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Volume 2, Number 2, April-June 1996

EMERGING INFECTIOUS DISEASES, VOLUME 2, NUMBER 2

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Editors

Editor
Joseph E. McDade, Ph.D., National Center for Infectious Diseases,
Centers for Disease Control and Prevention (CDC), Atlanta, Georgia, USA

Perspectives Editor
Stephen S. Morse, Ph.D.,The Rockefeller University,
New York, New York, USA

Synopses Editor
Phillip J. Baker, Ph.D., Division of Microbiology and Infectious Diseases,
National Institute of Allergy and Infectious Diseases,
National Institutes of Health, Bethesda, Maryland, USA

Dispatches Editor
Stephen Ostroff, M.D., National Center for Infectious Diseases,
CDC, Atlanta, Georgia, USA

Managing Editor
Polyxeni Potter, M.A., National Center for Infectious Diseases,
CDC, Atlanta, Georgia, USA


Liaison Representatives

Anthony I. Adams, M.D., Chief Medical Adviser,
Commonwealth Department of Human Services and Health,
Canberra, Australia

David Brandling-Bennett, M.D., Director,
Division of Communicable Diseases, Pan American Health Organization,
World Health Organization,
Washington, D.C., USA

Gail Cassell, Ph.D.,
Liaison to American Society for Microbiology,
University of Alabama at Birmingham,
Birmingham, Alabama, USA

Richard A. Goodman, M.D., M.P.H., Editor, MMWR,
CDC, Atlanta, Georgia, USA

William Hueston, D.V.M., Ph.D, Acting Leader,
Center for Animal Health Monitoring, Centers for Epidemiology and Animal
Health, Veterinary Services, Animal and Plant Health Inspection Service, U.S. Dept. of Agriculture
Fort Collins, Colorado, USA

James LeDuc, Ph.D., Advisor for Arboviral Diseases,
Division of Communicable Diseases, World Health Organization,
Geneva, Switzerland

Joseph Losos, M.D., Director General,
Laboratory Center for Disease Control,
Ontario, Canada

Gerald L. Mandell, M.D., Liaison to Infectious Diseases Society of America,
University of Virginia Medical Center,
Charlottesville, Virginia, USA

Phillip P. Mortimer, M.D., Director, Virus Reference Division,
Central Public Health Laboratory,
London, United Kingdom

Robert Shope, M.D., Director, Yale Arbovirus Research Unit,
Yale University School of Medicine,
New Haven, Connecticut, USA

Natalya B. Sipachova, M.D., Ph.D.
Scientific Editor
Russian Republic Information & Analytic Centre,
Moscow, Russia

Bonnie Smoak, M.D., Chief, Dept of Epidemiology, Division of Preventive
Medicine, Walter Reed Army Institute of Research,
Washington, D.C., USA

Robert Swanepoel, B.V.Sc., Ph.D., Head, Special Pathogens Unit,
National Institute for Virology,
Sandrinham 2131, South Africa

Roberto Tapia-Conyer, M.D.
Director General de Epidemiologa,
Direccin General de Epidemiologa,
Secretara de Salud,
Mxico


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Infectious Diseases.

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Maria T. Brito
Anne D. Mather, M.A.
Carol D. Snarey, M.A.

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Rita M. Furman, M.S.

Electronic Distribution
Carol Y. Crawford
David L. Smith
Cheryl G. Shapiro, M.S.

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The goals of Emerging Infectious Diseases (EID) are to promote the
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Begin each of the following sections on a new page and in this order: title
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not to exceed 40. Use of additional subheadings in the main body of the
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on experimental procedures should immediately follow the body of the text.
Photographs and illustrations are encouraged. Provide a short abstract (150
words) and a brief biographical sketch.

Dispatches: Provide brief updates on trends in infectious diseases or
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should be in a letter to the editor format and should not be divided into
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Table of Contents
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EMERGING INFECTIOUS DISEASES
Volume 2, Number 2, April-June 1996
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Perspectives

   * Globalization, International Law, and Emerging Infectious Diseases,
     David P. Fidler
   * On Epidemiology and Geographic Information Systems: A Review and
     Discussion of Future Directions, Keith C. Clarke, Sara L. McLafferty,
     and Barbara J. Tempalski
   * The Evolution and Maintenance of Virulence in Microparasites, Bruce R.
     Levin
   * The Infectious Diseases Impact Statement: A Mechanism for Addressing
     Emerging Infectious Diseases, Edward McSweegan

Synopsis

   * Emerging Disease Issues and Fungal Pathogens Associated with HIV
     Infection, Neil M. Ampel

Dispatches

   * An Outbreak of Ross River Virus Disease in Southwestern Australia,
     Michael Lindsay, Nidia Oliveira, Eva Jasinska, Cheryl Johansen, Sue
     Harrington, A.E Wright, and David Smith
   * Invasive Penicillin-Resistant Pneumococcal Infections: A Prevalence
     and Historical Cohort Study, Crystal B. Kronenberger, Richard E.
     Hoffman, Dennis C. Lezotte, and William M. Marine
   * Nosocomial Transmission of Multidrug-Resistant Mycobacterium
     tuberculosis in Spain, John V. Rulln, Dionisio Herrera, Rosa Cano,
     Victoria Moreno, Pere Godoy, Enrique F. Peir, Juan Castell, Consuelo
     Ibaez, Arturo Ortega, Leopoldo Snchez Agudo, and Francisco Pozo
   * Application of Pulsed-Field Gel Electrophoresis to an International
     Outbreak of Salmonella agona, E. John Threlfall, Michael D. Hampton,
     Linda R. Ward, and Bernard Rowe
   * Potential Risk for Dengue Hemorrhagic Fever: The Isolation of Serotype
     Dengue-3 in Mexico, Baltasar Briseo-Garca, Hctor Gmez-Dants, Enid
     Argott-Ramrez, RaulMontesano, Ana-Laura Vzquez-Martnez, Sergio
     Ibez-Bernal, Guillermina Madrigal-Ayala, Cuauhtmoc Ruz-Matus, Ana
     Flisser, and Roberto Tapia-Conyer
   * Improved Serodiagnostic Testing for Lyme Disease: Results of a
     Multicenter Serologic Evaluation, Robert B. Craven, Thomas J. Quan,
     Raymond E. Bailey, Raymond Dattwyler, Raymond W. Ryan, Leonard H.
     Sigal, Allen C. Steere, Bradley Sullivan, Barbara J.B. Johnson, David
     T. Dennis, and Duane J. Gubler
   * Emergence of Bartonella quintana Infection among Homeless Persons,
     Lisa A. Jackson, and David H. Spach
   * The Reemergence of Visceral Leishmaniasis in Brazil, Jorge R. Arias,
     Pedro S. Monteiro, and Fabio Zicker

Commentary

   * Molecular Epidemiology of Pneumocystis carinii Pneumonia, Charles B.
     Beard and Thomas R. Navin
   * Needed: Comprehensive Response to the Spread of Infectious Diseases,
     Harold M. Ginzburg

Letters

   * Transfusion-Associated Malaria, Frances Taylor
   * Reply to F. Taylor, Jane R. Zucker and S. Patrick Kachur
   * An Outbreak of Hemolytic Uremic Syndrome due to Escherichia coli
     O157:H-: Or Was It? P. N. Goldwater and K. A. Bettelheim
   * The Dilemma of Xenotransplantation, Claude E. Chastel
   * The Thucydides Syndrome: Ebola Dj Vu? (Or Ebola Reemergent?) P. E.
     Olson, C. S. Hames, A. S. Benenson, and E. N. Genovese

News and Notes

   * BSE Meeting at CDC
   * CDC Foundation Supports Emerging Infectious Disease Projects, Charlie
     Stokes

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Pages 77-84]

Perspectives

Globalization, International Law, and Emerging Infectious Diseases

David P. Fidler, J.D.
Indiana University School of Law, Bloomington, Indiana, USA
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Download Article

Abstract:
The global nature of the threat posed by new and reemerging infectious
diseases will require international cooperation in identifying,
controlling, and preventing these diseases. Because of this need for
international cooperation, international law will certainly play a role in
the global strategy for the control of emerging diseases. Recognizing this
fact, the World Health Organization has already proposed revising the
International Health Regulations. This article examines some basic problems
that the global campaign against emerging infectious diseases might face in
applying international law to facilitate international cooperation. The
international legal component of the global control strategy for these
diseases needs careful attention because of problems inherent in
international law, especially as it applies to emerging infections issues.

The growing literature on new and reemerging infectious diseases often
emphasizes the global nature of their threat; the U.S. Centers for Disease
Control and Prevention (CDC) defines these diseases as diseases of
infectious origin whose incidence in humans has increased within the past
two decades or threatens to increase in the near future" (1). The World
Health Organization has asserted that emerging infections represent a
global threat that will require a coordinated, global response (2). The
threat is global because a disease can emerge anywhere on the planet and
spread quickly to other regions through trade and travel. The global
challenge of emerging infections has serious consequences for national and
international law; a states ability to deal with them is eroded because
microbes do not respect internationally recognized borders (3). Experts
grappling with these diseases no longer consider that the pursuit of a
strictly national public health policy is adequate. The need for global
cooperation increases the importance of international law in the public
health arena. Part of the effort to create a global response to emerging
infections should be an understanding of the problems that may arise from
relying on international law in dealing with these diseases. This article
outlines issues that will have to be confronted in using international law
to combat emerging infections.

Globalization

The assertion that emerging infections are a global problem requiring a
global strategy echoes observations made in other spheres of public policy:
the traditional distinctions between national and international political,
social, and economic activities are losing their importance (4).
Globalization is eroding traditional distinctions between domestic and
foreign affairs. Globalization has been defined as the process of
denationalization of markets, laws, and politics in the sense of
interlacing peoples and individuals for the sake of the common good (5).
Globalization is distinguished from internationalization, which is defined
as a means to enable nation-states to satisfy the national interest in
areas where they are incapable of doing so on their own (5).
Internationalization involves cooperation between sovereign states, whereas
globalization refers to a process that is undermining or eroding
sovereignty.

Globalization arises from the confluence of something old and something new
in international relations. It involves the very old process of political
and economic intercourse among sovereign states. The new element is the
intensification and expansion of such intercourse made possibleby
technological advances in travel, communications, and computers.
Encouraging such intensification and expansion is liberal economic
thinking, which posits that economic interdependence makes all states
economically better off and builds order and peace in the international
system (6).

The changes wrought by new technologies unleashed in the receptive
international milieu created by liberal trade and economic policies have
led to the belief that these developments are undermining sovereignty.
Observers of international relations frequently note that governments no
longer have control over economic forces at work within their countries.
The speed and volume of international capital flows illustrate the
denationalization of economics occurring through the process of
globalization (7). Another example is the development of the global
companyan enterprise that can no longer be considered national because of
the global reach of its operations, financing options, markets, and
strategies (7). The globalization of finance and business has ramifications
for politics and law as leaders and legal systems adapt to the global era
(8).

In public health, a similar combination of old and new factors can be seen.
States have historically cooperated on infectious disease control, first
through international sanitary treaties and later through the World Health
Organization (WHO) (9). While international cooperation is not new, current
global circumstances confronting the control of infectious disease are.
Globalization is also at work in public health. The assertion that a
country cannot tackle emerging infectious diseases by itself demonstrates
that public health policy has been denationalized.

Globalization has affected public health in three ways. First, the
shrinking of the world by technology and economic interdependence allows
diseases to spread globally at rapid speed. Two factors contributing to the
global threat from emerging infections stem directly from globalization:
the increase in international travel (2, 10) and the increasingly global
nature of food handling, processing, and sales (2, 10). HIV/AIDS,
tuberculosis, cholera, and malaria represent a few infections that have
spread to new regions through global travel and trade (10). The beneficial
economic and political consequences of economic interdependence may have
negative ramifications for disease control. In the European Union, for
example, the free movement of goods, capital, and labor makes it more
difficult for member states to protect domestic populations from diseases
acquired in other countries (11).

Second, the development of the global market has intensified economic
competition and increased pressure on governments to reduce expenditures,
including the funding of public health programs, leaving states
increasingly unprepared to deal with emerging disease problems.
Industrialized as well as developing countries confront deteriorating
public health infrastructures (12). Referring to the United States, one
author described this deterioration as the thirdworldization of the
American health care system (13).

Third, public health programs have also gone global through WHO and
health-related nongovernmental organizations. Medical advances have spread
across the planet, improving health worldwide. The worldwide eradication of
smallpox in 1977 is a famous example. The global reach of health care
advances has, however, a darker side. The globalization of diseasecontrol
has contributed to the population crisis because people are living longer.
Overpopulation creates fertile conditions for the spread of disease:
overcrowding, lack of adequate sanitation, and overstretched public health
infrastructures (2). Further, the widespread use and misuse of antibiotic
treatments has contributed to the development of drug-resistant pathogens
(1, 2). Finally, the success of control efforts in previous decades caused
interest in infectious diseases to wane in the international medical and
scientific communities and is now hampering emerging infectious disease
control efforts (14).

International Solutions to Emerging Infections

International efforts are under way to respond to the threat of emerging
infectious diseases. WHO and CDC have drafted action plans that stress the
need to strengthen global surveillance of these diseases and to allow the
international community to anticipate, recognize, control, and prevent them
(1, 14, 15). WHO has also established a new unit to control and prevent
emerging infections by mobilizing resources rapidly at the first signs of
outbreaks (16). The Pan American Health Organization has also adopted a
regional plan for controlling emerging infections in the Americas (17).
Health authorities from Central American countries have adopted an
emergency plan to control the epidemics of dengue and dengue hemorrhagic
fever that recently swept through Central and South America (18).
Physicians in the European Union recognize the need for better surveillance
of infectious diseases (11). A U.S. government interagency working group
has underlined the importance of international cooperation in dealing with
the emerging infections threat (19). The U.S. Senate Labor and Human
Resources Committee held hearings in October 1995 on Emerging Infections:
A Significant Threat to the Health of the Nation (20). At the Halifax
Summit in 1995, the major industrialized countries adopted a pilot project
called Toward a Global Health Network designed to help governments deal
with emerging infections and other health problems (19) (Table 1). Clearly,
the emerging infections threat and the need for action are on the
international diplomatic and public health agendas.

   Table 1. Some common elements of
   global emerging-disease control
                plans

----------------------------------------
   Strengthen international
 surveillance networks to detect,
 control, and reduce emerging
 diseases.
   Improve the international public
 health infrastructure (e.g.,
 laboratories, research facilities,
 technology, and communications
 links.
   Develop better international
 standards, guidelines, and
 recommendations.
   Improve international capabilities
 to respond to disease outbreaks with
 adequate medical and scientific
 resources and expertise.
   Strengthen international research
 efforts on emerging diseases,
 particularly with regards to
 antibiotic-resistant strains of
 diseases.
   Focus attention and resources on
 training and supporting medical and
 scientific expertise.
   Encourage national governments to
 improve their public health care
 systems, devote resources to
 eliminating or controlling causes of
 emerging diseases and coordinate
 their public health activities with
 WHO and the international community.
---------------------------------------
 Sources: refs. 1, 14, 15, 19.

Although international control plans would involve private organizations
like universities and nongovernmental organizations, the primary actors on
the emerging infections stage are sovereign states. The action plans are
predominantly blueprints for cooperation among states and represent a call
for the internationalization of responses to a problem caused by
globalization. Put another way, the proposed solutions to the emerging
infections threat rely on the sovereign state, while the threat feeds off
the impotence of the state in addressing global disease problems. When it
comes to public health activities, globalization erodes sovereignty, but
the proposed solution makes sovereignty and its exercise critical to
dealing with the threat of emerging infections.

The consequences of the unavoidable emphasis on international cooperation
in the proposed action plans for emerging infections are troubling. To
achieve the desired objectives (Table 1), states will have to agree on many
issues and translate such agreement into guidelines or rules. International
law becomes important to the effort for emerging infections control.
Political leaders, diplomats, and scholars have long recognized the
weakness of international law in regulating state behavior. At first
glance, the prospect of having to rely on a notoriously weak institution of
international relations as part of the global effort to combat emerging
infections is unsettling.

International Law and Infectious Disease Control

We might have been less unsettled if our experience with international law
in controlling infectious diseases had been more positive. The success of
WHO in globalizing disease control programs might suggest that the defects
of international law have not hobbled its effectiveness in improving health
care worldwide. However, despite having the authority to do so, WHO has
been reluctant to use international law (21, 22). The International Health
Regulations administered by WHO represent the most important set of
international legal rules relating to infectious disease control, but the
regulations only apply to plague, yellow fever, and cholera (23). The
importance of health is mentioned in international declarations (for
example, see the Universal Declaration of Human Rights, art. 25 [1]) and
treaties (for example, see the International Covenant on Economic, Social
and Cultural Rights, art. 12), leading some legal scholars to argue that
international law creates a right to health (24); but this right does
not directly address the control of infectious diseases. WHO has refrained
from adopting rules on trade in human blood and organs, which does raise
issues of infectious disease control as illustrated by the sale of
HIV-contaminated blood in international commerce (25). Issues of disease
control also appear in specialized treaty regimes outside WHO, such as
treaties controlling marine pollution from ships (26). Other areas of
international public health law, for example, rules about infant formula
and guidelines on pharmaceutical safety, do not deal with the control of
infectious diseases (25).

The effectiveness of existing international law on infectious disease
control has been questioned. A 1975 WHO publication stated that the
International Health Regulations have not functioned satisfactorily at
times of serious disease outbreaks (27). More recently, WHOs efforts with
the International Health Regulations have been called a failure, and
noncompliance with these regulations has increased in connection with
reporting disease outbreaks (25). The HIV/AIDs crisis dramatically
illustrated the weaknesses of the health regulations. Since AIDs was not
originally (or subsequently) made subject to the regulations, states had,
and continue to have, no notification requirements in connection with this
new disease. Further, as HIV/AIDs spread globally, many states adopted
exclusionary policies that, according to experts, violated provisions of
the health regulations (25). In relation to one of the biggest disease
crises of this century, parts of the International Health Regulations were
irrelevant, and other parts were openly violated.

WHOs reluctance to apply international law has been attributed to its
organizational culture, which is dominated by scientists, doctors, and
medical experts. Perhaps the current weakness of international law on
infectious disease control reflects WHOs nonlegal strategy rather than the
inherent problems in international law itself. In connection with emerging
infections, however, WHO is advocating an international legal strategy by
recommending revision of the International Health Regulations (28). This
recommendation suggests that WHO acknowledges the need for international
legal agreement in dealing with emerging infections. The global threat
posed by these infections represents in many ways a test case for
international public health law.

The Challenge to International Law

The threat of emerging infectious diseases poses two challenges to
international law: first, theemerging infections problem exacerbates basic
weaknesses in the law. Second, these infections pose specific difficulties
in the law, which are related to the nature of disease and its prevention.

