 million went to tropical disease research, and

about one-fifth of that was given to all aspects of

malaria.

Even though the payoffs may not always be so immediately visible as those of the closely managed,

product-oriented AID program, the type of steady

support provided by NIAID assures the continuous

growth of a broad expanse of knowledge. Beyond

achieving their own breakthroughs, or even beyond

satisfying intellectual curiosity, such studies generate

a rich resource for more intense development projects.

TDR is sponsored jointly by the U.N. Development

Program, the World Bank, and WHO. It is funded by

contributions from its cosponsors, as well as from the

governments of more than 25 countries and from several businesses and foundations.

From its beginning in 1976 through March 1983,

TDR had received more than $117 million in contributions: $15 million from the United States, $22 million from Denmark, and $14 million from Sweden,

plus $9 million from the U.N. Development Program,

close to $5 million from the World Bank, and $7 million from WHO (118). For the 2 years, 1982 and 1983,

TDR had budgeted just over $61 million.

Research on all aspects of malaria (chemotherapy

and field applications as well as malaria immunology)

accounts for about 30 percent of TDR’s research and

development budget. In 1981, this amounted to $3.2

million. Of this, $1.36 million went to support 40

projects on malaria immunology and vaccines (see

chapter 3). Of these, 24 were in the United States (119).

Others were in Great Britain, France, Switzerland, and

Australia.

Malaria vaccine research within DOD is focused on

preventing disease in American troops stationed

abroad. Thus, DOD’s malaria vaccine efforts give

special emphasis to sporozoite and liver stage vaccines,

which have the potential of preventing the initial infection and symptoms. AID or TDR efforts are also

interested in blood stage vaccines as a means of curbing symptoms and interrupting the parasite’s life cycle in areas where the disease is endemic and transmission is heavy.

Within DOD, the Army is the lead service in malaria research, and its work is headquartered at the

Walter Reed Army Institute of Research. Navy activities are carried out by the Naval Medical Research

Institute.

Although DOD vaccine studies extend back 15

years, studies began to accelerate around 1980, when

the new biotechnology opened the possibility of obtaining antigen in purified form. The Army has collaborated with NIH in the effort to clone and characterize a sporozoite antigen; it is also working toward

the chemical synthesis of the sporozoite’s antigenic

epitopes. The Navy has pioneered work on the liver

stages of the parasite.

The Army and Navy malaria vaccine groups, each

of which employs about 20 persons, work closely together. Expenditures for each of the groups has been

estimated to be close to $1 million a year (1,30,45).

CDC of the U.S. Public Health Service participates

in malaria vaccine research at several levels. Its primate

resource center contains more than 100 New World

monkeys of value for research on P. falciparum and

P. vivax; several aspects of malaria immunology are

the subjects of in-house research projects; and CDC

serves as a reference center for the many strains of P.

falciparum. It also serves as a source of materials; as

of early 1984, CDC has supplied researchers at NYU

with 40 million P. vivax sporozoites, and was developing millions more. CDC also runs field units on three

continents; these may provide appropriate sites for

clinical testing.

CDC’S annual expenditure on malaria in recent years

has been estimated to be under $1 million (30). CDC

allocates approximately $160,000 to $200,000 a year

in direct support of malaria vaccine development (6).

The Rockefeller Foundation, through its Great Neglected Diseases of Mankind program, spends just

under $500,000 a year to fund several projects that are

directed toward, but not restricted to, malaria vaccines. The three main projects, which have been

funded for each of the last 8 years, are located at Harvard, Oxford, and the Walter and Eliza Hall Institute

in Melbourne. The group at NYU has also received

grants-in-aid, and is currently receiving funds for work

on synthetic vaccine development (125a).

Four industrial organizations are involved in vaccine research. The Burroughs-Wellcome Co. of Great

Britain, which funnels profits into research efforts

through the Wellcome Trust, has a long-standing commitment to tropical disease research. Since 1979,

Wellcome scientists have been working on a malaria

vaccine, primarily the genetic engineering of blood

stage preparations. Its current outlay is approximately

$500,000 a year. In Australia, the recently launched

Australian Biotechnical Corp. is now investing an estimated $1 to $2 million annually on malaria vaccine

projects. Roche Pharmaceuticals, a leader in the antimalarial drug field, recently initiated a collaboration

with the Swiss company Biogen to develop a malaria
238 . Status of Biomedical Research and Related Technology for Tropical Diseases

vaccine, also through genetic engineering. Support is

reported to be in excess of $5 million.

The Future of Malaria

Vaccine Research

Scientists have at hand an “antigen preparation” for

one stage of the parasite (the sporozoite) for two species of human malaria (P. falciparum and P. vivax),

and they are making plans leading to clinical trials.

The path leading from there to the successful control

of malaria through vaccination is long and uncharted.

The challenges ahead include: developing a polyvalent

(multiple component) vaccine, demonstrating that it

is safe and effective, conducting large field trials in developing countries, producing the vaccines in quantity, and delivering it to the populations at risk.

Developing a Polyvalent Vaccine

Antigens.—An ideal vaccine would likely combat

multiple forms of the parasite—sporozoite, blood

stage, and/or gamete, and perhaps liver stage, as well

as two or more types of malaria—. falciparum and

P. vivax, perhaps with F’. malariae or P. ovale. Alternatively, different preparations might be prescribed for

different populations. A sporozoite vaccine, for instance, might be appropriate for persons whose exposure is limited, such as tourists, whereas a merozoite

vaccine could be given to control disease symptoms

in an area where malaria is highly endemic; a gamete

preparation could be part of a public health campaign

to eliminate the disease.

A protective sporozoite antigen for P. falciparum

and P. vivax should soon be ready for testing, but it

is likely to take another 2 years or more to isolate and

produce pure antigens from the blood stages or gametes.

Work on liver stage antigens is still preliminary.

To counter the problems of antigenic variation, researchers am exploring a variety of possibilities. These

include presenting a parasite structure that is not normally antigenic to the host (but which is common to

all of the strains of a species) in such way that it becomes antigenic. Alternatively, they might be able to

identify parasite surface structures that play such an

important role (for instance, those that enable merozoites to attach to or invade red blood cells) that they

should be the same in every strain.

Fortunately, the protective antigen from the sporozoite does not appear to vary from strain to strain

(132). For a blood stage or gamete antigen to be useful

for a vaccine, however, one must be found that is common to all parasites of a given species, or at least have

limited variability. Current experimental work indicates that although a large number of unique geographically defined antigens exist, other antigens are

common to many strains. Researchers are now working to discover such antigens, and determine how they

might be manipulated for immunization purposes.

Adjuvant.—All experimental malaria vaccine preparations except the sporozoite have required the use

of an adjuvant. Unfortunately, the adjuvant that has

been most successful in animal studies, Freund’s complete adjuvant, produces side effects that make it unacceptable for human use.

Several alternatives are being explored. A bacterial

derivative and another substance have both successfully replaced Freund’s complete adjuvant in immunizing Aotus monkeys against P. falciparum (105,106);

however, their effects in humans are not known. Parasite-specific MAbs have also worked as an adjuvant,

enhancing the effects of blood stage vaccination of

mice (42).

Antigenic Variation.—Multiple strains of P. falciparum, each with unique antigens, have evolved. Different strains are found not only in different parts of

the world, but within given geographic areas. Moreover, some parasite populations appear capable of

changing antigens over the course of an infection.

Demonstrating Safety and Efficacy

A likely scenario for the trial of a candidate vaccine begins with several months of testing for toxicity

and carcinogenicity in mice and rabbits. Next the vaccine would be tested in primates to see that the preparation produces no discernible adverse effects and that

it does stimulate a protective immune response. Assuming that all is going well, the investigators will file

an Investigational New Drug application with FDA.

The first clinical tests (which might begin as soon

as 1985, under ideal conditions) will use healthy male

volunteers, recruited either from a university setting,

the military, or industry, and including some of the

scientists themselves (29). Again, the first round would

be to make sure that the preparation produces no

adverse effects. A second round, lasting 18 months,

would be designed to answer questions of efficacy:

Does it stimulate antibody production? Are these antibodies protective? How long does protection last? In

the case of an AID-sponsored vaccine, these volunteer

studies would then be replicated in endemic areas,

using local volunteers and personnel (29).

Conducting Field Trials

The pilot vaccine will first be tested in healthy males,

then healthy nonpregnant females, and then in pregnant females, and children, In addition to the basic
Case Study B—The Development of a Malaria Vaccine ● 239

issues of safety and efficacy, new questions will arise

in endemic areas. Will persons whose immune responses have been dampened by previous malaria infections respond in the same way as volunteers who

have never had the disease? Will nutrition affect the

body’s ability to respond? What about antimalarial

drugs?

Neither sites for field trials nor strategies have been

arranged. AID, which will work closely with WHO

in setting up, and to some extent funding, clinical

trials, had convened two planning meetings by mid1984 with representatives from DOD, CDC, FDA,

NIH, and the Pan American Health Organization, as

well as WHO. In March 1985, clinical trials will be

the focus of a meeting of the scientific working group

of WHO’s Malaria Immunology program.

For the results of the trial to be meaningful, epidemiologists will need to have mapped patterns of malaria transmission in the test area, and documented the

extent of preexisting immunity. The trials will need to

be carefully planned and closely supervised, conducted

by skilled personnel working in close cooperation with

the test population. Those people who are vaccinated

will have to be closely monitored and treated when

necessary.

Producing a Vaccine

The research institutions in which vaccines are being developed are not geared to produce large quantities of vaccine material; that formidable task will be

the concern of pharmaceutical or genetic engineering

companies.

The ensuing interrelationships among scientists/

universities, research sponsors such as AID and WHO,

and industry lead to a tangle of conflicting interests.

Who “owns” the discovery? Who should make a profit

from it? What are the incentives for genetic engineering/pharmaceutical companies to get involved? Who

will buy a vaccine—AID, WHO, DOD, philanthropic

foundations?

The issues took shape in 1981, following the Supreme Court’s ruling that biological are patentable.

NYU filed patents for the Nussenzweig group’s work

(presumably involving MAbs used in identifying and

cloning the circumsporozoite protein). When NYU

entered into negotiations with a genetic engineering

firm, Genentech, to produce the circumsporozoite protein, Genentech asked for exclusive license to market

the vaccine.

WHO, which had long supported the NYU work,

and which represents many developing countries, held

fast to its contractual requirements for public access

to work that it supports. AID, another Nussenzweig

sponsor, holds patent rights in the United States under

Federal law; AID asked NYU to submit the requisite

“petition for greater rights.”

The conflict dissipated in 1983 when NYU and

Genentech dropped their plans to work together.

Genentech said it was too busy with other projects;

NYU developed superior genetic engineering capabilities of its own. The issues raised, however, remain

unanswered,

To date, seven patents have reportedly been applied

for in the United States by four different laboratories

(30). Four involve sporozoite vaccines, and three are

related to merozoite vaccines.

In the meantime, AID is in the process of revising

the patent language in its contracts. The new U.S. patent law, which took effect in 1981, allows grant recipients to take out patents on Government-sponsored

work, providing the Government is allowed royaltyfree use of the invention. AID’s goal is to make sure

that any agreements struck between its contractors and

industry will not impede a vaccine’s getting to the market, AID would also like the vaccine to be available

to Third WorId countries and to the U.S. miIitary on

a cost-plus basis (58).

AID has held discussions with six domestic companies possibly interested in producing a vaccine. At

this time it seems likely that vaccine development in

this country will proceed under the Orphan Drug program of FDA (30). Overseas companies, particularly

those that have established working relationships in

the developing countries where malaria is prevalent,

have also expressed some interest. Parke-Davis, the

sole U.S. drug company to be involved in malaria vaccine development in the past, and which worked

through the AID program, pulled out of the field when

the company was sold in the late 1970s,

Delivering a Vaccine

In order for a malaria vaccine to alleviate disease

and prevent death, it must reach the people who inhabit those parts of the tropics, often impoverished

and remote, where the disease is prevalent. To accomplish this, it will be necessary to raise funds, build an

excellent logistical support system, and train personnel. The success of such an effort will depend on close

collaboration among international organizations, industry, philanthropic foundations, and national health

systems.

The history of the human battle against malaria has

been marked by a series of overly optimistic expectations followed by disappointments (41), and the hopes

for a vaccine may prove no exception. However, the

flood of progress has been so strong, and the possibilities created by the new technologies so vast, that re-
240 . Status of Biomedical Research and Related Technology for Tropical Diseases

searchers are resorting to phrases like “the most incredible time in the history of malariology. ”

Conclusions

Having watched the most pressing problems of the

197&-antigen supply and purity-give way before

the wonders of genetic engineering and protein chemistry, scientists are confident that, with ingenuity, they

will be able to meet today’s challenges. Perhaps it will

be possible to boost an antigen’s immunogenicity by

presenting it to the host in a new way, or to prolong

immunity by developing slow-releasing antigens, or

to “vaccinate” people by incorporating a parasite gene

into bacteria that normally inhabit the gastrointestinal

tract. Louis Miller of NIAID likens the search for solutions to standing next to a wall: “Suddenly someone

puts a hole in it and beyond are vistas we’ve never imagined” (73).

The problems of testing, production, and delivery

are no less imposing, but again the outlook is optimistic, and planners are pressing ahead. Moreover,

the liaisons and lessons of the WHO malaria eradication campaign should stand the vaccine effort in good

stead.

Just what form the first vaccine will take, and where

and when it will appear cannot be predicted, but,

according to Miller, “there is no question that vaccines

will be developed against malaria; vaccines have been

successful in every animal model tested” (73).

The sentiment is a venerable one in malaria research.

In 1897, Ronald Ross, in an attempt to prove that the

mosquito was the source of human infection, faced the

prospect of dissecting thousands of mosquitoes. Undaunted, he wrote: “The things are there and must be

found. It is simply a matter of hard work” (41).

Not even its most enthusiastic proponents expect a

vaccine, of itself, to subdue malaria. To be successful,

a vaccine must be complemented by both improved

vector control and better drugs—and fresh, creative

approaches to both are being explored. These include

mosquito-killing bacteria, mosquito-devouring fish,

and mosquito-debilitating micro-organisms, on the one

hand, and a Chinese herbal remedy, on the other. A

three-pronged attack, combining vaccine(s), drugs,

and vector control, provides the best chance yet of bettering the lives of millions.

Case Study B References

1. Beaudoin, R. L., Naval Medical Research Institute, National Naval Medical Center, Bethesda,

MD, personal communication, June 1984.

2. Brownf G. V., Anders, R. F., Mitchell, G. F., et

al., “Target Antigens of Purified Human Immu3.

4.

5.

6.

7.

8.

9.

10.

11.

12.

13.

14.

15.

16.

noglobulin Which Inhibit Growth of PZasmodium fa]ciparum in Vitro,” Nature 297:591-593,

1982.

Brown, K. N., and Brown, I. N., “Immunity to

Malaria: Antigenic Variation in Chronic Infections to Plasmodium knowlesi, ” Nature 208:

1286-1288, 1965.

Brown, K. N., Brown, I. N., and Hills, L. A.,

“Immunity to Malaria. I. Protection Against

Plasmod”um Jmowksi Shown by Monkeys Sensitized With Drug-Suppressed Infections or by

Dead Parasites in Freund’s Adjutant, ” Exp.

ParasitoZ. 28:304-317, 1970.

Bruce-Chwatt, L., “Malaria: From Eradication

to Control, ” New Scientist, pp. 17-20, Apr. 19,

1984.

Campbell, C. C., Center for Infectious Diseases,

Centers for Disease Control, Atlanta, GA, personal communication, June 1984.

Carter, R., and Chen, D. H., “Malaria Transmission Blocked by Immunization With Gametes

of the Malaria Parasite, ” Nature 263:57-60,

1976.

Carter, R., Miller, L. H., Rener, J., et al., “Target Antigens of Transmission-Blocking Immunity, ” Proc. Roy. Soc., in press.

Chapin, G., and Wasserstrom, R., “Pesticide

Use and Malaria Resurgence in Central America and India, ” Soc. Sci. Med. 17:270-293, 1984.

Clark, I. A., Virelizier, J.-L., Carswell, E. A., et

al., “Possible Importance of MacrophageDerived Mediators in Acute Malaria, ” Infect.

Immun. 32:1058-1066, 1981.

Clyde, D. F., “Immunisation of Man Against Falciparum and Vivax Malaria by Use of Attenuated Sporozoites, ” Am. J. Trop. Med. Hyg.

24:397-401, 1975,

Clyde, D. F., Most, H., McCarthy, V., et al.,

“Immunization of Man Against SporozoiteInduced Falciparum Malaria, ” Am. J. Med. Sci.

266:166-177, 1973.

Cochrane, A. H., Santoro, F., Nussenzweig, V.,

et al., “Monoclinal Antibodies Identify the Protective Antigens of Sporozoites of Plasm odium

knowlesi,” Proc. Nat. Acad. Sci. (USA) 79:5651,

1982.

Coggeshall, L. T., and Kumm, H. W., “Effect of

Repeated Superinfection Upon Potency of Immune Serum of Monkeys Harboring Chronic

Infections of Plasmodium knowlesi, ” J. Exp.

Med. 68:17-27, 1938.

Cohen, S., “Progress in Malaria Vaccine Development, ” Br. Med. Bull. 38:161-165, 1982.

Cohen, S., Butcher, G. A., and Crandall, R. B.,
Case Study B—The Development of a Malaria Vaccine • 241

17.

18.

19.

20.

21.

22.

23.

24.

25.

26.

27,

28.

29.

“Action of Malarial Antibody in Vitro, ” Nature

223:368-371, 1969.

Cohen, S., McGregor, I. A., and Barrington,

S. C., “Gamma Globulin and Acquired Immunity to Human Malaria, ” Nature 192:733-737,

1961.

Collins, W. E., and Contacos, P. G., “Immunization of Monkeys Against Plasmodium

cynomoZgi by X-Irradiated Sporozoites, ” Nature

236:176-177, 1972.

Coppel, R. L., Cowman, A. F., Lingelbach, K. R.,

et al., “Isolate-Specific S-Antigen of PZasmodium falciparum Contains a Repeated Sequence

of Eleven Amino Acids, ” Nature 306:751-756,

1983.

Cross, G., Rockefeller University, New York,

personal communication, June 1984.

Dame, J. B., Williams, J. L., McCutchan, T. F.,

et al,, “Structure of the Gene Encoding the

Immunodominant Surface Antigen on the

Sporozoite of the Human Malaria Parasite Plasmodium fala”parum, ” Science 225(4662):593-

599, 1984.

David, P. H., Hadley, T. J., Aikawa, M., et al.,

“Processing of a Major Surface Glycoprotein

Occurs During the Ultimate Stages of Differentiation in Plasmodium knowlesi Malaria, ” Mol.

Biochem. Parasitol., in press.

Deans, J. A., Alderson, T,, Thomas, A. W., et

al., “Rat Monoclinal Antibodies Which Inhibit

the in Vitro Multiplication of Plasmodium

knowlesi,” Clin. Exp. Immune]. 49:297-309,

1982.

Deans, J. A., and Cohen, S., “Immunology of

Malaria, ” Am. Rev. Microbiol. 37:25-49, 1983.

Desowitz, R. S., and Miller, L. H., “A Perspective on Malaria Vaccines, ” Bull. W.H. O. 58:897-

908, 1980.

Elliott, V., “The Funding of Tropical Disease Research, ” preliminary report prepared for the Office of Technology Assessment, U.S. Congress,

Washington, DC, March 1984,

Ellis, J,, Ozaki, L. S., Gwadz, R. W,, et al.,

“Cloning and Expression in E. coli of the Malarial Sporozoite Surface Antigen Gene From Plasmodium knowlesi, ” Nature 302:536-538, 1983.

Epstein, N., Miller, L. H., Kaushal, D. C., et al.,

“Monoclinal Antibodies Against a Specific Surface Determinant on Malaria (Plasmodium

knowksi) Merozoites B1ock Erythrocyte Invasion, ” J. Immune]. 127:212-217, 1981.

Erickson, J. M., “Malaria Vaccine Development:

An Integrated Management and Research Appreach, ” presentation to National Council for

30.

31.

32.

33.

34.

35.

36.

37.

38.

39

40.

41.

42.

International Health, Department of State,

Washington, DC, Apr. 26, 1984.

Erickson, J. M,, U.S. Agency for International

Development, Washington, DC, personal communication, April 1984.

Freeman, R. F., Trejdosieqic, A. J., and Cross,

G., “Protective Monoclinal Antibodies Recognizing Stage-Specific Merozoite Antigens of Rodent Malaria Parasites, ” Nature 284:366-370,

1980.

Freund, J., Thomson, K. O., Sommer, H. E., et

al., “Immunization of Monkeys Against Malaria

by Means of Killed Parasites With Adjutants, ”

Am. J. Trop. Med. 28:1-22, 1948.

Friedman, M. J., Roth, E. F., Nagel, R. L., et al.,

“Plasmodium falciparum: Physiological Interactions With the Human Sickle Cell, ” Exp.

Parasitol. 47:73-80, 1979.

Glysin, J., Barnwell, J., Schlesinger, D. H., et al.,

“Neutralization of the Infectivity of Sporozoites

From Plasmodium knowlesi by Antibodies to a

Synthetic Peptide,” J. Exp. Med., in press.

Godson, G. N., Ellis, J., Svec, P., et al., “Identification and Chemical Synthesis of a Tandemly

Repeated Immunogenic Region of P/asmodium

knowlesi Circumsporozoite Protein, ” Nature

305:29-33, 1983.

Grun, J. L., and Weidanz, W. O., “Immunity to

Plasmodium chabaudi adami in the T-Cell Deficient Mouse, ” Nature 290:143-145, 1981.

Gwadz, R. W., “Malaria: Successful Immunization Against the Sexual Stages of Plasmodium

gallinaceum, ” Science 193:1150-1151, 1976.

Gwadz, R. W., and Green, I., “Malaria Immunization in Rhesus Monkeys: A Vaccine Effective Against Both the Sexual and Asexual Stages

of Plasmodium knowlesi, ” J. Exp. Med. 148:

1311-1323, 1978.

Gwadz, R. W., Cochrane, A. H., Nussenzweig,

V., et al., “Preliminary Studies on Vaccination

of Rhesus Monkeys With Irradiated Sporozoites

of Plasmodium knowlesi and Characterization

of Surface Antigens of These Parasites, ” Bu/Z.

W.H. O. 57( Suppl,l):165-173, 1979,

Hadley, T. J., David, P. H., McGinniss, M. H.,

et al., “Identification of an Erythrocyte Component Carrying the Duffy Blood Group Fy-a

Antigen, “ Sa”ence 223:597-599, 1984.

Harrison, G., Mosquitoes, MaZaria & Man: A

History of the Hostilities Since 1880 (FJew York:

E.P. Dutton, 1978).

Harte, P. G., Cooke, A., and Playfair, J. H, L.,

“Specific Monoclinal IgM Isa Potent Adjuvant

in Murine Malaria Vaccination, ” Nature 302:
242 . Status of Biomedical Research and Related Technology for Tropical Diseases

43.

44.

45.

46.

47.

48.

49.

50.

51

52.

53.

54.

55,

256-257, 1983.

Haynes, J. D., Diggs, C. L., Hines, F. A., et al.,

“Culture of Human Malaria Parasites Plasmodium falciparum, ” Nature 263:767-769, 1976.

Heidelberger, M., Prout, C., Hindle, J. A., et al.,

“Studies in Human Malaria. II. An Attempt at

Vaccination of Paretics Against Blood-Borne Infections With Plasmodium vivax,”J. Immunol.

53:109-112, 1946.

Hockmeyer, W. T., Walter Reed Army Institute

of Research, Walter Reed Army Medical Center, Washington, DC, personal communication,

May 1984.

Holder, A. A., and Freeman, R. R., “Immunization Against Blood-Stage Rodent Malaria Using

Purified Parasite Antigens, ” Nature 294:361-

364, 1981.

Holder, A. A., and Freeman, R. R., “Biosynthesis and Processing of a IYasmodium falciparum

Schizont Antigen Recognized by Immune Serum

and a Monoclinal Antibody, ” J. Exp. Med.

156:1528-1538, 1982.

Hollingdale, M. R., Bio-Medical Research Institute, Rockville, MD, personal communication,

May 1984.

Hollingdale, M, R., Leef, J. L., McCullough, M.,

et al., ‘In Vitro Cultivation of the Exoerythrocytic Stage of PZasmodium berghei From

Sporozoites, ” Science 213:1021-1022, 1981.

Hollingdale, M. R., Leland, P., Leef, J. L., et al.,

“Entry of Z%smodium berghei Sporozoites Into

Cultured Cells, and Their Transformation Into

Trophozoites,“Am. J. Trop. Med. Hyg. 32:685-

690, 1983.

Hollingdale, M. R,, Nardin, E. H., Tharavanig,

S., et al., “Inhibition of Entry of FYasmodium

falcipamm and P. vivax Sporozoites Into Cultured Cells; An in Vitro Assay of Protective Antibodies, ” J. ImmunoZ. 132:909-913, 1984.

Hommel, M., “Malaria: Immunity and Prospects for Vaccination, ” West. J. Med. 135:285-

299, 1981.

Hommel, M., David, P. H., and Oligino, L. D.,

“Surface Alterations of Erythrocytes in Plasmodium falciparum Malaria: Antigenic Variation,

Antigenic Diversity and the Role of the Spleen, ”

1. Exp. Med. 157:1137-1148, 1983.

Hommel, M., David, P. H., Oligino, L. D., et al.,

“Expression of Strain-Specific Surface Antigens

on Plasmodium fa/ciparum-Infected Erythrocytes,” Parasite Immunol. 4:409-414, 1982.

Ifediba, T., and Vandenberg, J. P., “Complete in

Vitro Maturation of Falciparum Gametocytes, ”

Nature 294:364-366, 1981.

56.

57.

58.

59.

60.

61.

62.

63.

64.

65.

66,

67,

68.

Jayawardena, A. N., Targett, G. A. T., Carter,

R. L., et al., “The Immunological Response of

CBA Mice to P. yoelii. I. General Characteristics, the Effects of T-Cell Deprivation and

Reconstitution With Thymus Grafts,” Immunology 32:849-59, 1977.

Jensen, J. B., Boland, M. T., and Akod, M., “Induction of Crisis Forms in Cultured Plasmodium

falcipamm With Human Immune Serum From

Sudan,” Science 216:1230-1233, 1982.

Jordan, D., American Institute for Biological

Sciences, Arlington, VA, personal communication, May 1984.