Basic Weaknesses

The effectiveness of international law depends on the consent of states,
which means that sovereignty and its exercise determine the fate of
international legal rules (29). In adopting a legal strategy for its
emerging infectious disease action plan, WHO has to convince its member
states to take certain actions in response to disease emergence. The
sovereignty of states looms large in formulating a global response to
emerging infections, despite the fact that the process of globalization
undermines the sovereignty of the state to deal nationally with these
infections. In other words, the problem by-passes the state, but the
solution has to rely on the state through the medium of international law.
The central importance of the state and its sovereignty constitutes a basic
weakness in international law because international legal rules tend to
reflect the compromises necessary to achieve agreement and the
unwillingness of states to restrict their freedom of action through
international law. Part of the reason that the existing International
Health Regulations cover only a few diseases might be the unwillingness of
WHO member states to commit to more serious infectious disease control
measures. The vagueness and lack of specificity in the so-called right to
health also illustrate this problem. What is scientifically and medically
necessary to combat emerging diseases may not be what states are willing to
agree to undertake.

A second basic weakness follows from the sovereignty problemthe lack of
effective enforcement of international law. States often agree to an
international legal obligation without any serious intent of fulfilling it.
The alleged failure of the International Health Regulations may be due to
the failure of WHO member states to fulfill the duties they accepted.
Neither the regulations nor WHO has any power to enforce compliance (25).
An international legal regime on emerging diseases would also face this
enforcement problem.

Specific Difficulties

The very nature of the emerging disease threat poses special difficulties
for international law. The global scope of the problem necessitates
agreement by most states to control emerging diseases. If any major country
or group of countries does not participate, a gap in the global
surveillance and control network threatens the efficacy of the entire
effort. The negotiation of agreements involving many states is usually
difficult, because each state knows that its nonparticipation threatens the
success of the entire venture. This problem has occurred in international
environmental law, where global regimes have been needed to deal adequately
with environmental threats, such as ozone depletion.

A second specific difficulty arises from the extent of medical and
scientific resources needed to establish an effective global surveillance
and control network for emerging diseases. Fundamental aspects of the
proposed action plans involve improving surveillance networks, public
health infrastructures, scientific research, and medical and scientific
training (Table 1). Some states, particularly in the developing world, do
not have the medical, scientific, and financial resources to undertake such
measures. Unless more affluent countries provide theresources, developing
states may use the inequity of wealth in the international system as an
argument to complicate negotiating a global agreement. The so-called
North-South problem has made the negotiation of international
environmental agreements more difficult, as developing countries have
bargained for more lenient treatment or a transfer of resources from
affluent countries to help them improve environmental protection. A similar
dynamic may appear in any negotiations for a global emerging disease
effort. The U.S. interagency working group on emerging diseases has
observed that major U.S. contributions to developing countries for emerging
disease control purposes is not a likely prospect during this period of
deficit reduction and downsizing (19), which suggests that resource
availability will probably complicate international efforts in this area.

The problems associated with using international law in a global strategy
to combat emerging diseases raise the question whether international law
can provide an adequate foundation for the control of these diseases. The
uncomfortable position of having no choice but to rely on international law
when its weaknesses are substantial highlights the importance of thinking
through the international legal aspects of a global emerging disease plan
carefully.

WHOs Proposed Legal Strategy

WHO wants to revise the International Health Regulations as part of its
global emerging disease strategy (28). WHOs proposal deserves some
critical attention. It is not clear that the organization has adequate
authority to incorporate comprehensive emerging disease control measures
within the international regulations. Under Article 21 of the WHO
Constitution, the World Health Assembly can adopt binding regulations in
sanitary and quarantine requirements and other procedures to prevent the
international spread of disease (22). The World Health Assembly adopted the
International Health Regulations under Article 21. While Article 21 and the
regulations are relevant to emerging disease control efforts, it is
doubtful whether the regulations can serve as a foundation for a
comprehensive emerging disease control plan. The disease-outbreak
notification requirements in the regulations could be expanded to include
more diseases, but nothing in Article 21 gives the World Health Assembly
the authority to require WHO member states to strengthen public health
infrastructures, which is considered critical in the emerging disease
actions plans proposed to date (Table 1). It has been argued that
attempting to address such infrastructure problems is a solution which
cannot be obtained by an international instrument but only by the
improvement of the health conditions of the peoples of WHOs member states
(30). But, as the history of administering the International Health
Regulations has shown, notification requirements have not worked
satisfactorily and are weakened by the absence of adequate public health
resources. Further, Article 22 of the WHO Constitution makes regulations
promulgated under Article 21 automatically binding on WHO member states,
except for member states that reject such regulations or make reservations
thereto (31). Article 22 relates to the sovereignty problem and may deter
WHO member states from agreeing to serious revisions of the regulations.
Analysis of the regulations may question the wisdom of using the
regulations as the legal basis for dealing with emerging diseases.

The World Health Assembly has the power to adopt conventions or agreements
within WHOs competence (21). The Assembly could use this authority to
address aspects of the global emerging disease control strategy that cannot
be handled with a revision of the regulations.However, parceling up
emerging disease control measures between the International Health
Regulations and separate agreements would be legally complicated. Further,
WHO has not used this power to adopt conventions or agreements, which
explains its unwillingness to explore all legal options open to it.

Possible Alternative Legal Strategies

Alternative legal strategies to revising the International Health
Regulations range from reliance on the development of customary
international law to the adoption of multilateral treaties specifically on
emerging-disease control (Table 2). An issue related to these alternative
approaches is the substantive nature of the obligations contained in legal
documents. We have to ask not only how states might agree on control rules
but also what these states might agree to do. The proposed revision of the
regulations apparently would only apply the notification duties (currently
found in the regulations) to more diseases. As indicated earlier, WHO
cannot address in its revision of the regulations any of the improvements
in public health infrastructures, surveillance networks, scientific
research, or medical and scientific training at the heart of proposed
emerging disease action plans. Further, it is not clear whether WHO intends
to supplement expanded notification duties with any mechanism to monitor or
enforce such duties.
   Table 2. Alternative international legal strategies to
       revising the International Health Regulations

 Alternative legal                      Possible
 strategies        Possible advantages  disadvantages

 1. WHA            Integrates emerging  a. Emerging disease
 incorporates      disease control      control would
 emerging          measures into the     not be primary
  disease control  overall WHO          focus
 as part of the    approach to          b. World Health
  proposed World   international        Charter is likely
 Health Charter    health               to
  scheduled for    issues                be more
 initial                                aspirational than
 negotiations                            obligatory
  in 1997

 2. WHA adopts an  a. Avoids IHR model  a. WHA has no
 emerging          b. Has potential to  experience with
  disease-specific set out               using Article 19
 convention under   comprehensive       b. Large
  Article 19 of    global approach      multinational
 the WHO            to emerging         treaties tend
 Constitution      diseases              to contain general
                                        obligations
                                         rather than
                                        specific duties

 3. States         a. Takes emerging    a. WHO has to play
 negotiate a       disease control      central role in
 framework          out of WHO,          any emerging
  multilateral     eleminating          disease plan
 treaty on general  problem of WHO's    b.
                   reluctance to        Frameowrk-protocol
  emerging disease  use international   approach
 obligations,      law                   might not be
  accompanied by   b. Allows for new    appropriate model
 disease-specific  protocols to be       for emerging
 or                 adopted for new     disease control
  region-specific  diseases              because the
 protocols         c.                   emerging disease
 containing        Framework-protocol    problem differs
  detailed and     approach             from ozone
 specific           has been used with   depletion
 commitments       some success
  on emerging       in international
 disease control   environmental
                    law on ozone
                   depletion

 4. Encourage      a. Builds on strong  a. Emerging
 regional          regional systems     diseases require a
 arrangements       of cooperation and   global approach
  and integrate    coordination         not just a
 them into global  b. Offers "legal      regional approach
  regime over time laboraties to try    b. Amounts to
                    various approaches  emerging disease
                   to emerging           control for rich
                    disease control     regions, leaving
                   c. Avoids             many developing
                   diplomatic           countries
                   headaches             outside legal
                    involved in trying  regime
                   to negotiate         c. Risks
                    truly global legal  inconsistencies in
                   regimes              how
                                         emerging diseases
                                        are handled
                                         by different
                                        regions

 5. Encourage a    a. Gives states      a. Does not address
 bilateral         flexibility in       global nature of
 approach in        constructing legal   emerging disease
  which individual obligations          problem
 contries          b. Permits           b. Sanctions
  negotiate        possibility for      element is
 detailed and      sanctions            unrealistic
 specific           for failure to       and might be
  commitments on   live up to emerging  unfair to
 emerging                                developing
  diseases and      disease             countries lacking
 perhaps condition obligations           the resources
                                        necessary to
  trade benefits                         implement adquate
 and aid on                             emerging
 emerging                                disease control
  disease                               measures
 performance

 6. Incorporate    a. Links emerging    a. International
 emerging disease  disease control      "right to health"
  control as part   with larger,        has
 of international  powerful concepts     no definitive
  "right to         of human welfare    meaning or scope
 health," making   b. Builds on          and thus is a bad
 emerging          existing             foundation for
  diseases a human international         emerging disease
 rights issue       law on the "right   control
                   to health"           b. Human rights are
                                        inherently
                                         divisive in the
                                        international
                                         system; linkage
                                        with such a
                                         controversial area
                                        would hurt
                                         emerging disease
                                        control prospects

 7. Rely on        Customary            a. It will be
 customary         international norms  nearly impossible
 international     on                   to
  law to develop   emerging disease      develop general
 emerging          control would be     and uniform state
  disease-control  binding on all        practice
 norms             states except        recognized by
                   persistent           states as
                   objectors             legally binding in
                                        the emerging
                                         disease-control
                                        area
                                        b. Any customary
                                        norms that might
                                         form will probably
                                        be vague and
                                         hard to identify
                                        definitively
                                        c. Customary norms
                                        can take a very
                                         long time to
                                        develop

 WHA = World Health Assembly; WHO = World Health
 Organization; IHR = International Health Regulations.

International environmental law had to overcome some of the same obstacles
encountered by WHOs international legal effort for emerging disease
control. States realized that they could not handle global environmental
problems without international cooperation and rules (32). Further, states
knew that addressing environmental concerns would require changes for
governments and companies within states and that developing states might
have financial and technological difficulties implementing international
agreements (32). In developing international environmental law, states,
international organizations, and nongovernmental organizations did not rely
on old approaches but instead crafted new international legal rules to deal
with the global nature of the threats posed, the resource issue, and
compliance and enforcement problems (33). Whether international
environmental law has been successful is controversial; but it is important
that states have not been willing to admit that improving environmental
conditions within states is a solution that cannot be obtained by
international agreements. Models and precedents from international
environmental law are not in all respects helpful to the challenge of
emerging-disease control; but, at the very least, those grappling with an
international strategy for the emerging-disease threat could analyze
international environmental law and other innovative legal responses to
globalization to look for ways of making WHOs international legal strategy
on emerging diseases as effective as possible.

Those currently designing global emerging-disease control strategies will
eventually have to translate what is scientifically and medically needed to
combat these diseases into international agreement and cooperation through
international law. The movement from science and medicine into the realm of
international law will not be easy. Relying on the International Health
Regulations as the centerpiece of international law on emerging-disease
control may not be the most effective international legal strategy.
Whatever international legal approach is eventually taken will have to
confront somehow a fundamental paradox: globalization jeopardizes disease
control nationally by eroding sovereignty, while the need for international
solutions allows sovereignty to frustrate disease control internationally.
The combination of the process ofglobalization and the unavoidable need to
rely on international law produces a most unattractive medium in which to
wage potentially one of the most important medical and scientific endeavors
in history.

Dr. Fidler is associate professor of law at Indiana University School of
Law, Bloomington, where he teaches public and private international law. He
has a master of philosophy (M.Phil.) degree in international relations from
Oxford University, a J.D. from Harvard Law School, and a bachelor of civil
law degree from Oxford University. Before joining the faculty at Indiana
University School of Law, Dr. Fidler practiced law with Sullivan & Cromwell
in London, England, and with Stinson, Mag & Fizzell, P.C., in Kansas City,
Missouri.

Address for Correspondence:
David P. Fidler, J.D., Indiana University School of Law, Third Street and
Indiana Avenue, Bloomington, IN 47405-1001 USA; fax: 812-855-0555; e-mail:
davidfider@law.indiana.edu

Acknowledgments

I thank Jack Bobo, research assistant, for his valuable work during the
preparation of this article, and Lane Porter and Allyn Lise Taylor for
their helpful comments on earlier drafts of this article. I also thank my
colleagues on the faculty of Indiana University School of Law for their
helpful comments on an earlier draft presented at a faculty colloquium.

References

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Emerging Infectious Diseases Volume 2 * Number 2                    April-June 1996

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[Pages 85-92]

Perspectives

On Epidemiology and Geographic Information Systems: A Review and Discussion
of Future Directions

Keith C. Clarke, Ph.D., Sara L. McLafferty, Ph.D,. and Barbara J. Tempalski
Hunter College-CUNY, New York, New York, USA

Geographic information systems are powerful automated systems for the
capture, storage, retrieval, analysis, and display of spatial data. While
the systems have been in development for more than 20 years, recent
software has made them substantially easier to use for those outside the
field. The systems offer new and expanding opportunities for epidemiology
because they allow an informed user to choose between options when
geographic distributions are part of the problem. Even when used minimally,
these systems allow a spatial perspective on disease. Used to their optimum
level, as tools for analysis and decision making, they are indeed a new
information management vehicle with a rich potential for public health and
epidemiology.

Geographic information systems (GIS) are "automated systems for the
capture, storage, retrieval, analysis, and display of spatial data" (1).
Common to all GIS is a realization that spatial data are unique because
their records can be linked to a geographic map. The component parts of a
GIS include not just a database, but also spatial or map information and
some mechanism to link them together. GIS has also been described as the
technology side of a new discipline, geographic information science (2),
which in turn is defined as "research on the generic issues that surround
the use of GIS technology, impede its successful implementation, or emerge
from an understanding of its potential capabilities." Recently, GIS has
emerged as an innovative and important component of many projects in public
health and epidemiology, and this disciplinary crossover is the focus of
this review.

Few would argue that GIS has little to offer the health sciences. On the
other hand, like other new technologies, GIS involves concepts and analytic
techniques that can appear confusing and can lead to misunderstanding or
even overselling of the technology. In this article, we attempt to bridge
the gaps between the principles of geographic information science, the
technology of GIS, the discipline of geography, and the health sciences.
Our intent is to introduce to the epidemiologist a set of methods that
challenge the "visual" half of the scientist's brain.

Computers were first applied to geography as analytical and display tools
during the 1960s (3). GIS emerged as a multidisciplinary field during the
1970s. The discipline's heritage lies in cartography's mathematical roots:
in urban planning's map overlay methods for selecting regions and locations
based on multiple factors (4); in the impact of the quantitative revolution
on the discipline of geography; and in database management developments in
computer science.

Several factors combined in the 1970s to reinforce GIS development. First,
computers became more accessible and less costly. Second, mainframe
computers gave way to minicomputers and then workstations, which gave great
power to the user and included the access to networks that has led to its
own revolution in technology. Third, the types of user interface required
to operate technical software changed from batch, command-line, and remote
access to windowing systems and "point and click" graphic interaction. What
had been expensive, slow, and difficult has rapidly become inexpensive,
fast, and easy to use. A final but essential precondition to GIS
development was the broad availability of public domain digital map data,
in the form of maps of the landscape from the U.S. Geologic Survey and for
census areas from the U.S. Census Bureau. The current GIS World Sourcebook
(5) lists hundreds of system suppliers and sources of information and
catalogs system capabilities. In short, GIS has now come of age, to the
extent that the contributions of a growing number of parallel disciplines
have both influenced and been influenced by GIS. Other disciplines now
affecting GIS include forestry, transportation planning, emergency services
delivery, natural hazards planning, marketing, archeology, surveying, and
criminal justice. A wide array of capabilities and information awaits the
health scientist ready to pursue an interest in GIS.

In this article, we consider the functional capabilities of GIS and how
they can relate to epidemiology. We then review studies in epidemiology and
health science where GIS has already made a contribution and introduce the
technologic and analytic background. We review spatial analytic methods and
concepts of use in epidemiology and conclude by examining what the near
future holds for technologic changes and what these changes mean for the
study of emerging infectious diseases and other health applications.

GIS Functional Capabilities

GIS definitions usually focus on what tasks a GIS can do rather that what
it is. GIS functional capabilities follow the standard GIS definitions;
therefore, GIS can bring together the elements necessary for problem
solving and analysis.

Data capture implies that 1) data can be input into the GIS from existing
external digital sources; this is particularly the case when no data exist
for a project, and the base data must be assembled from other studies,
public domain datasets, and images. This usually means that GIS must be
able to import the most common data formats both for image-type (raster)
and line-type (vector) maps. 2) GIS can capture new map data directly; this
means either that the user can scan the map and input it into the GIS or
trace over a map's features using a digitizing tablet and enter them into
the GIS map database. 3) The GIS can accomplish everything that a regular
database system can, such as enter and edit data and update information in
the existing database.

Data storage implies storage of both map and attribute data. Attribute data
are usually stored in a relational database management system contained
within the GIS and accessed by a spreadsheet or query-driven user
interface. For storage, map data must be encoded into a set of numbers so
that the geometry of the map is available for query, but also so that the
map is stored digitally in one or more files. Image maps are usually stored
as gridded arrays. Line maps are encoded by any one of several systems, but
usually by using both the coordinate information and encoded topology, so
that the relationships between points, lines, and areas, such as the
adjacency of regions or the connectedness of lines, are known in advance.
The more efficient and flexible these data formats or structures, the more
operations can be performed on the map data without further processing.

Data records in GIS can be retrieved in one of two ways. The relational
database manager allows searching, reordering, and selecting on the basis
of a feature's attributes and their values. For example, the user may wish
to select out and order alphabetically the names of all health clinics that
had positive results in more than 10% of their tests. GIS also allows
spatial retrieval. The user could select all clinics by region, by their
latitude, or by their distance from the capital. The user could also select
all clinics that are more than 10 km from a major road and within 100 m of
a river or lake. In addition, combining searches is possible. There could
be several data "layers," for example vegetation, rivers, transportation,
and population of villages. A single retrieval could combine data from each
of these layers in a single query. Layers can also be weighted, so that
rivers, for example, are twice as important as roads in selecting villages
with a population under 500 surrounded by forest.

Display functions include predominantly the making of maps. Tools must
exist for constructing many types of maps, such as contours, symbols,
shading or choropleth, and sized symbols. Formal map display often follows
a series of more temporary map images, usually without a strict map
composition, and the result of a test, an analysis, or a query. In
addition, the GIS must be able to output finished format of maps to a
medium, such as PostScript, on a plotter or printer, or onto photographic
film.

Many tools exist to support field data collection. Tasks in which ancillary
demographic information needs to be input and coregistered are simple.
Habitat associated with a vector (e.g., a snail or a mosquito) may need
remotely sensed data, such as vegetation cover or weather data. If these
data are georegistered, integration is possible. One of the most useful
functions is called address matching, in which street addresses with house
numbers and street names are automatically placed into an administrative
unit or placed as a dot on the map. Thus a digital phone list or mailing
list of patients can be merged with the remainder of the data. In the
United States, the Census Bureau's TIGER files can usually match 70% to 80%
of unedited address records, and higher percentages if the address files
are proofed and/or the more detailed and up-to-date commercial street files
are used. In some field projects, the GIS's ability to make maps became the
mainstay of the effort, allowing planning of truck and jeep routes,
sequencing field clinics for optimal routes for visits, and even for local
navigation. The ability to display maps often goes far beyond their final
or use in the laboratory. Often a GIS image map is more accurate and up to
date than anything available locally.

Existing Applications of GIS in Epidemiology

Epidemiologists have traditionally used maps when analyzing associations
between location, environment, and disease (6). GIS is particularly well
suited for studying these associations because of its spatial analysis and
display capabilities. Recently GIS has been used in the surveillance and
monitoring of vector-borne diseases (7-9) water borne diseases (10), in
environmental health (11-13), modeling exposure to electromagnetic fields
(14), quantifying lead hazards in a neighborhood (15), predicting child
pedestrian injuries (12), and the analysis of disease policy and planning
(16).

In a recent study in Baltimore County, Maryland, GIS and epidemiologic
methods were combined to identify and locate environmental risk factors
associated with Lyme disease (7). Ecologic data such as watershed, land
use, soil type, geology, and forest distribution were collected at the
residences of Lyme disease patients and compared with data collected at a
randomly selected set of addresses. A risk model was generated combining
both GIS and logistic regression analysis to locate areas where Lyme
disease is most likely to occur.

GIS allows analysis of data generated by global positioning systems (GPS).
Combined with data from surveillance and management activities, GIS and GPS
provide a powerful tool for the analysis and display of areas of high
disease prevalence and the monitoring of ongoing control efforts. The
marrying of GIS and GPS enhances the quality of spatial and nonspatial data
for analysis and decision making by providing an integrated approach to
disease control and surveillance at the local, regional, and/or national
level.

GIS is being used to identify locations of high prevalence and monitor
intervention and control programs in areas of Guatemala for onchocerciasis
(9) and in Africa for trypanosomiasis (17). Spatial and ecologic data are
combined with epidemiologic data to enable analysis of variables that play
important roles in disease transmission. This integration of data is
essential for health policy planning, decision making, and ongoing
surveillance efforts. For example, as part of the guinea worm eradication
effort, the United Nation's Children's Emergency Fund placed pumps in
villages most infected with the disease to ensure access to a safe water
supply (18). GIS enabled researchers to locate high prevalence areas and
populations at risk, identify areas in need of resources, and make
decisions on resource allocation (16). Epidemiologic data showed a marked
reduction in prevalence in villages where pumps were introduced.

GIS was used in designing a national surveillance system for the monitoring
and control of malaria in Israel (19). The system included data on the
locations of breeding sites of Anopheles mosquitoes, imported malaria
cases, and population centers. The GIS-based surveillance system provided
means for administrative collaboration and a network to mobilize localities
in the case of outbreaks.

In 1985, the National Aeronautics and Space Administration (NASA)
established the Global Monitoring and Disease Prediction Program at Ames
Research Center in response to the World Health Organization's call for the
development of innovative solutions to malaria surveillance and control
(20). A major aspect of the program was to identify environmental factors
that affect the patterns of disease risk and transmission. The overall goal
of the program was to develop predictive models of vector population
dynamics and disease transmission risk using remotely sensed data and GIS
technologies.

Remotely sensed data have been used in many vector disease studies
(8,17,21-24). Remote sensing and GIS were used to identify villages at high
risk for malaria transmission in the southern area of Chiapas, Mexico (8).
An earth environmental analysis system for responding to fascioliasis on
Red River Basin farms in Louisiana was developed by integrating LANDSAT MSS
imagery with GIS (22). In Kwara State, Nigeria, a temporal analysis of
Landsat Thematic Mapper (TM) satellite data was used to test the
significance of the guinea worm eradication program based on changes in
agricultural production (21).

Spatial Analysis and GIS

GIS applications show the power and potential of such systems for
addressing important health issues at the international, national, and
local levels. Much of that power stems from the systems' spatial analysis
capabilities, which allow users to examine and display health data in new
and highly effective ways. Spatial analysis refers to the "ability to
manipulate spatial data into different forms and extract additional meaning
as a result" (25). It encompasses the many methods and procedures,
developed in geography, statistics, and other disciplines, for analyzing
and relating spatial information. Spatial relationships, those based on
proximity and relative location, form the core of spatial analysis.

Gatrell and Bailey (26) describe three general types of spatial analysis
tasks: visualization, exploratory data analysis, and model building. These
range in complexity from simple map overlay operations to statistical
models such as spatial interaction and diffusion models. The value of maps
for public health analysis has long been recognized; John Snow's now
classic maps of cholera cases in relation to the Broad Street pump are a
good example. However, with its extensive data management and display
capabilities, GIS offers much more than simple mapping. Map overlay
operations allow the analyst to compute new values for locations based on
multiple attributes or data "layers" and to identify and display locations
that meet specific criteria (27). For example, in targeting locations for
mosquito vector control, one might want to identify areas that have low
elevation, specific types of vegetation favored by mosquitoes, and are
within 100 m of ponds or other water bodies. Each of these attributes
comprises a distinct data layer. With GIS, one can create 100-m buffers
around water bodies and then select areas meeting all three criteria.
Display of these areas on a GIS-generated map has obvious benefits for
planning vector control strategies.

As indicated previously, this general class of procedures for weighing and
overlaying maps, also known as "suitability analysis," has been used in
diverse health applications. Typically the criteria and weights attached to
them are specified by the analyst based on expert knowledge or prior
research. Using the computational and visual display capabilities of GIS,
one can then explore the sensitivity of results to the weights and cutoff
values used. Another approach is to employ regression analysis to generate
the linear combination of factors that best explain spatial variation in
disease prevalence. The weights from the regression model are used to
create a composite index of risk which can then be mapped (7).

Visualization is also an important tool for showing the change in disease
patterns over time. Animation, embedded within a GIS, is highly effective
in depicting the spread or retreat of disease over space and time. A series
of animated maps were created to show the advance of the AIDS epidemic in
the United States as it moved from and within major cities (28). One could
imagine a similar animated map sequence showing the retreat and eventual
eradication of a disease like smallpox. Clearly much more research is
needed in this area, especially research that links animation to
theoretical models of disease diffusion, within a GIS environment.

Visualization can be used in novel ways to explore the results of
traditional statistical analysis. Displaying the locations of outlier and
influential values on maps and showing variation in values over space can
add a great deal to epidemiologic research. Although such tools are being
developed and explored, they would benefit greatly from a closer and more
seamless link between statistical packages and GIS (25).

The second general class of GIS methods addresses exploratory spatial
analysis. These methods allow the analyst to sift meaningfully through
spatial data, identify "unusual" spatial patterns, and formulate hypotheses
to guide future research (26). The quantity and diversity of spatial data
in GIS can be overwhelming: exploratory methods help the analyst make sense
of data and address "what if" questions. Advances in computing and graphics
technology have made this one of the most active areas in GIS/spatial
analysis research.

Among the most important exploratory methods for epidemiology and public
health are methods for identifying space-time clusters or "hot spots" of
disease. Openshaw's geographic analysis machine (GAM) was an early method
that worked completely within a hybrid GIS. The GAM's many applications
included an attempt to determine if spatial clusters of childhood leukemia
were located near nuclear facilities in Britain (29). The GAM works with
point data on disease cases and searches at regular intervals for
statistically significant clusters of disease prevalence. Maps display the
locations of significant clusters, showing the proximity of clusters to
hypothesized environmental threats such as nuclear facilities. Although
Openshaw's work was widely criticized on statistical grounds, it opened the
door for an active body of research on exploratory spatial analysis of
disease. Some of the new methods that have been developed as outgrowths of
Openshaw's approach have been published (30).

Exploratory methods are also valuable in searching for zones or districts
of high disease prevalence. Because areas may differ greatly in population
size, prevalence rates have different levels of variability and thus
reliability (31). Researchers have long used probability mapping to show
the statistical significance of prevalence rates (32); however, probability
mapping does not give a sense of the actual rates or the populations on
which they are based. An alternative method is to smooth rates towards a
regional or local mean value using empirical Bayes methods (33). Although
GIS and empirical Bayes methods have developed separately, there is much
scope for interaction. For example, GIS can be used to generate
geographically based regional or local means to which actual rates are
smoothed. These might be based on averaging rates for contiguous areas
(33,34); or they might rely on more complex, multivariate, spatial
clustering procedures that incorporate proximity as well as population
attributes.

Many methods for exploratory analysis of disease patterns are not
appropriate for infectious diseases because the methods are essentially
static and assume independence. For infectious diseases, cases clearly are
not independent and the diseases move through time and space. In these
situations, one can use spatial autocorrelation methods and space-time
correlograms to explore the spatial and temporal patterns of infectious
disease spread (35).

These methods provide a general sense of the speed and geographic pattern
of disease transmission. Although the methods have not typically been
incorporated in GIS, there is great potential for doing so, especially with
recent advances in computer animation.

Modeling, the final class of spatial analysis methods, includes procedures
for testing hypotheses about the causes of disease and the nature and
processes of disease transmission. In general, modeling involves the
integration of GIS with standard statistical and epidemiologic methods. GIS
can assist in generating data for input to epidemiologic models, displaying
the results of statistical analysis, and modeling processes that occur over
space. The first two points are evident in recent, regression-based
analyses of disease risk, such as the study of Lyme disease (7). There GIS
was used not only to integrate diverse datasets and calculate new
variables, such as slope and distance from forest, but also to map
geographic variation in disease risk, as predicted from a logistic
regression model.

Other GIS models are more explicitly spatial, expressing relationships or
flows between people and places. Spatial interaction and spatial diffusion
models are of particular relevance to the study of emerging diseases.
Spatial interaction models analyze and predict the movements of people,
information, and goods from place to place (36). The flows of people
between rural areas, villages, cities, and countries are all forms of
spatial interaction that are central to disease transmission. By accurately
modeling these flows, it is possible to identify areas most at risk for
disease transmission and thus target intervention efforts. Spatial
interaction models reflect two general principles: that interaction
decreases with distance and increases with population size or
"attractiveness." Given actual flow data, one can estimate values that show
the effects of distance and population size (or other "attractiveness"
factors) on interaction. The models can then be used to predict spatial
interaction patterns elsewhere. Although spatial interaction models and GIS
developed separately, some GIS now have spatial interaction modeling
capabilities (37).

Spatial diffusion models analyze and predict the spread of phenomena over
space and time and have been widely used in understanding spatial diffusion
of disease (38). Such models are quite similar to spatial interaction
models except that they have an explicit temporal dimension. By
incorporating time and space, along with basic epidemiologic concepts, the
models can predict how diseases spread, spatially and temporally, from
infected to susceptible people in an area (39) and aid in understanding the
emergence of infectious disease (40).

Data

Important technical and logistic innovations in data and data access for
GIS are under way and will come to fruition before the end of the century.
First, and by far the most important, have been increased access to the
Defense Department's global positioning systems (GPS), the availability of
inexpensive hand-held devices for using the system, and the addition of
direct-to-GIS data links to these systems. For a relatively modest
investment, field users can add geographic coordinates to their data
collection from anywhere in the world, at any time, and in any weather.
These systems are so flexible that their antennas can be placed on top of a
car, and the logger can be connected to a portable computer on the
dashboard, so that as the user drives along, the path of the vehicle is
permanently recorded in the GIS's own data format and displayed on screen
with a 1-s update. As these systems have become more common, they have also
gained in precision and accuracy. It is not uncommon for fixes to be
corrected using a process known as differential GPS, either after the fact
by computer software or in real time, so that each point is recorded to the
nearest meter on the ground. GPS and GIS together have permanently altered
the relationship between field data collection and data analysis. Data
collected in real time can be analyzed the same day and acted upon
immediately.

Similarly, various devices used for capturing overhead images and
photographs have undergone a similar revolution. First, technology has
improved, allowing images in the infrared, thermal, radar and other
wavelengths to be collected at higher and higher spatial resolutions.
Second, massive changes in policy have resulted from the end of the Cold
War. Formerly secret satellite data, such as the CORONA and Russian spy
imagery, are now broadly available, even searchable on the Internet. In the
United States, the National Air Photo program intends to remap the country
every 5 years at a scale of 1:12,000 with 1-m resolution and publish the
images as CD-ROMs. In addition, NASA's largest ever Mission to Planet Earth
and its Earth Observation System will begin to return unimaginable amounts
of information about the whole earth's geography and atmosphere well before
the end of the century. The data will be available to any Internet user and
distributed by a set of active archive centers.

Third, technical issues related to data transfer have been partially
eliminated. This has come about by the convergence toward sets of industry
standard formats such as GIF and TIF for images and new national and
international digital map data standards. In addition, efforts are now
under way to standardize reference information about datasets, termed
metadata, so that the equivalent of a Library of Congress cataloging will
be possible.

Finally, many datasets have become available that can form at least the
skeleton of a new GIS project almost anywhere in the world. By combining
public domain datasets, such as the Digital Chart of the World and
satellite imagery, with GPS and field data, the claim that data collection
and changes in format constitute 80% of the effort in a GIS project is
rapidly being eroded and replaced by a mere morning spent surfing the
Internet. Nevertheless, many of the world's nations are still poorly mapped
at the more detailed spatial scales required for local analysis.

Hardware

GIS hardware has continued to improve. On the high end, workstations have
both increased in power and dropped in price, making this platform the
choice for large, laboratory-based GIS projects. As the GIS software
packages have been modified for the workstation operating systems, most
commonly UNIX with X-Windows, operations that were impossible because of
computational complexity have now become commonplace. This trend will
continue to the extent that few technical constraints like memory and
central processing unit (CPU) power will exist for GIS. Some tasks, such as
skilled visual image identification and interpretation, have been partly or
wholly automated. On the low end, microcomputers have become immensely
powerful and fast, easily capable of performing basic GIS operations even
on portable computers. The theme of GIS mobility, added to satellite and
cellular telephone communications, has permanently transformed the ability
to operate with GIS in the field, and will lead to a new "data rich" era
for epidemiologic study.

In addition, the next generation of systems will depend on network
computing. Networks have allowed de facto parallel computing within a local
area network. By supporting personal multitasking, they have allowed data
to be held in a distributed way and retrieved for use on demand, and the
network has built an immensely powerful support structure for information
sharing. The World-Wide Web, for example, can deliver to a workstation-user
free GIS software, data, and information on how to install and use the
system, support for technical problems, and even an outlet to publish
scientific results.

Software

GIS software has improved remarkably in the latest generation and will
undergo still more changes. The basic tools of the computer programmer have
undergone a transition from first generation to object-oriented database
and programming languages, offering some benefits in program module
reusability, improved data handling, and ease of use as more and more
packages are rewritten to take advantage of these tools. The WIMP (windows,
icons, menus, and pointers) interfaces so common today owe their origins to
this technology. Today, the GIS research community suggests that as the
"desktop metaphor" becomes more commonly accepted, increasingly
sophisticated metaphors will take over for organizing computing, including
perhaps using maps themselves to manage the computer rather than vice
versa.

Some changes are far more practical but still of great value. Most software
systems now support context-sensitive help, electronic manuals, and
automatic installation and update procedures. Each of these could benefit
from intelligent software that uses an expert system base and continues to
tailor the system around the GIS operator's revealed use. Such software,
used over a network, has been termed an intelligent agent. Most GIS of the
future will use these methods to seek out new data over the network that
relate to your problem, alert you to mistakes in your data management and
analysis, and perhaps automatically compose maps and reports at the
completion of a project.

Multimedia and hypermedia are also rapidly becoming a component of GIS
software. Multimedia allow simultaneous use of text, sound, animation, and
graphics. GIS software has also developed the ability to interact in many
spoken languages, under different operating systems, and on many different
computers. The independence of the software and the tasks from particular
computer platforms, or even vendors, are a highly desirable element in a
distributed system.

GIS and Public Health

While it holds distinct promise as a tool in the fight against emerging
infectious diseases and other public health problems; it is not simply the
next widget to come into play. GIS can be seen as a new approach to
science, one with a history and heritage, a finite and well researched
suite of methods and techniques, and a research agenda of its own. It does
not fit neatly into the health scientist's toolbox. It requires rethinking
and reorganizing the way that data are collected, used, and displayed. It
requires expense, training, and a climb up a learning curve. It needs
maintenance and support and can be both overwhelming and threatening to the
uninitiated.

On the other hand, the base of research and scholarship using GIS in the
health sciences cannot be ignored. A first step would be to integrate
instruction on GIS into college curricula in public health. An admirable
body of experience in GIS education already exists, even a thoroughly
tested national curriculum that can be easily adapted to a new set of
demands (41). A second step would be to seek out more formal links between
the research communities working with GIS. There are astonishing
similarities for example in the field requirements for using GIS between
forestry, ecology, archeology and epidemiology that could provide
substantial benefits by the sharing of experiences and the pooling of
resources.

Above all, GIS should be seen as improving the set of tools to promote
public health. Good epidemiologic science and good geographic information
science go hand in hand. The future of GIS has already retained a role for
the geographically literate public health expert. Epidemiologists should
seize the opportunity to set their own agenda and influence the technology
and science toward the goal of public health.

Dr. Clarke is professor and chair, Department of Geology and Geography,
Hunter College, and on the faculty of the Earth and Environmental Sciences
Program at the Graduate School and University Center of the City University
of New York. Dr. Clarke's most recent research has been on environmental
simulation modeling, the impact of the Persian Gulf War on the technology
of cartography, and mapping to support disease control programs in Africa.

Address for correspondence:

Keith C. Clarke, Ph. D.,
Department of Geology and Geography
Hunter College-CUNY, 695 Park Avenue, New York, NY 10021, USA
fax: 212-772-5268; e-mail: kclarke@everest.hunter.cuny.edu

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EMERGING INFECTIOUS DISEASES
Volume 2, Number 2, April-June 1996
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[Pages 93-102]

Perspectives

The Evolution and Maintenance of Virulence in Microparasites

Bruce R. Levin, Ph.D.
Emory University, Atlanta, Georgia, USA

Abstract:
In recent years, population and evolutionary biologists have questioned the
traditional view that parasite-mediated morbidity and
mortalityvirulenceis a primitive character and an artifact of recent
associations between parasites and their hosts. A number of hypotheses have
been proposed that favor virulence and suggest that it will be maintained
by natural selection. According to some of these hypotheses, the
pathogenicity of HIV, Vibrio cholerae, Mycobacterium tuberculosis, the
Shigella, as well as Plasmodium falciparum, and many other microparasites,
are not only maintained by natural selection, but their virulence increases
or decreases as an evolutionary response to changes in environmental
conditions or the density and/or behavior of the human population. Other
hypotheses propose that the virulence of microparasites is not directly
favored by natural selection; rather, microparasite-mediated morbidity and
mortality are either coincidental to parasite-expressed characters
(virulence determinants that evolved for other functions) or the product of
short-sighted evolution in infected hosts. These hypotheses for the
evolution and maintenance of microparasite virulence are critically
reviewed, and suggestions are made for testing them experimentally.