Kemp, D. J., Coppel, R. L., Cowman, A. F., et

al., “Expression of Plasmodium falciparum

Blood-Stage Antigens in Escherichia coli: Detection With Antibodies From Immune Humans, ”

Proc. Nat. Acad. Sci. (USA) 80:3787-3791,

1983.

Kilejian, A., “Histidine-Rich Protein as a Model

Malaria Vaccine, ” Science 201:922-924, 1978.

Kilejian, A., “Characterization of a Protein Correlated With the Production of Knob-Like Protrusions on Membranes of Erythrocytes Infected

With Plasmodium falciparum, ” Proc. Nat.

Acad. Sci. (USA) 76:4650-4653, 1979.

Krotoski, W.A., Garnham, P. C. C., Bray, R. S.,

et al., “Observations on Early and Late PostSporozoite Tissues in Primate Malaria. I. Discovery of a New Latent Form (Hypnozoite), and

Failure of the Immunofluorescence Technique

To Detect Hepatic Forms Within the First 24

Hours After Infection,” Am. J. Trop. Med. Hyg.

31:24-35, 1982.

Lancet, “Epitaph for Global Malaria Eradication?” Lancet 15-16, 1975.

Laveran, C. L. A., “A Newly Discovered Parasite in the Blood of Patients Suffering From Malaria: Parasitic Etiology of Attacks of Malaria

(Classics in Infectious Diseases),” Rev. Infect.

Dis. 4:908-911, 1982.

Lucas, A. O., “New Knowledge is Accumulating Fast” (Interview), WHO Chronicle 36:26-36,

1982.

Majarian, W. R., Daly, T. M., Weidanz, W. P.,

et al., “Passive Immunization Against Murine

Malaria With an IgG 3 Monoclinal Antibody,”

J. Immunol., in press.

McBride, J. S., Walliker, D., and Morgan, G.,

“Antigenic Diversity in the Human Malaria

parasite Plasmodium falciparum, ” Science 217:

254-257, 1982.

McCutchan, T. F., Hansen, J. L., Dame, J. B., et

al., “Mung Bean Nuclease Cleaves PZasmodium
Case Study B—The Development of a Malaria Vaccine Ž 243

69.

70!

71,

72,

73.

74.

75.

76.

77.

78.

79.

80.

81

Genomic DNA at Sites Before and After Genes,”

Science 225:625-628, 1984.

Mendis, K. N., and Targett, G. A. T., “Immunization Against Gametes and Asexual Erythrocytic Stages of a Rodent Malaria Parasite, ” Nature 277:389-391, 1979,

Miller, L. H., “Distribution of Mature Trophozoites

and Schizonts of Hasnmdiurn faZciparurn in the

Organs of the Aotus trivirgatus, the Night Monkey,” Am. J. Trop. Med. Hyg. 18:860-865, 1969.

Miller, L. H., “Current Prospects and Problems

for a Malaria Vaccine,”J. Infect. Dis. 135:855-

864, 1977.

Miller, L. H., “Malaria,” Tropical and Geographical Medicine, K.S. Warren and A.A.F.

Mahmoud (eds. ) (New York: McGraw-Hill,

1984).

Miller, L. H., National Institute of Allergy and

Infectious Diseases, National Institutes of Health,

Bethesda, MD, personal communication, April

1984.

Miller, L. H., Aikawa, M., and Dvorak, J. A.,

“Malaria (Plasmodium knowZesi) Merozoites:

Immunity and the Surface Coat, ” J. Immunol.

114:1237-1242, 1975.

Miller, L. H., David, P. H., and Hadley, T. J.,

“Perspectives for Malaria Vaccination, ” Proc.

Roy. SW., in press.

Miller, L. H., Aikawa, M., Johnson, J. G., et al.,

“Interaction Between Cytochalasin b-Treated

Malarial Parasites and Erythrocytes, ” J. Exp.

Med. 149:172-184, 1979.

Mitchell, G. H., Butcher, G. A., and Cohen, S.,

“A Merozoite Vaccine Against Plasmodium

knowlesi Malaria,” Nature 252:311-313, 1974.

Mitchell, G. H., Butcher, G. A., Richard, W. H. G.,

et al., “Merozoite Vaccination of Douroucouli

Monkeys Against Falciparum Malaria, ” Lancet

1:1335-1338, 1977.

Mulligan, H. W., Russell, P. F., and Mohan,

B. N., “Active Immunization of Fowls Against

P. galZinaceum by Injections of Killed Homologous Sporozoites, ” J. Malaria Inst. India 4:25-

34, 1941.

Nardin, E. H., Nussenzweig, R. S., McGregor,

I. A., et al., “Antibodies to Sporozoites: Their

Frequent Occurrence in Individuals Living in

Areas of Hyperendemic Malaria, ” Science 206:

597-599, 1979.

Nardin, E. H., Nussenzweig, V., Nussenzweig,

R. S., et al., “Circumsporozoite Proteins of Human Malaria Parasites PZasmodium falciparum

and Phsmodium vivax, ” J. Exp. Med. 156:20-

29, 1982.

82.

83.

84.

85.

86.

87.

88.

89.

90.

91.

92.

93.

94.

Neva, F. A., “Looking Back for a View of the

Future: Observations on Immunity to Induced

Malaria,” Am. J. Trop. Med. Hyg. 26:211-215,

1977.

Neva, F. A., National Institute of Allergy and

Infectious Diseases, National Institutes of

Health, Bethesda, MD, personal communication, May 1984.

Nussenzweig, R. S., “Progress in Malaria Vaccine Development: Characterization of Protective Antigens, ” Scand. J. Infect. Dis. (Suppl. )

36:40-45, 1982.

Nussenzweig, R. S., New York University Medical Center, New York, personal communication, May 1984.

Nussenzweig, R. S., Vandenberg, J., and Most,

H., “Protective Immunity Produced by the Injection of X-Irradiated Sporozoites of PZasmodium berghei. IV. Dose Response, Specificity

and Humoral Immunity, ” Mi/it. Med. 134:1176-

1182, 1969.

Nussenzweig, R. S., Vandenberg, J., Most, H.,

et al., “Protective Immunity Produced by the Injection of X-Irradiated Sporozoites of PZasmodium berghei, ” Nature 216:160-162, 1967.

Nussenzweig, R. S., Vandenberg, J., Most, H,,

et al., “Specificity of Protective Immunity Induced by PZasmodium bezghei Sporozoites,” Nature 222:488-489, 1969.

Nussenzweig, R. S., Vandenberg, J., Spitalny,

G., et al., “Sporozoite-Induced Immunity in

Mammalian Malaria: A Review,” Am. J. Trop.

Med. Hyg. 21;722-728, 1972.

Nutter, J., National Institute of Allergy and Infectious Diseases, National Institutes of Health,

Bethesda, MD, personal communication, May

1984.

Ojo-Amaize, E. A., Vilcek, J., Cochrane, A. H.,

et al., “Plasmodium berghei Sporozoites Are

Mitogenic for Murine T-Cells, Induce Interferon,

and Activate Natural Killer Cells, ” J. Immunol.

133:1-5, 1984.

Orjih, A. U., Cochrane, A. H., and Nussenzweig, R. S., “Active Immunization and Passive

Transfer of Resistance Against SporozoiteInduced Malaria in Infant Mice, ” Nature 291:

331-332, 1981.

Perrin, L. H., and Dayal, R., “Immunity to

Asexual Erythrocyte Stages of P. falciparum:

Role of Defined Antigens in the Humoral Response,” Immunol. Rev. 61:245-269, 1982.

Perrin, L. H., Ramirez, E., Lambert, P. H., et al.,

“Inhibition of P. faZciparum Growth in Human

Erythrocytes by Monoclinal Antibodies,” Na-
244 . Status of Biomedical Research and Related Technology for Tropical Diseases

95.

96.

97.

98.

99.

100

101.

102.

103.

104.

105.

106.

107.

ture 289:301-303, 1981.

Peters, W., “Current Concepts in Parasitology:

Malaria,” N. Eng. J. Med. 297:1261-1264, 1977.

Peters, W., “Panel on Malaria: Introduction, ”

Scand. J. Infect. D“s. 36 (%ppl. ):24-25f 1982.

Potocnjak, P., Yoshida, N., Nussenzweig, R. S.,

et al., “Monovalent Fragments (Fab) of Monoclinal Antibodies to a Sporozoite Surface Antigen (Pb44) Protect Mice Against Malaria Infection,” J. Exp. Med. 151:1504-1512, 1980.

Richards, W. H. G., “Active Immunisation of

Chicks Against PZasmodium gallinaceum by Inactivated Homologous Sporozoites and Erythrocytic Parasites, “Nature 212:1392-1394, 1966.

Richards, W. H. G., “Antigenic Studies of the

Class Sporozoa With Particular Reference to

Species of F’lasmod”um, ” Ph.D. thesis, University of London, 1969.

Rieckmann, K. H., Carson, P. E., Beaudoin,

R. L., et al., “Sporozoite Induced Immunity in

Man Against an Ethiopian Strain of F!!asmodium

[alciparum,” Trans. Roy. Soc. Trop. Med. Hyg.

68:258-259, 1974.

Schmidt-Ullrich, R., Lightholder, J., and Monroe, M. T. M., “Protective PZasmodium knowZesi

Mr 74,000 Antigen in Membranes of SchizontInfected Rhesus Erythrocytes, ” J. Exp. Med.

158:146-158, 1983.

Schwartz, A. L., Fridovich, S. E., Knowles, B. B.,

et al., “Characterization of the Asialoglycoprotein Receptor in a Continuous Hepatoma

Line,” J. Biol. Chem. 256:8878-8881, 1981.

Sergent, E., and Sergent E., “Sur I’immunite

clans le paludisme des oiseaux, Conservation in

vitro des sporozoites de P]asmodium rehctum.

Immunite relative obtenue par inoculation de ces

sporozoites, ” Comptes rendues de l’Academie

des Sciences, Paris 151:407-409, 191o.

Siddiqui, W. A., “An Effective Immunization of

Experimental Monkeys Against a Human Malaria Parasite, Plasmodium falciparum,” Science

197:388-389, 1977,

Siddiqui, W. A., Kan, S.-C., Kramer, K., et al.,

“Use of a Synthetic Adjuvant in an Effective

Vaccination of Monkeys Against Malaria, ” Nature 289;64-66, 1981.

Siddiqui, W. A., Taylor, D. W., Kan, S.-C., et

al., “Vaccination of Experimental Monkeys

Against Plasmo&”um faZciparum: A Possible

Safe Adjuvant,” Science 210:1237-1239, 1978.

Smith, G, L,, Godson, G. N., Nussenzweig, V.,

et al., “PZasmodium knoudesi Sporozoite Antigen: Expression by Infectious Recombinant Vaccinia Virus,” Science 224:397-399, 1984.

108.

109.

110.

111.

112.

113.

114.

115.

116.

117.

118.

119.

120.

Smith, G. L., Mackett, M., and Moss, B., “Infectious Vaccinia Virus Recombinant That Express Hepatitis B Surface Antigen, ” Nature

302:490-495, 1983.

Spitalny, G. L., and Nussenzweig, R. S., “Effect

of Various Routes of Immunization and Methods of Parasite Attenuation on the Development

of Protection Against Sporozoite Induced Rodent Malaria,” Proc. Helminth. Soc. Wash. 30

(Special Issue) :506-514, 1972.

Spitalny, G. L., Verhave, J. P., Meuswissen,

J. H. E. T., et al., “Plasmodium berghei: T-Cell

Dependence of Sporozoite-InducedImmunit y in

Rodents,” Exp. Parasitol. 42:73-81, 1977.

Stanley, H. A., and Reese, R. T., “In Vitro Inhibition of Intracellular Growth of Plasmodium

falciparum by Immune Sera, ” Am. J. Trop.

Med. Hyg. 33:12-16, 1984.

Tapchaisri, P., Chomcharan, Y., Poonthong,

C., et al., “Antisporozoite Antibodies Induced

by Natural Infection, ” Am. J. Trop. Med. Hyg.

32:1203-1208, 1983.

Targett, G. A. T., and Fulton, J. D., “Immunization of Rhesus Monkeys Against Plasmodium

knondesi Malaria,” Exp. Parasite]. 17:180-193,

1965.

Thomson, K. J., Freund, J., Sommer, H. E., et

al., “Immunization of Ducks Against Malaria

by Means of Killed Parasites With or Without

Adjutants,’’ Am. J. Trop. Md. Hyg. 27:70-105,

1947.

Trager, W., and Jensen, J. B., “Human Malaria

Parasites in Continuous Culture,” Science 193:

673-675, 1976.

Trager, W., and Jensen, J. B., “Cultivation of

Malarial Parasites,” Nature 273:621-622, 1978.

Udeinya, I. J., Miller, L. H., McGregor, I. A., et

al., “Plasmod”um falciparum Strain-Specific Antibody Blocks Binding of Infected Erythrocytes

to Amelanotic Melanoma Cells, ” Nature 303:

429-431, 1983.

U.N. Development Program/World Bank/

WHO, Special Programme for Research and

Training in Tropical Diseases, Sixth Programme

l?eport: 1 July 1981-31 December 1982 (Geneva:

WHO, 1983).

U.S. Agency for International Development, A

Profile of Selected Biomedical Research Efforts

into Diseases of Major Public Health Importance

to PeopZe of Developing Countries (Washington, DC: AID, 1982).

U.S. Department of Health and Human Services, National Institutes of Health, National

Institute of Allergy and Infectious Diseases,
Case Study B—The Development of a Malaria Vaccine ● 245

121.

122.

123.

124.

125.

125a.

NIAIDAwards and Projects, 1967-1983 (Washington, DC: DHHS, 1984).

Vandenberg, J. P., Nussenzweig, R. S., and Most,

H., “Protective Immunity Produced by the Injection of X-Irradiated Sporozoites of PZasrnodium berghei. V. In Vitro Effects of Immune Serum on Sporozoites, ” Milit. Med. 134:1183-

1190, 1969.

Vandenberg, J. P., Nussenzweig, R. S., and Most,

H., “Protective Immunity Produced by the Bite

of X-Irradiated Mosquitoes Infected With Plasmodium berghei, ” J. Parasit. 56:350-351, 1970.

Vandenberg, J. P., Nussenzweig, R. S., Sanabria,

Y., et al., “Stage Specificity of Anti-Sporozoite

Antibodies in Rodent Malaria and Its Relationship to Protective Immunity, ” Proc. HeZminth.

Soc. Wash. 39:514-525, 1972.

Vogel, R. J,, University of Arizona, Tucson, AZ,

personal communication, Jui-te 1984.

Ward, R. A., and Hayes, D. E., “Attempted Immunization of Rhesus Monkeys Against Cynomolgi Malaria With Irradiated Sporozoites, ”

Proc. Helminth. Soc. Wash. 39:525-529, 1972.

Warren, K. S., Rockefeller Foundation, New

York, personal communication, 1984.

126,

127.

128.

129.

130.

131.

132.

Wigzell, H., “Malaria Immunology, ” Scand. J.

Infect. Dis. 36(suppl. ):37-39, 1982.

World Health Organization, “Development of

Malaria Vaccines: Memorandum From a USAID/

WHO Meeting,” Bull. W.H. O. 61:81-92, 1983.

Wyler, D. J., “Malaria-Resurgence, Resistance,

and Research, ” iVew E’ng. J. Ikled. 303:875-878,

934-940, 1983.

Yoshida, N., Nussenzweig, R. S., Potocnjak, P.,

et al., “Hybridoma Produces Protective Antibodies Directed Against the Sporozoite Stage of

Malaria Parasite,” Science 207:71-73, 1980.

Yoshida, N., Potocnjak, P., Nussenzweig, V.,

et al., “The Protective Antigen of Sporozoites

of Plasmodium berghei, ” J. Exp. Med. 154:1225,

1236, 1981.

Zavala, F., Cochrane, A. H., Nardin, A. H., et

al., “Circumsporozoite Proteins of Malaria Parasites Contain a Single Immunodominant Region With Two or More Identical Epitomes, ” J.

Exp. Med. 157:1947-1957, 1983.

Zavala, R., Masuda, A., and Nussenzweig,

R. S., “Species-Specific Epitope Is Identical in

Phenotypically Different Circumsporozoite Proteins of Human Malaria, ” Fed. Proc. 43:1808

(Abst. ), 1984.
Appendixes
Appendix A.—Acknowledgments and

Health Program Advisory Committee

OTA would like to thank the members of the advisory panel and the contractors who provided material

for this assessment. In addition, OTA acknowledges the following individuals for their assistance in reviewing

drafts or furnishing information:

Nina Agabian

University of Washington

Anthony J. Allison

Syntex Research

Ronald Anthony

University of Maryland School of Medicine

Fausto Araujo

Palo Alto Medical Research Foundation

I. Asher

U.S. Agency for International Development

William Bancroft

Walter Reed Army Institute of Research

Paul F. Basch

Stanford University School of Medicine

William R. Beisel

U.S. Army

David Bishop

University of Alabama at Birmingham

A. Bloom

U.S. Agency for International Development

Barry R. Bloom

Albert Einstein College of Medicine

John C. Boothroyd

Stanford University School of Medicine

D. Boykin

National Institutes of Health

Thomas M. Buchanan

Pacific Medical Center (Seattle)

Gary H. Campbell

Centers for Disease Control

Craig Canfield

Walter Reed Army Institute of Research

Pat Carney

Naval Medical Research and Development Command

Clint Carter

Vanderbilt University

John Caulfield

Brigham and Women’s Hospital

S. D. Chaparas

National Institutes of Health

A. J. Clayton

Health and Welfare Canada

Gloria Coe

Pan American Health Organization

Gerald R. Cole

University of Maryland School of Medicine

Daniel G. Coney

Veterans Administration Medical Center (Nashville)

William E. Collins

Centers for Disease Control

Peter G. Contacos

International Medical Consultations, Inc.

Mary E. Corning

Consultant

George A. M. Cross

The Rockefeller University

Joel Dalrymple

U.S. Army

Gustave J. Dammin

Harvard Medical School

Thomas M. Daniels

University Hospitals (Cleveland)

Carter Diggs

Walter Reed Army Institute of Research

John E. Donelson

University of Iowa

Wilbur G. Downs

Yale School of Medicine

C. Draasbeck

Pan American Health Organization

Elizabeth Dragon

CODON Corp.

Herbert DuPont

University of Texas Medical School

James Dvorak

National Institutes of Health

Dennis Dwyer

National Institutes of Health

Henry Ehrlich

Cetus Corp.

James M. Erickson

U.S. Agency for International Development

Marion J. Finkel

U.S. Food and Drug Administration

249
250 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Jorge Flores

National Institutes

Arlene Fonaroff

National Institutes

Thomas P. Gillis

of Health

of Health

Marshall University School of Medicine

Mayer Goren

National Jewish Hospital (Denver)

M. Greene

National Academy of Sciences

M. Groves

U.S. Department of Defense

Richard L. Guerrant

University of Virginia School of Medicine

Donald Ham

Brigham and Women’s Hospital

Eugene Hayunga

Uniformed Services University of the Health Sciences

F. Herder

U.S. Agency for International Development

Wayne T. Hockmeyer

Walter Reed Army Institute of Research

Donald Hopkins

Centers for Disease Control

H. Hornbeak

National Institutes of Health

Tamotsu Imaeda

New Jersey Medical School

Silas Jackson

National Library of Medicine

Leon Jacobs

Gorgas Memorial Institute

Geoffrey M. Jeffrey

Consultant

J. Jennings

Pharmaceutical Manufacturers Association

William S. Jordan, Jr.

National Institutes of Health

Robert L. Kaiser

Centers for Disease Control

Paul Kaufman

Pharmaceutical Manufacturers Association

W. M. Kemp

Texas A&M University

Jay J. Kingham

Pharmaceutical Manufacturers Association

Charles Kirkpatrick

National Jewish Hospital (Denver)

Thomas Klei

Louisiana State University

Paul Knopf

Brown University

Dennis Knudson

Yale University School of Medicine

Dennis Kopecko

Walter Reed Army Institute of Research

Richard D. Kreutzer

Youngstown State University

Raymond Kuhn

Wake Forest University

David Lanar

National Institutes of Health

James W. LeDuc

U.S. Army Medical Research Institute of Infectious

Diseases

F. Luelmo

Pan American Health Organization

Austin J, MacInnis

University of California at Los Angeles

A. A. F. Mahmoud

University Hospitals (Cleveland)

Jerry Manning

University of California

Joseph Marr

University of Colorado Health Science Center

Lore McNicol

National Institutes of Health

Louis H. Miller

National Institutes of Health

Brian Murphy

National Institutes of Health

Neal Nathanson

University of Pennsylvania School of Medicine

Franklin Neva

National Institutes of Health

John Newland

Uniformed Services University of the Health Sciences

Nadia Nogueira

The Rockefeller University

Howard Noyes

Walter Reed Army Institute of Research

John E. Nutter

National Institutes of Health

Allison O’Brien

Uniformed Services University of the Health Sciences
App. A—Acknowledgments and Health Program Advisory Committee ● 251

Eric Ottesen

National Institutes of Health

John Parkhurst

National Institutes of Health

Curtis Patton

Yale University School of Medicine

Clifford A. Pease, Jr,

Consultant

C. J. Peters

U.S. Army Medical Research Institute of Infectious

Diseases

Elmer Pfefferkorn

Dartmouth Medical School

Mario Philipp

New England Biolab

S. Michael Phillips

University of Pennsylvania School of Medicine

Winy Piessens

Harvard School of Public Health

D. Praeger

The MacArthur Foundation

Diane Pratt

Brigham and Women’s Hospital

J. G. Randolph

Centers for Disease Control

J. Ristroph

U.S. Department of Defense

Donald H. Rubin

University of Pennsylvania School of Medicine

Phillip Russell

Walter Reed Army Institute of Research

Richard B. Sack

Baltimore City Hospital

David L. Sacks

National Institutes of Health

Dorothea L. Sawicki

Medical College of Ohio

A. Faye Schrater

Smith College

Philip Scott

National Institutes of Health

John Scovill

U.S. Army Medical Research and Development

Command

Alan Sher

National Institutes of Health

Larry Simpson

University of California at Los Angeles

Mette Strand

Johns Hopkins University School of Medicine

Ellen Strauss

California Institute of Technology

Kenneth Stuart

Issaquah Health Research Institute

Herbert B. Tanowitz

Albert Einstein College of Medicine

Grace M. Thorne

New England Medical Center Hospital

C. W. Todd

Harvard School of Public Health

Dennis Trent

Centers for Disease Control

J. Vorismarti

U.S. Department of Defense

Craig K. Wallace

Fogarty International Center

J. Warren

Pharmaceutical Manufacturers Association

R. H. Watten

Gorgas Memorial Laboratory

Thomas H. Weller

Harvard University School of Public Health

Karl A. Western

National Institutes of Health

Roy Widdus

Institute of Medicine

Phillip E. Winter

U.S. Army (retired)

Dyann Wirth

Harvard School of Public Health

Ming M. Wong

University of California at Davis

David J. Wyler

Tufts University School of Medicine

Martin D. Young

University of Florida College of Veterinary Medicine
252 ● Status of Biomedical Research and Related Technology for Tropical Diseases

HEALTH PROGRAM ADVISORY COMMITTEE

Sidney S. Lee, Committee Chair

President, Milbank Memorial Fund

New York, NY

H. David Banta

WHO Consultant and Director of

Project on Future Health Technologies

The Netherlands

Robert Evans

Professor

Department of Economics

University of British Columbia

Rashi Fein

Professor

Department of Social Medicine and Health Policy

Harvard Medical School

Boston, MA

Harvey V. Fineberg

Dean

School of Public Health

Harvard University

Boston, MA

Melvin Glasser*

Director

Health Security Action Council

Washington, DC

Patricia King

Professor

Georgetown Law Center

Washington, DC

Joyce C. Lashof

Dean

School of Public Health

University of California-Berkeley

Berkeley, CA

Alexander Leaf

Professor of Medicine

Harvard Medical School

Massachusetts General Hospital

Boston, MA

‘Until October 1983.

Frederick Mosteller

Professor and Chair

Department of Health Policy and Management

School of Public Health

Harvard University

Boston, MA

Norton Nelson

Professor

Department of Environmental Medicine

New York University Medical School

New York, NY

Robert Oseasohn

Associate Dean

University of Texas-San Antonio

San Antonio, TX

Nora Piore

Senior Fellow and Advisor to the President

United Hospital Fund of New York

New York, NY

Dorothy P. Rice

Regents Lecturer

Department of Social and Behavioral Sciences

School of Nursing

University of California-San Francisco

San Francisco, CA

Richard K. Riegelman

Associate Professor

George Washington University School of Medicine

Washington, DC

Walter L. Robb

Vice President and General Manager

Medical Systems Operations

General Electric Co.

Milwaukee, WI

Frederick C. Robbins

President

Institute of Medicine

Washington, DC

Rosemary Stevens

Professor

Department of History and Sociology of Science

University of Pennsylvania

Philadelphia, PA
Appendix B.— Method for Analysis of

Tropical Disease Research Funding

Description of Methodology

Much of the information included in OTA’S analysis of tropical disease research funding in chapter 3 was

taken from documents made available by the organizations that fund biomedical research in tropical diseases. These documents were supplemented, in many

cases, by discussions with relevant officials of the organizations. Some organizations assembled information

specifically for OTA’S analysis.

Limitations of the Data

One difficulty in developing information about

funding for research on tropical diseases is that the

researcher can never be sure that every relevant funding source has been identified. The sources of funding

described in OTA’S analysis may not include every

funding source, but do include the great majority of

organizations engaged in tropical disease research

funding in the United States.

A second difficulty in developing information about

tropical disease research funding is that organizations

vary in the definitions they use for biomedical research

activities. Because of the attention that has been given

to the six tropical diseases included in the Special Program for Research and Training in Tropical Diseases

(TDR), many organizations identify research efforts

in these diseases specifically. In some cases, however,

research activities that may be directly relevant to one

of these diseases maybe allocated to another research

type when data are reported.

TDR, for example, reports disease-specific information for malaria, schistosomiasis, trypanosomiasis,

leishmaniasis, filariasis, and leprosy, but has additional

separate categories for transdisease research. Transdisease research, such as investigations in vector biology and control, may or may not be directly concerned

with any one of the six TDR diseases. Thus, without

reviewing the research project proposals or protocols

for each transdisease investigation, a researcher may

not be able to determine how funds are allocated

among the six diseases. The figures for TDR in OTA’S

analysis exclude TDR projects funded under the transdisease categories, except in the sections of the analysis concerned with research objectives and recipients.

The basis of the classification scheme used to collect and store information about biomedical research

activities at the National Institute of Allergy and Infectious Diseases (NIAID) is the causative organism.

NIAID compiles data on research in the six diseases

of TDR for its own reporting purposes, and these data

are used in OTA’S analysis. However, NIAID research

on diarrheal diseases and acute respiratory infections

(ARIs) is identifiable only by causative agent, and for

the most part lies outside the areas identifiable as tropical disease research.

Information about the U.S. Agency for International

Development’s (AID) funding program is difficult to

break down beyond broad categories of expenditure,

because funds are allocated to categories which may

include several grants and contracts concerned with

similar objectives but with different timetables. Lists

of awards made through AID’s Small Grants Program

are available from the Office of the Science Advisor

and the National Research Council’s Board on Science and Technology for International Development

(BOSTID). However, these lists identify the date of

the award, but do not identify the funding level for

each award by year. For this reason, OTA used the

total amount of grant funds committed in the year in

which the grant was awarded.

The information about the U.S. Department of Defense (DOD) in chapter 3 was provided by officials

at the U.S. Army Medical Research and Development

Command, who were kind enough to compile the data

according to the study criteria. The officials reported

to OTA that DOD data are not routinely collected by

disease type, but rather by managerial funding category. The officials also pointed out that the precise

amount of funds for each category may vary as priorities are reassessed and changes made in allocations.

Data about the tropical disease research activities

of the Centers for Disease Control (CDC) were assembled for OTA by the Centers’ Financial Management

Office.

The Rockefeller Foundation and the Edna McConnell Clark Foundation publish a list of awards made

each year. By reviewing this list, OTA was able to allocate awards to the appropriate categories of interest.

Pharmaceutical companies do not make data available detailing their commitment to biomedical research. The information in OTA’S analysis about pharmaceutical companies, therefore, is not complete and

is largely anecdotal.

253
Appendix C.— Glossary of Acronyms and Terms

Glossary of Acronyms

AID —U.S. Agency for International

Development

AIDS —acquired immunodeficiency syndrome

BCG —Bacillus Calmette-Guerin (vaccine)

BOSTID —Board on Science and Technology for

BTI

CATT

CDC

CDD

cDNA

CF

CIE

COA

COPT

CRL

DDT

DEC

DHF

DHHS

DNA

DOD

DPT

ELISA

EPI

FCA

FDA

FIC

GMI

GML

GND

HI

International Development (NAS)

—Bacillus thuringiensis israeliensis

—Card agglutination test for

trypanosomiasis

–Centers for Disease Control (DHHS)

—Program for Control of Diarrheal

Diseases (WHO)

—complementary DNA

—complement fixation (test)

—counterimmunoelectrophoresis

—coagglutination (test)

—circumoval precipitin test

—Cholera Research Laboratory

(Pakistan)

—dichloro-diphenyl-trichloro-ethane

—diethylcarbamazine

—dengue hemorrhagic fever

—U.S. Department of Health and

Human Services

—deoxyribonucleic acid

—U.S. Department of Defense

—diphtheria/pertussis/tetanus (vaccine)

—enzyme-linked immunosorbent assay

—Expanded Program on Immunization

(WHO)

—Freund’s complete adjuvant

—Food and Drug Administration

(DHHS)

—Fogarty International Center for

Advanced Study in the Health

Sciences (NIH)

—Gorgas Memorial Institute of Tropical

and Preventive Medicine

—Gorgas Memorial Laboratory

–Great Neglected Diseases of Mankind

(program) (Rockefeller Foundation)

—hemagglutination inhibition (test)

ICDDR/B—International Center for Diarrheal

Disease Research, Bangladesh

ICMRT —International Center for Medical

Research (Calcutta)

IFA —indirect fluorescent antibody (test)

IgG

IgM

IHA

ILRAD

IMMAL

IND

INH

IOM

1PM

LA

LRTI

MAbs

mRNA

NAMRU

NAS

NIAID

NIH

NRC

NYU

ORT

PAHO

PAS

PMA

PPD

P/s

PSTC

RIA

RNA

RPHA

RSV

RVF

TDR

T/S

UNDP

UNICEF

URTI

VSG

WHO

WRAIR

—immunoglobulin G

—immunoglobulin M

—indirect hemagglutination assay

—International Laboratory for Research

on Animal Diseases

—Scientific Working Group on the

Immunology of Malaria (TDR)

—investigational new drug

—isoniazid

–Institute of Medicine (NAS)

—integrated pest management

–latex agglutination (test)

—lower respiratory tract infection

—monoclinal antibodies

—messenger RNA

—U.S. Naval Medical Research Unit

—National Academy of Sciences

—National Institute of Allergy and

Infectious Diseases (NIH)

–National Institutes of Health (DHHS)

—National Research Council

—New York University

—oral dehydration therapy

—Pan American Health Organization

—para-aminosalicylic acid

—Pharmaceutical Manufacturers

Association

—purified protein derivative

—pyrimethamine/sulfadoxine

—Program in Science and Technology

Cooperation (AID)

—radioimmunoassay

—ribonucleic acid

—reverse passive hemagglutination (test)

—respiratory syncytial virus

—Rift Valley fever

—Special Program for Research and

Training in Tropical Diseases (U.N.

Development Program/World

Bank/World Health Organization)

—trimethoprim/sulfamethoxazole

—U.N. Development Program

—U.N. International Children’s

Emergency Fund

—upper respiratory tract infection

—variant surface glycoprotein

—World Health Organization

—Walter Reed Army Institute of

Research

254
App. C—Glossary of Acronyms and Terms • 255

Glossary of Terms

Acquired immunity: Disease resistance in a person or

animal acquired after birth. Such immunity may be

active or passive.

Acquired immunodeficiency syndrome: See AIDS.

Active immunity: Disease resistance in a person or animal due to antibody production or other responses

of the immune system after exposure to a diseasecausing agent or a vaccine. (Compare passive immunity. )

Acute: Having a sudden onset, sharp rise, and short

course. (Compare chronic. )

Acute respiratory infections (ARIs): A group of infections of the respiratory tract caused by viruses, bacteria, or mycoplasma, or very rarely, by fungi,

protozoa, and the secondary effects of worms. Examples are pertussis, influenza, diphtheria, and

measles. For clinical purposes, ARIs are divided between the upper and lower respiratory tracts. Upper respiratory tract infections occur around the

teeth, gums, sinuses, throat, tonsils, epiglottis, middle ear, larynx, and trachea. Lower respiratory tract

infections, in the lungs and bronchi, include various types of pneumonia and bronchitis.

Adenovirus: One of a group of viruses that causes upper respiratory disease and pneumonia.

Adjuvant: A substance added to the main ingredient

of a prescription or solution to increase its effectiveness.

Aerosolized vaccine: A vaccine administered through

the nose and mouth by inhaling a vapor, rather than

through injection or ingestion.

African sleeping sickness: Infection by Trypanosoma

brucei gambiense (chronic form found in west

Africa) or by T. b. rhodesiense (acute form found

in east Africa). Also called “African trypanosomiasis.” (See also trypanosomiasis. )

African trypanosomiasis: See African sleeping sickness.

Agglutination: The process by which cells or particles

adhere and form visible clumps, a process which

is exploited in many diagnostic tests.

AIDS (acquired immunodeficiency syndrome): A disorder characterized by an acquired (not inherited)

deficiency of the immune system probably caused

by a retrovirus known as HTLV-111, which is complicated by a rare type of cancer (Kaposi’s sarcoma)

or infections caused by micro-organisms that usually do not produce infections in healthy individuals.

Amebiasis: Infection of the colon with amebae.

American trypanosomiasis: See Chagas’ disease.

Animal reservoir: See reservoir, host.

Antibiotic: A chemical substance produced by a microorganism that is administered to fight infections,

usually bacterial infections, in humans or animals.

Examples are pencillin, tetracycline, and erythromycin.

Antibody: A serum protein (immunoglobulin) molecule produced by white blood cells in response to

exposure to a specific antigen, and characterized by

specific reactivity with that antigen. At present, five

classes of antibodies are distinguishable. Most of

the antibodies that circulate in the blood are immunoglobulin G (IgG); the others are IgM, IgA,

IgD, and IgE. Antibodies are responsible for “humoral immunity. ”

Antigen: A substance, usually a protein or complex

carbohydrate, which, when introduced into the

body of a human or other animal, stimulates the

production of an antibody that reacts specifically

with it.

Antigen probe: A sequence of DNA that is used to detect the presence of a particular nucleotide base sequence.

Antimalarial: A drug, such as chloroquine, that prevents or suppresses malaria infection.

Antiserum: Blood serum containing antibodies to a

specific antigen.

Arboviral infections: A group of infections caused by

arboviruses. Important human arboviral infections

include yellow fever, dengue fever, oropouche fever, chickungunya, Japanese encephalitis, other viral encephalitides, and hemorrhagic fevers.

Arbovirus: One of a large number of viruses transmitted by or thought to be transmitted by arthropods (mosquitoes, ticks, etc.). The arboviruses do

not comprise a natural taxonomic category of related organisms, but are grouped together because

of their mode of transmission. Among them, however, are several groups of viruses that are closely

related.

Arenavirus: One of a group of viruses traditionally

grouped with the arboviruses, probably incorrectly.

Arenaviruses probably are not transmitted by arthropods, though their life cycles still are not fully

known.

Arthropod: An invertebrate animal belonging to the

phylum Arthropoda, which includes insects, ticks,

spiders, and crustaceans.

Attenuated vaccine: A vaccine made of whole pathogenic organisms that are treated with chemicals,

radioactivity, or other means to render them incapable of producing infection.

Autoantibody: An antibody that is formed by an individual against the individual’s own tissues.
256 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Autoimmunity: An immune state in which antibodies are produced by an individual against his or her

own tissue (autoantibodies).

Avirulent; nonvirulent: Capable of causing only a mild

or inapparent infection.

Bacillus (pi., bacilli): Any of various rod-shaped, aerobic bacteria belonging to the genus Bacillus.

Bacillus Calmette-Guerin (BCG) vaccine: A vaccine

prepared from attenuated Mycobacterium bovis, an

agent that infects cattle, and used to immunize humans against infection with M. tuberculosis, the related bacterium that causes human tuberculosis.

Bacteremia: A pathologic state characterized by the

presence of bacteria in the blood.

Bactericide: An agent capable of killing bacteria.

Bacteriology: The scientific study of bacteria.

Bacteriophage: One of a group of viruses that infects

and replicates in certain bacteria. Also called

“phage.”

Bacteriostatic: Capable of slowing or halting the

growth of bacteria without killing them.

Bacterium (pl., bacteria): Any of a group of one-celled

micro-organisms having round, rodlike, spiral, or

filamentous bodies that are enclosed by a cell wall

or membrane and lack fully differentiated nuclei.

Bacteria may exist as free-living organisms in soil,

water, organic matter, or as parasites in the bodies

of plants and animals. Some, but not all, bacteria

can cause disease.

Base pairs (of nucleic acids): Nucleotide bases that pair

across the double helix of the DNA or RNA molecule in a very specific way: adenine with thymine

(or uracil in RNA), cytosine with guanine.

Biological control: The control of insects or other organisms necessary for the development or transmission of disease, through measures such as the introduction of natural predators or pathogens of the

target organism, the use of naturally produced

chemicals, and the introduction of sterile insects to

the breeding population. (Compare environmental

control. )

Bionomics: The study of the relationship of organisms

to the environment.

Biotechnology: Techniques that use living organisms

or substances from such organisms to make or modify a product. In this report, biotechnology refers

to recombinant DNA techniques and other sophisticated tools relying on the ability to harness and

manipulate genetic material.

B-lymphocytes: See lymphocytes.

Carrier: A person or animal who, without apparent

symptoms, harbors a pathogen and may serve as

a source of infection to others.

Case fatality rate: The proportion of individuals with

a specific diagnosis who die from the disease during a specified period of time.

Cell culture: Growth in the laboratory of cells isolated

from multicellular organisms. Each culture is usually of one type.

Cell-mediated immunity: Immunity resulting from increase of activity by living cells in the blood and

other tissues (e.g., T-lymphocytes, natural killer

cells) that directly and nonspecifically destroy foreign material. (Compare humoral immunity. )

Chagas’ disease: Infection by Trypanosoma cruzi,

transmitted by reduviid bugs. The disease was discovered and described by Carlos Chagas of Brazil.

It is characterized by an acute course in children

with fever, encephalitis, and inflammation of the

heart muscle (often life-threatening or fatal), and

a chronic course in adults leading to heart disease

and heart failure. Also called “American trypanosomiasis.” (See also trypanosomiasis.)

Chemoprophylaxis: The prevention of disease by the

use of drugs or chemicals.

Chemotherapy: The use of specific chemical agents to

arrest the progress of, or eradicate, disease in the

body.

Cholera: A severe diarrheal disease caused by the bacterium Vibrio cholerae.

Chromosomes: The rodlike structures of a cell’s nucleus that store and transmit genetic information;

the physical structures that contain genes.

Chronic: Lingering, lasting, as opposed to acute.

Clone: A group of genetically identical cells or organisms produced asexually from a common ancestor.

Cold chain: The means whereby vaccines can be transferred from the manufacturer to the physician in

the field at a sufficiently low temperature to ensure

the effectiveness of the vaccines. Some of the important childhood vaccines require continuous

refrigeration.

Complement: A protein complex in plasma that causes

the lysis of bacteria and other pathogens that react

with antibody.

Complementary DNA (cDNA): DNA that is complementary to messenger RNA; used for cloning or

as a probe in DNA hybridization studies.

Congenital: Existing at birth.

Cross-reactivity: The property of an organism able to

provoke an immunologic reaction against a different organism. The tuberculosis vaccine BCG, for

example, is an attenuated strain of Mycobacterium

bovis (a bovine tuberculosis) that provokes the immune reaction against M. tuberculosis, the cause

of human tuberculosis.
App. C—Glossary of Acronyms and Terms ● 257

DDT (dichloro-diphenyl-trichloro-ethane): An insecticide used to control mosquitoes and other vectors

of tropical diseases.

Dengue fever: An acute febrile disease caused by an

arbovirus, transmitted by mosquitoes of the genus

Aedes, and characterized by fever, severe pains in

the head, eyes, muscles, and joints, and a skin

eruption.

Diagnostic technology: A technology used to determine the presence or absence of disease, and/or to

characterize the extent of the disease.

Diarrheal diseases: Diseases characterized by the passage of loose watery stools, usually at more frequent

than normal intervals. The dehydration that accompanies diarrhea is the cause of great morbidity and

mortality, particularly among infants and children.

DNA (deoxyribonucleic acid): A nucleic acid, containing the sugar ribose, that is found in cell nuclei and

is the carrier of genetic information. (Compare

RNA .)

DNA-DNA hybridization: See nucleic acid hybridization.

DNA probe: A sequence of DNA that is used to detect the presence of a particular nucleotide sequence

in a sample of DNA.

DNA sequence: The order of nucleotide bases in the

DNA helix.

Dysentery: Inflammation of the intestine characterized

by pain, intense diarrhea, and the passage of mucus and blood.

Electrolytes: Any compound that dissociates into

charged ions in solution and can conduct a current

of electricity. A balance of these in body fluids is

necessary for the body to function normally.

Elephantiasis: The enormous swelling of a limb, usually a leg, as a result of lymphatic obstruction by

filarial worms, followed by thickening of the skin

and subcutaneous tissues.

Emesis: Vomiting.

Encephalitis: Inflammation of the tissues of the brain.

Endemic: Constantly present or persistent within a

given geographic area, a term used to refer to a human disease or an infectious agent. (Compare enzootic, epidemic. )

Enteric: Pertaining to the intestine.

Enterotoxigenic: Capable of producing a toxin within

the intestine. Some strains of E. coli cause disease

by producing such a toxin.

Entomology: The scientific study of insects.

Environmental control: The control of disease through

measures involving the alteration of the physical

environment to eliminate the conditions necessary

for the survival of the insects or other organisms

that transmit or harbor the agents of disease.

Enzootic: The constant presence or persistence of an

animal disease or infectious agent within a given

geographic area; the animal counterpart of endemic.

(Compare endemic, epizootic.)

Enzyme: Any of a group of catalytic proteins that are

produced by living cells and that mediate and promote the chemical processes of life without themselves being altered or destroyed.

Epidemic: A sudden increase in the incidence rate of

a human illness, affecting large numbers of people

and spread over a wide area. (Compare endemic,

epizootic. )

Epidemiology: The scientific study of the distribution

and occurrence of diseases and health conditions,

and their determinants.

Epitope: A structural part of an antigen that is responsible for an antibody response against that antigen.

Also called “antigenic determinant. ”

Epizootic: Affecting many animals in one region simultaneously; the animal counterpart of epidemic.

(Compare enzootic, epidemic. )

Erythrocytes: Red blood cells.

Escherichia coli: A species of rod-shaped bacteria that

inhabit the normal intestinal tract of vertebrates.

Some strains cause intestinal disease and diarrhea

in humans through at least three mechanisms: enterotoxigenic E. coli produces toxins that cause excessive fluid production in the intestine; enteroinvasive E. coli invades the cells of the intestinal

wall; enteropathogenic E. coli produces a toxin that

causes disease in infants. Many nonpathogenic

strains of E’. coli are used as hosts in recombinant

DNA technologies.

Etiologic: Causative.

False negative: A negative test result in an individual

with the disease in question, i.e., the patient is incorrectly diagnosed as not having a particular

disease.

False positive: A positive test result in an individual

who does not have the disease in question, i.e., the

individual is incorrectly diagnosed as having a particular disease.

Fertility rate: The annual number of live births per

1,000 women of child bearing age (15 to 49 years)

in a defined population.

Filaria: Parasitic nematode worms, named for their

threadlike appearance.

Filariasis: A disease in humans due to infection with

filarial worms, such as Wuchereria bancrofti, Brugia malayi (transmitted by mosquitoes), and Onchocerca vulvulus (transmitted by blackflies).

Adults of W. bancrofti and B. malayi live in the

human lymphatic system and connective tissues,

where they may cause obstruction. The immature
258 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

worms (microfilariae) migrate to the host’s bloodstream. Completion of the parasite’s life cycle is dependent on passage through a mosquito.

Fluke: The common name for a large number of species of parasitic flatworms that form the class

Trematoda.

Gamete: A mature germ cell with one set of chromosomes, capable of fusing with another gamete and

forming a new, genetically distinct, individual.

Gametocyte: A life cycle stage of Plasmodium, the

malarial agent; this stage infects mosquitoes after

the mosquito bites an infected human (or other

mammal), and gives rise to the gamete, or, sexual

stage, of the parasite.

Gene: The basic unit of heredity; an ordered sequence

of nucleotides comprising a segment of DNA. A

gene contains the sequence of DNA that encodes

one polypeptide chain (via RNA).

Genome: The genetic endowment of an organism or

individual.

Genus (pi., genera): A taxonomic category that includes groups of closely related species of plants or

animals.

Glycoprotein: A protein with attached sugar groups.

Helminth: Any parasitic worm.

Hemorrhagic fever: A severe complication of some viral diseases that involves internal or external bleeding. Several arboviruses sometimes cause epidemic

outbreaks of hemorrhagic fever.

Host: 1. In the context of parasitology, a living organism that harbors a parasite. Definitive hosts harbor the adult or sexual stage of a parasite; intermediate hosts harbor the larval or asexual stages

of a parasite. 2. In the context of recombinant DNA

technology, the organism into which a scientist inserts foreign DNA.

Humoral immunity: Immunity associated with antibodies that circulate in the blood. (Compare cellmediated immunity. )

Hybridoma: Product of fusion between a myeloma cell

(which divides continuously in culture and is “immortal”) and a lymphocyte (antibody-producing

cell); the resulting cell grows in culture and produces

the specific antibody produced by the parent

lymphocyte (a monoclinal antibody).

Hypernatremia: An abnormally high concentration of

sodium ions in the blood.

Hypnozoite: Forms of some species of Plasmodium,

the cause of malaria, that remain dormant in liver

cells, sometimes for many years, retaining their ability to activate an infection and cause acute malaria.

Hyponatremia: An abnormally low concentration of

sodium ions in blood. Hyponatremia often accompanies severe diarrhea.

Immune: Protected against disease by innate or acquired immunity.

Immune response: The reaction of an organism to invasion by a foreign substance. Immune responses

are often complex, and may involve the production

of antibodies from special cells (B-lymphocytes), as

well as a varying set of physical and chemical responses from other cells of the immune system.

Immune serum: Blood serum that contains antibodies

and can be used to confer passive immunity to a

variety of diseases.

Immune system: A specialized group of body cells and

cell products that respond to foreign organisms and

substances in the body. The cell products are largely

immunoglobulins (antibodies). Some lymphocytes

and various other cells of the immune system

directly attack foreign organisms.

Immunity: A living organism’s condition of being able

or capacity to resist a particular disease. Innate immunity is natural or inherited. Acquired immunity

may be active or passive. Active immunity results

from previous exposure to the disease-causing agent

or vaccination; passive immunity is the result of

transfer of preformed antibodies in immune serum

or from mother to fetus. (See also cell-mediated immunity, humoral immunity. )

Immunization: The deliberate introduction of an antigenic substance (vaccine) or antibodies into an individual, with the aim of producing active or passive immunity to a disease. (Compare vaccination. )

Immunization technology: Interventions designed to

render individuals resistant to disease if they are exposed to a disease-causing agent. Vaccines are the

most important tools of immunization.

Immunoassay: The use of antibodies to identify and

quantify substances. The binding of antibodies to

antigen, the substance being measured, is often accompanied by tracers such as radioisotopes.

Immunodiagnosis: A process whereby specified immunologic characteristics of cells, serum, or biologic

specimens are determined for the purpose of diagnosing disease.

Immunogenic: Able to cause an immune response.

Immunoglobulin: Any of a set of serum glycoprotein

molecules that have the ability to bind other

molecules with a high degree of specificity. Immunoglobulins include all the antibodies.

Immunology: The scientific study of immunity, induced sensitivity, and allergy.

Immunosuppression: Suppression of the immunologic

response.

Incidence: The frequency of new occurrences of disease within a defined time interval in a defined population. Incidence rate is the number of new cases
App. C—Glossary of Acronyms and Terms ● 259

of specified disease divided by the number of people in a population over a specified period of time,

usually 1 year.

Incubation: The time between infection by a diseasecausing organism and the appearance of disease.

Infant mortality rate: Number of deaths among children less than 1 year old as a fraction of the total

number of live births in a year.

Infection: The entry and proliferation of any pathogenic organism in another organism.

Innate immunity: Immunity that is genetically determined at birth.

Inoculate: To introduce immunologically active material (e.g., an antibody or antigen) into, especially

in order to treat or prevent a disease,

Insecticide: A substance capable of killing insects.

Insect vector: An insect that can transmit a diseaseproducing organism from one human or animal to

another.

Integrated pest management (1PM): The use of a combination of biological, chemical, environmental

measures to control vectors that transmit tropical

diseases to humans or other animals.

Intermediate host: See host.

In vitro: Literally, in glass; pertaining to a biological

process or reaction taking place in an artificial environment, usually a laboratory. Sometimes the term

is used to include the growth of cells from multicellular organisms under cell culture conditions.

In vivo: Literally, in the living; pertaining to a biological process or reaction taking place in a living

cell or organism.

Kinetoplastid: Characteristic structure at the base of

the flagellum in certain protozoa (e.g., Leishmania

spp. and Trypanosoma spp. ).

Larvicide: A substance capable of killing insect larvae.

Leishzmania: A genus of flagellated parasitic protozoans

that cause leishmaniasis.

Leishmaniasis: Any of several infections caused by

Leishmania spp., transmitted by sandflies. Cutaneous leishmaniasis is a skin ulcer caused by L. mexicana (New World) or L. tropica (Old World). Mucocutaneous leishmaniasis is an ulceration of the nose

and throat caused by L. brazdiensis, occurring in

tropical America. Visceral leishmaniasis, also called

“kala-azar,” is a generalized and internal disease

caused by L. donovani (New and Old World).

Lepromatous leprosy: See leprosy.

Leprosy: A chronic, infectious, granulomatous disease

of humans caused by the bacillus Mycobacterium

Ieprae. The disease occurs almost exclusively in

tropical and subtropical regions, and ranges in

severity from localized, spontaneously remitting lesions (tuberculoid leprosy) to malignant lesions with

progressive anesthesia, paralysis, ulceration, nutritive disturbances, gangrene, and mutilation (lepromatous leprosy). Also called “Hansen’s disease. ”

Lesion: A wound, injury, or one of the individual

points or patches of a multifocal disease.

Lower respiratory tract infection (LRTI): See acute respiratory infections.

Lymphocytes: Specialized white blood cells involved

in the body’s immune response. B-lymphocytes

originate in the bone marrow and when stimulated

by antigen produce circulating antibodies (humoral

immunity). T-lymphocytes originate in the thymus

and engage in a type of defence that does not depend directly on antibody attack (cell-mediated immunity).

Lyophilized: Freeze-dried.

Microphage: A type of large, ameba-like cell, found

in the blood and lymph, which consumes foreign

particles, including bacteria and parasites.

Malaria: Any of a group of human febrile diseases

caused by infection of red blood cells by protozoan

parasites of the genus Plasmodium, transmitted by

mosquitoes of the genus Anopheles. Four species

of Plasmodium cause malaria in humans: P. falciparum, P. vivax, P. malariae, and P. ovale. P. vivax

and P. ovale have a persistent stage in the liver that

causes relapses. Many other species of Plasmodium

infect monkeys, rodents, birds, and reptiles.

Merozoite: A life cycle stage of the malarial agent Plasmodium; this stage develops in the vertebrate host’s

liver, then enters the circulatory system and infects

red blood cells.

Messenger RNA (mRNA): RNA that serves as the template for protein synthesis; it carries the transcribed

genetic code from DNA and directs protein synthesis.

Microfilariae: Slender, motile prelarval forms of filarial

nematodes, the parasites that cause filariasis.

Micro-organism: A minute, microscopic, or submicroscopic living organism. Examples are bacteria, mycoplasma, viruses, and protozoa.

Molecular biology: The study of biology at the level

of individual molecules, such as proteins and DNA.

Molluscicide: Any chemical agent used to kill mollusks; in the context of tropical diseases, snails necessary in the life cycles of schistosomes are the most

important targets.

Monoclinal antibodies (MAbs): Homogeneous antibodies derived from clones of a single cell. MAbs

recognize only one chemical structure. They are useful in a variety of industrial and medical capacities

since they have remarkable specificity.