How much of the emergence and reemergence of infectious diseases is due to
evolution, rather than ecological, technical, and social change (1)? Under
what conditions will attenuated vaccine organisms become virulent? Are
hospitalized and immunocompromised hosts reservoirs for the evolution of
virulent pathogens (2)? The answers to these and related questions require
an understanding of the ecological conditions and genetic processes
responsible for the evolution and maintenance of parasite-mediated
morbidity and mortality in infected hostsvirulence, as we shall define it
here.

At least since Darwins time (3), evolutionary biologists have been
interested in infectious diseases, but primarily with respect to the role
of these diseases in the adaptation and evolution of humans and other
species (4). A bit more than 15 years ago, this interest in infectious
disease took a new turn, a focus on the microbes responsible for these
diseases and the evolution and maintenance of their virulence. Here I offer
a relatively brief and personal review of current theories of the evolution
and maintenance of virulence in the bacteria, viruses, protozoa, and single
cell fungi, microparasites (to use the term employed by population
biologists), responsible for infectious diseases. I consider how these
theories fit, what is known about the epidemiology of microparasite
infections and the mechanisms of pathogenesis, and discuss procedures to
test hypotheses derived from these theoretical considerations of the
population biology and evolution of microparasites. For other recent
reviews of this subject, see (5-8).

The Conventional Wisdom

At one time, virulence was almost universally considered an artifact of
recent associations between parasites and their hosts (9, 10), and to a
fair extent, it still is (11). In accord with this view, which Bob May and
Roy Anderson called conventional wisdom (12), parasite-host coevolution
is necessarily in the direction of commensalism or, nicer yet, mutualism.
The logic behind this view is pleasing to human sensibilities. A fully
evolved parasite would not harm the host it needs for its survival,
proliferation, and transmission. Indeed, the appeal of this view of nature
of parasite-host coevolution wassufficient for its corollary to also be
assumed valid. That is, pathogenesis is often taken as evidence of recent
associations between parasites and their hosts.

Many observations are consistent with conventional wisdom about
parasite-host coevolution. This is particularly so for most of the
so-called emerging diseases. For example, Legionnaires disease, Lyme
disease, and pneumonia caused by hantavirus are consequences of human
infection by parasites and/or commensals of other species, rather than by
organisms that have had a long association with humans. In fact, for these
emerging diseases and some older microparasitic diseases, like Rocky
Mountain spotted fever, anthrax, and rabies, humans play no (or at best a
negligible) role in the transmission of the parasite and, in that sense,
are an evolutionary dead end. While HIV is transmitted between humans, its
association with our species is almost universally considered recent (13,
14).

Other observations can be interpreted as inconsistent with conventional
wisdom. For some virulent pathogens, like Shigella and Neisseria
gonorrhoeae, humans appear to be either the unique or the dominant host and
vector for infectious transmission (15). For other lethal microparasitic
diseases like malaria and tuberculosis (TB), there is evidence that these
microparasites have had a long history in human populations and that humans
play a major if not unique role in their infectious transmission. However,
for the pathogens involved in both these diseases, animal origins have been
implicated, and it is difficult to find clear evidence of their existence
(or that of other extant pathogens) before the origins of agriculture
(16-18). [The existence of genetic polymorphisms, like sickle cell,
thalassemia, and glucose 6-phosphate dehydrogenase (G6PH) deficiency (19,
20), Duffy-negative blood groups (21), and specific HLA alleles (22)
maintained by Plasmodium-mediated selection can also be interpreted as
evidence for malarias long association with humans. There is evidence for
inherited resistance to TB among mammals (23), and arguments that TB
epidemics have selected for inherited resistance in humans (16). However,
that evidence is not as compelling as that for malaria.] One can always
rescue conventional wisdom from these inconsistent observations by assuming
that long is not long enough for these microparasites to evolve or
coevolve with humans to a more amenable relationship. Then again, it may
well be that some microparasites responsible for new infections in human
hosts will evolve to become increasingly virulent human pathogens and be
readily transmitted between human hosts.

Conventional wisdom is not based on hypotheses that can be readily tested
and rejected. Microparasites that lead to the extinction of their only host
face the same fate as the host. On the other hand, evolving and becoming
gentle and prudent in treating their hosts (when natural selection
operating at the level of individual microparasites favors profligate
behavior like virulence) require some form of group-level or kin selection
(8), and/or a host evolutionary response that unilaterally converts an
otherwise virulent microparasite into a commensal. Conventional wisdom does
not account for the actual mechanisms responsible for the evolution of
benign associations between microparasites and their hosts.

Epidemiologic Models and the Enlightenment

In the early 1980s, at least among evolutionary biologists, conventional
wisdom gave way to what, in an earlier consideration of this subject
Catharina Svanborg and I satirically (but sympathetically) referred to as
the enlightenment (24). In accord with this new view, natural selection
could favor the evolution and maintenance of virulenceas well as commensal
and symbiotic associations between microparasites and their hosts. In other
words, virulence could be the evolved as well as the primitive stage of
these associations. The direction of natural selection in any given
situation depends on the epidemiology and ecology of the microparasite and,
in particular, the relationship between its virulence and its rate of
infectious transmission in the host population. [In at least the
mathematical theory, the morbidity component of microparasite virulence is
not treated explicitly (25). The symptoms and pain resulting from infection
are implicitly incorporated in the rates of disease-associated mortality,
recovery, and transmission. Moreover, while acknowledging the existence of
microparasite-mediated selection and evolution in the host population, for
the most part, the enlightened view of microparasite-host coevolution has
concentrated on the changes in the microparasite population. The idea is
that because of the relatively longer generation times, the rate of
evolution in the host population is going to be low.] This can be seen in
the equation for the finite rate of increase of a directly transmitted
microparasite in a wholly susceptible host population (12, 25, 26), 

[Image not available in ASCII version]

where  is the rate constant of infectious transfer of the microparasite,
N, the density of the susceptible host population, a, the rate of
microparasite-induced mortality (virulence), b, the rate of
microparasite-independent mortality, and v, the rate of recovery. R0 is the
number of secondary infections caused by a single primary infection and
serves as a measure of the fitness (here and elsewhere in a Darwinian
sense) of the parasite in this naive host population. At any given host
density, N, this measure of fitness of the parasite is directly
proportional to its transmissibility, , and the term of its persistence in
an infected host, the reciprocal of a + b + n.

If the parameters of the R0 equation were independent of each other, the
predictions derived from this equation would be consistent with
conventional wisdom: benign parasites would evolve. That is, natural
selection would favor highly transmissible [Image not available in ASCII 
version], incurable [Image not available in ASCII version],
commensals [Image not available in ASCII version], or symbionts 
[Image not available in ASCII version]. On the other hand, if
transmission and virulence, the parameters  and a in the R0 equation, were
positively coupled, natural selection could favor the evolution and
maintenance of some level of virulence, [Image not availiable in ASCII 
version], in the microparasite population.

In accord with the epidemiologic perspective implicit in the R0 equation,
an understanding of the evolution of virulence in microparasites comes down
to elucidating the relationship between the rate at which the microparasite
is transmitted between hosts and the rate of parasite-mediated mortality in
individual infected hosts. If that relationship is positive, then some
level of virulence may be favored. And, since the first statements of this
new view of parasite-host coevolution (12, 26, 27), much of the research on
the evolution of virulence has focused on the association between these two
components of parasite fitness.

The most cited, and to me the single most compelling, evidence in support
of this new interpretation of microparasite-host coevolution comes from the
experiments using myxoma virus to control European rabbit populations in
Australia and Europe (26, 28, 29). Within a relatively short time after the
release of highly virulent myxoma, the viruses recovered from the then
decimated and sometimes more resistant wild rabbitpopulations were less
virulent and had lower rates of disease-induced mortality on control
laboratory rabbits than those initially released. However, the extent to
which myxoma virus from the wild became attenuated was substantially less
than that which could be achieved experimentally (29). This was interpreted
as evidence for a positive coupling between the rates of infectious
transmission and rates of virus-induced mortality, a trade-off between
virulence and transmission. Highly virulent forms of the virus had a
disadvantage because they killed the rabbits too quickly and thus reduced
the time available for them to be picked up by the insect (mosquito or
flea) vectors required for their infectious transmission. Viruses that were
too attenuated had a disadvantage because they generated fewer skin lesions
and had lower densities of circulating virions, which presumably would
reduce the rate at which they would be bitten by these insect vectors, the
likelihood of biting vectors picking up myxoma, and the number of virions
picked up at any given bite. Thus, in contrast to conventional wisdom and
in accord with the enlightened interpretation, natural selection could
favor and maintain the virulence of microparasites. This results when there
is a positive coupling between a parasites virulence and its capacity for
infectious transmission.

The myxoma story is particularly compelling because the quantitative
relationship between virulence and transmissibility inferred from the
epidemiologic data and models was independently tested and demonstrated
experimentally (30, 31). The myxoma story remains the only one for the
microparasites of eukaryotic hosts where the predictions about transmission
and virulence made from an interpretation of epidemiologic observations
were tested experimentally. With few exceptions (32), inferences about the
relationship between transmission and virulence and the trade-offs between
these two attributes of a microparasites association with its host have
been derived from comparative evolution studies or retrospective
interpretations of epidemiologic data. In some cases, these inferences are
reasonably strong, e.g., in the study by Alan Herre (33) on fig wasps and a
nematode parasite and by Deiter Ebert (34) on a planktonic crustacean with
a protozoan parasite. The latter study is particularly convincing because
it includes independent, experimental evidence of a positive correlation
between the density of spores in infected hosts and the virulence and
transmissibility of this protozoan parasite.

The enlightened view on the virulence of microparasites sometimes takes the
positive association between the virulence of a microparasite and its
transmissibility as axiomatic; therefore, it assumes that a microparasites
virulence is constrained solely by the need to keep the host alive to
facilitate its transmission to new hosts. [One way to experimentally augment
the virulence of a microparasite, as, for example, measured by declines in
its LD50, is to artificially pass that microbe between hosts (15). From one
perspective, this result is consistent with the trade-off hypothesis, as
the effect of passage is to make the parasites transmission independent of
the hosts survival, thereby allowing it to become more virulent without
compromising its need to be transmitted to other hosts. However, increased
virulence in a passage experiment is not sufficient evidence for that
trade-off. (It may well be that the parasites capacity for infectious
transmission is impaired as a consequence of whatever increased its
capacity for infectious transmission.) I know of no experiments that
demonstrate that the increase in virulence generated during a passage
experiment is also reflected as increasedtransmissibility, as is necessary
for the trade-off interpretation. Indeed, it may well be that an increase
in the case-mortality rate or a reduction in the LD50 of a microparasite
will be reflected as a reduction in its natural transmissibility.] This is
implicit in much of Paul Ewalds writing on this subject (6, 35) and is the
basis of his main thesis that changes in rates of infectious transmission
will select for microparasite strains or species with different levels of
virulence.

By assuming a necessarily positive relationship between a microparasites
capacity for infectious transmission and the extent of morbidity and rate
of mortality it causes in infected hosts, a positive trade-off
(relationship) between transmissibility and virulence, Paul Ewald has
generated scenarios for the evolution of virulence and changes in virulence
for a diverse array of microparasites, including those responsible for
cholera, influenza, dysentery, and AIDS (6, 35). While the details of
Ewalds stories may differ, the plot is almost always the same: increases
in the rates of transmission favor increases in virulence, and the reverse.
For example, Ewald has postulated that the virulence of HIV observed in
contemporary human populations, AIDS, is in large part due to evolution in
this retrovirus responding to the increases in human-human transmission
rate resulting from more promiscuous sexual behavior. [I quote: Severe
immunodeficiency could develop in an old association [between a sexually
transmitted virus or SIV and its host] as a result of increases in sexual
partner rates causing evolution of increased virulence (p. 143, reference
6).] If rates of unprotected sexual contact decline, so should the
virulence of HIV (p. 144, reference 6). However, even when a direct
relationship between the virulence and transmission rate of HIV is assumed,
a deeper consideration of the epidemiology and course of this sexually
transmitted disease shows that this simplistic conclusion about evolution
and the virulence of HIV is chock full of caveats (36, 37). The relative
contributions of transmission and virulence (as measured by the time before
the onset of AIDS) to the fitness of HIV in the population of hosts depends
on whether the disease is in an epidemic or endemic phase. Moreover, as I
consider later, there are other, very different, hypotheses for the
evolution of the virulence of this retrovirus and other pathogenic microbes
that do not require the necessarily positive association between infectious
transmission and virulence upon which Ewald has based his arguments for the
evolution and maintenance of virulence in microparasites.

A corollary of the hypothesis of a positive trade-off between
transmissibility and virulence is that if all else were equal, increases in
the degree of vertical (e.g., from a mother to a fetus) transmission of a
parasite, relative to its horizontal (infectious) transmission would favor
reductions in its virulence (38). There is compelling, experimental
evidence to support this corollary. However, the evidence is restricted to
experiments with E. coli and its phage, f1, which can be transmitted
vertically, in the course of cell division, or horizontally, by infecting
susceptible, uninfected bacteria (39). While some, like me most of the
time, may believe in the adage what is true for E. coli is true for
elephants, but only more so, other, less coli-centric souls, may want to
see more experiments of this type with microparasites and vertebrate hosts.
I certainly do. 

Within-Host Population Dynamics and Virulence of
Microparasites

There is a dearth of experimental investigations of the quantitative
relationship between the transmission and virulence of microparasites.
During the past few years, however, there has been a flurry of theoretical
studies of the within-host population dynamics of microparasites that have
specifically considered the relationship between the virulence and
transmission rates of microparasites and their densities and/or rates of
replication in infected hosts (40-44). In the simplest models developed in
these theoretical studies of the within-host population dynamics of
microparasites, the virulence of the microparasite, as measured by either
the rate at which it kills its host or its LD50, is assumed to be directly
proportional to its rate of proliferation in that host, and its rate of
infectious transmission is directly proportional to its within-host density
(41). Under these conditions, in the absence of superinfection or mutation,
selection favors microparasites with intermediate rates of within-host
replication, i.e., intermediate levels of virulence. More complex
situations, like the coexistence of microparasite lineages with different
levels of virulence, result when virulence is proportional to the
within-host growth rate of the parasite and single hosts can be infected
with parasites of different growth rates (43) or when there are high rates
of mutation to different levels of virulence within a host (45). Moreover,
with superinfection and mutation, the theory developed in these two reports
predicts that the average level of virulence of a parasite in an infected
host can exceed that anticipated from models that do not allow for
superinfection and/or assume that the parasites level of virulence in an
infected host remains invariant.

The Convergence of Theories

The predictions that can be made on the basis of the current view of the
evolution of virulence differ from predictions that might follow
conventional wisdom because the new view allows for natural selection in
the parasite population to favor the evolution and maintenance of some
level of virulence. Moreover, even when there is a positive association
between a parasites virulence and its transmissibility, under the
conditions described in the following paragraph, the predictions of new
methods can still converge with those of conventional wisdom.

If the density of the sensitive host population is regulated by the
parasite, an extension of the enlightened theory predicts that natural
selection in the microparasite population can lead to continuous declines
in the level of virulence, possibly to immeasurable values (46). Although
not stated in this general way, the same conclusion about declining
virulence can be drawn from models of the epidemiology of HIV/AIDS (36,
37). During the epidemic phase of a microparasitic infection, when the host
population is composed primarily of susceptible hosts, selection favors
parasites with high transmission rates and thus high virulence. As the
epidemic spreads, the proportion of infected and immune hosts increases and
the density of susceptible hosts declines. As a result, the capacity for
infectious transmission becomes progressively less important to the
parasites Darwinian fitness and persistence in the host population.
Selection now favors less virulent parasites that take longer to kill their
host and, for that reason, are maintained in the host population for more
extensive periods. Analogous arguments have been made for the latent period
of a bacteriophage infection (47), the evolution of lysogeny (48), the
tradeoff between vertical and horizontal transmission (49, 50), and the
advantages of microparasite latency in general (40).

Alternative Models for the Evolution of Microparasite Virulence

For any microparasite, the rate of transmission between hosts will always
be a significant component of fitness, and, if all else is equal, parasites
transmitted at higher rates in the host population have a selective
advantage over less transmissible forms. On the other hand, there is no
reason to assume that in general a microparasites rate of infectious
transmission will be positively associated with its virulence. Moreover,
even when there is no relationship or a negative relationship between
transmission and virulence, there are at least two ways by which natural
selection can lead to the evolution and maintenance of virulence,
coincidental evolution (24) and short-sighted within-host selection (51).

Coincidental Evolution

According to the coincidental evolution hypothesis, parasite-mediated
morbidity andmortality are what Gould and Lewontin (52) likened to the
spandrels of gothic churches. While these structural necessities may frame
the frescos and paintings within, that is not the reason for their
existence. They are architectural constraints. Analogously, the factors
responsible for the virulence of a microparasite in an infected host may
have evolved for some purpose other than to provide the parasite an
advantage within a host or its transmission to other hosts.

It would be difficult to account for the evolution of botulism toxin by
selection favoring Clostridium botulinum that kill people who eat
improperly canned food. The same argument could be made for the toxins of
C. tetanae and possibly for those produced by other free-living Clostridia.
Although these organisms may proliferate in humans, they are soil bacteria,
and the effects of the toxin may not contribute to their capacity to
colonize, proliferate, and be maintained in humans or to their capacity to
be transmitted between human hosts. How many other microparasite-induced
symptoms, and the resulting host morbidity and mortality, provide no
advantage to that microbe in (or on) a host or its transmission between
hosts? Did the lipopolysaccharides and other components of bacterial cell
walls and cell membranes evolve because the fitness of bacteria expressing
them is enhanced by endotoxininduced overresponse of the immune system
responsible for the morbidity and mortality of sepsis (53)? Do the toxins
confer an advantage on E. coli O157 or Staphylococcus aureus (or the
plasmids and phages that code for these toxins) because they produce,
sometimes lethal, symptoms in infected hosts, hemolytic uremic and toxic
shock syndromes, respectively? An earlier paper on this subject (24) argued
that the adhesins produced by the E. coli responsible for the morbidity of
symptomatic urinary tract infections evolved and are maintained to
facilitate colonization of the gut. The painful symptoms of urinary tract
infections generated by an inflammatory response to these adhesins may
confer no advantage for the E. coli expressing them in the urinary tract
and may in fact lead to the clearance of those bacteria (24).

Each of the symptom-inducing toxins and adhesins described above, as well
as many other so-called virulence determinants (54) may indeed facilitate
the microparasites ability to colonize, proliferate, or be maintained in
infected hosts, and/or be transmitted between hosts. This certainly sounds
reasonable for many virulence determinants, e.g., the somatic cell
invasiveness mechanisms of Shigella, the capsules of Streptococcus, the
diarrhea-inducing toxins produced by Vibrio cholerae, and the sneezing and
coughing induced by rhinoviruses. On the other hand, it is necessary to
formally test this hypothesis that these symptoms have that effect and
reject the alternative, that the morbidity and mortality generated by the
expression of a specific virulence determinant provides neither a within-
or between-host (infectious transmission) advantage to the parasite.

Short-Sighted Evolution

Natural selection is a local phenomenon. Characters that confer a survival
or replication advantage on the individual organisms that express them at a
given time or in a given habitat will be favored and evolve at that time
and in that habitat. Whether the expression of those temporally or locally
favored characters will increase or reduce the fitness of that organism at
other times or in other habitats is irrelevant. Also irrelevant is whether
a locally favored character makes the population better or less adapted to
its environment at large or augments the likelihood of its survival in the
future. This myopia is a fundamental premise of the theory of evolution by
natural selection and the basis of theshort-sighted evolution hypothesis
for microparasite virulence (51).

Within an infected vertebrate host, microparasite populations go through
many replication cycles and may achieve very high densities. They may also
reside and proliferate in many different subhabitats (tissues and cells)
and confront a variety of different and ever-changing constitutive and
inducible host defenses which may, sequester, kill, or in other ways
inhibit their proliferation. As a consequence of classic mutation,
transposition, and recombination, genetic variability will be continually
generated in the populations of infecting microbes. Mutant or recombinant
microparasites that are better able to 1) avoid being done in or inhibited
by the hosts defenses; 2) proliferate in the host; or 3) invade and
replicate in novel habitats, tissues, and cells where there is less
competition from members of its species would have an advantage in that
host. This would occur even when the expression of the characters
responsible for that local advantage reduces likelihood of the transmission
to other hosts. Stated another way, the morbidity or mortality caused by a
microparasite infection could be the result of the within-host evolution
that is short-sighted because that virulence actually reduces the rate at
which that parasite is transmitted to other hosts.

Three examples of microparasite virulence that could be products of this
mode of evolution can be considered (51). For two of these examples,
bacterial meningitis and poliomyelitis, many human hosts are infected by
the responsible microparasites, primarily Haemophilus influenzae, Neisseria
meningitidis, and Streptococcus pneumoniae for meningitis and poliovirus
for poliomyelitis, but very few manifest the symptoms of these infections.
In the case of meningitis, the neurologically debilitating and sometimes
fatal symptoms of the infection are a consequence of an inflammatory
response against the bacteria entering and proliferating in the cerebral
spinal fluid. These meningitis-causing bacteria normally reside in the
nasopharyngeal passages and are transmitted by droplet infection. The
cerebrospinal fluid is, at least with respect to their infectious
transmission, a dead end. On the other hand, bacteria capable of invading
and proliferating in that habitat could have a local advantage as there are
no other competing populations and only modest defenses. An analogous
argument can be put forth for poliovirus. Symptomatic infections with this
virus are caused by their invasion of, and proliferation in, the neurologic
tissue of the central nervous system. Poliovirus normally replicates in the
mucosal cells of the mouth, throat, and intestines and is transmitted by
the oral-fecal route. Poliovirus virions proliferating in the central
nervous system would almost certainly not be transmitted. The evidence in
support of short-sighted evolution for the virulence of these specific
microparasites is mostly circumstantial (51). On the other hand,
short-sighted evolution for the virulence of specific microparasites is a
hypothesis that can be tested. If the hypothesis is valid, the
microparasites responsible for the symptoms would be genetically different
from their ancestors that infected the host and better adapted for
proliferation in the site of the symptoms than the ancestors themselves.

The third example of short-sighted evolution of virulence considered, HIV,
is different from the other two in that virtually every human infected with
this retrovirus that does not die of other causes, eventually manifests and
succumbs to AIDS. However, although the case mortality of HIV infection may
approach unity, as measured by the rate of mortality (deaths per unit of
time), from an epidemiologic perspective, HIV is not a very virulent virus.
There is substantial variation in the time between infection and the onset
of AIDS. On average in industrialized countries, the term of this infection
is 8 to 10 years (55). During the early phase of an HIV epidemic, most
transmission of the virus occursduring the initial viremia, probably before
seroconversion and certainly before the onset of AIDS (37, 56). It is not
at all clear how the transmissibility of HIV virions during this early
phase of the infection is related to the time of onset of AIDS. HIVs that
are more transmissible early in the infection may lead to an earlier onset
of AIDS. If this is the case and all else were equal, increasing
opportunities for transmission during the epidemic phase would favor
increases in HIV virulence (36, 37). However, there may be no association
between HIVs capacity to be transmitted early and the time of onset of
AIDS, or the time until the onset of AIDS may increase with the
transmissibility of the virus during the early phase of the infection.
Under either of these conditions, selection during the epidemic phase of
the disease would favor more transmissible but less virulent HIVs.

In the course of HIV infection, the HIV population undergoes continuous
genetic changes. In fact, in a number of hypotheses of HIV pathogenesis,
AIDS is a consequence of mutation and selection in the HIV population that
occurs during the course of the infection in individual hosts (57-60),
i.e., short-sighted, within-host evolution. Albeit different in their
details, all of these hypotheses are consistent with what is known about
HIV infection, and all can account for the course of these infections and
variable time of onset of AIDS.

Experimental Evolution Meets Experimental Epidemiology

Results of recent studies by population and evolutionary biologists predict
at least three ways by which the virulence of microparasites can be favored
and will be maintained by natural selection. 1) Direct selection: there is
a positive relationship between the parasites virulence and its rate of
infectious transmission; 2) coincidental evolution: the parasites
virulence is due to character(s) favored and maintained by selection for
some other function and the expression of those virulence determinants in
an infected host does not confer a net advantage or disadvantage in the
parasite population at large; and 3) short-sighted, within-host, evolution:
the parasites responsible for the morbidity and mortality of an infection
are selected for within the host because of a local advantage, and that
evolution reduces the rate at which that locally adapted parasite is
transmitted between hosts.

At this time, these predictions are based almost entirely on general theory
and retrospective interpretations of epidemiologic and other observations
about specific microparasites. Although this theory and these
interpretations may be appealing, in a formal Popperian sense (61), almost
all the mechanisms postulated for the evolution of virulence of specific
microparasites are no more than untested hypotheses. However, unlike most
evolutionary hypotheses, those about the evolution of microparasite
virulence can be tested and rejected with prospective, experimental studies
with laboratory animal and plant hosts. These tests could be at two levels;
first, tests of the validity of the assumptions behind these models of the
evolution of virulence and second by tests of the predictions made from the
consideration and analysis of these models.

For the direct selection hypothesis, it is essential to demonstrate a
positive relationship between a microparasites virulence and its rate of
(or capacity for) infectious transmission. For mammalian hosts, protocols
exist for determining this relationship (30-32). The object would be to
estimate the densities of microbes at the sites of transmission (e.g.,
feces, nasal passages) during the entire course of the infection. Moreover,
it would be useful to separately test the colonization ability and
virulence of the microbes fromthese sites. According to the coincidental
evolution hypothesis, it is possible that the virulence determinant
responsible for morbidity and mortality in the host provides a local
advantage to the parasite expressing it; whether it does or not could be
tested with competition experiments between strains of that microparasite
that are isogenic save for that virulence determinant. The genetic basis of
many virulence determinants are known, and it should be possible to
construct these strains. However, unlike in the direct selection
hypothesis, in coincidental evolution, microbes expressing the virulence
determinants should not be over represented at the sites of infectious
transmission. Under the short-sighted evolution hypothesis, microbes
isolated from the tissues and organs responsible for the symptoms of the
infection (e.g., in the cerebrospinal fluid) should be better adapted for
proliferation in those organs and tissues than the originally infecting
strain from which they were derived. This could be tested with pairwise
competition experiments between the original and potentially evolved
strains injected at the site of the symptoms with a common, genetically
marked competitor of that parasite. Here, too, it is necessary to
demonstrate that the strain responsible for the symptoms is not
overrepresented at the site of infectious transmission.

To test the prediction of the direct selection hypothesis and to exclude
that mechanism in tests of the coincidental and short-sighted alternatives,
it is necessary to study the epidemiology of the microparasites as well as
their within-host properties. For bacteriophages and bacteria this is a
relatively easy task, e.g., testing Abedons hypothesis (47) about the
direct relationship between the density of sensitive bacteria and selection
for latent period length (a measure of virulence) and burst size (a measure
of transmissibility). For eukaryotic hosts, this kind of study is going to
be more difficult and, at this time, may not be possible. The basic
protocols for experimental studies of the epidemiology of bacterial and
viral infections of laboratory mice were developed and >successfully
employed a long time ago (32, 62). Greenwood and colleagues (32), studied
microparasites with different transmissibilities (infectivity) and
virulence.In one replica of their study of pasteurellosis (due to
infections with a bacterium they call Pasturella muriseptica in
experimental populations of mice, they report the appearance of a variant
that had gained infectivity and retained, or perhaps increased, its
original virulence. However, in general, their study and Fenners (62)
provide little information about the direction of natural selection in
these microparasite populations. With respect to evolutionary questions,
these were wait-and-see experiments. Only one strain of microparasite was
introduced into each population, and it was necessary to wait for mutations
that changed their virulence or transmissibility. More information about
the direction of selection and a better test of these evolutionary
hypotheses could be obtained in these types of experiments if two or more
genetically marked strains of microparasites with different virulence and
transmissibility were introduced simultaneously and were allowed to
compete. However, experiments of these types are costly, labor-intensive,
and time-consuming, and because of concerns about animal rights, it may be
difficult to get permission to do these experiments with mice or other
higher vertebrates. On the other hand, experiments of this type with
insects and other invertebrate animal hosts as well as plants would be
tenable and valuable as tests of the general theory, albeit less
immediately relevant to the evolution and maintenance of virulence in human
pathogens.

Acknowledgments

I thank Marc Lipsitch, Deiter Ebert, Jim Bull, David Thaler, and Tomoko
Steen for reading this manuscript and for many insightful comments and
suggestions, most of which I agreed with and a few of which Ive
incorporated. This endeavor was supported by a grant from the National
Institutes of Health, GM33782.

Dr. Levin is professor of biology at Emory University and director of the
Graduate Program in Population Biology Ecology and Evolution. He is a
population and evolutionary biologist, who, like a number of others of his
ilk, recently discovered infectious disease. Currently he and the
postdoctoral fellows and students working with him are doing theoretical
(mathematical modeling) and experimental research on the within-host
population dynamics of bacterial infections and their treatment, and the
epidemiology, population genetics, and evolution of antibiotic resistance.

Address for correspondence: Bruce R. Levin, Ph. D.,
Emory University, EcLF, 1510 Clifton Road, N.E., Atlanta, GA 30322, USA;
fax: 404-727-2880;
e-mail: biobrl@emuvm1.cc.emory.edu

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     paradigm: a critique of the adaptationist programme. Proc R Soc Lond B
     Biol Sci 1979; 205:581-98.
 53. Whitnack E. Sepsis. In: Schaechter M, Medhoff G, Eisenstein BI,
     editors. Mechanisms of microbial disease. Baltimore: Williams &
     Wilkins, 1993: 770-8.
 54. Finlay BB, Falkow S. Common themes in microbial pathogenicity.
     Microbiol Rev 1989; 52:210-30.
 55. Fauci AS. Multifactorial nature of human immunodeficiency virus
     disease: implications for therapy. Science 1993; 262:1008-11.
 56. Jacques A, Koopman JS, Simon CP, Longini IM. The role of primary
     infections in epidemics of HIV infections in gay cohorts. J Acquir
     Immune Defic Syndr 1994;7:1169-84.
 57. Nowak MA, Anderson RM, McLean AR, Wolfs TFW, Goudsmit J, May RM.
     Antigenic diversity thresholds and the development of AIDS. Science
     1991; 254:963-9.
 58. McLean AR. The balance of power between HIV and the immune system.
     Trends Microbiol 1993; 1:9-13.
 59. Mittler JM, Antia R, Levin BR. Population dynamics of HIV
     pathogenesis. Trends in Ecology and Evolution 1995; 10:224-7.
 60. Mittler JM, Levin BR, Antia R. T-cell homeostasis, competition and
     drift: AIDS as HIV-accelerated senescence of the immune repertoire. J
     Acquir Immune Defic Syndr Hum Retrovirol (in press).
 61. Popper KR. The logic of scientific discovery. New York: Harper, 1965:
     479.
 62. Fenner F. The epizootic behaviour of mousepox (infectious ectromelia
     of mice) II. The course of events in long-continued epidemics. J
     Hygiene 1948; 46:383-93.

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

---------------------------------------------------------------------------
[Pages 103-108]

Perspectives

The Infectious Diseases Impact Statement: A Mechanism for Addressing
Emerging Diseases

Edward McSweegan, Ph.D.
National Institute of Allergy and Infectious Diseases, National Institutes
of Health, Bethesda, Maryland, USA
---------------------------------------------------------------------------

Abstract:
The use of an Infectious Diseases Impact Statement (IDIS) is proposed for
predictive assessments of local changes in infectious diseases arising from
human-engineered activities. IDIS is intended to be analogous to an
Environmental Impact Statement. The drafting of an IDIS for specific
activities, particularly in developing nations, would provide a formal
mechanism for examining potential changes in local health conditions,
including infected and susceptible populations, diseases likely to
fluctuate in response to development, existing control measures, and
vectors likely to be affected by human activities. The resulting survey
data could provide a rational basis and direction for development,
surveillance, and prevention measures. An IDIS process that balances
environmental alterations, local human health, and economic growth could
substantially alter the nature of international development efforts and
infectious disease outbreaks.

A 1995 report by Aksoy et al. (1) describing the GAP (Turkish acronym for
the Southeastern Anatolia Irrigation Project) irrigation project in Turkey
suggests that anticipating the emergence or expansion of vector-borne and
zoonotic diseases in a limited environment is a useful exercise. According
to the report, a number of diseases (e.g., leishmaniasis, malaria, and
schistosomiasis) are likely to increase in direct response to the expansion
of irrigation and the increases in under water acreage and human population
in the GAP region. The succinct overview of the disease and vector
conditions in the GAP area could serve as a starting point for creating
what will be referred to in this article as an Infectious Diseases Impact
Statement (IDIS), a document that would be analogous to the Environmental
Impact Statement (EIS) routinely used in the United States to assess the
likely effects of construction, irrigation, agriculture, and similar
activities on a local environment or region. An IDIS, however, would not
assess the environment directly, but rather would predict changes in local
disease patterns resulting from changes to the local environment.

Like an EIS, an IDIS would be a predictive and proactive assessment.
Drafting an IDIS for a particular region or microenvironment would provide
a formal mechanism for asking (and attempting to answer) specific questions
about future changes in local health conditions. For example, what are the
diseases and vectors in the given area? How are the proposed changes to the
environment (e.g., dam-building, forest-clearing) likely to change the
incidence and the prevalence of those diseases and vectors? What actions
should be taken during the course of a given project and in the future to
prevent potential increases in disease and vector populations? If an
increase in human disease is likely, is the expense of the proposed project
warranted? Will the economic benefits of a particular development or
agriculture project be offset by increased costs in health care,
vaccination, and vector control?

The 1969 National Environmental Policy Act was designed to provide a legal
mechanism in the United States for evaluating potential impact to the
environment from development activities and for permitting the public to
participate in the evaluation process at the earliest stages
(i.e.,scoping). A Council of Environmental Quality in the executive
branch of the federal government was established as a monitor, and the EIS
process was implemented to inform decision makers and the public of
potential environmental problems and reasonable alternatives to proposed
actions. Environmental Protection Agency (EPA) requirements for
environmental assessments are outlined in the Code of Federal Regulations
(CFR). An EIS is intended to prospectively examine impact upon the quality
of the human environment of the United States and, in appropriate cases,
upon the environment of the global commons outside the jurisdiction of any
nation (46 CFR sec. 504.7). The requirements of an EIS typically include
descriptions of human populations in the designated area, current land use
patterns, air quality, noise levels, locations of wetlands and coastal
zones, sites of historical or cultural value, and non-point source
pollution. The protection of human health is implied in the EIS process,
but this concern is usually assumed to pertain to the location of
industrial plants and dumps and to exposure to toxic chemicals, heavy
metals, ionizing radiation, and pesticides. The EIS process contains no
explicit reference to infectious diseases or disease vectors affecting
human health in response to deliberate environmental changes. In 1995, the
only published EIS references to diseases, infectious or otherwise, were
for proposed control measures at two California plant nurseries. Yet past
events suggest that attention should be directed toward changes in
infectious disease patterns directly attributable to human-engineered
events.

For example, the construction of the Aswan Dam in Egypt is widely believed
to have precipitated the appearance of Rift Valley fever (RVF) in Egypt
during the 1970s (2). Tens of thousands of RVF cases and hundreds of deaths
followed. Similarly, completion of the Diama Dam in Senegal, in 1987, led
to epidemics of malaria and RVF (3); impoundment of the Volta Lake in
Ghana, in 1968, led to an explosive outbreak of schistosomiasis (4).
Increased agriculture on the Argentine pampas and along the edges of
Bolivian forests has contributed to frequent hemorrhagic fever outbreaks
caused by Junin and Machupo viruses, respectively (5). Mining operations in
the Brazilian jungles have led to outbreaks of malaria (6). Road-building
projects under way in Papua New Guinea are likely to bring large numbers of
susceptible human hosts into contact with rare and yet-to-be-discovered
viruses. These epidemics and encounters with new diseases are the
unforeseen consequences of critically altering the local environment. As a
consequence, development and agriculture projects initiated to improve the
lives of local populations can have the opposite effect by increasing
disease prevalence and causing new epidemics. Embedding an IDIS requirement
into the planning and execution of large-scale projects likely to alter
local environments could prevent new epidemics and reduce infectious
disease associated morbidity and mortality.

How Would an IDIS Work?

An IDIS would first need to be established as an integral component of any
activity likely to affect the health of a local population. In tropical and
developing regions of the world, that would include a variety of national
and international development activities. The area designated for
large-scale alteration would be surveyed for current disease vectors, and
the local populations would be examined for diseases likely to be affected
by the project in question. The quality of the surveillance and the
extrapolation of expected changes brought on by a particular activity would
vary, depending on the knowledge of diseases, vectors, local host immunity,
and other factors. Although the variables increase the margin of
uncertainty in such extrapolations, theseestimates would be expected to
improve as the state of field and laboratory research improves and
experience with preparing an IDIS increases. A retrospective examination of
earlier projects in similar environments would also provide information for
developing an IDIS. The standards of existing credible scientific
evidence and reasonably foreseeable impact that current environmental
impact assessments rely on could also be applied to the early stages of the
IDIS process.

The resulting preproject assessment would provide a snapshot of conditions
in a defined area, including the following: diseases likely to fluctuate in
response to project activities, numbers of infected and susceptible hosts,
existing control measures, and vectors likely to be affected by project
activities. Such baseline data are frequently absent from development and
agriculture activities (7). Knowing what diseases are already present, and
how they might be changed, allows one to ask how anticipated changes in
disease prevalence and distribution might be prevented or controlled
through changes in the proposed project, improved case finding and
treatment, changes in local sanitation and housing, increased vaccination
or prophylaxis, or pest management programs. Some or all of the above
health maintenance measures could then become components of the overall
planning, budgeting, and execution of any major development or agricultural
activity in the area. Health and health maintenance would become factors in
the overall design and cost of the project. In many instances, local
disease surveillance would become an ongoing part of the project, with
supplemental assessments being made to refine the original IDIS.