Monocytes: Phagocytic, large white blood cells, containing one nucleus.
260 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

Morbidity: The condition of being diseased.

Morphology: The study of the configuration or structure of organisms.

Mortality rate: The death rate, often made explicit for

a particular characteristic, e.g., age, sex, or specific

cause of death. A mortality rate contains three essential elements: 1) the number of people in a population group exposed to the risk of death; 2) a time

factor; 3) the number of deaths occurring in the exposed population during a certain time period.

Mycobacterial diseases: A group of human and animal diseases caused by species of the bacterial genus Mycobacterium. Important human mycobacterial diseases are tuberculosis and leprosy.

Mycology: The scientific study of fungi.

Mycoplasma: Micro-organisms similar to bacteria, but

lacking a rigid cell wall.

Nagana: A disease of cattle and other livestock in

Africa, caused by Trypanosoma brucei, the cause

of African sleeping sickness in humans.

Necrosis: Death of tissue.

Nematodes: Elongated, cylindrical worms, also called

roundworms, many of which are parasites. Hookworm and the worms that cause trichinosis are

nematodes. The filarial worms also belong to this

group.

Nucleic acid: Macromolecules composed of sequences

of nucleotides. There are two kinds of nucleic acids:

DNA, which contains the sugar deoxyribose, and

RNA, which contains the sugar ribose.

Nucleic acid hybridization: Matching of either DNA

or RNA (depending on the organism) from an unknown organism with DNA or RNA from a known

organism. This method is used in tropical disease

research for identifying species and strains of

pathogens.

Nucleotide: A structural unit of nucleic acid, consisting of a base, a sugar, and a phosphate molecule.

Oligonucleotides: Short segments of DNA or RNA,

made up of a few (2 to 10) nucleotides.

Onchocerciasis: An infection of humans with the filarial worm Onchocerca vovulus, transmitted by the

bite of blood-sucking blackflies, The disease is generally characterized by skin nodules that can become fibrous and calcified. Also called “African

river blindness, ” for the blindness that occurs when

the worms invade the eye.

Oral dehydration therapy (ORT): The treatment or

prevention of dehydration due to diarrhea by a specific water solution of electrolytes and glucose (salts

and sugar) taken by mouth.

Oropouche fever: An arboviral disease transmitted by

biting midges (Culicoides spp.). Symptoms include

anorexia, rash, and joint and muscle pain.

Orphan Drug Act: Public Law 97-414, which charges

the U.S. Government with identifying and promoting orphan products, defined as drugs and devices

for rare diseases.

Pandemic: Worldwide epidemic.

Parasite: An organism living in or on another living

organism, obtaining from the host organism part

or all of its organic nutriment.

Parasitemia: The presence of parasites in the blood.

Parasitic disease: A disease caused by a parasite. Examples are malaria, schistosomiasis, trypanosomiasis, leishmaniasis, and filariasis.

Parasitology: The scientific study of the relationship

between parasites and their hosts.

Passive immunity: Immunity that is the result of the

transfer of preformed antibodies in immune serum

or from mother to fetus. (Compare active immunity. )

Pathogen: A specific causative agent (e.g., a virus or

bacterium) of a disease.

Pathogenesis: The mode of origin and development of

a disease process.

Pathogenicity: The condition of causing disease.

Pathology: The scientific study of the cause of disease,

and the associated structural and functional changes

that result from disease.

Pertussis: An infectious inflammatory respiratory disease of children caused by the bacterium Bordetella

pertussis. The disease is characterized by explosive

attacks of coughing ending in an inspiratory whoop.

Also known as “whooping cough.”

Phage: See bacteriophage.

Phagocytosis: Consumption of foreign particles (e.g.,

bacteria) by cells that use ameboid movement to

surround the particle and then digest it.

Phenotype: The observable characteristics of a strain

or species.

Phlebotomine sandflies: Insect vectors of Leishmania

spp., the agents of leishmaniasis.

Plasmid: An extrachromosomal, self-replicating, circular segment of DNA. Plasmids can be used as

“vectors” for cloning foreign DNA in bacterial

“host” cells.

Plasmodium: The genus of protozoans that cause malaria in humans and other animals.

Pneumonia: An acute or chronic inflammation of the

lungs, which can be caused by a variety of microorganisms.

Polypeptide: A sequence of amino acids (at least three)

joined in a chain.

Prevalence rate: The number of existing cases of a disease in a defined population at a particular time,

or over a specified time period.

Probe: See DNA probe.
App. C—Glossary of Acronyms and Terms ● 261

Prophylaxis: The prevention of disease.

Protozoa: A phylum of one-celled animals, a few of

which cause disease in humans. Examples are the

causes of malaria, leishmaniasis, and trypanosomiasis.

Reagent: A substance that takes part in a chemical reaction.

Recombinant DNA: The hybrid DNA produced by

joining pieces of DNA from different sources together in vitro.

Recombinant DNA techniques: Techniques that allow

specific segments of DNA to be isolated and inserted

into a bacterium or other host (e. g., yeast, bacteria) in a form that will allow the DNA segment to

be replicated and expressed as the cellular host multiplies.

Recrudescence: The reappearance of a morbid process or its symptoms after a period of improvement.

Reduviid bug: A blood-sucking bug in the Reduviid

family that is the vector of Trypanosoma cruzi, the

agent of Chagas’ disease.

Reservoir: Any person, animal, arthropod, plant, soil,

or substance (or combination of these) in which an

infectious agent lives in such manner that it can be

transmitted to a susceptible host.

Respiratory syncytial virus (RSV): The most important cause of lower respiratory disease (pneumonia

and bronchiolitis) in children under 2 years of age.

Restriction enzymes: Bacterial enzymes that cut DNA

at specific nucleotide sequences.

Rhinovirus: One of many virus families that cause upper respiratory disease, Rhinovirus is a cause of the

“common cold. ”

Rickettsia: A group of rod-shaped micro-organisms

which may be transmitted by arthropods and are

responsible for some human diseases such as Rocky

Mountain spotted fever and epidemic typhus,

River blindness: See onchocerciasis.

RNA (ribonucleic acid): A nucleic acid, containing the

sugar ribose, that is found in cytoplasm and some

cell nuclei and is associated with the control of cellular chemical activities. In its three forms—messenger RNA, transfer RNA, and ribosomal RNA—

RNA assists in translating the genetic message of

DNA into the finished protein. (Compare DNA. )

RNA-RNA hybridization: See nucleic acid hybridization.

Rotavirus: Any of a group of viruses (round in shape)

which are the major cause of diarrhea] disease in

infants and children,

Salmonella: A genus of bacteria that can cause diarrheal disease.

Schistosoma: The genus of blood flukes that cause

schistosomiasis.

Schistosomiasis: A chronic, debilitating infection by

worms of the genus Schistosoma (“blood flukes”).

The three most important species in humans are:

S. mansoni, S. haematobium, and S. japonicum.

Sensitivity: The ability of a diagnostic test accurately

to diagnose a disease or condition when the disease

or condition is present. High sensitivity means few

false negatives. (Compare specificity. )

Serotype: A group of closely related micro-organisms

that are distinguished by their possession of a common set of antigenic characteristics. The term also

refers to the antigen set characteristic of such a

group.

Serum: The clear liquid which separates in the clotting of blood and which contains the antibodies that

were present in the whole blood.

Shigella: A genus of bacteria, some of which can cause

diarrheal disease.

Species: A taxonomic category that includes closely

related, morphologically similar individuals that actually or potentially interbreed; the principal subdivision of a genus.

Species complex: A group of two or more closely related species that can only be differentiated by

cytogenetic analysis or cross-breeding experiments.

Specificity: The ability of a diagnostic test correctly

to determine that an individual does not have a specific disease or condition. High specificity means

few false positives. (Compare sensitivity. )

Sporozoite: A life cycle stage of the malarial agent

Plasmodium; this is the stage injected by the mosquito vector into the vertebrate host’s bloodstream.

Strain: A group of organisms of the same species having a distinctive quality or characteristic (biochemical, pathogenic, or other) that can be differentiated,

but are not different enough to constitute a separate species.

Subunit vaccine: A vaccine that contains only portions

of an antigenic molecule of a pathogen. Subunit

vaccines can be prepared by using recombinant

DNA technology to produce all or part of the antigenic molecule or by artificial (chemical) synthesis

of short peptides.

Surveillance: Constant observation of an area to determine the level of disease activity.

TDR: The acronym for the Special Program for Research and Training in Tropical Diseases, sponsored

jointly by the U.N. Development Program, the

World Bank, and the World Health Organization.

TDR diseases: The six diseases singled out for attention by the Special Program for Research and Training in Tropical Diseases (TDR): malaria, schistosomiasis, trypanosomiasis, filariasis, Ieishmaniasis,

and leprosy.
262 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Therapeutic technology: A technology that cures or

relieves the symptoms of a disease or other medical condition.

Titer: The lowest concentration (highest dilution) of

an active substance (e.g., antibody in serum) that

causes a discernible reaction with another substance.

T-lymphocytes: See lymphocytes.

Toxin: A substance, produced in some cases by microorganisms, which is toxic to other living organisms.

Transcription: The synthesis of messenger RNA on a

DNA template; the resulting RNA sequence is complementary to the DNA sequence. This is the first

step in gene expression. (Compare translation. )

Translation: The process in which the genetic code

contained in the nucleotide base sequence of messenger RNA directs the synthesis of a specific order of amino acids to produce a protein. (Compare

transcription. )

Transmission: The passage of a pathogen from one

host to another host, or from vector to host.

Trematode: Any of a group of parasitic flatworms, including the flukes, of the phylum Platyhelminthes.

Schistosomes are important human trematode parasites.

Trypanosoma: A genus of slender, polymorphic, parasitic protozoans that cause trypanosomiasis.

Trypanosomiasis: Any of several diseases caused by

infection with species of the genus Trypanosoma.

The important human diseases are African sleeping sickness (also called African trypanosomiasis)

and Chagas’ disease (also called American trypanosomiasis). African sleeping sickness is caused by T.

brucei rhodesiense in east Africa or T. b. gambiense

in west Africa, both transmitted by the tsetse fly.

Chagas’ disease is caused by T. cruzi, transmitted

by blood-sucking reduviid bugs.

Tsetse flies: Any of several two-winged flies of the genus Glossina that occur in Africa south of the Sahara; medically important as vectors of African

trypanosomiasis.

Tubercle bacillus: A bacillus causing tuberculosis; usually refers to Mycobacterium tuberculosis, the principal cause of human tuberculosis.

Tuberculoid leprosy: See leprosy.

Tuberculosis: A chronic infectious disease of humans

and animals caused by any of several species of

mycobacteria. It usually begins with lesions in the

lung, but can spread to other parts of the body.

Upper respiratory tract infection (URTI): See acute respiratory infections.

Vaccination: The deliberate introduction of an antigenic substance (vaccine) into an individual, with

the aim of producing active immunity to a disease.

(Compare immunization.)

Vaccine: A preparation of living, attenuated, or killed

bacteria or viruses, fractions thereof, or synthesized

antigens identical or similar to those found in the

disease-causing organisms, that is administered to

produce or increase immunity to a particular

disease.

Vaccinia virus: The organism that causes cowpox; its

injection into humans results in immunity to the related smallpox virus.

Vector: A transmission agent: 1. In the context of

medicine, a carrier of disease; usage commonly

refers to arthropods (e.g., mosquitoes, sandflies,

ticks) or rodents. 2. In the context of recombinant

DNA technology, the DNA molecule used to introduce foreign DNA into host cells; vectors include

plasmids, bacteriophages, and other forms of DNA.

Vector bionomics: The study of the habits (feeding,

resting, and breeding) of vectors of disease and variations among different strains and in different

locales.

Vector-borne disease: A disease transmitted by an insect or other vector. Such diseases include malaria,

trypanosomiasis, and arboviral infections.

Vector control technology: A technology aimed at

controlling the vectors that transmit disease or other

organisms (e.g., snails) that are not true vectors,

but serve as intermediate hosts of human or other

animal disease organisms.

Vibrio cholerae: The bacterium that causes cholera.

Virology: The study of viruses.

Virulence: The degree and severity with which a pathogen is able to infect a population and cause disease.

Virus: Any of a large group of submicroscopic agents

infecting plants, animals, and bacteria and characterized by a total dependence on living cells for reproduction and by a lack of independent metabolism. A fully formed virus consists of nucleic acid

(DNA or RNA) surrounded by a protein or protein

and lipid coat.

Water-borne disease: A disease transmitted through

contaminated water. Most diarrheal diseases can be

water borne.

Whooping cough: See Pertussis.

Wild-type: The most frequently encountered phenotype in natural breeding populations.

Xenodiagnosis: A technique in which an intermediate

host or vector is used to diagnose the presence of

parasites in humans; e.g., reduviid bugs are permitted to feed on someone suspected of having

Chagas’ disease, and later, the bugs are examined

for the presence of T. cruzi parasites.
App. C—Glossary of Acronyms and Terms ● 263

Yellow fever: An acute febrile disease caused by an in monkey reservoir hosts; urban yellow fever refers

arbovirus that is transmitted by mosquitoes. Symp- to transmission of the same virus to humans.

toms include a high fever, jaundice, black vomit, Zoonosis: A disease primarily of animals that is transand anuria (absence of urine excretion). The virus missible to humans under natural conditions.

that causes jungle/sylvan yellow fever is maintained
References
References

1.

2.

3.

4.

5.

6.

7.

8.

9.

10,

11.

12

13.

Abdel-Hafez, S. K., Phillips, S. M., and Zodda,

D. M., “Schistosoma mamsoni: Detection and

Characterization of Antigens and Antigenemia by

Inhibition Enzyme-Linked Immunosorbent Assay

(IELISA),” Exp. Parasit. 55:219-232, 1983.

Abe, M., Minagawa, F., Yoshino, Y., et al.,

“Fluorescent Leprosy Antibody Absorption (FLAABS) Test for Detecting Subclinical Infection

With Mycobacterium leprae, ” Int. J. Leprosy

48(2):109-119, 1980.

Aikawa, M., Miller, L. H., Johnson, J., et al.,

“Erythrocyte Entry by Malarial Parasites: A

Moving Junction Between Erythrocyte and Parasite,” J. Cell Biol. 77:72-82, 1978.

Amler, R. W., Orenstein, W. A., and Bart, K. J.,

“The Immune Response to Aerosolized Measles

Vaccine,” J. A.M.A. 251:2408, 1984.

Anderson, L. J., Patriarca, P. A., Hierholzer, J. C.,

et al., “Viral Respiratory Illnesses, ” Med. CJinics

N. Amer. 67:1009-1030, 1983.

Antczak, D. F., “Monoclinal Antibodies: Technology and Potential Use, ” J. Am. Veterin. Med.

ASSOC. 181:1005-1010, 1982.

Aperia, A., Broberger, O., Thodenius, K., et al.,

“Development of Renal Control of Salt and Fluid

Homeostasis During the First Year of Life, ” Acta

Paediat. Scand. 64:393, 1975.

Araujo, F. G., Chiari, E., and Dias, J. C. P., “Demonstration of Trypanosoma cruzi Antigen in Serum From Patients With Chagas’ Disease, ” Lancet 1:246-249, 1981.

Araujo, F. G., Handman, E., and Remington,

J. S., “Use of Monoclinal Antibodies To Detect

Antigens of Toxoplasma gondii in Serum and

Other Body Fluids, ” Infect. Immun. 30:12-16,

1980.

Araujo, F. G., Sharma, S. D., Tsai, V., et al.,

“Monoclinal Antibodies to Stages of Trypanosoma cruzi: Characterization and Use for Antigen Detection, “ Infect. Immun. 37:344-349, 1982.

Asher, I., Office of Science Advisor, U.S. Agency

for International Development, Washington, DC,

personal communication, March 1984.

Austrian, R., “Pneumococcal Infection and Pneumococcal Vaccine” (editorial), N. E’ng. J. Med.

297:938-939, 1977.

Austrian, R., and Gold, J., “PneumococcalBacteremia With Especial Reference to Bacteremic

Pneumococcal Pneumonia, ” Ann. Int. Med.

60:759-776, 1964.

14.

15.

16.

17.

18.

19.

20.

21.

22.

23,

24.

25.

26.

27

Aziz, M. A., Diop, I. M., Diallo, S., et al., “Efficacy and Tolerance of Ivermectin in Human Onchocerciasis, ” Lancet 2:171-173, 1982.

Barnett, B., “Viral Gastroenteritis, ” Med. Clinics

N. Amer. 67:1031-1058, 1983.

Basch, P., “The Role of Biotechnology in Tropical Disease Research, ” typescript, contract report

prepared for the Office of Technology Assessment, U.S. Congress, Washington, DC, 1984.

Basta, S. S., and Churchill, A., Iron Deficiency

Anemia and the Productivity of Adult Males in

Indonesia, World Bank Staff Working Paper No.

175 (Washington, DC: World Bank, 1974).

Baxter, J. D., “Genetic Engineering and Its Impact

on Medicine, ” Ann. Acad. Med. Singapore

12:311-325, 1983.

Beaudoin, R. L., Strome, D. P. A., Tubergen,

T. A., et al., “Plasmodium berghei: Irradiated

Sporozoites of the ANKA Strain as Immunizing

Agents in Mice, “ Exp. Parasit. 39:438-443, 1976.

Behrman, J, N., Tropical Diseases: Responses of

the Pharmaceutical Companies (Washington and

London: American Enterprise Institute, 1980).

Benenson, A.S. (cd.), Control of Communicable

Diseases in Man, 13th ed. (Washington, DC:

American Public Health Association, 1981).

Bernards, A., Michels, P. A.M., Lincke, C. R., et

al., “Growth of Chromosome Ends in Multiplying Trypanosomes, ” Nature 303:592-597, 1983.

Black, R. E., Brown, K. H., Becker, S., et al.,

“Longitudinal Studies of Infectious Diseases and

Physical Growth of Children in Rural Bangladesh. II. Incidence and Etiology of Diarrhea, ”

Am. J. Epidemiol. 115:315-324, 1982.

Black, R. E., Levine, M. M., Ferreccio, C., et al.,

and the Chilean Typhoid Committee, “Salmonella typhi Ty21a Vaccine Trials in Santiago,

Chile, ” Abstract #268, in Abstracts of the 1984

ICAAC.

Boothroyd, J. C., Paynter, C. A., Coleman, S. L.,

et al., “Complete Nucleotide Sequence of Complementary DNA Coding for a Variant Surface

Glycoprotein From ~rypanosoma brucei, ” J.

Mol. Biol. 157:547-556, 1982.

Bordier, C., Garavito, R. M., and Armbruster,

B., “Biochemistry and Structural Analyses of Microtubules in the Pellicular Membrane of Leishmania tropica, “J. Protozool. 29:560-565, 1982.

Bourne, P. G., New Directions in International

Health Cooperation: A Report to the President

267
268 Ž Status of Biomedical Research and Related Technology for Tropical Diseases

28,

29,

30!

31,

32.

33.

34.

35.

36.

37.

38.

39.

40.

41.

42<

(Washington, DC: U.S. Government Printing Office, 1978).

Braude, A.I. (cd.), Medical Microbiology anclh

fectious Diseases (Philadelphia: W.B. Saunders,

1981).

Brener, Z. “Recent Developments in the Field of

Chagas’ Disease, ” Bull. W.H.0. 60(4):463-473,

1982.

Brennan, P. J., and Barrow, W. W., “Evidence for

Species-Specific Lipid Antigens in Mycobacterium leprae, ” Int. J. Leprosy 48:382, 1980.

Bruce-Chwatt, L. J., Essential Malariology (London: Camelot Press Ltd., 1980).

Bruce-Chwatt, L.J. (cd.), Chemotherapy ofiVlalaria, 2d ed. (Geneva: World Health Organization, 1981).

Bruce-Chwatt, L. J., “Malaria: From Eradication

to Control,” New Scientist 102(1406):17-20, 1984.

Bryceson, A., and Pfaltzgraf, R. E., Leprosy

(Edinburgh: Churchill Livingston, 1979).

Bulla, A., and Hitze, K. L., “Acute Respiratory

Infections: A Review,” Bull. W.H.O. 56:481-496,

1978.

Burkot, T. R., Zavala, F., Gwadz, R. W., et al.,

“Identification of Malaria-Infected Mosquitoes by

a Two-Site Enzyme-Linked Immunosorbent Assay,” Am. J. Trop. Med. Hyg. 33(2):227-231,

1984.

Burnet, M., Natural History of Infectious Diseases (Cambridge, England: Cambridge University Press, 1972).

Butler, T., Linh, N. N., Arnold, K., et al., “Therapy of Antimicrobial-Resistant Typhoid Fever, ”

Antimicr. Agents & Chemother. 11:645-650,

1977.

Butterworth, A. E., Taylor, D. W., Veith, M. C.,

et al., “Studies on the Mechanism of Immunity

in Human Schistosomiasis, ” lmmunolog. Rev.

61:5-39, 1982.

Buynak, E. B., Weibel, R. E., Carlson, A. J., et al.,

“Further Investigations of Live Respiratory Syncytial Virus Vaccine Administered Parenterally, ”

Proc. Soc. Exp. Biol. Med. 160:272-277, 1979.

Caldwell, H. D., Kirchheimer, W. F., and Buchanan,

T. M., “Identification of a A4ycobacterium Zeprae

Specific Protein Antigen(s) and Its Possible Application for the Serodiagnosis of Leprosy, ” ht.

J. Leprosy 47:477, 1979.

Campbell, C. C., Collins, W. E., Nguyen-Dinh,

P., et al., “Plasmod”um faZciparum Gametocytes

From Culture in Vitro Develop to Sporozoites

That Are Infectious to Primates, ” Science

217:1048-1050, 1982.

43,

44,

45,

46.

47.

48.

49.

50.

51.

52.

53.

54.

Campbell, G. H., Griswold, S., Cain, G., et al.,

“Monoclinal Antibodies to Antigens of TWpanosoma rhodesiense, ” Monoclinal Antibodies and

T-CeZl Hybridomas, G.J. Hammering, U. Hammering, and J.F. Kearney (eds. ) (Amsterdam:

Elsevier-North Holland Biomedical Press, 1981).

Capron, A., Dessaint, J.-P., Capron, M., et al.,

“Effecter Mechanisms of Immunity to Schistosomes and Their Regulation, ” Behring Institute

Mitteilungen 71:51-70, 1982.

Carpenter, C. C., and Sack, R. B., “Infectious Diarrheal Syndromes, ” Update I, Harn”son’s Principles of Internal Medicine, 9th cd., K.I. Isselbacker, R.D. Adams, E. Braunwald, et al. (eds. )

(New York: McGraw Hill, 1981).

Carter, R., and Chen, D. H., “Malaria Transmission Blocked by Immunization With Gametes of

the Malaria Parasite, ” Nature 263:57-60, 1976.

Cash, R. A., Music, S.1., Libonati, J.P, et al., “Response of Man to Infection With Vibrio cholerae.

II. Protection From Illness Afforded by Previous

Disease and Vaccine, ”J. Infect. Dis. 130:325-333,

1974.

Changusuphajaisiddhi, T., Sabchareon, A., and

Attanath, P., “Treatment of Quinine Resistant

Falciparum Malaria in Thai Children,” Southeast

Asian J. Trop. Med. Pub. Hlth. 14:357-362, 1983.

Chanock, R. M., and Murphy, B. R., “Use of

Temperature-Sensitive and Cold-Adapted Mutant Viruses in Immunoprophylaxis of Acute Respiratory Tract Disease, ” Rev. Infect. Dis. 2:421-

432, 1980.

Chanock, R. M., and Tully, J. G., “Mycoplasma,”

Microbiology, 3d cd., R. Davis, H.N. Dulbecco,

B. D. Eisen, et al. (eds. ) (New York: Harper &

Row, 1980).

Chanock, R. M., Kapikian, A. Z., Perkins, J. C.,

et al., “Vaccines for Non-Bacterial Respiratory

Diseases Other Than Influenza,” Pan. Am. Hlth.

Org. Sci. Pub. 26:101-116, 1971.

Chao, R. K., Fishaut, M., Schwartzman, J. D., et

al., “Detection of Respiratory Syncytial Virus in

Nasal Secretions From Infants by Enzyme-Linked

Immunosorbent Assay, ” J. Infect. Dis. 139:483-

486, 1979.

Chaulet, P., Benachenhou, A., Virot, J. P., et al.,

“Results of the International Co-operative Inquiry

Into the Cost of Anti-Tuberculosis Drugs, ” BuJ1.

Intern. Union Against Tuberculosis 53:247-253,

1978.

Check, W. A., “Renewed Control Efforts Emphasize ‘Unrecognized’ Influenza Threat, ” J. A.M.A.

251(20):2629-2637, 1984.
References Ž 269

55.

56.

57.

58.

59.

60.

61.

62.

63.

64.

65.

66.

67.

68.

Cherubin, C. E., “Antibiotic Resistance of Sahnonella in Europe and the United States, ” Rev. 1rIfeet. Dis. 3:1105, 1981.

Cho, S., Yanagihara, D. L., Hunter, S. W., et al.,

“Serological Specificity of Phenolic Glycolipid I

From Mycobacteriurn leprae, ” Infect. hnmun.

41:1077-1083, 1983.

Chretien, J., Holland, W., Macklen, P,, et al.,

“Acute Respiratory Infections in Children, ” N.

Eng. J. Med. 310:982-984, 1984.

Chu, K, Y., Vanderburg, J. A., and Klumpp, R. K.,

“Techniques for Estimating Densities of Br.dinus

truncatus roldfsi and Its Horizontal Distribution

in Volta Lake, Ghana, ” Bull. W.H.0. 54:411-416,

1976.

Ciolli, D., “Immune Protection Against Schistosoma mansoni in Permissive and Nonpermissive

Hosts, ” Scripta Varia 47:79-90, 1982.

Clements, M. L., Betts, R. F., and Murphy, B. R.,

“Advantage of Live Attenuated Cold-Adapted Influenza A Virus Over Inactivated Vaccine for

A/Washington/80 (H3N2) Wild-Type Virus Infection, ” Lancet 1:705-708, 1984.

Clyde, D. F., “Immunization of Man Against Falciparum and Vivax Malaria by Use of Attenuated

Sporozoites, ” Am. J. Trop. Med. Hyg. 24:397-

401, 1975.

Clyde, D. F., Most, H., McCarthy, V. C., et al.,

“Immunization of Man Against Sporozoite-Induced

Falciparum Malaria, “ Am, J. Med. Sci. 266:169-

177, 1973.