Who Would Request an IDIS, and Who Would Respond to the Request?

Initial candidates would likely be donor organizations (the U.S. Agency for
International Development and the World Bank, for example) that provide
funding and oversight. In the absence of federal or international statutes,
these organizations have the stature and financial capability to make
infectious disease control an integral part of their development projects.
Indeed, they should have an urgent interest in doing so because increases
in diseases or new epidemics increase financial demands on them for
medicines, vaccines, and pest control. In the end, more money would be
spent beating back the outbreaks and epidemics that foresight might have
prevented. National health ministries, state and territorial health
departments, and local medical communities in developing countries might
also request or initiate an IDIS. The practice of drafting an IDIS and
implementing its recommendations might also rejuvenate underfunded areas of
international health, vector biology, parasitology, and medical entomology
as professionals in these fields are called on to conduct infectious
disease assessments of development activities. The peer-reviewed literature
and electronic services such as ProMED, Outbreak, and the World Health
Organization (WHO) and Centers for Disease Control and Prevention
World-Wide Web sites could provide the public scoping role that posting
in the Federal Register and allowing a period for public comment provide in
the EIS process in the United States.

The first application of an IDIS to a large-scale development or
environmental activity could come from western donor organizations working
in the developing world. The successful demonstration of an IDIS could
encourage other organizations, national health officials, and health
activists to push for the routine integration of public health with
national development.This could happen in the United States, as well. The
United States recently experienced the emergence of Sin Nombre virus in the
Southwest and is theoretically open to the introduction of five
vector-borne diseases: malaria, Rift Valley fever, yellow fever, dengue,
and arbovirus encephalitides (15). Public health officials and citizen
activists could initiate independent IDIS for activities perceived to
threaten the balance between health, the environment, and domestic
productivity.

What are the Strengths and Limitations of the IDIS Process?

A project-embedded IDIS would not be the same as an environmental
management program, which seeks to control disease vectors through
environmental modification and manipulation and through reduced human
contact with vectors (8). An IDIS would, in fact, precede environmental
management control measures by first postulating the likely emergence of
specific pathogens and vectors. The usefulness of an IDIS lies in its
ability to provide a conceptual framework for identifying potential disease
problems, and, indeed, preventing them by altering or curtailing the very
activities that could lead to disease emergence.

In an activist sense, an IDIS could be wielded as a tool of caution or
prevention, much as an EIS is wielded in the United States to alter or halt
some activity perceived to be a threat to the environment. That ability to
influence potential changes and to affect health could be vital; public
health concerns connected with agricultural and developmental projects are
usually a low priority among foreign ministries, international donor
organizations, and engineers (9); neglecting them can leave the full
benefits of development unrealized.

Lest anyone imagines that an IDIS could be used solely as a tool of the
political Left, as a kind of liberation microbiology, it is important to
point out that the same IDIS could be used to justify the use of pesticides
and other organized control measures, including the relocation of local
populations. Recently, for example, pesticide use has come under attack by
various environmental groups, and donor organizations have become
increasingly reluctant to fund such activities (10). In the United States,
EPAs Endangered Species Act has also tended to thwart the use of
pesticides because of potentially adverse impact on some birds and mammals
(11). However, an IDIS describing the probable emergence of important
disease vectors could be used to justify such use. Thus, a health care
issue could be twisted into a health scare by either the political Right or
the Left. The recent ratification of the North American Free Trade
Agreement (NAFTA) was preceded in the United States by an effort to stall
the treaty with an EIS requirement. If an IDIS had predicted new disease
outbreaks from increased border trade and traffic, that concern might have
had greater impact on the public imagination than more abstract concerns
about atmospheric particulates in the border region and could have been
effectively used by anti-NAFTA forces. An IDIS should be not a political
tool but rather a valuable information source that helps guide economic
development and land use.

How Can an IDIS Complement Existing Surveillance Systems?

Almost half of the planets five billion people are at risk for one or more
vector-borne diseases (12, 13). Surveillance remains a key tool for
monitoring these diseases and identifying new cases and outbreaks. Four
types of surveillance are used in the control of vector-borne diseases:
1)recording human cases, 2) determining vector distribution and
infectivity, 3) monitoring vertebrate reservoirs, and 4) tracking weather
patterns to predict vector distribution (14). But throughout the developing
world and across tropical boundaries, effective and continuous surveillance
is extremely difficult, if not impossible. Cases are missed; outbreaks go
unreported. Effective case reporting and continuous field monitoring are
best conducted in limited, well-defined areas. Within the microenvironments
of human activities, an IDIS could provide valuable baseline surveillance
data before changes to that area occur and affect disease and vector
distributions. This information could provide a rational basis and
direction for ongoing monitoring and corrective measures (e.g.,
vaccination, relocation, pest control). Focusing on a limited area and a
limited number of diseases in that area may also expand the use of
promising but underutilized technologic methods such as remote sensing and
geographic information systems (GIS).

Haines et al. (15) noted the importance of vector-borne disease monitoring
and recommended that remote sensing and GIS be used to detect changes in
ecosystems and vector populations. To a large extent, however, the
advantages of satellite imagery and GIS have not been realized, in part,
because of the frequent absence of ground truth (data on diseases,
vectors, and other factors in the area) and of having to wait to observe
natural environmental changes likely to affect disease and disease
transmission (16-18). Satellite imagery for much of the planet has been
collecting in databases since 1972 (16). By 1998, accumulated satellite
data will be in the petabyte (1,000 terabyte) range, 1,000 times larger
than the contents of the Library of Congress (B. Montgomery, NASA, pers.
comm.). High-resolution, multispectral, multiyear images for many potential
development and agricultural sites are available. Using a preproject IDIS
to ground truth the projects environment with current satellite imagery,
it may be possible to more completely describe local disease and vector
conditions and make more accurate predictions about their plasticity during
periods of construction, flooding, or farming. The result would be a firmer
linkage of ground surveillance and satellite imagery to monitor public
health changes within a well-defined and limited environment.

In recent years, the sudden emergence of rare or forgotten diseases such as
Ebola virus infection, dengue, yellow fever, plague, and hantavirus (Sin
Nombre virus) infection has attracted the attention of the public and
inspired renewed commitments to surveillance and control. WHO recently
formed a rapid response unit (the Division of Emerging, Viral and Bacterial
Diseases Surveillance and Control) to deal with outbreaks of new and
reemerging infections (19). Similarly, nine Southeast Asian countries held
a meeting on emerging diseases and concluded that each country should also
develop rapid response teams for epidemics (20). However, these disease
control efforts are almost entirely passive, with staff, equipment, and
budgets idling in anticipation of something eventually happening somewhere.
It is difficult to maintain a high degree of public and financial support
for such wait-and-see approaches to disease control. The United States has
suffered a serious decline in national surveillance and outbreak
investigations, in part, because of decreased support for passive
monitoring programs (11).

Is an IDIS Really Needed When the Existing EIS Statutes Already Cover Human
Health and Safety Concerns?

In the United States the need is not clear. Infectious diseases caused by
environmentalmanipulation may be assumed to fall under the general EIS
category of human health. However, infectious diseases have not often been
considered in the past, and it is easy to imagine that if they were a
factor in the EIS process, an environmental/infectious disease issue could
be smothered under the weight of government regulations and the adversarial
legal system. EPA operates under 16 federal statutes and 70 congressional
committees and subcommittees and is engaged in some 600 lawsuits at any
given time (21). Moreover, emerging infectious disease issues could bring
EPA and the EIS process into conflict with the missions of federal agencies
and state and local health departments. Outside the United States, beyond
federal statutes and informed public debate, the need for an IDIS is
clearer. In the developing world, epidemics and substandard health care are
common, and the national goals of healthy environment and healthy economy
are usually at odds. An IDIS process that balances environmental
alterations, local human health, and economic development could
substantially alter the nature of international development efforts and
infectious disease outbreaks.

To the ancient Greeks, the past appeared in front of them, real and
visible; the future was behind them, unseen and unknowable. With that
perspective, they were always glancing nervously backward, looking for a
future that usually managed to creep up and tap them on the shoulder. In a
sense, we have the same perspective for disease surveillance and control
that the ancient Greeks had for time. Past epidemics and our responses to
them are readily apparent; it is that unexpected tap on the shoulder by a
hantavirus or an Ebola virus that is always so startling. We cannot know
when and where such pathogens will emerge. Their appearance is often a
chance event initiated by unpredictable changes in weather or the
accidental encounter of a single person with a mysterious vector. These
taps on the shoulder are an affront to our sense of control and
understanding of disease. Moreover, it is unsettling to the publics sense
of security and its faith in medical research. Although we cannot expect to
eliminate the surprises of emergent pathogens in the near future, we can
take control of situations in which our own actions directly lead to the
emergence of diseases. Generating an IDIS in areas where human activities
are likely to disrupt endemic-disease patterns would be an important step
in controlling future outbreaks. Routine application of a preproject IDIS
could improve local surveillance and health care planning by 1) providing
baseline data on endemic-disease and vector prevalence and competence; 2)
embedding projected health maintenance costs into the planning and cost of
any project or activity likely to influence the environment and public
health; and 3) providing a mechanism for instituting project alterations
and health care measures to offset adverse effects on the health of local
populations.

Acknowledgments

Special thanks to Dr. Michael Gottlieb, National Institute of Allergy and
Infectious Diseases, Parasitology and International Programs Branch, and
Donald C. Baur, Esq., Perkins-Cole, Washington, D.C., for critical comments
and suggestions.

Dr. McSweegan is a member of the Parasitology and International Programs
Branch at the National Institute of Allergy and Infectious Diseases
(NIAID), National Institutes of Health, Bethesda, Md. He has a
long-standing interest in international science and health, and emerging
diseases. He served in the U.S. State Department as an AAAS Fellow before
joining NIAIDs Tropical Medicine and International Research Office in
1988.

Address for correspondence:

Edward McSweegan, Ph.D.
National Institute of Allergy and INfectious Diseases
National Institutes of Health
Solar Bldg., Rm. 3A34
Bethesda, MD 20892-7630, USA
fax: 301-402-0659; e-mail: em8p@nih.gov

References

  1. Aksoy S, Ariturk S, Armstrong MYK, Chang KP, Drtbudak Z, Gottlieb M,
     et al. The GAP project in southeastern Turkey: the potential for
     emergence of diseases. Emerging Infectious Diseases 1995; 1:62-3.
  2. Meegan JM, Shope RE. Emerging concepts on Rift Valley fever. In:
     Pollard M, editor. Perspectives in virology. New York: Alan R. Liss,
     1981.
  3. Lederberg J, Shope RE, Oaks SC, editors. Emerging infections:
     microbial threats to health in the United States. Washington, DC:
     Institute of Medicine, National Academy Press, 1992;71-2.
  4. Scott D, Senker K, England EC. Epidemiology of human Schistosoma
     haematobium infection around Volta Lake, Ghana, 1973-75. Bull WHO
     1982;60:89-100.
  5. Morse SS. Emerging viruses: defining the rules for viral traffic.
     Perspect Biol Med 1991;34:387-409.
  6. de Andrade ALSS, et al. High prevalence of asymptomatic malaria in
     gold mining areas in Brazil. Clin Infect Dis 1995;20:475.
  7. Service MW. Rice, a challenge to health. Parasitol Today 1989;5:162-5.
  8. Ault SK. Environmental management: a re-emerging vector control
     strategy. Am J Trop Med Hyg 1994;50(Suppl):35-49.
  9. Silver GA. 1995. International Health Organization Policy Watch. The
     Federation of American Scientists.
     (http://www.clark.net/pub/gen/fas/ihm).
 10. Arata AA. Difficulties facing vector control in the 1990s. Am J Trop
     Med Hyg. 1994;50(Suppl):6-10.
 11. Longstreth J, Wiseman J. Human health. In: Smith JB, Tirpak DA,
     editors. The potential effects of global climate change on the United
     States: Appendix G, Health. Washington, DC: U.S. Environmental
     Protection Agency, 1989.
 12. Beck LR, Rodriguez MH, Dister SW, Rodriguez AD, Rejmankova E, Ulloa,
     et al. Remote sensing as a landscape epidemiologic tool to identify
     villages at high risk for malaria transmission. Am J Trop Med Hyg
     1994;51:271-80.
 13. Knudsen AB, Sloof R. Vector-borne disease problems in rapid
     urbanization: new approaches to vector control. Bull WHO 1992;70:1-6.
 14. Consortium for International Earth Sciences Information Network
     (CIESIN) Thematic Guides. Provisional Release. 1995. Programs for
     surveillance, treatment, and control of vector-borne diseases.
 15. Haines A, Epstein PR, McMichael AJ. Global health watch: monitoring
     impacts of environmental change. Lancet 1993;342:1464-9.
 16. Washino RK, Wood RL. Application of remote sensing to arthropod vector
     surveillance and control. Am J Trop Med Hyg. 1994;50Suppl:134-44.
 17. Rogers DJ, Williams BG. Monitoring trypanosomiasis in space and time.
     Parasitology 1993;106:S77-S92.
 18. Barinaga M. Satellite data rocket disease control efforts into orbit.
     Science 1993; 261:31-2.
 19. World Health Organization. WHO establishes new rapid-response unit to
     combat growing world-wide threat of emerging diseases. WHO/75. Press
     release, 17 October 1995.
 20. Plianbangchang S. Southeast Asia intercountry consultative meeting.
     Emerging Infectious Diseases 1995;1:158.
 21. Environmental Protection Agency. The common sense initiative. Pub. No.
     EPA100F94004. Washington, DC: Environmental Protection Agency.

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Pages 109-116]

Synopsis

Emerging Disease Issues and Fungal Pathogens Associated with HIV Infection

Neil M. Ampel, M.D.
University of Arizona College of Medicine, Tucson Veterans Affairs Medical
Center, Tucson, Arizona, USA

Abstract:
Fungal diseases are increasing among patients infected with human
immunodeficiency virus (HIV) type 1. Infections due to Candida and
Cryptococcus are the most common. Although mucocutaneous candidiasis can be
treated with oral antifungal agents, increasing evidence suggests that
prolonged use of these drugs results in both clinical and microbiologic
resistance. The optimal therapy for cryptococcal meningitis remains
unresolved, although initial treatment with amphotericin B, followed by
life-long maintenance therapy with fluconazole, appears promising. Most
cases of histoplasmosis, coccidioidomycosis, and blastomycosis occur in
regions where their causative organisms are endemic, and increasing data
suggest that a significant proportion of disease is due to recent
infection. Aspergillosis is increasing dramatically as an opportunistic
infection in HIV-infected patients, in part because of the increased
incidence of neutropenia and corticosteroid use in these patients.
Infection due to Penicillium marneffei is a rapidly growing problem among
HIV-infected patients living in Southeast Asia. Although the advent of oral
azole antifungal drugs has made primary prophylaxis against fungal diseases
in HIV-infected patients feasible, many questions remain to be answered
before the preventive use of antifungal drugs can be advocated.

Over the last decade, the incidence of fungal infections has increased
dramatically. The human immunodeficiency virus (HIV) type 1 epidemic
accounts for a large share of this increase. This article is not a general
guide to the diagnosis or treatment of fungal diseases but rather a review
of emerging disease issues in regard to these infections among HIV-infected
patients. The infections discussed include candidiasis; cryptococcosis; the
endemic mycoses histoplasmosis, coccidioidomycosis, and blastomycosis;
aspergillosis; and penicilliosis. In addition, the role of preventive
therapy for fungal infections in HIV-infected patients is assessed. Fungal
pathogens and their most common manifestations in HIV-infected patients are
listed separately (Table).

------------------------------------------------------------------
    Table. Common fungal pathogens in HIV
  infection and their most frequent clinical
                  syndromes
------------------------------------------------------------------
     Organism          Clinical syndrome
------------------------------------------------------------------
 Candida          Thrush, vaginal
 albicans         candidiasis, esophageal
                  candidiasis

 Cryptococcus
 neoformans       Meningitis

 Histoplasma      Disseminated infection
 capsulatum       with fever and weight loss

 Coccidioides     Diffuse and focal
 immitis          pulmonary disease

                  Localized pulmonary
 Blastomyces      disease and disseminated
 dermatitidis     infection, including
                  meningitis

 Aspergillus      Pulmonary disease with
 fumigatus        fever, cough, and
                  hemoptysis

                  Fever alone or with
 Penicillium      pulmonary infiltrates,
 marneffei        lymphadenopathy, or
                  cutaneous lesions
------------------------------------------------------------------

Candidiasis

Mucocutaneous candidiasis is one of the most common manifestations of HIV
infection. In one prospective study, 84% of HIV-infected patients had
oropharyngeal colonization by Candida species on at least one occasion, and
55% developed clinical thrush (1). While other yeasts may occasionally
cause clinical disease, Candida albicans is the organism isolated from most
patients (1, 2). Candida species normally colonize the gastrointestinal
tract of healthy adults, and most infections in HIV-infected patients are
endogenously acquired. In some cases, candidal strains can be transmitted
from person to person (1). During the course of HIV infection, patients
appear to be colonized with one or a few dominant strains, which tend not
to change over time. Powderly and colleagues isolated the same strain of C.
albicans in 11 of 17 patients with recurrent yeast infection, by DNA probe
analysis (2). In another study, using contour-clamped homogeneous electric
field electrophoresis, Sangeorzan et al. found that 60% of patients were
colonized with one dominant strain of C. albicans. In 74% of these
patients, recolonization with the same strain occurred after antifungal
therapy (1). Using biotyping and restriction fragment length polymorphism
analysis of 25S ribosomal DNA, Whelan and colleagues found that strains of
C. albicans isolated from 24 patients with AIDS were not significantly
different from strains from 23 patients without HIV infection (3). Thus,
the candidal strains causing disease in patients with HIV infection appear
to be the same as those colonizing patients without HIV infection and, in
most patients, do not change over time.

In HIV-infected patients, candidiasis is virtually always mucocutaneous,
involving the oropharynx, the esophagus, and the vagina. HIV infection by
itself is not associated withthe syndrome of disseminated candidiasis,
which is characterized by candidemia, endophthalmitis, and multiple organ
involvement. The precise immunologic processes that control candidal
infection in HIV-infected patients are not known. However, mucocutaneous
candidiasis is clearly related to the development of clinical cellular
immunodeficiency. In fact, oropharyngeal candidiasis is an independent
predictor of immunodeficiency in patients with AIDS (4). Moreover, a CD4
lymphocyte count < 200/ l is a major risk factor for the development of
clinical thrush in HIV-infected persons (1).

Although oropharyngeal candidiasis is frequent in men, recurrent vaginal
candidiasis is a common early manifestation of HIV infection in women. The
location and severity of candidiasis in women with HIV infection appear to
be closely associated with the degree of cellular immunodeficiency, based
on the peripheral blood CD4 lymphocyte count. In a study of 66 women,
mucocutaneous candidiasis developed in more than half of the women over a
median of 14 months of follow-up; vaginal candidiasis, with a mean CD4
lymphocyte count of 506/l, developed only in 10, while oropharyngeal
candidiasis, with a mean count of 230/l developed in 16, and esophagitis,
with a mean count of 30/l developed in nine (5).