Cochrane, A. H., Nussenzweig, R. S., and Nardin,

E. H., “Immunization Against Sporozoites, ” Malaria, vol. 3, J.P. Kreier (cd. ) (New York: Academic Press, 1980).

Cochrane, A. H., Santoro, F., Nussenzweig, V,,

et al., “Monoclinal Antibodies Identify the Protective Antigens of Sporozoites of Plasmodium

knowksi,” Proc. Nat. Acad. Sci. (USA) 79:5651-

5655, 1982.

Cohen, S., and Mitchell, G. H., “Prospects for Immunization Against Malaria, ” Curr. Topics

Microbiol. Immunol. 80:98-137, 1978.

Cohen, S., and Warren, K.S. (eds. ), The Immunology of Parasitic Infections (Oxford: Blackwell

Scientific, 1984).

Cohen, S., Butcher, G. A., and Mitchell, G. H.,

“Immunization Against Erythrocytic Forms of

Malaria Parasites,” Adv. Exp. Med. Biol. 93:89-

112, 1977.

Cohen, S., Butcher, G. A., and Mitchell, G. H.,

et al., “Acquired Immunity and Vaccine in Malaria, ” Am. J. Trop. Med. Hyg. 26(suppl.):223-

232, 1977.

69.

70!

71.

72.

73.

74.

75.

76.

77.

78.

79.

80.

81.

82.

Collins, F. H., Zavala, F., Graves, P. M., et al.,

“First Field Trial of the Immunoradiometric Assay To Detect Sporozoites in the Gambia, ” Am.

J. Trop. Med. Hyg. 33(4):538-543, 1984.

Collins, F. M., “The immunology of Tuberculosis,” Am. Rev. Resp. Dis. 125:42-49, 1982.

Constantine, N. T., and Anthony, R. L., “Antigenic Differentiation of the Kinetoplastids Leishmania braziliensis and Trypanosoma cruzi by

Means of Monoclinal Antibodies, ” J. Protozool.

30:346-350, 1983.

Convit, J., Aranzazu, N., Zuniga, M., et al., “Immunotherapy and Immunoprophylaxis of Leprosy, ” Leprosy Rev. Special Issue:47-60, 1983.

Cook, J. A., “Sources of Funding for Training and

Research in Parasitology, ” The Current Status

and Future of Parasitology, K.S. Warren and E.F.

Purcell (eds. ) (New York: Josiah Macy Jr. Foundation, 1981).

Cook, J. A., Jordan, P., and Warren, K. S., “The

St. Lucia Experiment and the Evolution of a Contemporary Global Strategy for the Control of

Schistosomal Disease, ” typescript, Rockefeller

Foundation, New York, 1981.

Corning, M. E., A Review of the United States

Role in International Biomedical Research and

Communications: International HeaZth and Foreign Policy, NIH Publication No. 80-1638 (Bethesda, MD: National Institutes of Health, 1980).

Couch, R. B., and Kasel, J. A., “Immunity to Influenza in Man,” Ann. Rev. MicrobioZ. 37:529-

549, 1983.

Coulaud, J. P., Lariviere, M., Aziz, M. A,, et al.,

“Ivermectin in Onchocerciasis, ” Lancet 2:526-527,

1984.

Craig, P. S., Hocking, R. E., Mitchell, G. F., et al.,

“Murine Hybridoma-Derived Antibodies in the

Processing of Antigens for the Immunodiagnosis

of Hydatid (Echinococcus granulosus) Infection

in Sheep,” Parasitology 83:303-317, 1981.

Craig, P. S., Mitchell, G. F., Cruise, K. M., et al.,

“Hybridoma Antibody Immunoassay for the Detection of Parasitic Infection: Attempts To Produce an Immunodiagnostic Reagent for a Larval

Taeniid Cestode Infection,” Austra/. J. Exp. Biol.

Med. Sci. 58:339-350, 1980.

Crane, M. S. J., and Dvorak, J. A., “Vertebrate

Cells Express Protozoan Antigen After Hybridization,” Science 208:194-196, 1980.

Cross, G. A.M., “The Role of Molecular Biology

in Parasitology, ” Parasitology: A Global Perspective, K.S. Warren and J.Z. Bowers (eds. ) (New

York: Springer Verlag, 1983).

Cruickshank, J. K., and Mackenzie, C., “Immu-
270 ● Status of Biomedical Research and Related Technology for Tropical Diseases

nodiagnosis in Parasitic Disease, ” Br. Med. J.

283:1349-1350, 1981.

83. Cruise, K. M., Mitchell, G. F., Garcia, E. G., et al.,

“Hybridoma Antibody Immunoassay for the Detection of Parasitic Infection: Further Studies on

a Monoclinal Antibody With Immunodiagnostic Potential for Schistosomiasis japonica, ” Acta

Tropica 38:437-447, 1981.

84. Cruise, K. M., Mitchell, G. F., Garcia, E. G., et al.,

“Sj23, the Target Antigen in Schistosorna japonicum Adult Worms of an Immunodiagnostic Hybridoma Antibody,” Parasite Immunol. 5:37-45,

1983.

85. Cruise, K. M., Mitchell, G. F., Tapales, F. P., et

al., “Murine Hybridoma-Derived Antibodies Producing Circumoval Precipitation (COP) Reactions

With Eggs of Schistosoma japonicum, ” Austral.

J. Exp. Biol. Med. Sci. 59:503-514, 1981.

86. Curlin, G., Levine, R., Aziz, K. M. A., et al.,

Proc. Ilth Joint US-Japan Cholera Conf., New

Orleans, pp. 314-329 (Bethesda, MD: U.S. Department of Health, Education, and Welfare,

1976).

87. Current, W. L., “Cryptosporidiosis” (response to

letters to the editor), N. Eng. J. Med. 309:1326-27.

88. Cutting, J. W., “A Survey of Intestinal Parasitism in a Community on the Pan American Highway Route in Eastern Panama, ” PAHO BuIZ.

9(7):13-18, 1975.

89. Danforth, H. D., Campbell, G. H., Leef, M. F., et

al., “Production of Monoclinal Antibodies by

Hybridomas Sensitized to Sporozoites of Plasmodium berghei, ” J. ParasitoZ. 68:1029-1033,

1982.

90. Dean, A. G., Ching, Y, C., Williams, R. B., et al.,

“Test for Escherichia coli Enterotoxin Using Infant Mice: Application in a Study of Diarrhea in

Children in Honolulu,’r J. Infect. Dis. 125:407-11,

1972.

91. de Ibarra, A. A. L., Howard, J. G., and Snary, D.,

“Monoclinal Antibodies to Leishmania tropica

major: Specificities and Antigen Location, ”

Parasitology 85:523-531, 1982.

92. De Quadros, C. A., “More Effective Immunization, ” Proc. Roy. Soc. London 13 (209):111-118,

1980.

93. Des Moutis, I., Ouassi, A., Grzych, J. M., et al.,

“Onchocerca VOZVUIUS: Detection of Circulating

Antigen by Monoclinal Antibodies in Human

Onchocerciasis, ’’Am. J. Trop. Med. Hyg. 32:533-

542, 1983.

94. Diamond, J. M., “Taxonomy by Nucleotides, ”

Nature 305:17-18, 1983.

95. diRaddo, J., and Warden, W., “Innovation and

Availability in the United States of Drugs for

Tropical Diseases, ” Pharmaceuticals in Developing Countries (Washington, DC: National Academy of Sciences, 1979).

96. Dissous, C., Grzych, J. M., and Capron, A.,

“Schistosoma mansoni Surface Antigen Defined

by a Rat Monoclinal Ig2a,” J. Immune/.

129:2326-2328, 1982.

97. Dowries, B. A., and Ellner, D. P., “Comparison

of Sputum Counterimmunoelectrophoresis and

Culture in Diagnosis of Pneumococcal Pneumonia,” J. C’lin. Microbiol. 10:662-665, 1979.

98. Draper, C. C., “The Use of Counterimmunoelectrophoresis in Immunodiagnosis, ” Trans. Roy.

Soc. Trop. Med. Hyg. 70:93-97, 1976.

99. Dresden, M. H., Sung, C. K., and Deelder, A. M.,

“A Monoclinal Antibody From Infected Mice to

a Schistosoma mansoni Egg Proteinase, ” J. lmmuno]. 130:1-3, 1983.

100. Dutary, B. E., and LeDuc, J. W., “Transovarial

Transmission of Yellow Fever Virus by a Sylvatic

Vector, Haemagogus equinus,” Trans. Roy. Soc.

Trop. Med. Hyg. 75(1):128, 1981.

101, East African/British Medical Research Council

Study, “Controlled Clinical Trials of Five ShortCourse (4-Month) Chemotherapy Regimens in

Pulmonary Tuberculosis,” Am. Rev. Resp. Dis.

123:165-170, 1981.

102. Echeverria, P., Blacklow, N. R., and Sanford,

L. B., “A Prospective Study of Travelers’ Diarrhea

Among American Peace Corps Volunteers in Rural Thailand: Infections With Multiresistant Enteric Pathogens, ” Symposium on Cholera, Gifu,

Oct. 6-8, 1980; Proceedings of the 16th Joint Conference U.S.-Japan Cooperative Medical Science

Program ChoZera Panel, S. Kuwahara and Y.

Zinnaka (eds.) (Tokyo: Fuji Printing, 1980).

103. Echeverria, P., Seriwatana, J., Leksomboon, U.,

et al., “Identification by DNA Hybridization of

Enterotoxigenic Escherichia coli in Homes of Children With Diarrhea, ” L.ancet 1:63-66, 1984.

104. Eddy, G. A., and Peters, C. J., “The Extended

Horizons of Rift Valley Fever: Current and Projected Immunogens, ” New Developments With

Human and Veterinary Vaccines, A. Mizrahi

(cd. ) (New York: Alan R. Liss, Inc., 1980).

105. Edna McConnell Clark Foundation, Annual Report, 1981, New York, 1982.

106. Edna McConnell Clark Foundation, Annual Report, 1982, New York, 1983.

107. Elliott, R., “The Influence of Vector Behavior on

Malaria Transmission,” Am. J. Trop. Med. Hyg.

21:755-763, 1972.

108. Elliott, V., and Contacos, P., “A Profile of

Selected Biomedical Research Efforts Into Diseases of Major Public Health Importance to Peo-
References ● 271

109.

110.

111.

112.

113.

114<

115,

116.

117.

118.

119.

120.

121.

122.

123

ple in Developing Countries, ” typescript, prepared

for the U.S. Agency for International Development, Washington, DC, November 1982.

Ellis, J., Ozaki, L. S., Gwadz, R. W., et al., “Cloning and Expression in l?. coli of the Malarial

Sporozoite Surface Antigen Gene From Plasmodium knowlesi, ” Nature 302:536-38, 1983.

Erickson, J., Division Chief, Office of Health,

U.S. Agency for International Development, Washington, DC, personal communication, March

1984.

Esser, K. M., Schoenbechler, M. J., and Gingrich,

J. B., “~~panosomarhoc/esiense Blood Forms Express All Antigen Specificities Relevant to Protection Against Metacyclic (Insect Form) Challenge,” J. Immunol. 129:1715-1718, 1982.

Falkow, S., “Plasmids, Transposons and Gene

Cloning, ” Modern Genetic Concepts and Techniques in the Study of Parasites, F. Michal (cd. )

(Basel: Schwabe & Co. AG, 1981).

Fekety, R., “Recent Advances in Management of

Bacterial Diarrhea,” Rev. Infect. Dis. 5:246-247,

1983.

Findlay, G. M., Recent Advances in Chemotherapy, 3d cd., vol. 1 (Philadelphia: Blakiston, 1950).

Fisher, G. U,, Gordon, M. P., Lobel, H. O., et al.,

“Malaria in Soldiers Returning From Vietnam, ”

Am. J. Trop. Med. Hyg. 19:27-39, 1970.

Fitts, R., Diamond, M., Hamilton, K., et al.,

“DNA-DNA Hybridization Assay for Detection

of Salmonella spp. in Foods, ” App. Environ.

Microbiol. 46:1146-1151, 1983.

Flores, J,, Purcell, R. H., Perez, I. W. R. G., et al,,

“A Dot Hybridization Assay for Detection of

Rotavirus, ” Lancet 1:555-559, 1983.

Foege, W. H., “The Global Elimination of Measles, ” Publ. Hlth. Rep. 97:402-405, 1982.

Foster, S.0., “Participation of the Public in

Global Smallpox Eradication, ” PubJ. Mth. Rep.

93(2):147-149, 1978.

Fox, J. P., “Is a Rhinovirus Vaccine Possible?”

Am. J. Epidemiol. 103:345-354, 1976.

Fox, W., and Nunn, A. J., “The Cost of Antituberculous Drug Regimens, ” Am. Rev. Resp.

Dis. 120:503-509, 1979.

Franzen, L., Westin, G., Shabo, R., et al., “Analysis of Clinical Specimens by Hybridisation With

Probe Containing Repetitive DNA From Plasmodium falciparum,” Lancet 2:525-528, 1984.

Fulton, R. E., and Middleton, P. J., “Comparison

of Immunofluorescence and Isolation Techniques

in the Diagnosis of Respiratory Viral Infections

of Children, ” Infect. Immun. 10:92-101, 1974.

124.

125.

126,

127,

128,

129,

1304

131,

132,

133,

134,

135,

136<

137<

138,

Gaidamovich, S. Y., and Melnikova, E, E., “Passive Haemolysis-in-Gel With Togaviridae Arboviruses, ” Intervirology 13:16-20, 1980.

Gardner, P. S., and McQuillin, J., Rapid Virus

Diagnosis (London: Butterworth, Inc., 1980).

Germanier, R., “Vaccination Against Cholera

and Typhoid Fever, ” Behring Institute Mitteilungen 71:43-50, 1982.

Gilman, R. H., Hornick, R. B., Woodard, W. E.,

et al., “Evaluation of a UDP-glucose-4-epimeraseless Mutant of Salmonella typhi as a Live Oral

Vaccine,” J. Infect. Dis. 136:717-723, 1977.

Gilman, R. H., Terminel, M., Levine, M. M., et

al., “Comparison of Trimethoprim-Sulf amethoxazole and Arnoxicillin in Therapy of Chloramphenicol-Resistant and Chloramphenicol-Sensitive Typhoid Fever, ”J. Infect. Dis. 132:630-636,

1975.

Ginsberg, H. S., “Picornaviruses,” Microbiology,

3d cd., B.D. Davis, R. Dulbecco, H.N. Eisen, et

al. (eds. ) (New York: Harper & Row, 1980).

Gitter, B. D., and Damian, R. T., “Murine Alloantigen Acquisition by Schistosomula of Schistosoma mansoni: Further Evidence for the Presence

of K, D, and I Region Gene Products on the

Tegumental Surface, ” Parasite Immunology 4:383-

393, 1982.

Glass, R. I., Huq, I., Alim, A. R. M. A., et al.,

“Emergence of Multiply Antibiotic-Resistant

Vibrio cholerae in Bangladesh, ” J. Infect. Dis.

142:939-942, 1980.

Glover, D. M., Genetic Engineering: Cloning

DNA (London: Chapman & Hall, 1980).

Godal, T,, and Negasse, K., “Subclinical Infection in Leprosy, “ Br. Med. J. 3:557-559, 1973.

Godson, G. N., Ellis, J., Svec, R., et al., “Identification and Chemical Synthesis of a Tandemly

Repeated Immunogenic Region of Plasmodium

knowlesi Circumsporozoite Protein, ” Nature

305:29-33, 1983.

Goldsmith, M. F., “ ‘Imported’ Leprosy in the

U.S., ” J. A.M.A. 251(1):18, 1984.

Goodman, H., “Potential Applications of Immunology Research for the Control of Tropical Diseases, ” Behring Institute Mitteilungen 71:23-28,

1982.

Goodman, L. S., and Gilman, A., Pharmacological Basis of Therapeutics, 5th ed. (New York:

MacMillan Publishing, 1975).

Green, M. S., Kark, J. D., Witzum, E., et al., “Frozen Stored Leishmania tropica Vaccine: The Effects of Dose, Route of Administration and Storage on the Evaluation of the Clinical Lesion. Two
272 ● Status of Biomedical Research and Related Technology for Tropical Diseases

139,

140.

141.

142.

143.

144.

145.

146.

147.

148.

149.

150.

151.

152.

153

Field Trials in the Israel Defense Forces, ” Trans.

Roy. Soc. Trop. Med. Hyg. 77:152-159, 1983.

Greenblatt, C. L., Slutzky, G. M., de Ibarra,

A. A. L., et al., “Monoclinal Antibodies for Serotyping Leishmania Strains, ” J. CZin. MicrobioZ.

18:191-193, 1983.

Groves, M., U.S. Army Medical Research and

Development Command, Fort Detrick, MD, personal communication, March 1984.

Guerrant, R. L., Kirchoff, L, V., Shields, D. S., et

al., “Prospective Study of Diarrheal Illnesses in

Northeastern Brazil: Patterns of Disease, Nutritional Impact, Etiologies, and Risk Factors, ” J.

Infect. Dis. 148:986-997, 1983.

Gwadz, R. W., “Successful Immunization Against

the Sexual Stages of FYasrnodium gallinaceum, ”

Science 17:1150-1151, 1976.

Halstead, S. B., “Dengue Hemorrhagic Fever—A

Public Health Problem and a Field for Research, ”

Bull. W.H.0. 58(1):1-21, 1980.

Halstead, S. B., “WHO Fights Dengue Hemorrhagic Fever, ” WHO Chron. 36(2):65-67, 1982.

Handman, E., and Hocking, R. E., “Stage-Specific, Strain-Specific, and Cross-Reactive Antigens of L.eishmania Species Identified by Monoclinal Antibodies, ” Infect. Immun. 37:28-33,

1982.

Handman, E., and Remington, J. S., “Serological

and Irnrnunochemical Characterization of Monoclinal Antibodies to Toxoplasma gondii, ” Immurzo]ogy 40:579, 1980.

Hansman, D., and Bullen, M. M., “A Resistant

Pneumococcus” (letter), Lancet 2:264-265, 1967.

Harats, N,, Ackerman, A., and Shalit, M,, “Quinine-Related Hypoglycemia, ” A/. Eng. J. Med.

310:1331, 1984.

Harboe, M., Closs, O., Bjune, G., et al., “Mycobactefium leprae Specific Antibodies Detected by

Radioimmunoassay,“ Scand. J. Immunol. 7:111-

120, 1978.

Harrison, G., Mosquitoes, Malaria, and Man: A

History of the Hostilities Since 1880 (New York:

E.P. Dutton, 1978).

Henderson, D. A., “The Eradication of Smallpox, ” Scientific American 235(4):25-33, 1976.

Herder, F. R., Deputy Director for Health and

Population, U.S. Agency for International Development, Washington, DC, personal communication, March 1984.

Hewitt, J., Coates, A. R. M., Mitchison, D. A., et

al., “The Use of Murine Monoclinal Antibodies

Without Purification of Antigen in the Serodiagnosis of Tuberculosis, ” J. Immunol. Methods

55:205211, 1982.

154.

155.

156

157.

158.

159<

160.

161.

162.

163.

164.

165.

166.

167.

168.

Hierholzer, J. C., and Hirsch, M. S., “Croup and

Pneumonia in Human Infants Associated With

a New Strain of Respiratory Syncytial Virus, ” J.

Infect. Dis. 140:826-828, 1979.

Hill, H. R., and Matsen, J. M., “Enzyme-Linked

Immunosorbent Assay and Radioimmunoassay

in the Serologic Diagnosis of Infectious Diseases, ”

J. Infect. Dis. 147:258-262, 1983.

Hirumi, H., Doyle, J. J., and Hirumi, K., “African Trypanosomes: Cultivation of Animal-Infective Trypanosoma brucei in Vitro, ” Science

196:992-994, 1977.

Hoffman, D. B., The Strategic Plan: Schistosomiasis Research (New York: Edna McConnell Clark

Foundation, 1983).

Hollingdale, M. R., Leef, J. L., McCullough, M.,

et al., “In Vitro Cultivation of the Exoerythrocyt ic Stage of PZasmodium berghei From

Sporozoites, ” Science, 213:1021-1022, 1981.

Hommel, M., David, P. H., and Oligino, L. D.,

“Surface Alterations of Erythrocytes in Plasmodium falciparum Malaria: Antigenic Variation,

Antigenic Diversity and the Role of the Spleen, ”

J. Exp. Med. 157:1137-1148, 1983.

Hommel, M., David, P. H., Oligino, L. D., et al.,

“Expression of Strain-Specific Surface Antigens

on Plasmodium falciparum-Infected Erythrocytes,” Parasite Immunol. 4:409-419, 1982.

Hopkins, D., Centers for Disease Control, Atlanta,

GA, personal communication, August 1984.

Hornbeak, H., Tropical Medicine Officer, National Institute of Allergy and Infectious Diseases,

National Institutes of Health, Bethesda, MD, personal communication, March 1984.

Hoskins, T. W., Davies, R. R., Smith, A. J., et al.,

“Assessment of Inactivated Influenza A Vaccine

After Three Outbreaks of Influenza A at Christ’s

Hospital, ” Lancet 1:33-35, 1979.

Houba, V. (cd.), Immunological Investigation of

Tropical Parasitic Diseases (Edinburgh: Churchill

Livingstone, 1980).

Hubbard, W., “Introductory Comments for the

Panel on Problems and Constraints, ” Pharmaceuticals in Developing Countries (Washington, DC:

National Academy of Sciences, 1979).

Hudson, L., “Immunobiology of Trypanosoma

cruzi Infection and Chagas’ Disease, ” Trans. Roy.

Soc. Trop. Med. Hyg. 75:493-498, 1981.

Hull, B. P., Spence, L., Basset, D., et al., “The

Relative Importance of Rotavirus and Other

Pathogens in the Etiology of Gastroenteritis in

Trinidadian Children, ” Am. J. Trop. Med. Hyg.

31:142-148, 1982.

Hunter, G. W., Swartzwelder, J. C., and Clyde,
References ● 273

169.

170.

171.

172.

173.

174.

175.

176.

177.

178.

179.

180,

D. F., Tropical Medicine, 5th ed. (Philadelphia:

W.B. Saunders, 1976).

Iarotski, L. S., and Davis, A., “The Schistosomiasis Problem in the World: Results of a WHO

Questionnaire Survey, “ Bull. J“V.H.0, 99(1):115-

127, 1981.

Imaeda, T., Kirchheimer, W. F., and Barksdale,

L., “DNA Isolated From A4ycobacten”um leprae:

Genome Size, Base Ratio, and Homology With

Other Related Bacteria as Determined by Optical DNA-DNA Reassociation, ” J. Bacteriol.

150:414-417, 1982.

Jaffe, C. L., and McMahon-Pratt, D., “Monoclinal Antibodies Specific for Leishmania tropica: I. Characterization of Antigens Associated

With Stage- and Species-Specific Determinants, ”

J. Immunol. 131:1987-1993, 1983.

Jahrling, P. B., Peters, C. J., and Stephen, E. L.,

“Enhanced Treatment of Lassa Fever by Immune

Plasma Combined With Ribavirin in Cynomolgus

Monkeys,” J. Infect. Dis. 149:420-427, 1984.

Jelnes, J. E., “Simple Equipment for Enzyme Electrophoresis of Schistosoma, Z?iomphalaria, and

Bulinus Species, ” Zeitschrift fur Parasitenkunde

66:117-119, 1981.

Jelnes, J. E., “Enzyme Profiles of Biomphalaria

and BuJinus Species (Mollusca, Pulmonata ),”

Malacologia 22:45-47, 1982.

John, T.J. “Diagnosis and Epidemiology of Acute

Respiratory Infections in Children, ” typescript,

presented at Workshop on Acute Respiratory

Diseases Among Children of the World, Chapel

Hill, NC, May 18-19, 1983.

Jordan, P., and Rosenfield, P. L., “Schistosomiasis Control: Past, Present, and Future, ” Ann.

Rev. Publ. H7th. 4:311-334, 1983.

Kagan, I., “Standardization and Quality Control

of Immunodiagnostic Methods, ” Behring Institute

MitteiZungen 71:10-22, 1982.

Kaper, J. B., Lockman, H., Baldini, M. M., et al.,

“Recombinant Nontoxinogenic Vibrio cholerae

Strains as Attenuated Cholera Vaccine Candidates, ” Nature 308:655-658, 1984.

Kapikian, A. Z., Cline, W. L., Kim, H. W., et al.,

“Antigenic Relationships Among Five ReovirusLike (RVL) Agents by Complement Fixation (CF)

and Development of New Substitute CF Antigens

for the Human RVL Agent of Infantile Gastroenteritis, ” Proc. Soc. Exp. Biol. Med. 152:535-

539, 1976.

Kapikian, A. Z., Mitchell, R. H., Chanock, R. M.,

et al., “An Epidemiologic Study of Altered Clinical Reactivity to Respiratory Syncytial (RS) Virus Infection in Children Previously Vaccinated

181.

182.

183.

184.

185,

186<

187<

188.

189.

190.

191.

192.

With an Inactivated RS Virus Vaccine, ” Am. J.

Epidemiol. 89:405-421, 1969.

Kapikian, A, Z., Wyatt, R. G., and Levine, M. M,,

“Oral Administration of Human Rotavirus to

Volunteers: Induction of Illness and Correlates

to Resistance, ” J. Infect. Dis. 147:95-106, 1983.

Kapikian, A. Z., Wyatt, R. G., Levine, M. M., et

al., “Studies in Volunteers With Human Rotaviruses. International Symposium on Enteric Infections in Man and Animals: Standardization of

Immunological Procedures, Dublin, Ireland, ” Develop. Biol. Standard 53:209-218, 1983.

Kardjito, T., Handoyo, I., and Grange, J. M.,

“Diagnosis of Active Tuberculosis by Immunological Methods 1. The Effect of Tuberculin Reactivity and Previous BCG Vaccination on the Antibody Levels Determined by ELISA, ” Tubercle

63:269-274, 1982.

Kaul, A., Scott, R., Gallagher, M., et al., “Respiratory Syncytial Virus Infection: Rapid Diagnosis in Children by Use of Indirect Immunofluorescence, ” Am. J. Dis. Child. 132:1088-1090,

1978.

Keusch, G., “Shigellosis,” Critical Reviews in

Tropical Medicine, vol. 1, R.K. Chandra (cd. )

(New York: Plenum Press, 1982).

Kim, H. W., Canchola, J. G., Brandt, C. D., et al.,

“Respiratory Syncytial Virus Disease in Infants

Despite Prior Administration of Antigenic Inactivated Vaccine, ” Am. J. Epidemiol. 89:422-434,

1969.

Kobayashi, K., Kitaura, T., and Goto, N., “Studies on Multiple Drug-Resistant SalmoneJ]a typhi

Isolated From Two Independent Patients Treated

With Chloramphenicol, ” Microbiol. Immunol.

23:423426, 1979.

Kohbata, S., Takahashi, M., and Yabuuchi, E.,

“Lactose Fermenting, Multiple Drug-Resistant

Salmonella t~hi Strains Isolated From a Patient

With Postoperative Typhoid Fever, ” J. C/in.

Microbiol. 18:920, 1983.

Krupp, M. A., and Chatton, M. J., Current Medical Diagnosis and Treatment (Los Altos, CA:

Lange Medical Publications, 1984).

Kuberski, T. T., and Rosen, L., “A Simple Technique for the Detection of Dengue Antigen in

Mosquitoes by Immunofluorescence, ” Am. J.

Trop. Med. Hyg. 26:533-537, 1977.

Kurathong, S., Srichaikul, T., Isarangkra, P., et

al., “Exchange Transfusion in Cerebral Malaria

Complicated by Disseminated Intravascular Coadulation, ” Southeast Asian J. Trop. Med. Pub.

Hlth. 10:389-391, 1979.

Kutsuzawa, T., Konno, T., Suzuki, H., et al.,
274 ● Status of Biomedical Research and Related Technology for Tropical Diseases

“Isolation of Human Rotavirus Subgroups 1 and

2 in Cell Culture, ”J. C’/in. Microbio/. 16:727-730,

1982.

193. Lancet, “Epitaph for Global Malaria Eradication?” Lancet 2:15-16, 1975.

194. Lancet, “New Faces Among the Campylobacters, ” Lancet 2:662, 1983.

195. Laveissiere, C., Couret, D., and Challier, A.,

“Description and Design Details of a Biconical

Trap Used in the Control of Tsetse Flies Along

the Banks of Rivers and Streams, ” WHO/VBC/

79, 746, 1979.

196. Leary, J. J., Brigati, D. J., and Ward, D. C., “Rapid

and Sensitive Calorimetric Method for Visualizing Biotin-Labeled DNA Probes Hybridized to

DNA or RNA Immobilized on Nitrocellulose:

Bio-Blots,” Proc. Nat. Acad. Sci. (USA) 80:4045-

4049, 1983.

197. LeDuc, J. W., Chief, Department of Epidemiology, U.S. Army Medical Research Institute of Infectious Diseases, Fort Detrick, MD, personal

communication, August 1984.

198. Lemette, E. H., “Viral Respiratory Diseases: Vaccines and Antivirals, ” Bu21. W.H.0. 59:305-324,

1981.

199. Lepes, T., “Status of the Global Malaria Eradication Program—Progress So Far Made and Problems Encountered, ” Abstr. Ninth Int. Congr.

Trop. Med. Malaria 1:308-310, 1973.

200. Levine, M. M., “Immunity to Cholera as Evaluated in Volunteers, ” Cholera and Related Diarrheas, O. Ouchterlony and J. Holrngren (eds. )

(Basel: S, Karger, 1980).

201. Levine, M. M., Nalin, D. R., Craig, J. P., et al.,

“Immunity of Cholera in Man: Relative Role of

Antibacterial Versus Antitoxic Immunity, ”

Trans. Roy. Soc. Trop. Med. Hyg. 73:3-9, 1979.

202. Lewin, R., “Genetic Probes Become Ever

Sharper, ” Science 221:1167, 1983.

203. Lobel, H. O., and Kagan, I. G., “Seroepidemiology of Parasitic Diseases, ” Ann. Rev. Microbiol.

32:329-347, 1978.

204. Lobmann, M., Charlier, P., Delem, A., et al.,

“Challenge Experiments in Colostrum-Deprived

Piglets Previously Immunized With Human Type

2 and Bovine RIT 4237 Rotavirus Strains: Evidence of Homologous and Heterologous Protection, ” Abstracts of the Fifth International Congress of Virology, Strasbourg, France, August

1981.

205. Lobmann, M., Charlier, P., Delem, A., et al.,

“Cross-Protection Studies in Piglets Artificially

Infected With the Bovine Rotavirus Strain RIT

4237, and Challenged With Human Rotavirus

Type 2 and Type 3,” Proceedings of the International Symposium Recent Advances in Enten”c Infections, Brugge, Belgium, September 1981.

206. Lovborg, U., Monoclinal Antibodies: Production and Maintenance (London: William Heinemann Medical Books, 1982).

207. Luelmo, F., Pan American Health Organization,

Washington, DC, personal communication,

March 1984.

208. Lumsden, W. H. R., Kimber, C. D., Dukes, P., et

al., “Field Diagnosis of Sleeping Sickness in the

Ivory Coast. I. Comparison of the Miniature

Anion-Exchange/ Centrifugation Technique With

Other Protozoological Methods, ” Trans. Roy.

Soc. Trop. Med. Hyg. 75(2):242-250, 1981.

209. Lupton, H. W., Peters, C. J., and Eddy, G. A.,

“Rift Valley Fever: Global Spread or Global Control?” Proceedings of the 86th Annual Meeting

of the United States Animal Health Association,

Nashville, TN, 1982.

210. Mackenzie, D., “Leprosy: The Beginning of the

End,” New Scientist 102(1408):30-33, 1984.

211. Maiztequi, J. I., Fernandez, N. J., and deDamilano,

A. J., “Efficacy of Immune Plasma in Treatment

of Argentine Hemorrhagic Fever and Association Between Treatment and a Late Neurological Syndrome, ” Lancet 2:1216, 1979.

212. Mandell, G. L., Douglas, R. G., and Bennett, J.E.

(eds. ), principles and Practice of Infectious Diseases (New York: John Wiley & Sons, 1979).

213. Mansfield, J.M. (cd.), Parasitic Diseases, vol. 1

(New York: Marcel Dekker, Inc., 1981).

214. Markell, E. K., and Voge, M., Medical Parasitology (Philadelphia: W.B. Saunders, 1981).

215. Mata, L. J., “The Children of Santa Maria Cauque,”

A Prospective Field Study of Health and Growth

(Cambridge, MA: MIT Press, 1978).

216. Mata, L. J., Kronmal, R. A., and Villegas, H., “Diarrhea Disease: A Leading World Health Problem, ” Cholera and Related Diarrheas, O.

Ouchterlony and J. Holmgren (eds. ) (Basel: S.

Karger, 1980).

217. Maugh, T. H., “Leprosy Vaccine Trials To Begin

Soon,” Science 215:1083-1086, 1982.

218. McBride, J. S., “Monoclinal Antibodies and Applications to Parasitisms, ” Molecular Biology

of Parasites, J. Guardiola, L. Luzzato, and W.

Trager (eds. ) (New York: Raven Press, 1983).

219. McCormick, J., Centers for Disease Control,

Atlanta, GA, personal communication, 1984.

220. McLaren, D. J., “The Role of Eosinophils in Tropical Disease, ” Seminars in Hematology 19(2):100-

106, 1982.

221. McNamara, M. K., Ward, R. E., and Kohler, H.,
References • 275

222

223.

224.

225.

226,

227<

228<

229.

230.

231.

232.

233.

234.

“Monoclinal Idiotope Vaccine Against Str-eptococcus pneurnoniae Infection, ” %~ence 226:1325-

1326, 1984.

Mehra, V., Brennan, P. J., Rada, E., et al.,

“Lymphocyte Suppression in Leprosy Induced by

Unique M. leprae Glycolipid,” Nature 308:194-

197, 1984.

Mekalanos, J. J., Swartz, D, J., Pearson, G. D. N,,

et al., “Cholera Toxin Genes: Nucleotide Sequence Deletion Analysis and Vaccine Developmerit, ” Nature 306:551-557, 1983.

Melsom, R., Naafs, B., Marboe, M., et al., “Antibody Activity Against Mycobacterium leprae

Antigen 7 During the First Year of DDS Treatment in Lepromatous (BL-LL) Leprosy, ” Leprosy

Rev. 49:17-29, 1978,

Meyer, H. M., Jr., Hopps, H. E., Parkman, P. D.,

et al., “Review of Existing Vaccines for Influenza, ”

/. Clin. Pathol. 70:146-152, 1978.

Mhalu, F. S., Mmari, P. W., and Ijumba, J., “Rapid

Emergence of El Tor Vibrio cholerae Resistant to

Antimicrobial Agents During First Six Months of

Fourth Cholera Epidemic in Tanzania, ” Lancet

1:345-347, 1979.

Michelson, E. H., and Dubois, L., “An Isoenzyme

Marker Possibly Associated With the Susceptibility of Z3iomphalaria glabrata Populations to

Schistosoma mansoni, ” Acta Trop. 38:419-426,

1981.

Migasena, S,, “Hypoglycemia in Falciparum Malaria,” Ann. Trop. Med. Parasit. 77:323-324,

1983.

Miles, M. A., “The Epidemiology of South American Trypanosomiasis: Biochemical and Immunological Approaches and Their Relevance to

Control,” Trans. Roy. Soc. Trop. Med. Hyg.

77:5-23, 1983.

Miller, J. A., “Surprising Problems Crop Up in

Cholera Vaccine Work,” Science News, p. 263,

Apr. 28, 1984.

Miller, L. H., “Current Prospects and Problems

for a Malaria Vaccine,” J. Infect. Dis. 135:855-

864, 1977.

Miller, R. A., Dissanayake, S., and Buchanan,

T. M., “Development of an Enzyme-Linked Immunosorbent Assay Using Arabinomannan From

Mycobacterium smegmatis: A Potentially Useful

Screening Test for the Diagnosis of Incubating

Leprosy,” Am. J. Trop, Med. Hyg. 32:555-564,

1983.

Milstein, C., “Monoclinal Antibodies,” Scientific

American 243:66-74, 1980.

Mitchell, G. F., “Effecter Cells, Molecules and

Mechanisms in Host-Protective Immunity to Parasites, ” Immunology 38:209-223, 1979.

235.

236

237<

238.

239.

240.

241.

242.

243

244.

245.

246.

Mitchell, G. F., “Hybridoma-Derived Antibodies

and Immunodiagnosis of Parasitic Infection, ”

Monoclinal Antibodies and T-Cell Hybridomas,

G.J. Hammering, U. Hammering, and J.F. Kearney (eds. ) (Amsterdam: Elsevier-North Holland

Biomedical Press, 1981).

Mitchell, G. F., “Hybridomas in Immunoparasitology, ” Monoclona] Hybn”doma Antibodies:

Techniques and Applications, G.R. Hurrell (cd. )

(Boca Raton, FL: CRC Press, 1982).

Mitchell, G. F., and Cruise, K. M., “Monoclinal

Antiparasite Antibodies: A Shot-in-the-Arm for

Immunoparasitology,” Monoclinal Antibodies

and T-CelJ Hybridomas, G.J. Hammering, U.

Hammering, and J.F. Kearney (eds. ) (Amsterdam: Elsevier-North Holland Biomedical Press,

1981).

Mitchell, G. F., Cruise, K, M., Garcia, E. G., et al.,

“A Hybridoma-Derived Antibody With Immunodiagnostic Potential for Schistosomiasis Japonica,” Proc. Nat. Acad. Sci. (USA) 78:3165-3169,

1981.

Mitchell, G. F., Garcia, E. G., and Cruise, K. M.,

“Competitive Radioimmunoassays Using Hybridoma and Anti-Idiotype Antibodies in Identification of Antibody Responses to, and Antigens of, Schistosoma japon.icum,” Austral. J. Exp.

Biol. Med. Sci. 61:27-36, 1983.

Mitchell, G. F., Premier, R. R., Garcia, E. G., et

al., “Hybridoma Antibody-Based Competitive

ELISA in Schistosoma japonicum Infection, ” Am.

!. Trop. Med. Hyg. 32:114-117, 1983.

Mitchell, G. H., Butcher, G. A., and Cohen, S.,

“Merozoite Vaccination Against Plasmodium

knowlesi Malaria, ” Immunology 29:397, 1975.

Mitchell, G. H., Butcher, G. A., Richards, W. H. G.,

et al., “Merozoite Vaccination of Douroucouli

Monkeys Against Falciparum Malaria, ” Lancet

1:1335-1338, 1977.

Morley, D. C., “Why Do Children in the USA

Survive?” New Developments in Tropical Medicine 11, T.W. Simpson, G .T. Strickland, and

M.A. Mercer (eds. ) (Washington, DC: National

Council for International Health, 1983).

Morrow, J. F., “Recombinant DNA Techniques, ”

Methods in Enzymology 68:3-24, 1979.

Moseley, S. L., Echeverria, P., Seriwatana, J., et

al., “Identification of Enterotoxigenic Escherichia

coli by Colony Hybridization Using Three Enterotoxin Gene Probes, ” J. Infect. Dis. 145:863-

69, 1982.

Moseley, S. L., Huq, I., Alim, A. R. M. A., et al.,

“Detection of Enterotoxigenic Escherichia coli by

DNA Colony Hybridization, ” J. Infect. Dis.

142:892-898, 1980.
276 ● Status of Biomedical Research and Related Technology for Tropical Diseases

247. Mott, K. E., and Dixon, H., “Collaborative Study

on Antigens for Immunodiagnosis of Schistosomiasis,” Bull. W.H.O. 60:729-753, 1982.

248. Mufson, M. A., “Pneumococcal Infections, ”

J. A.M.A. 246:1942-1948, 1981.

249. Nalin, D. R., and Cash, R. A., “The Optimal Oral

Therapy Formula for Cholera and Cholera-Like

Diarrheas,” Proceedings of the Sixth Annual International Epidemiologic Association Scientific

Meeting (Belgrade: Savremena Administracya,

1971).

250. Nalin, D. R., Cash, R. A., Islam, R., et al., “Oral

Maintenance Therapy for Cholera in Adults, ”

Lancet 2:370-373, 1968.

251. Nardin, E. H., and Nussenzweig, R. S,, “StageSpecific Antigens on the Surface Membrane of

Sporozoites of Malaria Parasites,” Nature 274:55-

57, 1978.

252. National Academy of Sciences, Tropical Health:

A Report on a Study of Needs and Resources

(Washington, DC: National Academy Press, 1962).

253. National Academy of Sciences, Board on Science

and Technology for International Development,

Manpower Needs and Career Opportunities in

the Field Aspects of Vector Biology (Washington,

DC: National Academy Press, 1984).

254, National Research Council, Pest Control; An

Assessment of Present and Alternative Technologies, vol. 5 (Washington, DC: National Academy Press, 1976).

255. National Research Council, Board on Science and

Technology for International Development, “Grants

Approved Through January 1984, ” Washington,

DC, mimeo, 1984.

256, Nelson, G, S., “Issues in Filariasis—A Century of

Inquiry and a Century of Failure, ” Acta Tropica

38:197-204, 1981.

257. Nichols, B. L., and Soriano, H. A., “A Critique

of Oral Therapy of Dehydration Due to Diarrheal

Syndrome,” Am. J. C]in. Nutr. 30:1457-1472,

1977.

258. Nogueira, N., and Coura, J. R., “American Trypanosomiasis (Chagas’ Disease), ” Tropical and

GeographicaZMedicine, K. Warren and A. Mahmoud (eds. ) (New York: McGraw Hill, 1984).

259. Nogueira, N. C., ChapIan, S., Tydings, J. D., et

al., “Trypanosoma cruzi: Surface Antigens of

Blood and Culture Forms,”J. Exp. Med. 153:629-

639, 1981.

260. Noriki, H., “Evaluation of Toxoid Field Trial in

the Phillipines, ” Proceedings of the Izth Joint

U.S.-Japan Cholera Conference (Sapporo), H.

Fukmi and Y. Zinnaka (eds.) (Tokyo: Fuji, 1977).

261. Nussenzweig, R, S., Vandenberg, J., Most, H., et

al., “Protective Immunity Produced by the Injection of X-Irradiated Sporozoites of Plasmodium

berghei,” Nature 216:16@162, 1967.

262. Nussenzweig, R. S., Vandenberg, J., Spitalny, G.,

et al., “Sporozoite-Induced Immunity in Mammalian Malaria: A Review, ” Am. J. Trop. Med.

Hyg. 21:722-728, 1972.

263. Nutter, J. E., Chief, Office of Program Planning

and Evaluation, National Institute of Allergy and

Infectious Diseases, National Institutes of Health,

Bethesda, MD, personal communication, April

1984.

264. Pan American Health Organization, Acute Respiratory Infections in Children, Ref: RD 21/3

(Washington, DC: PAHO, 1982).

265. Pan American Health Organization, Health Conditions in the Americas: 1977’-1980, PAHO Scientific Publication No. 427 (Washington, DC:

PAHO, 1982).

266. Pan American Health Organization, Report of

the Director: Quadrennial 1978-81, Annual 1981,

Official Document No. 183 (Washington, DC:

PAHO, 1982).

267. Pappas, M. G., Hajkowski, R., and Hockmeyer,

W., “DOT-Enzyme-Linked Immunosorbent Assay (DOT-ELISA): A Microtechnique for the

Rapid Diagnosis of Visceral Leishmaniasis, ” J.

Immunol. Methods 64:205-214, 1983.

268. Paula, A. S. V., “Prevalencia, Morbidade e Mortalidade da doenca de Chagas em Minas Gerais,

Belo Horizonte, ” Monograph of ESMG, Minas

Gerais, Brazil, 1978.

269. Peacock, R., and Poynter, D., “Field Experience

With a Bovine Lungworm Vaccine, ” Vaccines

Against Parasites. Symposia of the British Societyfor Parasitology, vol. 18, A.E. R. Taylor and

R. Muller (eds. ) (Oxford: Blackwell Scientific

Publications, 1980).

270. Pearson, T. W., “Antigens of African Trypanosomes: Studies Using Monoclinal Antibodies, ”

Monoclinal Antibodies and T-Cell Hybridomas,

G.J. Hammering, U. Hammering, J.F. Kearney

(eds. ) (Amsterdam: Elsevier-North Holland Biomedical Press, 1981).

271, Pereira, M. S., “Problems in the Control of Epidemic Influenza by Vaccination,” Ann. Clin. Res.

14:220-224, 1982.

272. Perkins, F. T., “Vaccines Used in the WHO Expanded Programme on Immunization, ” Behring

Institute Mitteilungen 71:29-33, 1982.

273. Perlmannf P., “Immunology and Parasitic Diseases, ” Parasitology: A Global Perspective, K.S.
References ● 2 7 7

274.

275.

276.

277<

278

279,

280.

281,

282.

283.

284.

285,

286.

Warren and J.Z. Bowers (eds. ) (New York: Springer

Verlag, 1983).

Peters, W., Chemotherapy and Drug Resistance

in Malaria (London: Academic Press, 1970).

Peters, W., “Chemotherapy of Malaria, ” MaIan”a, J.P. Krier (cd. ) (New York: Academic Press,

1980).

Phillips, R. A., “Water and Electrolyte Losses in

Cholera, ” Fedn. F’roc. 23:705, 1964, as cited in

D.R. Nalin, R.A. Cash, R. Islam, et al., “Oral

Maintenance Therapy for Cholera in Adults, ”

L.ancet 2:370-373, 1968.

Pinder, M., and Hewett, R. S., “Monoclinal Antibodies Detect Antigenic Diversity in Theileria

parva Parasites, ” J. Immunol. 124:1000-1001,

1980.

Pitkanen, T., “Traveller’s Diarrhea Caused by

Campylobacter jejuni, ” Ann. Clin. Res. 14:111-

113, 1982.

Potocnjak, P., Yoshida, N., Nussenzweig, R. S.,

et al., “Monovalent Fragments (Fab) of Monoclinal Antibodies to a Sporozoite Surface Antigen (Pb 44) Protect Mice Against Malarial Infection, ” J. Exp. Med. 151:1504-1513, 1980.

Potocnjak, P., Zavala, F., Nussenzweig, R., et

al., “Inhibition of Idiotype-Anti-Idiotype Interaction for Detection of a Parasite Antigen: A New

Immunoassay, ” Science 215:1637-1639, 1982.

Powell, K. E., Meador, M. P., and Farer, L. S.,

“Recent Trends in Tuberculosis in Children, ”

J. A.M.A. 251:1289-1292, 1984.

Prager, D., Program Officer, MacArthur Foundation, Chicago, IL, personal communication,

March 1984.

Pratt, D. M., and David, J. R., “Monoclinal Antibodies That Distinguish Between New World

Species of Leishmania, ” Nature 291:581-583,

1981.

Prentice, M. A., Jordan, P., Bartholomew, R. K.,

et al., “Reduction in Transmission of Schistosoma

mansoni by a Four-Year Focal Mollusciciding

Programme Against Biomphalaria giabrata in

Saint Lucia, ” Trans. Roy. Soc. Trop. Med. Hyg.

75:789-798, 1981.

Preston, S. H., Keyfits, N., and Schoen, R.,

“Causes of Death: Life Tables for National Populations, ” Guidelines for Acute Respiratory Infections Research and Program Development

(Geneva: WHO, 1983).

Prince, G, A., Suffin, S. C., Prevar, D. A., et al.,

“Respiratory Syncytial Virus Infection in Owl

Monkeys: Viral Shedding, Immunological Response, and Associated Illness Caused by WildType Virus and Two Temperature-Sensitive Mutants, ” Infect. Immun. 26:1009-1013, 1979.

287.

288.

289.

290.

291.

292.

293.

294.

295.

296.

297.

298.

299.

300.

Puffer, R. R., and Griffith G. W., “Patterns of Urban Mortality: Report of the Interamerican Investigation of Mortality, ” PAHO Scientific Publication No. 151 (Washington, DC: Pan American

Health Organization, 1967).

Randolf, J. G., Budget Analyst, Financial Management Office, Centers for Disease Control,

U.S. Department of Health and Human Services,

Atlanta, GA, personal communication, April

1984.

Reeves, W. C., Dillman, L., Quiroz, E., et al.,

“Opportunities for Studies of Children’s Respiratory Infections in Panama, ” Pediatr. Res. 17:

1045-1049, 1983.

Rice-Ficht, A. C., Chen, K. K., and Donelson,

J. E., “Sequence Homologies Near the C-Termini

of the Variable Surface Glycoproteins of Trypanosoma brucei, ” Nature 294:53-57, 1981.

Richards, W. H. G., Mitchell, G. H., Butcher,

G. A., et al., “Merozoite Vaccination of Rhesus

Monkeys Against P2asmodium knowlesi: Immunity to Sporozoite (Mosquito-Transmitted) Challenge,” Parasitology 74:191-198, 1977.

Riley, I. D., and Douglas, R. M., “An Epidemiologic Approach to Pneumococcal Disease, ” Rev.

Infect. Dis. 3:233-245, 1981.

Rockefeller Foundation, Report to the Rockefeller Foundation Trustees on the Health Program,

New York, December 1982.

Ross, S., Controni, G., and Khan, W., “Resistance of Shigellae to Ampicillin and Other Antibiotics,” J. A.M.A. 221:45-47, 1972.

Rowe, D. S., “The Role of Monoclinal Antibody

Technology in Immunoparasitology, ” Immunol.

Today 1:30-33, 1980.

Rowe, D. S., and Hirumi, H. E., The in Vitro Culture of the Pathogens of Tropical Diseases, Proceedings of a Workshop, Nairobi, Kenya, Feb.

4-9, 1979 (Basel: Schwabe & Co. AG., 1980).

Ruiz-Palacios, G. M., “Etiologic Agents of Acute

Diarrhea: Bacterial and Parasitic, ” Acute Diarrhea: Its Nutritional Consequences in Children,

J.A. Bellanti (cd.) (New York: Nestle Vevey/Ramen

Press, 1983).

Sabin, A. B., “Immunization Against Measles by

Aerosol, ” Rev. Infect. Dis. 5:514-523, 1983.

Sabin, A. B., Fernandez de Castro, J., Flores

Arechiga, J., et al., “Clinical Trial of Inhaled Aerosol of Human Diploid and Chick Embryo Measles Vaccine, ” Lancet 2:604, 1982.

Sabin, A. B., Flores Arechiga, A., Fernandez de

Castro, J., et al., “Successful Immunization of

Children With and Without Maternal Antibody

by Aerosolized Measles Vaccine: I. Different Results With Undiluted Human Diploid Cell and
278 ● Status of Biomedical Research and Related Technology for Tropical Diseases

301.

302.

303.

304.

305.

306.

307.

308

309.

310.

311.

312.

313.

Chick Embryo Fibroblast Vaccines, ” J. A.h4.A.

249:2651-2662, 1983.

Sabin, A. B., Flores Arechiga, A., Fernandez de

Castro, J., et al., “Successful Immunization of Infants With and Without Maternal Antibody by

Aerosolized Measles Vaccine: II. Vaccine Comparisons and Evidence for Multiple Antibody Response,” J. A.M.A. 251:2363-2371, 1984.

Sack, D. A., and Sack, R. B., “Test for Enterotoxigenic Escherichia coh’ Using Y-1 Adrenal Cells in

Miniculture,” Infect. Immun. 11:334-36, 1975.

Sacks, D., and Sher, A., “Evidence That AntiIdiotype Induced Immunity to Experimental African Trypanosomiasis Is Genetically Restricted

and Requires Recognition of Combining SiteRelated Idiotypes, ” J. Immunol. 131:1511-1515,

1983.

Salk, J., “Future Prospects for Vaccination

Against Virus Diseases, ” Behring Institute Mittei]ungen 71:43-52, 1982.

Samples, J. R., and Buettner, H., “Ocular Infection by a Biological Insecticide, ” J. Infect. Dis.

148:614, 1983.

Sarett, L. H., The United States Pharmaceutical

Industry, Pharmaceuticals in Developing Countries (Washington, DC: National Academy Press,

1979).

Schauf, V., “East-West Efforts Key Into Leprosy

Research, ” J. A.M.A. 251(1):15-18, 1984.

Schechter, M., Flint, J. E., Voller, A., et al., “Purified Trypanosoma cruzi Specific Glycoprotein

for Discriminative Serological Diagnosis of South

American Trypanosomiasis (Chagas’ Disease), ”

Lancet 2:939-941, 1983.

Schiffler, R. J., Mansur, G. P., Navin, T. R., et al.,

“Indigenous Chagas’ Disease (American TrYpanosomiasis) in California, ’’J.A.M.A. 251(22):2983-

2984, 1984.

Schild, G. C., and Assaad, F., “Vaccines: The

Way Ahead,” World Health Forum 4:353-357,

1983.

Schmaljohn, A. L., Johnson, E. D., Dalrymple,

J. M., et al., “Non-Neutralizing Monoclinal Antibodies Can Prevent Lethal Alphavirus Encephalitis,” Nature 297:70-72, 1982.