Mucocutaneous candidiasis can be treated either topically or with systemic
antifungal agents (1, 6), but such therapy does not eradicate colonization
(1). Recently, several reports have noted the failure of azole drugs,
particularly fluconazole, to treat recurrent cases of oropharyngeal
candidiasis (1, 7). While factors such as diminishing cellular immunity,
drug interactions, or decreased drug absorption may account for some of
these treatment failures, increasing evidence suggests that Candida
organisms are developing drug resistance.

In the past, a lack of consensus on the methods for performing antifungal
susceptibility tests made it difficult to establish whether clinical
failure of antifungal therapy was due to resistance of the organism.
However, the National Committee for Clinical Laboratory Standards (NCCLS)
has now developed reference methods allowing for uniform testing of yeast
isolates (8). Using an NCCLS method, Sangeorzan et al. found that the MIC
of fluconazole for Candida isolates increased over time among patients who
had received fluconazole compared with those who received clotrimazole.
Clinical resistance to fluconazole was associated with an increased MIC
required by the isolate as well as the patients low CD4 lymphocyte count
(1). These data strongly suggest that continued use of antifungal agents,
particularly fluconazole, leads to both clinical treatment failure and
antifungal resistance, especially in highly immunodeficient patients.

Cryptococcosis

A rare disease before the HIV epidemic, cryptococcosis was identified very
early in the epidemic as one of the most common life-threatening infections
in AIDS patients (9). However, issues regarding its epidemiology and
therapy remain unresolved.

A single species, Cryptococcus neoformans, is responsible for virtually all
clinical cases of cryptococcosis. The species exists in two varieties,
neoformans and gattii, which inhabit different ecologic niches. C.
neoformans var. neoformans has been isolated in many parts of the world
from numerous sites, most frequently from soil containing high amounts of
dried bird excreta, particularly of pigeons and chickens. It has long been
presumed that inhalation of soil contaminated with such excreta is the most
likely source of cryptococcal infection. However, few data support this
hypothesis. In contrast, the only known environmental source of C.
neoformans var. gattii is the river red gum tree (Eucalyptus
camaldulensis), which grows in rural Australia. Although infection due to
var. neoformans is worldwide, cases due to var. gattii have only been
identified in tropical and subtropical regions, including areas where E.
camaldulensis is not found (10).

Virtually all instances of cryptococcosis among HIV-infected persons have
been caused by var. neoformans. The ubiquity of var. neoformans in the
environment may be making exposure and subsequent infection likely;
however, no clear link has ever been established between environmental
sources of C. neoformans and the development of cryptococcosis in patients
with HIV infection (10). In a recent study, Varma and colleagues, using
genomic probe analysis, could not find a direct link between environmental
sources of C. neoformans and infection in patients with AIDS or a unique
strain of C. neoformans that was infecting these patients (11). Suppression
of cellular immunity appears to be a critical factor in the development of
cryptococcosis in HIV-infected patients, with the development of disease
relating directly to the risk for AIDS and to the CD4 lymphocyte count (12,
13).

The appropriate therapy for cryptococcal meningitis in HIV-infected
patients is unsettled at this time. The combination of amphotericin B plus
flucytosine for 4 to 6 weeks has been considered standard for patients
without HIV infection. However, concern about increased toxicity and
decreased efficacy (12) has led to a reconsideration of this regimen in
HIV-infected patients.

Several studies have examined the use of oral fluconazole in lieu of
amphotericin B for initial therapy of cryptococcal meningitis in patients
with HIV infection. Larsen and colleagues studied 21 patients and found
that amphotericin B plus oral flucytosine resulted in fewer clinical
failures and faster cryptococcal clearance of the cerebrospinal fluid than
fluconazole alone (14). A large collaborative trial compared results with a
relatively low dose of intravenous amphotericin B to those with oral
fluconazole and found no significant difference in the overall mortality
rate (15). However, in this trial, the early mortality rate, defined as
death within the first 2 weeks of therapy, was slightly higher, and time to
first negative CSF culture was somewhat longer among patients in the
fluconazole group.

The recurrence of cryptococcosis, even after initial therapy has rendered
the CSF culture sterile, is extremely common in HIV-infected patients (16);
continued antifungal therapy appears to reduce the risk for recurrence (12,
16). Fluconazole in daily doses has been shown to be effective in
preventing relapse (16) and is superior to amphotericin B in weekly doses
(17).

Current information suggests that therapy for cryptococcal meningitis in
HIV-infected patients should begin with amphotericin B, with or without
flucytosine. Suppressive therapy with fluconazole should be given
subsequently to prevent a relapse. When available, the results of a study
sponsored by the Mycoses Study Group and AIDS Clinical Treatment Group of
the National Institute of Allergy and Infectious Diseases, National
Institutes of Health, should clarify these issues.

Histoplasmosis, Coccidioidomycosis, and Blastomycosis

Unlike candidiasis and cryptococcosis, infections caused by Histoplasma
capsulatum, Coccidioides immitis, and Blastomyces dermatitidis are acquired
in specific geographic regions. Infections due to these fungi were not
initially associated with HIV infection because the HIV epidemic in the
United States began in the large urban areas of the East and West Coasts,
outside the areas in which these fungi are endemic. As HIV infection spread
to the Midwest, where histoplasmosis and blastomycosis are endemic, and to
the Southwest, where coccidioidomycosis occurs, these fungi became
recognized as major opportunistic agents.

Histoplasmosis

H. capsulatum var. capsulatum causes infection worldwide and is the
organism most associated with disease in HIV-infected patients. A few cases
of histoplasmosis in HIV-infected patients have been due to H. capsulatum
var. duboisii (18), which is found in tropical Africa. In the Western
Hemisphere, infection is concentrated in the easternUnited States but is
also found in the Caribbean as well as in Central and South America. H.
capsulatum var. capsulatum is typically isolated from soil contaminated
with avian or bat excreta, and a number of epidemics among persons without
HIV infection have been associated with disruption of contaminated soil.

Most cases of histoplasmosis in patients with HIV infection have occurred
within the recognized area for endemic H. capsulatum in North America, the
Ohio and Mississippi River valleys. However, within that area, great
variability has been seen in the incidence of disease, with most cases
being reported from a single city, Indianapolis, Indiana (19, 20). Since
1978, Wheat has recorded several outbreaks of histoplasmosis in that city,
mostly centered in areas where active construction led to soil disruption.
In the most recent outbreak, patients with AIDS accounted for more than 50%
of the culture-proven cases of histoplasmosis (21); these data suggest that
many of these cases represent new infection due to recent exposure rather
than reactivation of latent disease.

Cases of histoplasmosis and HIV-infection have also been reported well
outside the recognized histoplasmosis-endemic areas (22, 23). In some
instances, these cases represent reactivation of infection acquired during
residence in or travel to disease-endemic regions. In other instances, they
appear to represent acute infection after disruption of microfoci of H.
capsulatum that exist outside the recognized disease-endemic areas (19).

Patients with progressive disseminated histoplasmosis, the most common form
of disease among HIV-infected persons, usually have fever, malaise, and
weight loss over a period of weeks. Diagnosis is often established by
isolating the fungus from respiratory secretions, blood, or bone marrow,
but detecting Histoplasma capsulatum var. capsulatum polysaccharide antigen
(HPA) in the serum or urine is also helpful (19).

Amphotericin B therapy is usually effective for progressive disseminated
histoplasmosis in patients with HIV infection. Itraconazole may be useful
for less severe cases (24). However, relapses are extremely common when
therapy is stopped (19, 25), and maintenance therapy with intermittent
amphotericin B (25) or with itraconazole (26) is required to prevent this.
Fluconazole is less effective but can be used by those who cannot tolerate
amphotericin B or itraconazole (27). Urine and serum HPA levels decline
with successful therapy and can be useful in determining a patients
response to therapy as well as assessing a patient for relapse (26-28).

Coccidioidomycosis

Within the disease-endemic area (U.S. Southwest), coccidioidomycosis is one
of the most frequent opportunistic infections in persons with AIDS (29). In
a prospective study in Tucson, Arizona, active coccidioidomycosis developed
in 25% of a cohort of HIV-infected patients over a 41-month period (30).
The major risk for developing disease was immunosuppression, as manifested
by a CD4 lymphocyte count below 250/l, a diagnosis of AIDS, or anergy
indicated on control skin tests. Length of time in the disease-endemic
area, a history of prior coccidioidomycosis, and a positive coccidioidal
skin test were not associated with the development of active
coccidioidomycosis. These data suggest that most coccidioidomycosis cases
among HIV-infected persons in a disease-endemic area are recently acquired
and not due to reactivation of latent infection. However, as with
histoplasmosis, in a small number of HIV-infected patients, previously
acquired infection is reactivated, and clinical coccidioidomycosis develops
while the patient is residing outside the disease-endemic area (29).

Despite recent outbreaks of coccidioidomycosis in the San Joaquin Valley
and in Northridge, California (31, 32), most cases of coccidioidomycosis
among HIV-infected patients have been reported from Arizona, particularly
the metropolitan areas of Phoenix and Tucson (30, 33). Whether this
represents underreporting of disease in California or a true increase in
incidence in Arizona is unknown.

Most HIV-infected patients with coccidioidomycosis seek treatment for
pulmonary disease. In many, chest radiographs show a diffuse,
reticulonodular pattern. Approximately 70% of patients with this pattern
die within 1 month despite antifungal therapy (30, 33). In fact, this
radiographic pattern may mimic that seen with Pneumocystis carinii
pneumonia (34). Sites of dissemination frequently seen in patients without
HIV infection, such as skin, soft tissue, bone, joint, and meninges, appear
less common among patients with HIV infection (30, 33). Serologic tests can
be useful in diagnosing coccidioidomycosis in HIV-infected patients,
although they are more likely to have negative results than patients
without HIV infection (35). On the other hand, a positive coccidioidal
complement-fixation serologic test result in an HIV-infected patient, even
in the absence of clinical illness, predicts impending active
coccidioidomycosis (36).

Comparative trials regarding the therapy of coccidioidomycosis in
HIV-infected patients have not been carried out. For severe disease, such
as for diffuse, reticulonodular pneumonia, amphotericin B is recommended.
However, for less fulminant infection, fluconazole has been effective. As
with cryptococcosis and histoplasmosis, life-long maintenance therapy with
an oral azole is recommended, although relapses have occurred despite this.

Blastomycosis

Blastomycosis is the least common of the three endemic mycoses in North
America among patients with HIV infection; fewer than 25 cases have been
reported. Only recently has an environmental link been established for
infection, when the organism was isolated from riverbank soil in
association with an outbreak (37).

Pappas and colleagues have published the largest series of cases associated
with HIV infection, consisting of 15 patients from 10 medical centers (38).
Ten of the 15 cases were reported from sites within the known
disease-endemic area, the midwestern and southeastern United States, and
the other five patients had resided within the disease-endemic area at some
time before the diagnosis. Most patients were clinically immunodeficient at
the time of diagnosis and had either chronic pulmonary infection or disease
disseminated beyond the lungs, including meningitis. In all but one case,
the diagnosis was established by culture of B. dermatitidis, whereas
results of serologic tests, when performed, were uniformly negative. Both
amphotericin B and ketoconazole were used successfully as therapy in the
series.

Aspergillosis

Although disseminated aspergillosis was originally listed as an infection
at least moderately predictive of AIDS, it was removed from the list in
1984 because only three cases had been reported among 1,762 patients (39).
However, over the last 5 years, the number of cases of invasive
aspergillosis among HIV-infected patients has dramatically increased
(40-44); more than 75 cases now have been documented. The largest series,
containing 33 patients, was reported by Lortholary and colleagues from
France (43). All patients had AIDS and a median CD4 lymphocyte count of
27/l. About half the patients had the traditional risk factors for
invasive aspergillosis (neutropenia or corticosteroid use) at the time of
diagnosis. Culture of fluid from bronchoalveolar lavage was positive in all
cases in which it was done; A. fumigatus grew in 29 of 31 patients. In this
study, isolating the fungus from bronchoalveolar lavage fluid correlated
with histologic evidence of invasive aspergillosis in 14 of 15 patients.

In all series, the death rate has been extremely high despite therapy.
Intravenous amphotericin B has been used most often for treatment, but
itraconazole has been tried in some instances (40, 43). In a recent study
of the use of itraconazole for the treatment of invasive aspergillosis in a
variety of patients, therapy was unsuccessful in all 16 patients with AIDS
and aspergillosis (45). In seven of these, either toxicity to the drug
developedor clinical symptoms worsened while they were receiving
itraconazole; nine died, two directly of aspergillosis and seven of other
causes.

If the incidence of aspergillosis is increasing among HIV-infected
patients, the factors associated with this increase remain unclear. The
identified risk factors for aspergillosis have no doubt increased among
HIV-infected patients in the last decade with the use of such drugs as
zidovudine and ganciclovir, which are associated with neutropenia, and
corticosteroids for the treatment of severe Pneumocystis carinii pneumonia
and other conditions. Others have postulated that prior pneumonia, which
may result in diminished macrophage function (43) and contaminated air,
inhaled during marijuana smoking (40), may also play a role. Further
studies are needed to clarify this.

Penicilliosis

Piehl and colleagues reported the first case of infection due to
Penicillium marneffei in an AIDS patient in 1988 when they described a
patient in Chicago with persistent fever, anorexia, and a papular skin
rash. The patients travel history was not given. Blood, bone marrow,
sputum, and skin biopsy specimen cultures all grew P. marneffei. The
patient responded to therapy with amphotericin B, but relapsed once the
therapy was discontinued (46).

Since that report, the number of reported cases among HIV infected patients
has risen rapidly, particularly in association with the HIV epidemic in
Thailand. In a series of 80 patients reported by Supparatpinyo and
colleagues (47), most patients were men from the Chiang Mai province of
northern Thailand. The most common characteristic in these patients was a
generalized papular rash; some of the lesions had central ubilication
reminiscent of molluscum contagiosum. Diagnosis was usually established by
examining Wright-stained samples of bone marrow aspirates or touch smears
of skin biopsy specimens.

Treatment with amphotericin B has been successful in most patients.
Itraconazole and fluconazole may also be useful. The mortality rate appears
most related to a delay in diagnosis. Relapse is common once therapy is
stopped (48); therefore, antifungal therapy should be life-long in the
HIV-infected patient with penicilliosis.

In northern Thailand, penicilliosis is now the third most common
opportunistic infection (after tuberculosis and cryptococcosis) among
HIV-infected persons (48). Given that P. marneffei appears endemic in
Southeast Asia, penicilliosis can be expected to become an even larger
problem as the HIV epidemic continues to expand in this part of the world.

Prevention of Fungal Infections in HIV-Infected Patients

With the rise of fungi as opportunistic pathogens among patients infected
with HIV and with the success of preventive therapy for other opportunists,
such as P. carinii, primary prevention of fungal disease in HIV-infected
patients should be pursued. The goal of any such prevention would be to
increase both the length and the quality of the patients life.

Since the availability of the oral azoles, using antifungal agents to
prevent fungal diseases in HIV-infected patients has become a promising
approach. However, several questions must be considered before antifungal
chemoprophylaxis is used (49). Is the prevalence of disease high enough to
make the drug useful in most patients? Is the efficacy of the drug in
preventing disease sufficient? Does the therapy induce the development of
drug resistance? Is the cost of the drug reasonable? Finally, does the drug
have an acceptable toxicity profile, and does it interact or interfere with
the metabolism of other drugs?

Few answers to these questions are available. In a recently published
prospective, randomized study comparing fluconazole to topical clotrimazole
(13), fluconazole was significantly better than clotrimazole in preventing
oropharyngeal and esophageal candidiasis as well as cryptococcal
meningitis. The benefit of fluconazole was greatest among patients whose
CD4 lymphocyte count was < 50/l. However, 10% of the patientshad at least
one episode of candidiasis while taking fluconazole, which suggests drug
resistance. Moreover, the overall and fungal-disease-related death rates
were no different between the two groups. Given the present data, further
studies will be needed before the issue of primary prevention of fungal
disease through chemoprophylaxis is settled (50).

Dr. Ampel is an associate professor of medicine at the University of
Arizona College of Medicine and director of HIV Clinical Services at the
Tucson Veterans Affairs Medical Center. His research focuses on the immune
response in coccidioidomycosis.

Address for correspondence:

Neil M. Ampel, M.D.
Medical Service (111), Veterans Affairs Medical Center, 3601 S. Sixth
Avenue, Tuscon, AZ 85713, USA
fax: 520-629-1861
e-mail: nampl@aol.com

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Pages 117-120]

Dispatches

An Outbreak of Ross River Virus Disease in Southwestern Australia

More than 540 serologically confirmed cases of Ross River (RR) virus
disease have been reported from the southwest region of Western Australia
since November 1995 (Figure 1). Most affected by the mosquito-borne disease
are communities on the Swan Coastal Plain south of Perth. Cases have also
been reported from towns farther south or inland and from Perth itself.
These regions were foci of RR virus activity during previous southwest
outbreaks in 198889 and 199192 (1,2); however, the current outbreak
differs somewhat in the timing and location of virus activity. This article
is a preliminary overview of the incidence of disease, mosquito and virus
activity, and environmental conditions before and during the outbreak.

Monitoring of the incidence of human disease provided no indication of
abnormally high levels of RR virus activity until mid-December 1995 when
the number of reported cases began to rise sharply. In contrast, monitoring
of mosquito breeding sites, adult mosquito populations, and environmental
conditions in late October and November 1995 showed a potential for high
levels of virus transmission.

[Image not available in ASCII version]
Figure 1. Seriologically confirmed cases of Ross River virus disase, by
month of onset, in the southwest of Western Australia, July 1995 to
February 1996, as reported by doctors to the Health Department of Western
Australia (when possible, case follow-up questionnaires were administered
by environmental health officers from relevant local authorities). Only a
small number of cases diagnosed by state and private laboratories, although
the patient was not notified, have been included. Consequently, the number
of cases shown is almost certainly an underestimate of the true number of
serologically confirmed cases. Almost 65% of cases have dates of onset in
January 1996. However, further notifications and analysis of follow-up
questionnaires that have not yet been carried out for many January/February
cases may alter this pattern. Previous southwest outbreaks also peaked in
January or February but were considerably less acute.

The areas affected most, in terms of numbers of cases and attack rates (not
shown), are coastal towns and communities around the Leschenault Inlet
(including the city of Bunbury), between 165 and 190 km south of Perth and
in the shires of Capel and Busselton, as well as on the coast between 190
and 245 km south of Perth (Table 1). These regions are popular tourist
destinations during the summer holidays. It appears that many
holiday-makers from elsewhere in the southwest, as well as local residents,
were exposed to infected mosquitoes in these regions during the
ChristmasNew Year period.