Seriwatana, J., Echeverria, R., Escamilla, J., et

al., “Identification of Enterotoxigenic Escherichia

coli in Patients With Diarrhea in Asia With Three

Enterotoxigenic Gene Probes,” Infect. 1mmun.

42:152-155, 1983.

Shaw, P. K., Brodsky, R. E., Lyman, D. O., et al.,

“A Community Outbreak of Giardiasis With Evidence of Documented Transmission by a Munici314.

315.

316

317,

318,

319,

320.

321,

322,

323,

324,

325<

326

pal Water Supply,” Ann. Int. Med. 87:426-432,

1977.

Shinnick, T. M., Sutcliffe, J. G., Green, N., et al.,

“Synthetic Peptide Immunogens as Vaccines, ”

Ann. Rev. Microbiol. 37:425-446, 1983.

Simasathein, S., Duangmani, C., and Echeverria, P., “Haemophilus influenza Type B Resistant to Ampicillin and Chloramphenicol in an

Orphanage in Thailand, ” Lancet 2:1214-1217,

1980.

Smith, A., ‘The Cuban Experience, ” Lancet

2:616, 1983.

Smith, C. B., Purcell, R. H., Bellanti, J. A., et al.,

“Protective Effect of Antibody to Parainfluenza

Type Virus, ” N. Eng. J. Med. 275:1145-1152,

1966.

Smith, C. E., “Major Disease Problems in the

Developing World, ” Conference Proceedings:

Pharmaceuticals for Developing Countries (Washington, DC: Institute of Medicine, National

Academy of Sciences, 1979).

Smith, G. L., Murphy, B. R., and Moss, B., “Construction and Characterization of an Infectious

Vaccinia Virus Recombinant That Expresses the

Influenza Hemagglutinin Gene and Induces Resistance to Influenza Virus Infection in Hamsters, ”

Proc. Nat. Acad. Sci. (USA) 80:7155-7159, 1983.

Smith, M. A., Clegg, J, A., Snary, D., et al., “Passive Immunization of Mice Against Schistosoma

mansoni With an IgM Monoclinal Antibody, ”

Parasitology 84:83-91, 1982.

Smrkovski, L., Buck, R. L., ALcantara, A. K., et

al., “In Vitro Mefloquine Resistant Plasmodium

falciparum From the Philippines, ” L.ancet 2:322-

323, 1982.

Snary, D., “7’rypanosoma cruzi: Antigenic Invariance of the Cell Surface Glycoprotein, ” Exp.

Parasit. 49:68-77, 1980.

Snary, D., Ferguson, M. A., Scott, M. T., et al.,

“Cell Surface Antigens of Trypanosoma cruzi:

Use of Monoclinal Antibodies To Identify and

Isolate an Epimastigote Specific Glycoprotein, ”

Molec. Biochem. Parasitol. 3:343-356, 1981.

Snyder, J. D., and Merson, M. H., “The Magnitude of the Global Problem of Acute Diarrhoeal

Disease: A Review of Active Surveillance Data, ”

Bull. W.H. O. 60:605-613, 1982.

Soenarto, Y., Sebodo, T., Ridho, R., et al.,

“Acute Diarrhea and Rotavirus Infection in Newborn Babies and Children in Yogyakarta, Indonesia From June 1978 to June 1979, ” J. Clin.

lkficrobiol. 14:123-129, 1981.

Soulsby, E. J. L., “The Application of the Immune
References Ž 279

327

328.

329<

330,

331

332.

333.

334.

335.

336

337.

338,

Response in Protozoal Infections to Immunoprophylaxis in Man and Animals, ” Behring Institute

Mitteilungen 71:104-113, 1982.

Spencer, H. C., Kariuki, D. M., and Koech, D. K.,

“Chloroquine Resistance in Z%smodium falc@arum From Kenyan Infants, ” Am. J. Trop. Med.

Hyg. 32:922-925, 1983.

Spencer, H. C., Kipinger, T,, Agure, R., et al.,

“~lasmodium falciparum in Kisumu, Kenya ;

Differences in Sensitivity to Amodiaquine and

Chloroquine in Vitro,”J. Infect. Dis. 148:732-736,

1983,

Spielman, A., Department of Tropical Public

Health, Harvard School of Public Health, Boston, MA, personal communication, 1984.

Stead, W. W., and Dutt, A. K., “Chemotherapy

for Tuberculosis Today, ” Am. Rev. Z?esp. Dis.

125:94, 1982.

Stintzing, G,, Back, E., Tufveson, B., et al., “Seasonal Fluctuations in the Occurrence of Enterotoxigenic Bacteria and Rotavirus in Pediatric Diarrhea in Addis Ababa, ” Bull. W.H. O. 59:67-73,

1981.

Strand, M., McMillan, A., and Pan, X., “Schistosoma mansoni: Reactivity With Infected Human

Sera of Monoclinal Antibody Characterization

of a Glycoprotein in Different Developmental

Stages,” Exp. Parasit. 54:145-156, 1982.

Strickland, G. T., Hunter’s Tropica]Medicine, 6th

ed. (Philadelphia: W,B. Saunders, 1984).

Strickland, G. T., and Hunter, K. W., “The Use

of Immunopotentiators in Malaria, ” Int. J. Nucl.

Med. Bio]. 7:133-140, 1980.

Sundaram, K., and Castellino, J.B. (eds. ), “Nuclear Methodology and Techniques in the Study

of Parasitic Disease in Humans: Proceedings of

the International Atomic Energy Agency Advisory Group Meeting, ” Int. J. Nucl. Med. Biol.

7( Special Issue No. 2), 1980.

Taylor, D. W., and Butterworth, A. E., “Monoclinal Antibodies Against Surface Antigens of

Schistosomula of Schistosoma mansoni, ” F’arasito)ogy 84:65-82, 1982.

Taylor, J. O., Sachar, D. B., Kinzie, J. L., et al.,

Symposium on Cho~era, R.S. Gordon (cd.), Palo

Alto, CA, 1967, as cited in D.R. Nalin, R.A.

Cash, R. Islam, et al., “Oral Maintenance Therapy for Cholera in Adults, ” L.ancet 2:370-373,

1968.

Taylor, M. G., “Vaccination Against Trematodes, ” Vaccines Against Parasites. Symposia of

the British Society [or Parasitology, vol. 18,

A.E.R. Taylor and R. Muller (eds. ) (Oxford:

Blackwell Scientific Publications, 1980).

339<

340,

341

342.

343.

344.

345.

346.

347.

348.

349.

350.

351.

352.

Teixeira, A. R. L., “Immunoprophylaxis of Chagas’

Disease,” Advances Exp. Med. Biol. 93:243-280,

1977.

Tesh, R. B., “A Method for the Isolation and Identification of Dengue Viruses, Using Mosquito Cell

Cultures, ’’Am. J. Trop. Med. Hyg. 28:1053-1059,

1979.

Thakur, C. P., Kumar, M., Singh, S. K., et al.,

“Comparison of Regimens of Treatment With Sodium Stibogluconate in Kala-Azar, ” Br. Med. J.

228:895-897, 1984.

Thirumoorthi, M. C., and Dajani, A. A., “Comparison of Staphylococcal Coagglutination, Latex

Agglutination, and Counterimmunoe] ectrophoresis

for Bacterial Antigen Detection, ” J. C/in.

Microbiol. 9:28-32, 1979.

Trager, W,, and Jensen, J. B., “Human Malaria

Parasites in Continuous Culture, ” Science

193:673-675, 1976.

Trager, W., and Jensen, J. B., “Cultivation of

Erythrocytic Stages, “ Bu]l. W.H. O. 55(2-3):363-

365, 1977.

Tugwell, P., and Greenwood, B. M., “Pneumococcal Antigen in Lobar Pneumonia, ” J. Clin.

Path. 28:118-123, 1975.

Tuow, J., Langendijk, E. M. J., Stoner, G. L., et

al., “Humoral Immunity in Leprosy: Immunoglobulin G and M Antibody Responses to Mycobacterium leprae in Relation to Various Disease

Patterns,” Infect. Immune]. 36:885-892, 1982.

Tyrrell, D. A., Schild, G. C., Dowle, W. R., et al.,

“Development and Use of Influenza Vaccines, ”

Bull. W.H. O. 59:165-173, 1981.

Ubell, R. N., “High-Tech Medicine in the Caribbean. 25 Years of Cuban Health Care,” N. Eng.

J. Med. 309:1471-1472, 1983.

U.N. Development Program/World Bank/WHO,

Special Programme for Research and Training in

Tropical Diseases, Report of the IMMLEP Subcommittee Meeting on the Planning of Leprosy

Vaccine Trials, Geneva, Feb. 12-14, 1980, TDR/

IMMLEP(SUB-TRIALS) /80.3 (Geneva: WHO,

1980).

U.N. Development Program/World Bank/WHO,

Special Programme for Research and Training in

Tropical Diseases, Sixth Report on Steering Committee Meetings of the IMMLEP Scientific Working Group, TDR/IMMLEP/81.l (Geneva: WHO,

1981).

U.N. Development Program/World Bank/WHO,

Special Programme for Research and Training in

Tropical Diseases, Facts and Figures, No. 6, Summary Tables (Geneva: WHO, January 1982).

U.N. Development Program/World Bank/WHO,
280 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Special Programme for Research and Training in

Tropical Diseases, Country Profi]e–USA (Geneva: WHO, 1983).

353. U.N. Development Program/World Bank/WHO,

Special Programme for Research and Training in

Tropical Diseases, Sixth Programme Report: I

July 1981-31 December 2982 (Geneva: WHO,

1983).

354. U.S. Agency for International Development, Office of the Science Advisor, “Program in Science

and Technology Cooperation, ” mimeo, Washington, DC, July 11, 1983.

355. U.S. Congress, General Accounting Office, Issues

Affecting Continuation of United States Funding

of the Gorgas A4emori”al Institute, GAO/NSIAD83-38 (Washington, DC: GAO, 1983).

356. U.S. Congress, Office of Technology Assessment,

A Review of Selected Federal Vaccine andlmmunization Policies Based on Case Studies of Pneumococcal Vaca”ne, OTA-H-96 (Washington, DC:

U.S. Government Printing Office, September

1979).

3s7. U.S. Congress, Office of Technology Assessment,

Impacts of Applied Genetics: Micro-Organisms,

Plants, and AnimaZs, OTA-HR-132 (Washington,

DC: U.S. Government Printing Office, April

1981).

358. U.S. Congress, Office of Technology Assessment,

Cost-Effectiveness Analysis of Inactivated Influenza Vaccine, OTA-H-152 (Washington, DC:

U.S. Government Printing Office, December

1981).

359. U.S. Congress, Office of Technology Assessment,

World Population and Fertility Planning Technologies: The Next 20 Years, OTA-HR-157

(Washington, DC: U.S. Government Printing Office, February 1982).

360. U.S. Congress, Office of Technology Assessment,

Quality and Relevance of Research and Related

Activities at the Gorgas Memorial Laboratory—

A Technical Memorandum, OTA-TM-H-18 (Washington, DC: U.S. Government Printing Office,

August 1983).

361. U.S. Congress, Office of Technology Assessment,

Commercial Biotechnology: An International

Analysis, OTA-BA-218 (Washington, DC: U.S.

Government Printing Office, January 1984).

362. U.S. Congress, Office of Technology Assessment,

Medical Technology and Costs of the Medicare

Program, OTA-H-227 (Washington, DC: U.S.

Government Printing Office, July 1984).

363. U.S. Congress, Office of Technology Assessment,

Africa Tomorrow: Issues in Technology, Agriculture, and U.S. Foreign Aid—A Technical

Memorandum, OTA-BP-F-31 (Washington, DC:

U.S. Government Printing Office, December

1984).

364. U.S. Department of Commerce, U.S. Direct Investment Abroad (Washington, DC: DOC, 1977).

365. U.S. Department of Defense, Washington Headquarters Services, Directorate for Information,

Operations, and Reports, Selected Manpower

Statistics for FY 1983 (Washington, DC: DOD,

1983).

366. U.S. Department of Health and Human Services,

Centers for Disease Control, “Surveillance Survey: Measles Encephalitis—United States, 1962-

1979, ” Morbidity and Mortality Weekly Report

30(29):362-364, 1981.

367. U.S. Department of Health and Human Services,

Centers for Disease Control, Annual Summary

1981: Morbidity and Mortality Weekly Report,

HHS Publication No. (CDC) 82-8241, October

1982.

368. U.S. Department of Health and Human Services,

Centers for Disease Control, “African Trypanosomiasis, ” Morbidity and Mortality Weekly Report 32(8):112-113, 1983.

369. U.S. Department of Health and Human Services,

Centers for Disease Control, “Tuberculosis and

Leprosy Control in Developing Countries,” Morbidity and Mortality Weekly Report 32(18):234-

241, 1983.

370. U.S. Department of Health and Human Services,

Centers for Disease Control, “Surveillance of

Acute Respiratory Infections: Meeting of the

Technical Advisory Group.” Morbidity and Morta]ity WeekZy Report 32(21):275-281, 1983.

371. U.S. Department of Health and Human Services,

Centers for Disease Control, “Diarrheal Diseases

Control Program: Rotavirus Diarrhea, ” Morbidity and Mortality Weekly Report 32(24):311-317,

1983.

372, U.S. Department of Health and Human Services,

Centers for Disease Control, “Refugees, ’’Annual

Summary, 1982: Morbidity and Mortality Weekly

Report, HHS Publication No. (CDC) 84-8241,

December 1983.

373. U.S. Department of Health and Human Services,

Centers for Disease Control, “Yellow Fever Vaccine, ” Morbidity and Mortality Weekly Report

32(52):679-688, 1984.

374. U.S. Department of Health and Human Services,

Centers for Disease Control, “ACIP: Prevention

and Control of Influenza, ” Morbidity and Mortality Weekly Report 33(19):253-266, 1984.
References ● 281

375. U.S. Department of Health and Human Services,

Centers for Disease Control, “Update: Acquired

Immunodeficiency Syndrome (AIDS) —United

States,” Morbidity and Mortality Weekly Report

33(47):661-664, 1984.

376. U.S. Department of Health and Human Services,

Centers for Disease Control, Health Information

for International Travel, HHS Publication No.

(CDC) 840-8280 (Washington, DC: U.S. Government printing Office, 1984).

377. U.S. Department of Health and Human Services,

Centers for Disease Control, Atlanta, GA, personal communication, 1984.

378. U.S. Department of Health and Human Services,

National Center for Health Statistics, Health—

United States, 1983, HHS Publication No. (PHS)

84-1232 (Washington, DC: U.S. Government

Printing Office, 1983).

379. U.S. Department of Health and Human Services,

National Institutes of Health, “International Cooperation by the National Institute of Allergy and

Infectious Diseases, FY 1981, ” mimeo, Bethesda,

MD, no date.

380. U.S. Department of Health and Human Services,

National Institutes of Health, “International Cooperation by the National Institute of Allergy and

Infectious Diseases, FY 1982, ” mimeo, Bethesda,

MD, no date.

381, U.S. Department of Health and Human Services,

National Institutes of Health, “International Cooperation by the National Institute of Allergy and

Infectious Diseases, FY 1983, ” mimeo, Bethesda,

MD, no date.

382. U.S. Department of Health and Human Services,

National Institutes of Health, IVIH Data Book

1982 (Washington, DC: U.S. Government Printing Office, 1982).

383. U.S. Department of Health and Human Services,

National Institutes of Health, personal communication, Mar, 22, 1984.

384. Vandenberg, J. P., Nussenzweig, R. S., and Most,

H., “Protective Immunity Produced by Injections

of X-Irradiated Sporozoites of Plasmodium berghei.

V. In Vitro Effects of Immune Serum on Sporozoites, ” A4ilit. Med. 134 (suppl. ):1183-1190, 1969.

385. Vesikari, T., Isolauri, E., Delem, A., et al., “Immunogenicity and Safety of Live Oral Attenuated

Bovine Rotavirus Vaccine Strain RIT 4237 in

Adults and Young Children, ” L.ancet 2:807-811,

1983.

386. Vesikari, T., Isolauri, E., D’Hondt, E., et al.,

“Protection of Infants Against Rotavirus Diarrhea

by RIT 4237 Attenuated Bovine Rotavirus Strain

Vaccine, ” Lancet 1:977-980, 1984.

387. Vischer, E., and OberhoIzer, R., “The European

Pharmaceutical Industry, ” Pharmaceuticals for

Developing Countries (Washington, DC: National Academy of Sciences, 1979).

388. Voller, A., and De Savigny, D., “Diagnostic Serology of Tropical Parasitic Diseases, ” ]. 1rnmunol. Meth. 46:1-29, 1981.

389. Voller, A., Bidwell, D. E., and Bartlett, A., The

Enzyme Linked Immunosorbent Assay (ELISA)

(Guernsey, England: Dynatech Europe, 1979).

390. Waddy, B. B., “Research Into the Health Problems of Manmade Lakes, With Special Reference

to Africa, ” Trans. Roy. Soc. Trop. Med. Hyg.

69(1):39-50, 1975.

391. Wahdan, M. H., Serie, C., Germanier, R., et al.,

“A Controlled Field Trial of Live Oral Typhoid

Vaccine Ty21a, ” Bu]i. W.H.O. 58:469, 1980.

392. Walsh, J. A., and Warren, K. S., “Selective Primary Care: An Interim Strategy for Disease Control in Developing Countries, ” N. Eng, J. Med.

301:967-974, 1979.

393. Walton, B. C., Harper, J., and Neal, R. A., “Effectiveness of Allopurinol Against Leishmania

braziliensis panamensis in Aotus trivergatus, ”

Am. J. Trop. Med. Hyg. 32(1):46-50, 1983.

394. Ward, J., and Koornhof, H., “Antibiotic Resistant Pneumococci, in Current Clinical Topics in

Infectious Diseases (New York: McGraw-Hill,

1980).

395. Warren, D. A., Looareesuwan, S., Warren, M. J.,

et al., “Dexamethasone Proves Deleterious in Cerebral Malaria, ” N. Eng. J. Med. 306:313-319,

1982.

396. Warren, K. S., and Mahmoud, A. A. F., Tropical

and Geographical Medicine (New York: McGraw

Hill, 1984).

397. Watson, J. D., Tooze, J., and Kurtz, D. T., Recombinant DNA: A Short Course (New York:

Scientific American Books, 1983).

398. Wayne, L. G., “Microbiology of Tubercle Bacilli,”

Am. Rev. Respirat. Dis. 125:31-41, 1982.

399. Webbe, G., and Lambert, J. D. H., “Plants That

Kill Snails and Prospects for Disease Control, ”

Nature 302:754, 1983.

400. Weller, T. H., “Training Grant Programs in Tropical Medicine, ” Am. J. Trop. Med. Hyg. 23(4):

821-827, 1974.

401. White, N. J., Looareesuwan, S., Warren, D. A.,

et al., “Quinine Pharmacokinetics and Toxicity

in Cerebral and Uncomplicated Falciparum Malaria,” Am. J. Med. 73:564-572, 1982.

402. White, N. J., Looareesuwan, S., Warren, D. A.,

et al., “Quinine Loading Dose in Cerebral Malaria,” Am. J. Trop. Med. Hyg. 32:1-5, 1983.
282 ● Status of Biomedical Research and Related Technology for Tropical Diseases

403.

404.

405.

406.

407.

408.

409.

410.

411.

412.

413.

414.

415.

416.

417.

White, N. J., Warren, D. A., Chanthavanich, P.,

et al., “Severe Hypoglycemia and Hyperinsulinemia in Falciparum Malaria, ” IV. Eng. -1. Med.

309:61-66, 1983.

Wiernick, A., Jarstrand, C., and Tunvall, G.,

“The Value of Immunoelectroosmophoresis (IEOP)

for Etiological Diagnosis of Acute Respiratory

Tract Infections Due to Pneumococci and Mycoplasma Pneumoniae, ” Scand. J. Infect. Dis.

10:173-176, 1978.

Wignall, F. S., U.S. Navy, Gorgas Memorial Laboratory, Panama City, Panama, personal communication, July 1983.

Wilcocks, C., and Manson-Bahr, P. E. C., Manson’s Tropical Diseases (London: Bailliere Tindall, 1972).

Winkelstein, W., Jr., “Epidemiological Considerations Underlying the Allocation of Health and

Disease Care Resources, ” Int. J. Epidemiol.

1(1):69-74, 1972.

Winter, P., U.S. Department of Defense, Washington, DC, personal communication, July 1984.

Wirth, D. F., and Pratt, D. M., “Rapid Identification of Leishmania Species by Specific Hybridization of Kinetoplast DNA in Cutaneous Lesions, ”

Proc. Nat. Acad. Sci. (USA) 79:6999-7003, 1982.

Wood, J. N., Hudson, L., Jessell, T. M., et al., “A

Monoclinal Antibody Defining Antigenic Determinants on Subpopulations of Mammalian Neurones and T~panosoma cruzi Parasites, ” Nature

296:34-38, 1982.

Woode, G. N., Bridger, J. C., Jones, J. M., et al.,

“Morphological and Antigenic Relationships Between Viruses (Rotaviruses) From Acute Gastroenteritis of Children, Calves, Piglets, Mice and

Foals, ” Infect. Immune]. 14:804-810, 1976.

World Bank, Health: Sector Policy Paper (Washington, DC: World Bank, 1980).

World Development Forum, “Hooray for the

Plumbers,” World Development Forum 2(10),

May 31, 1984.

World Health Organization, “Tuberculosis Prevention Trial, ” Bull. W.H. O. 57:819-827, 1979.

World Health Organization, BCG Vaccination

Policies, Tech. Report Series, No. 651 (Geneva:

WHO, 1980).

World Health Organization, Proposed Program

Budget for the Financial Period 1982-1983 (Geneva: WHO, 1980).

World Health Organization, Vaccination Against

Tuberculosis, Tech. Report Series, No. 651 (Geneva: WHO, 1980).

418.

419.

420.

421.

422.

423.

424.

425.

426.

427.

428,

429,

430<

431

432

World Health Organization, Rapid Laboratory

Techniques for the Diagnosis of Viral Infections,

Technical Report Series, No. 661 (Geneva:

WHO, 1981).

World Health Organization, Oral E’nteric Vaccines, Report of a WHO Working Group, EURO

Reports and Studies, No. 63 (Copenhagen: WHO,

1982).

World Health Organization, Report of the Second Meeting of the Scientific Working Group on

Viral Diarrheas: Microbiology, Epidemiology,

Immunology and Vaccine Development (Geneva:

WHO, 1982).

World Health Organization, “Rapid Laboratory

Viral Diagnosis, “ Bull. W.H.O. 61:43-44, 1983.

World Health Organization, “Development of

Malaria Vaccines: Memorandum From a USAID/

WHO Meeting, ” Bull. W.H.O. 61:81-92, 1983.

World Health Organization, “Live Oral Typhoid

Vaccine,” Bull. W.H.O. 61:251, 1983.

World Health Organization, “Rotavirus Diarrhea,” Bull. W.H.O. 61:251-253, 1983.

World Health Organization, Program for Control of Diarrheal Diseases, DCC/TAG/84.2A

(Geneva: WHO, 1984).

World Health Organization, Programme for

Control of Diarrheal Diseases, “Live Oral Typhoid Vaccine Ty21a—Report of the Meeting of

a Subcommittee of the Scientific Working Group

on Bacterial Enteric Infections, ” unpublished document, WHO/CDD/82.6 (Geneva: WHO, 1982).

World Health Organization, Programme for

Control of Diarrheal Diseases, Third Programme

Report, 1981-1982, WHO/CDD/83.8 (Geneva:

WHO, 1983).

Wright, W. H., Forty Years of Tropical Medicine

Research: A History of the Gorgas Memorial Institute of Tropical and Preventive Medicine, Inc.

and The Gorgas Memorial Laboratory (Washington, DC: Reese Press, 1970).

Wyatt, R. G., James, S. D., Bohl, E. H., et al.,

“Human Rotavirus Type 2: Cultivation in Vitro,”

Science 207:189-191, 1980.

Wyler, D. J., “Malaria-Resurgence, Resistance,

and Research, ” N. Eng. J. Med. 308:875-878,

1983.

Yoshida, N., Nussenzweig, R. S., Potocnjak, P.,

et al., “Hybridoma Produces Protective Antibodies Directed Against the Sporozoite Stage of Malaria Parasite, ” Science 207:71-73, 1980.

Yoshida, N., Potocnjak, P., Nussenzweig, V., et

al., “Biosynthesis of Pb44, the Protective Anti-
References ● 283

433.

434.

435.

gen of Sporozoites of Plasmodium berghei, ” J.

Exp. Med. 154:1225-1236, 1981.

Young, D. B., and Buchanan, T. M., “A Serological Test for Leprosy With a Glycolipid Specific 436.

for Mycobacterium leprae, ” Science 221:1057-

1059, 1983.

Zavala, F., Cochrane, A. H., Nardin, E. H., et al.,

“Circumsporozoite Proteins of Malaria Parasites 437.

Contain a Single Immunodeficient Region With

Two or More Identical Epitomes, ” J. Exp. Med.

157:1947-1957, 1983.

Zavala, F., Gwadz, R. W., Collins, F. H., et al.,

“Monoclinal Antibodies to Circumsporozoite

Proteins Identify the Species of Malaria Parasite

in Infected Mosquitoes, ” Nature 299:737-738,

1982.

Zodda, D. M., and Phillips, S. M., “Monoclinal

Antibody-Mediated Protection Against Schistosoma mansoni Infection in Mice, ” J. Immunol.

129:2326-2328, 1982.

Zola, H., and Brooks, D., “Techniques for the

Production and Characterization of Monoclinal

Hybridoma Antibodies, ” Monoclinal Hybrjdoma Antibodies: Techniques and Applications,

G.R. Hurrell (cd. ) (Boca Raton, FL: CRC Press,

1982).

Index

Acute respiratory infections (ARIs), 15-16

bronchiolitis, 146

bronchitis, 15

causative agents

adenoviruses, 146, 175

bacteria, 89-90, 147-148

Bordetella pertussis, 90, 148, 175, 194

coronaviruses, 147, 175

coxsackieviruses, 147

echoviruses, 147

fungi, 196

Haemophilus influenza, 175, 176

Mycoplasma, 147-148, 176

respiratory syncytial virus (RSV), 146, 174

rhinoviruses, 147, 175

Streptococcus, 175

viruses, 89, 145

control, 90-91

diagnostic techniques, 173-176

diphtheria, 15

immunology, 145, 146

incidence, 89

influenza, 15, 145, 174-176

in the United States, 7, 22

measles, 15, 89, 147

natural history, 88-89

parainfluenza, 146, 174

pneumonia, 15, 146, 147, 148

prevalence, 89

serology, 176

treatment, 90, 173, 193-196

type of infection, 60

vaccines, 15, 90, 145, 146, 147, 148

whooping cough, 15, 148, 175

Adenovirus, 146, 175

Aedes aegyptir 8, 93

Aeromonas, 192

African sleeping sickness, 13, 67-71. See also

Trypanosomiasis

control, 70-71

diagnosis, 165-166

incidence, 69

natural history, 69

symptoms, 69

treatment, 70, 186

vaccine, 138-140

variant surface glycoprotein (VSG), 138

African trypanosomiasis, See African sleeping

sickness

Agency for International Development (AID), 17,

18

Bureau of Science and Technology, 52

funding by, 44-45, 52, 236

oral dehydration therapy, 216, 217

Program in Science and Technology Cooperation (PSTC), 44

Agglutination assays, 156, 160

AID. See Agency for International Development

Allopurinol, 48, 78

Ambrosia maritima, 123

Amebiasis, 193

American Society for Tropical Medicine and

Hygiene, 31

American trypanosomiasis. See Chagas’ disease

Ancylostoma, 85

A. duodenale, 193

Anopheles mosquito, 12, 61, 227

A. minimus flavirostris, 116

A. nuneztovari, 116

Antibiotics, 87, 181-182, 184

Antibodies, 108-112, 129-149, 155-161, 228. see

also Immunity; Immunology

Antigens, 108-112, 129-149, 154, 158-161, 162, 170

malaria, 228, 238

Antimalarial drugs. See Drugs; Therapeutic agents,

malaria

Antimicrobial. See Antibiotics

Antiviral drugs. See Drugs

Aotus monkey, 135, 136

Arboviral and related infections, 16, 59, 91-95.

causative agents, 60, 91

Arenaviridae, 91-92

Bunyaviridae, 91

Togaviridae, 91

chikungunya, 16

dengue fever, 16, 93, 95, 149

diagnostic techniques, 176-177

encephalitis, 16, 59, 148-149

hemorrhagic fever, 16, 149

Lassa fever, 94

natural history, 91

Oropouche fever, 16, 93

Rift Valley fever, 93

symptoms, 92

vaccine, 16

viral encephalitides, 16

yellow fever, 16, 59, 92-93, 100, 148

ARIs. See Acute respiratory infections

Ascaris, 85

A. lumbricoides (roundworm), 88

ASTMH. See American Society for Tropical Medicine and Hygiene

Bacillary dysentery, 144

Bacillus Calmette-Guerin vaccine. See BCG vaccine

Bacillus sphaericus, 119

287
288 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Bacillus thuringiensis israeliensis (BTI), 119

Bacteria, 15, 60, 86-87. See also Acute respiratory

infections; Arboviral and related viral infections; Diarrheal and enteric diseases

antibiotic resistant, 86-87

infections, 172-173, 175-176

pathogenic, 119

Balantidium coli, 193

BCG (Bacillus Calmette-Guerin) vaccine, 15, 85,

141, 142, 172

Biological insecticides, 119-120

Biomphalaria, 123

Bionomics, 115-116

Biotechnology. See Deoxyribonucleic acid; Hybridoma; Monoclinal antibodies

Blackflies, 80, 119

Blood cells, See Erythrocytes; Eosinophils

Board on Science and Technology for International

Development (BOSTID), 44-45, 52, 55

Bordetella pertussis. See Whooping cough

BOSTID. See Board on Science and Technology

for International Development

Bronchiolitis, 146

Brugia malayi, 14, 78-79, 140, 168

BTI. See Bacillus thuringiensis israeliensis

Campylobacter, 85, 86, 88, 173, 192

Capillaria philippinensis, 193

Card agglutination test, 165

CATT. See Card agglutination test

CDC. See Centers for Disease Control

CDD. See World Health Organization, Program

for Control of Diarrheal Diseases

Cellognost test, 165

Centers for Disease Control (CDC), 17

funding by, 42-43, 44, 237

Chagas’ disease, 13, 71-74

animal model, 139

control, 72-74, 102

diagnosis, 155-156, 166-167

incidence, 72

in the United States, 8

natural history, 71

symptoms, 71-72

treatment, 72, 187

vaccine, 138-140

Childhood diseases, 89-90

Chlamydia, 60, 90

Chloroquine, 12, 63-64

Cholera, 144, 172, 173. See also Vibrio cholerae;

Diarrhea] and enteric diseases

CIE. See Counterimmunoelectrophoresis

Circumoval precipitin test (COPT), 157

Clostridium

Cl. difficile, 192

Cl. perf.ringes, 192

Coagglutination (COA) test, 160, 175

Coelomomyces, 120

Computed tomography, 154

Control of disease. See Disease control

Copper sulfate, 123

COPT. See Circumoval precipitin test

Corynebacterium diphtheria, 90, 194

Counterimmunoelectrophoresis (CIE), 156

Cryptosporidium, 85, 88, 193

Cuba, 27-28

Cuzicinomyces clavorsporus, 119-120

Cytogenetic studies, 121

DDT (dichloro-diphenyl-trichloroethane), 10, 63,

116, 117, 121

resistance to, 65, 117, 226

substitutes, 117

toxicity, 117

Delta-toxin, 119

Dengue fever, 8, 16, 93

Deoxyribonucleic acid (DNA), 87, 88, 105, 106,

107-108, 132, 136, 139, 140, 141, 143, 154

diagnostic technologies, 154, 162, 164, 166-167,

168, 171, 172, 173

immunization technologies, 132, 136, 139, 140,

141, 143

malaria, 231, 234-235

Department of Defense (DOD), 17, 18

funding, 45-46, 52, 54-55, 237

Department of Health and Human Services

(DHHS), 17, 18, 41. See also National Institute of Allergy and Infectious Diseases;

National Institutes of Health; Centers for

Disease Control; Fogarty International

Center for Advanced Study in the Health

Sciences

Department of Health, Education, and Welfare.

See Department of Health and Human

Services

DHF (dengue hemorrhagic fever). See Dengue

fever

DHHS. See Department of Health and Human

Services

Diagnostic technologies, 153-154

acute respiratory infections (ARIs), 173-176

antibiotics, 173, 174

arboviral and related viral infections, 176-177

bacterial infections, 172-173, 175-176

Bordetella pertussis, 175

deoxyribonucleic acid (DNA), 162, 163, 164,

166-167, 168, 171, 172, 173, 226

diarrheal and enteric diseases, 172-173

direct examination, 155

electron microscopy, 172
Index ● 2 8 9

filariasis, 168-169

genetic tools, 161-162

Haemophilus influenza, 175, 176

influenza, 174

leishmaniasis, 167-168

leprosy, 169-170

malaria, 163-164

monoclinal antibodies, 161-162, 163-164, 165,

166, 168, 169, 170, 171, 177

Mycoplasma pneumonia, 175

parasites, 164, 169

precision, 153

predictive value, 1s3

schistosomiasis, 156, 164-165

sensitivity, 153

serology, 155-161, 163, 170, 172, 174, 175, 176,

177

specificity, 153

sporozoite diagnosis, 163-164

Streptococcus, 175

trypanosomiasis, 165-167

tuberculosis, 170-172

viral infections, 172

Diagnostic technologies, conventional diagnostic

techniques, 153-161

acute respiratory infections (ARIs), 172-173,

173-174

arboviral infections, 176-177

diarrheal and enteric diseases, 172

filariasis, 168

leishmaniasis, 167

leprosy, 169

malaria, 163

schistosomiasis, 164

trypanosomiasis, 165

tuberculosis, 170-171

Diagnostic technologies, tests and assays

agglutination assays, 156, 160

“Biken” gel diffusion test, 173

card agglutination test (CATT), 165

Cellognost test, 165

circumoval precipitin tests (COPT), 1s7

coagglutination (COA) test, 160, 175

complement fixation (CF) test, 155-156, 163,

166, 174, 175, 176

computed tomography, 154

counterimmunoelectrophoresis (CIE) test, 156,

172, 175, 176

cross-reactivity, 153

direct agglutination test, 156

ELISA (enzyme-linked immunosorbent assay),

157, 163, 165, 168, 170, 171, 172, 173,

174, 175, 176, 177

hemagglutination inhibition (HI) test, 156, 174,

176, 177

immunoelectrophoresis, 156

indirect fluorescent antibody (IFA) test, 157,

163, 170, 172, 174, 175, 176, 177

indirect hemagglutination (IHA) test, 156, 163,

175

labeled immunodiagnostic reagent assays,

156-157, 160, 163

Lancefield precipitation test, 175

latex agglutination (LA) test, 175, 176

lepromin (Mitsuda) test, 169

lymphocyte transformation test, 169-170

Mantoux test, 171

neutralization test, 156

nucleic acid hybridization probes, 162

Quellung test, 176

radioimmunoassay (RIA), 157, 170, 172, 175

reverse passive hemagglutination (RPHA), 160,

176

thin-layer immunoassay (TIA), 157

Tine test, 171

tuberculin skin text, 85

Vollmer patch test, 171

xenodiagnosis, 154

X-rays, 171

Diarrheal and enteric diseases, 15, 59, 85-88,

142-144, 201

ad libitum treatment, 210-21I

amebiasis, 193

bacterial infections, 143, 203

causes, 15, 60, 86-88, 201, 203

cholera, 144, 172, 173, 191, 210

coliform infection, 143

dehydration, 201, 212-213, 215

diagnostic techniques, 172-173

electrolytes (salts), 202, 207-210, 213-214, 215

Escherichia coli. See E. coli

incidence, 85-86, 205

infant diarrhea, 201-224

in the United States, 7, 205

malnutrition, 204

natural history, 85-86

Rotavirus, 85, 86, 143, 172, 203

Salmonella, 15, 85, 87, 143, 172, 173, 192

shigellosis, 191-192

treatment, 11, 15, 85, 190-193, 205, 215-216. See

also Oral dehydration therapy

typhoid fever, 144

vaccine, 142-144

viral infections, 143, 191, 203

Dichlorobromophenol, 123

Dichloro-diphenyl-trichloroethane. See DDT
290 • Status of Biomedical Research and Related Technology for Tropical Diseases

DNA. See Deoxyribonucleic acid

DOD. See Department of Defense

DPT vaccine, 148

Drugs, 12, 20. See also Therapeutic agents

Duffy antigens, 228

Echinococcus granulosus (tapeworm), 196

Elephantiasis, 14

ELISA. See Enzyme-linked immunosorbent assay

Encephalitis, 8

Entamoeba, 85, 88, 192-193

Enteric diseases. See Diarrheal and enteric diseases

Enzyme-linked immunosorbent assay, 78, 85, 121.

See also Diagnostic technologies, tests and

assays, ELISA

Eosinophils, 137-138

Equine encephalitis, 8

Erythrocytes, 61, 65, 136, 233-235

Escherichia

E. coli, 85, 86, 87, 136, 143, 148, 172-173, 192,

195, 203

E. histolytica, 88

Expanded Program on Immunization (EPI), 16, 105

FDA. See Food and Drug Administration

Fertility rate, 204

FIC. See Fogarty International Center for Advanced Study in the Health Sciences

Filaria

Brugia malayi, 78-80, 140, 168, 188

Loa lea, 188

Onchocerca volvulus, 78, 140, 168, 169, 188

Wuchereria bancrofti, 14, 78-80, 116, 140, 168,

187

Filariasis, 14

African eyeworm disease, 188

blackflies, 80, 119

causative agent, 60, 78, 80

diagnostic techniques, 168-169

incidence, 80, 81

natural history, 78

prevalence, 80

symptoms, 80

treatment, 188

Fogarty International Center for Advanced Study

in the Health Sciences (FIC), 43-44

Food and Drug Administration (FDA), 21

Foundations, 46-47

Edna McConnell Clark Foundation, 46-47, 54,

165

MacArthur Foundation, 47

Rockefeller Foundation, 47, 49, 54, 101-102, 237

Funding, 37-ss. See also Research

distribution, 52-54

recipients of, 52, 53-55

sources, 37-49, 236-238

types of research, 49-51

Fungi, 119-120, 196

Gambusia, 120

Gastroenteritis. See Diarrheal and enteric diseases

Giardia, 85, 88, 192-193

Giardiasis. See Giardia

Glossina. See Tsetse flies,

Gorgas Memorial Institute of Tropical and Preventive Medicine (GMI), 43

Growth hormones, 120

Haemophilus influenza, 90, 148, 175, 176, 194,

195

Hansen’s disease. See Leprosy

Helminths, 29, 60, 193, 196

Hemagglutination inhibition (HI) test, 156

Hookworm, 193

Hybridoma, 109-111, 139, 233

Hypernatremia, 212-213, 214

Hyponatremia, 212-213, 214

IHA. See Indirect hemagglutination test.

ILRAD. See International Laboratory for Research

on Animal Diseases

Immunity, 129-131, 228-230

Immunization, 52, 103-105, 131. See also Expanded Program on Immunization

Immunization technologies, 129-149

acute respiratory infections (ARIs), 144-148

arboviral and related infections, 148-149

diarrheal and enteric diseases, 142-144

filariasis, 140

influenza, 145

leishmaniasis, 140

leprosy, 140-141

malaria, 134-137

schistosomiasis, 137-138

trypanosomiasis, 138-140

tuberculosis, 142

vaccines, 131-149

Immunoelectrophoresis, 156

Immunoglobulins, 155

Immunology, 137, 145, 155-161, 228-230. see also

Immunization; Immunization technologies

Indirect hemagglutination (IHA) test, 156

Infant mortality, 6, 7, 201-205

Influenza, 174

Insecticides, 13, 117-120. See also DDT, resistance

to

International Health Research Act of 1960 (Public

Law 86-610), 23, 30-31
Index ● 2 9 1

International Laboratory for Research on Animal

Diseases (ILRAD), 138

1PM. See Vector control, integrated pest

management

Isoopora belli, 193

Kala azar. See Leishmaniasis

Kebsiella pneumoniae, 90, 195

LA. See Latex agglutination test

Labeled immunodiagnostic reagent

Latex agglutination (LA) test, 160,

Leishmania, 14, 74, 140

L. braziliensis, 74, 167, 187

L. donovani, 167, 187

L. mexicana, 74, 167, 187

L. tropica, 74, 167, 187

life cycle, 76

Leishmaniasis, 14, 74-78

causative agent, 60

control, 78

cutaneous leishmaniasis, 187

diagnostic techniques, 167-168

immunology, 78, 140

incidence, 75, 77-78

kala azar, 14, 74, 167, 187

assays, 156, 160

175, 176

mucocutaneous leishmaniasis, 167, 187

natural history, 74, 76

symptoms, 74, 140

treatment, 48, 78, 187

type of infection, 60

vaccination, 140

visceral leishmaniasis, 14, 74, 167, 187

Leprosy (Hansen’s disease), 14, 80-84, 140

causative agent, 60, 80

diagnostic techniques, 169-170

immunology, 80, 82, 84, 140-141

incidence, 82

Iepromatous leprosy, 80, 140, 170, 188

lepromin test, 169

Mitsuda test, 169

Mycobacterium leprae, 80, 170, 188. See also

Mycobacterium

symptoms, 80, 140

treatment, 83, 188-189

tuberculoid leprosy, 80, 170, 188

vaccine, 140-141

Lower respiratory tract infections (LRTIs), 89-90,

194

Lymphocyte transformation test, 169-170

MAbs. See Monoclinal antibodies

Malaria, 12. See also Plasmodium

antibody, 229-230

antigens, 229-230, 233, 234

antimalarial, 183-185

causative agent, 60, 116, 225, 226-227

control, 10, 61-64, 115-116, 118-119, 120, 226

diagnostic techniques, 163-164

immunology, 64, 134-137, 228-230

incidence, 61, 63-64

in the United States, 8

Malaria Eradication Campaign, 118-119

mosquitoes, 10, 115-116, 120, 226, 227

natural history, 59-62, 225, 227, 233, 234, 235,

236

prevalence, 61

research, 22, 48, 65, 134-137, 225, 230-233

symptoms, 226

treatment, 48, 182-185

tropical splenomegaly syndrome (TSS), 137

vaccine, 4, 11, 20, 65, 121, 134-137225-245

Malnutrition, 7

Marisa cornuarietis, 123

Medication. See Drugs; Therapeutic agents

Melioidosis, 195

Metarhizium anisopliae, 120

Molluscicides, 122-123

Monoclinal antibodies (MAbs)

diagnostic technologies, 154, 155, 163, 165, 166,

168, 171

disease control, 106-112, 121

immunization technologies, 131, 132, 136, 138,

139, 140, 142, 149

malaria, 226, 232-233, 234, 235

Mosquitoes, 10, 16, 115-116, 120, 227

Mycobacterium

M. avium-intracellulare, 171

M. leprae, 14, 80, 141, 169, 170, 171, 188

M. smegmatis, 172

M. tuberculosis, 15, 84, 171, 189

Mycoplasma, 15, 60, 90, 175, 195

M. pneumonia, 148, 176

Nagana, 138

NAS. See National Academy of Sciences

National Academy of Sciences (NAS), 31-33

National Institute of Allergy and Infectious Diseases (NIAID), 17, 18

funding, 41-42, 52-54, 236-237

National Institutes of Health (NIH), 17. See also

Fogarty International Center for Advanced

Study in the Health Sciences

National Research Council (NRC), 31

Neutralization test, 156

NIAID. See National Institute of Allergy and Infectious Diseases

Niclosamide, 123

NIH. See National Institutes of Health

NRC. See National Research Council
292 ● Status of Biomedical Research and Related Technology for Tropical Diseases

Onchocerca volvulus, 14, 60, 78, 140, 168, 169

Oncomelania, 123

Oral dehydration therapy (ORT), 3-4, 11-12, 15,

85, 190-191, 201-224. See also Diarrheal

and enteric diseases

advantages, 207

clinical aspects of treatment, 211-213

composition of solutions, 213-216

dehydration, prevention of, 215

economy, 218

effectiveness, 216, 217-218

limitations, 213

vomiting, 212

Organotin compounds, 123

Orphan Drug Act of 1983 (Public Law 97-414),

20-21

ORT. See Oral dehydration therapy

PAHO. See Pan American Health Organization

Pan American Health Organization (PAHO),

216-217

Paragonimus westermani (fluke), 196

Pertussis. See Whooping cough

Pharmaceutical Manufacturers’ Association (PMA),

47

Phytolacca dodecandra, 123

Plague. See Yersinia pestis

Plasmodium, 226-227. See also Malaria

antimalarial, 183-185, 225-245

culture, 135

life cycle, 61, 134, 182, 183, 227

P.

P.

P.

P,

P.

P.

P.

P.

P.

P.

P.

berghei, 134, 135, 136, 227

chabaudi, 135, 136, 227

falciparum, 61, 63, 134, 135, 136, 137, 164,

182, 183-184, 225, 226, 227, 228, 234, 238

ga]linaceum, 226-227

knowlesi, 134, 135, 136, 227, 234

lophurae, 227

malariae, 61, 134, 182, 184, 226, 227

ovale, 61, 134, 182, 184, 227

vinckei, 227

vivax, 8, 59, 134, 136, 182, 184, 226, 227,

228, 238

yoelli, 134, 135, 227, 234

Pleisiomonas, 192

PMA. See Pharmaceutical Manufacturers’ Association

Pneumococci, 175. See also Streptococcus

Pneumocystis carinii, 195

Pneumonia, 90, 146, 148, 194, 195

Praziquantel, 12, 13. See also Schistosomiasis

Presbytis

P. cristata, 140

P. melalophos, 140

Program in Science and Technology Cooperation

(PSTC). See Agency for International Development

Protozoa, 13, 14, 15, 29, 59, 60, 85, 192-193,

195-196

Pseudomonas pseudomallei, 90, 195

PSTC. See Agency for International Development,

Program in Science and Technology

Cooperation

Public Health Service Act, 30

Radioimmunoassay (RIA), 121, 157

Reduviid bug (kissing bug), 8, 121

Respiratory infections. See Acute respiratory infections

Respiratory syncytial virus (RSV), 146, 174

Reverse passive hemagglutination (RPHA) test, 160

Rhinovirus, 147, 175

RIA. See Radioimmunosassay

Ribavirin, 12

Rifampicin, 85

Rotavirus, 85, 86, 143, 172, 203

RPHA. See Reverse passive hemagglutination test

RSV. See Respiratory syncytial virus

Salmonella, 15, 85, 87, 143, 172, 173

S. typhi, 87, 144, 172, 192

Sandflies, 14, 16, 75

Sarcocystis, 193

Schelia, 102

Schistosoma, 12-13, 65

S. haematobium, 65, 68, 137, 164, 186

S. japonicum, 65, 68, 137, 164, 186, 196

S. mansoni, 65, 68, 101, 137, 138, 164, 186, 196

S. mekongi, 65

Schistosomiasis, 12-13, 101-102

causative agent, 60, 65

control, 67, 101-102, 122-123

diagnostic techniques, 156, 164-165

incidence, 67-68

life cycle, 65-67, 137

natural history, 65

St. Lucia experiment, 101-102

treatment, 185-186

vaccine, 138

Serology, 154, 158-161, 170, 172, 174. see also Diagnostic technologies, tests, and assays

Shigella, 15, 85, 86, 87, 142, 143, 144, 172

S. dysenteriae, 87, 144

S. flexneri, 87, 144

S. sonnei, 87, 144

Shigellosis, 144

Simulium. See Blackflies

Skin tests, 154
Index ● 2 9 3

Smallpox, 29, 100

Snails, 101-102, 122-123

Special Program for Research and Training in

Tropical Diseases (TDR), 17, 18, 22,

37-38, 169

funding, 42, 44, 45-46, 50-51, 52, 237

research and development, 39

Sporozoite diagnosis, 163

Staphylococcus, 192

S. aureus, 90, 160, 194, 195

Streptococcus

S. pneumonia, 90, 111, 147-148, 175, 194-195

S. pyogenes, 90, 148, 175, 194, 195

Strongyloides stercoralis (threadworms), 193

Taeneorhynchus, 120

TDR. See Special Program for Research and Training in Tropical Diseases

Tests. See Diagnostic technologies

Therapy. See Oral dehydration therapy; Therapeutic technologies

Therapeutic technologies

acute respiratory infections (ARIs), 193-196

antibiotics, 181-182, 184

arboviral and related viral infections, 196

diarrheal and enteric diseases, 191-193

filariasis, 188

leishmaniasis, 187-188

leprosy, 188-189

malaria, 182-185

resistance to, 182, 183, 190, 191-192

schistosomiasis, 185-186

symptomatic treatment, 181

trypanosomiasis, 186-187

tuberculosis, 189-190

Thin-layer immunoassay (TIA), 157, 159

Treatment. See Therapeutic technologies

Trematode, 12, 186. See also Schistosoma

Trichuris trichura (whipworms), 193

Tropical splenomegaly syndrome (TSS), 137

Trypanosoma, 13, 67

T. brucei, 13, 67, 69, 139, 166

T. brucei gambiense, 13, 69-70, 138, 165, 166,

186

T. brucei rhodesiense, 13, 69-70, 111, 138, 165,

166, 186

T. cruzi, 13, 67, 71-74, 102, 138, 139, 166, 187

Trypanosomiasis, 13, 67-71. See also African

sleeping sickness; Chagas’ disease

causative agent, 60, 67

control, 71-72, 102

diagnostic techniques, 165-167

incidence, 69, 71

in the United States, 8

natural history, 69, 71

symptoms, 69, 71-72

treatment, 13, 70, 74, 186-187

vaccine, 138-140

Tsetse flies, 13, 69, 121-122

TSS. See Tropical splenomegaly syndrome

Tuberculosis, 15, 84-85

control, 84, 142

diagnostic techniques, 170-172

incidence, 84

in the United States, 7-8

Mycobacterium tuberculosis, 84, 142, 189. See

also Mycobacterium

natural history, 84

symptoms, 84

treatment, 84, 189-190

tubercle bacilli, 170, 171

tuberculin skin test, 85

vaccine (BCG vaccine), 85, 142, 172

Typhoid fever, 144, 172

UNICEF. See U.N. International Children’s Emergency Fund

U.N. International Children’s Emergency Fund

(UNICEF), 216, 217

Upper respiratory tract infections (URTIS), 89-90,

194

Vaccination, 10-11, 52

Vaccines, 103-105, 131-149

acute respiratory infections, 90, 144-148

arboviral infections, 59, 148-149

development, 20, 104, 136-137

diarrheal and enteric diseases, 142-144

filariasis, 140

Ieishmaniasis, 140

leprosy, 141

malaria, 65, 134-137, 225-245

schistosomiasis, 138

trypanosomiasis, 138-140

tuberculosis, 15, 85, 140, 142, 172

Variant surface glycoprotein (VSG), 69, 71, 72,

138

Vector control, 10, 63, 70, 103

arthropods, 117-121

biological control methods, 120, 123

bionomics, 115-116, 124

filariasis, 14

fungi, 119-120

growth hormones, 120

identification of disease organisms, 121

insecticides, 13, 117-120

integrated pest management (1PM), 123, 125

leishmaniasis, 14
294 ● Status of Biomedical Research and Related Technology for Tropical Diseases

malaria, 12

molluscicides, 122

mosquitoes, 10, 16, 115-116, 120, 227

predators, 120

reduviid bugs, 122

research needs, 124

schistosomiasis, 12-13

snails, 122-123

species complexes, 120

trypanosomiasis, 13

tsetse flies, 121-122

Venezuelan equine encephalitis. See Equine encephalitis

Vibrio cholerae, 86, 87-88, 143, 144, 172, 173, 191

Virus, 15, 60, 86, 89, 172, 194. see also Acute

respirator infections; Arboviral and related viral infections; Diarrhea] and enteric

diseases

VSG. See Variant surface glycoprotein

WHO. See World Health Organization

Whooping cough (pertussis), 90, 148, 175, 194

World Health Organization (WHO), 18-19, 28, 38,

40

funding by, 38, 40, 50, 52, 237 See also Expanded Program on Immunization (EPI)

Malaria Eradication Campaign, 118-119, 236

Program for Control of Diarrheal Diseases

(CDD), 38, 40, 52, 216

Program in Acute Respiratory Infections, 38

research, 22

Wuchereria bancrofti, 14, 78-80, 116, 140, 168

Xenodiagnosis, 154

X-rays, 154, 171

Yellow fever, 16, 59, 92-93

Yersinia pestis, 85, 88, 192, 195