Many of the Perth cases are from semirural, outlying suburbs, but some are
from suburbs closer to the city center, often near the Swan and Canning
rivers or fresh water wetlands and lakes. Follow-up questionnaires indicate
that a considerable proportion of metropolitan cases were in persons
exposed in the southwest, particularly in the Leschenault, Capel, and
Busselton regions. However, many cases from Perth also appear to have been
locally acquired. Locally acquired cases were also present during the two
previously reported outbreaks.

Many cases have also been reported from the Peel region, 70 km to 130 km
south of Perth, surrounding the Peel Inlet and Harvey Estuary. However,
considerably more cases had been reported by late February in the Peel
region during the 198889 and 199192 outbreaks. Also, during 1988 and 1991
virus activity in the Peel outbreaks commenced earlier than in the
Leschenault and Capel-Busselton regions; this is apparently not the case
during the current outbreak (Table 1). The reasons for these differences
are not yet clear, but extensive control of saltmarsh mosquito breeding has
been carried out in the Peel region this season.

Large saltmarshes and brackish wetlands in the Peel, Leschenault, Capel,
and Busselton regions provide an ideal breeding habitat for Ae.
camptorhynchus mosquitoes (5,6). This species is the major vector of RR
virus in the southwest of Western Australia. Surveillance during previous
outbreaks has clearly shown that the risk for RR virus transmission in
coastal regions of the southwest increases markedly if large populations of
adult Ae. camptorhynchus persist into late spring and summer (1,2). Adult
mosquito populations and RR virus activity are monitored routinely by our
laboratory at up to 40 sites between Rockingham and Dunsborough (50 km to
260 km south of Perth) each fortnight through spring and summer. In
addition, saltmarshmosquito breeding sites are regularly monitored by local
authorities and the health department.
--------------------------------------------------------------------------
     Table 1. Cases of Ross River virus disease, by month of onset and
   geographic region, in the southwest of Western Australia, July 1995 to
                               February 1996a
--------------------------------------------------------------------------
 Southwest region      Jul   Aug  Sep  Oct   Nov  Dec  Jan   Feb   Totals
--------------------------------------------------------------------------
 Metro. area              1    1               6    23   99   17       147

 Peel                          1     3    1    1     8   34    9        57

 Leschenault              2               1    3     9  114   30       159

 Capel/Busselton                          2    1    26   73   20       122

 Inland/south coast            2          1    2     3   30   27        65

 North/east of Perth                 1    4    2          5    3        15
--------------------------------------------------------------------------
 Totals                   3    4     4    9   15    69  355  106       565
--------------------------------------------------------------------------
 a Cases are recorded by region in which exposure most likely occurred,
 where available (from case follow-up questionnaires), or by region of
 residence. Data are incomplete. Case follow-up questionnaires are
 available for many January/February cases but have not yet been analyzed,
 and most pathology laboratory reports (nonnotified serologically
 confirmed patients (i.e., figures represent underestimate of true number
 of confirmed cases) are not included.
--------------------------------------------------------------------------

Widespread breeding of Ae. camptorhynchus (larvae) was observed in the
CapelBusselton region in late October 1995 and prompted a health
department warning of an increased risk for RR virus transmission in the
southwest. However, almost no activities to control mosquito larvae were
carried out in the worst-affected regions. The adult mosquito monitoring
program subsequently showed that extremely large populations of Ae.
camptorhynchus survived through November and December. (Figures 2 and 3).
The number of mosquitoes collected per trap per night in the
CapelBusselton region during November and December 1995 (up to 10,000
mosquitoes per trap at some sites) is unprecedented in the 5 years of
surveillance in the region. Similar results were obtained in the
Leschenault region where the number of Ae. camptorhynchus mosquitoes
collected during December 1995 and January 1996 were similar to those
observed during the 199192 outbreak. These observations, along with the
expected seasonal exodus of city dwellers to these areas during the
Christmas holidays, prompted a second warning by the health department in
December 1995.

Fourteen isolates of RR virus were obtained from Ae. camptorhynchus
mosquitoes collected at a major wetland west of Busselton on December 7,
1995 (Figure 3). Large populations of potential vertebrate hosts (western
gray kangaroos, Macropus fuliginosus) were also observed in close proximity
to this site throughout the spring and summer. Case follow-up
questionnaires indicate thata large percentage of Busselton patients were
exposed in this locality. The last time RR virus was isolated from this
site was during the 1991-92 outbreak in the region.

Mosquito populations in the CapelBusselton region (Figure 3) and most
areas of the Leschenault region dropped rapidly by mid-January 1996.
However, further cases with dates of onset in February have been reported
(Table 1), which suggests a high infection rate in the remaining adults.
The results of processing of adult mosquitoes collected in February are,
therefore, eagerly awaited.

[Image not available in ASCII version]
Figure 2. Mean number of adult mosquitoes (total population and dominant
species) and isolations of Ross River virus from mosquitoes at
CapelBusselton region, wetland site, January 1994 to April 1995.

[Image not available in ASCII version]
Figure 3. Mean number of adult mosquitoes (total population and dominant
species) and isolations of Ross River virus from mosquitoes at
CapelBusselton region, wetland site, January 1995 to January 1996.

Several isolates of RR virus were also obtained from mosquitoes collected
in the Peel region (Table 2). The recent isolations from Ae. vigilax are of
particular concern. This species is regarded as the major vector of RR
virus in coastal areas of northern and eastern Australia (3,4), but until
now it has had little or no role in transmitting RR virus in the southwest
(1,2). Ae. vigilax has become the dominant species in the Peel region
between December and March since the opening of the Dawesville Channel.
This mosquito is a vicious biter, even during the day if weather conditions
are suitable, and is known to disperse considerable distances from breeding
sites. Thus, the potential for interaction between infected mosquitoes and
humans in the Peel region may be greater and occur over a wider area than
originally thought.
-----------------------------------------------
 Table 2 Isolations of Ross River (RR) virus
    from mosquitoes collected in the Peel
            region: 1995-96 season
-----------------------------------------------
                                   Isolates
    Date           Species           of RR
                                     virus
-----------------------------------------------
 14-Sep-95   Ae. camptohynchus         1

 24-Oct-95   Ae. camptohynchus         2

 7-Dec-95    Ae. camptohynchus         1

 27-Dec-95   Ae. camptohynchus         5

 15-Jan-96   Ae. camptohynchus         2

 15-Jan-96   Ae. vigilax               3
-----------------------------------------------

Analyses of environmental conditions before and during the outbreak are not
yet complete. However, record-high daily rainfall was recorded in October
at numerous centers in the southwest. Above-average rainfall occurred in
November in Perth and Mandurah and in December in Mandurah, Banbury, and
CapelBusselton. These were accompanied by above-average October and
November temperatures at many southwest centers. A series of extremely high
tides was also recorded along the PeelLeschenault region coast around
December 20. This resulted from unusually early cyclonic activity (three
cyclones) along the north and west coasts of Western Australia during
December. Clearly, a combination of some or all of these factorsenabled
widespread breeding and survival of vector mosquito species. Late spring
and summer rains, a short-term rise in sea level (accompanied by higher
tides), and mild spring and summer temperatures were predisposing factors
during previous outbreaks in the southwest (2,4).

Preliminary analysis of the location of virus activity (measured as either
human cases or isolations from mosquitoes) shows that activity is far less
likely in regions in which virus activity was detected in the previous
season. Thus, length of time since the previous outbreak also appears to be
a predisposing factor for higher levels of virus activity in the southwest.
The reason for this is not yet known but may be due to higher levels of
immunity in recently infected populations of enzootic or amplifying
vertebrate hosts. This may help explain the comparatively reduced numbers
of cases in the Peel region this season following moderate levels of virus
activity last year, which coincided with the opening of the Dawesville
Channel.

A small number of cases of Barmah Forest virus infection have been
diagnosed during the current outbreak. Numerous cases of a RR virus-like
disease have also been reported, as in the 1988-89 and 1991-92 outbreaks.
Serum from these patients has been tested for IgM antibody to RR and Barmah
Forest viruses but is negative for both viruses. Some of these cases may be
in persons that had not seroconverted at the time of the first blood
sample. However, many have since provided further samples, all of which
have had negative test results. Sera from these patients are being tested
against a wide range of other Australian arboviruses, and more blood
samples will be sought to ensure that the phenomenon is not due to an
extremely delayed immunologic response to RR virus.

Michael Lindsay,* Nidia Oliveira, Eva Jasinska,* Cheryl Johansen, Sue
Harrington, A.E Wright, and David Smith

*University of Western Australia, Queen Elizabeth II Medical Centre,
Nedlands, Western Australia; University of Queensland, Brisbane,
Queensland; Health Department of Western Australia, Mt. Claremont, Western
Australia;  Western Australian Centre for Pathology and Medical Research,
Queen Elizabeth II Medical Centre, Nedlands, Western Australia

References

  1. Lindsay MD, Condon R, Mackenzie JS, Johansen C, DErcole M, Smith D. A
     major outbreak of Ross River virus infection in the south-west of
     Western Australia and the Perth metropolitan area. Communicable
     Disease Intelligence 1992;16:290-4.
  2. Lindsay MD, Latchford JA, Wright AE, Mackenzie JS. Studies on the
     ecology of Ross River virus in the southwest of Western Australia.
     Arbovirus Research in Australia 1989;5:28-32.
  3. Mackenzie JS, Lindsay MD, Coelen RJ, Broom AK, Hall RA, Smith DW.
     Arboviruses causing human disease in the Australasian zoogeographic
     region. Arch Virol 1994:136:447-67.
  4. Russell RC. Ross River virus: disease trends and vector ecology in
     Australia. BullSoc Vector Ecol 1994;19:73-81.
  5. Wright AE. Report on the mosquito eradication campaign: survey of
     mosquitoes in the Bunbury region, Western Australia. Health Department
     of Western Australia, 1988.
  6. Wright, AE. Report on the mosquito eradication campaign: survey of
     mosquitoes in the Mandurah region, Western Australia. Health
     Department of Western Australia, 1988.

---------------------------------------------------------------------------

Emerging Infectious Diseases * Volume 2 * Number 2            April-June 1996

---------------------------------------------------------------------------
[Pages 121-124]

Dispatches

Invasive Penicillin-Resistant Pneumococcal Infections:
A Prevalence and Historical Cohort Study

More than 25 years ago, isolates of Streptococcus pneumoniae were uniformly
susceptible to penicillin. However, since a penicillin-resistant
pneumococcus was first identified in 1967 (1), the incidence of
penicillin-resistant S. pneumoniae (PRSP) strains has been gradually
increasing. In certain areas of the United States, PRSP strains have become
widespread; Alaska has the highest reported prevalence, 26% (2); a recent
study conducted in Atlanta, Georgia, found a 25% prevalence of PRSP (3).
Outside the United States, an even higher (33%-58%) prevalence of PRSP has
been reported (2).

Pneumococcal infections are a leading cause of morbidity and mortality in
the United States. S. pneumoniae causes more than 500,000 cases of
pneumonia, 55,000 cases of bacteremia, and 6,000 cases of meningitis
annually, which result in 40,000 deaths (4). The death rate from
pneumococcal bacteremia approaches 30%, despite the use of appropriate
antimicrobial therapy (5). Reports of refractory illness due to resistant
pneumococci demonstrate the clinical relevance of these strains (6,7).
Identifying risk factors in the development of PRSP infections is important
for both the prevention and treatment of these infections.

The prevalence of invasive infections due to PRSP was previously studied in
Denver, Colorado (8,9). We undertook the study described here to determine
the prevalence of invasive PRSP infections in the Colorado Front Range and
to determine whether invasive PRSP infections have increased in
metropolitan Denver since the earlier studies. In addition, we studied a
cohort of patients who had invasive pneumococcal disease during 1994 in
metropolitan Denver to ascertain risk factors for invasive PRSP infections.

Twenty-six hospital microbiology laboratories in the Colorado Front Range,
which comprises the 10 largest counties in Colorado and 80% of the states
population (10), submitted to the Colorado Department of Public Health and
Environment reports of all blood and cerebrospinal fluid (CSF) isolates of
S. pneumoniae that were tested for penicillin susceptibility during 1994.
(Penicillin susceptibility testing on invasive pneumococcal isolates was
standard practice for the laboratories and did not depend on a clinicians
request.) For the part of the study that assessed prevalence, penicillin
resistance was defined as a minimum inhibitory concentration (MIC) of >=
0.12 g/ml or an oxacillin zone of inhibition < 20 mm. Isolates that were
tested by an MIC method were further classified as intermediately (0.12
g/ml-1.0 g/ml) or highly (>=2.0 g/ml) penicillin resistant. For the part
of the study in which the cohort was analyzed, only isolates that were
confirmed as penicillin resistant (i.e., MIC >= 0.12 g/ml) by either broth
dilution or the E test (AB Biodisk, North America, Inc., Culver City,
California) were included. Data for the cohort study were collected by
chart review by the principal investigator, and telephone interviews of
patients were conducted by trained interviewers in the Health Statistics
Survey Research Unit of the Colorado Department of Public Health and
Environment. For patients under 18 years of age, a parent or legal guardian
was interviewed. When patients had died, a relative of the patient (if
available) was interviewed.

Invasive pneumococcal infections were found in 363 patients in the Colorado
Front Range; 49 (13%) of the infections were resistant to penicillin. In
metropolitan Denver, 29 (14%) of the invasive pneumococcal infections were
penicillin-resistant, of which 20 (69%) were intermediately penicillin-
resistant (i.e., MIC 0.12-1.0 g/ml), and 9 (31%) were highly
penicillin-resistant (i.e., MIC >= 2.0 g/ml). This prevalence rate of
invasive PRSP infections is significantly higher than the previously
reported rates in Denver of 1% (8) and 7% (9). Previous surveillance of
invasive PRSP isolates showed that one region in the United States, which
included Colorado, had a significantly higher rate of penicillin resistance
among pneumococcal isolates than other U.S. regions (11).

Half of the PRSP strains in the cohort part of the study that were tested
for cephalosporin susceptibility were resistant to an extended-spectrum
cephalosporin. Our results are similar to the recent Atlanta study which
found that 34% to 54% of PRSP infections were resistant to an
extended-spectrum cephalosporin (3). These rates are much higher than other
reported rates of cephalosporin resistance among PRSP isolates of 27% in
Kentucky and 25% in Tennessee, (12). This has important implications for
the management of invasive PRSP infections, especially in meningitis, where
MICs of -lactam antibiotics in the cerebrospinal fluid may be less than
the MICs of -lactam antibiotics in the blood (13). The Centers for Disease
Control and Prevention recommends that in areas where pneumococcal
resistance to cephalosporins is high, empiric therapy with vancomycin plus
an extended-spectrum cephalosporin should be considered in all cases of
meningitis potentially caused by S. pneumoniae, until the results of
culture and susceptibility testing are available (14).

A number of studies have addressed the clinical relevance of PRSP
infections and have attempted to identify predictive factors for the
development of these infections. In our analysis of the demographic and
clinical characteristics of the study population (Table), we found that
day-care attendance by a member of the patients household in the 3 months
before the patients illness was associated with invasive PRSP infections.
Indeed, 26% of patients with PRSP infections, compared with 7% of patients
with penicillin-sensitive infections, had at least one member in their
household, excluding the patient, who had been attending a day-care center
before becoming ill.

  Table. Demographic and clinical characteristics of patients with
                   invasive penicillin-resistant
 S. pneumoniae (PRSP) and penicillin-sensitive S. pneumoniae (PSSP)
                             infections
---------------------------------------------------------------------------
                              PRSP;       PSSP;       Odds ratio
                             n = 29    n = 180 (%)       (95%
                               (%)                    confidence
                                                       interval)
---------------------------------------------------------------------------
 Demographic
 characteristics

  Age, years

    < 5                      11 (38) 39 (22)        1.9 (0.4-9.1)

    5-64                      8 (28) 93 (52)        Reference

    > 64                     10 (34) 48 (27)        1.9 (0.5-7.1)

  Sex

    Male                     12 (41) 104 (58)       Reference

    Female                   17 (59)  76 (42)       1.8 (0.6-5.4)

  Racea

    White                    16 (55) 100 (56)       Reference

    Nonwhite                 11 (38)  68 (38)        1.4 (0.5-3.9)

 Clinical characteristics

  Site of infection

    Blood                    26 (90) 171 (95)       Reference

    CSF                       3 (10)   9 (5)        0.8 (0.1-5.1)

  Underlying medical
   condition                 14 (48) 108 (60)       1.0 (0.3-3.2)

  Antibiotic useb            14 (52)  56 (39)       2.5 (0.9-7.1)

  No history of
   penicillin allergyc       26 (90) 163 (91)       0.7 (0.1-3.4)

  Previous hospitalizationd   5 (19)  33 (23)       0.3 (0.1-1.5)

  Day-care attendance

    Patients < 11 yearse      5 (50)  19 (50)       1.1 (0.4-2.7)

    Household member(s)f      7 (26)  10 (7)        8.1 (2.2-30.7)

  Residence in a long-term
  care facility

  (patients > 64 years)g      2 (20)  13 (27)       0.7 (0.1-3.6)

  Hospital-acquired
   infection                  1 (3)    8 (4)        0.4 (0.04-4.9)

  Outcome

    Survived                 25 (86) 156 (87)       Reference

    Died                      4 (14)  24 (13)       1.6 (0.4-6.3)
---------------------------------------------------------------------------
 a Missing information on 2 PRSP and 12 PSSP patients.
 b Includes patients who had taken an antibiotic in the 3 months
 before illness; missing information on 2 PRSP and 36 PSSP
 patients.
 c Missing information on 1 PSSP patient.
 d Patients who were hospitalized in the 3 months before illness;
 missing information on 2 PRSP and 34 PSSP patients.
 e Children < 11 years of age who attended day care in the 3 months
 before illness; children < 11 years of age, N = 56 (PRSP = 12,
 PSSP = 44); missing information on 2 PRSP and 6 PSSP patients.
 f Patients with at least 1 child < 11 years of age (excluding the
 patient) in the household who attended day care in the 3 months
 before illness; missing information on 2 PRSP and 39 PSSP
 patients.
 g Adults > 64 years of age who resided in a LTC facility in the 3
 months before illness; adults > 64 years of age, N = 58 (PRSP =
 10, PSSP = 48).
---------------------------------------------------------------------------

This study is unique in that, to our knowledge, day-care attendance among
household members of patients has not been studied. Even though most
studies have not specifically considered family members as a potential mode
of PRSP transmission, rates of nasopharyngeal carriage of PRSP are
significantly higher in family contacts of children colonized with PRSP who
were attending day-care centers (7, 15).

Our finding that patients with PRSP infections were more likely to have had
a child in their household who had attended day-care during the months
before their illness suggests that day-care settings may serve as foci for
spreading resistant pneumococcal strains. Antibiotics are extensively used
to treat upper respiratory infections that children attending day-care
centers often have; the practice of administering a prolonged course of
prophylactic antibiotics to children with recurrent otitis media who attend
day-care centers (16) may promote the selection of resistant bacteria in
these settings (6, 15). These children may subsequently transmit resistant
S. pneumoniae to susceptible persons in their households. Thus, patterns of
antibiotic treatment of children who attend day-care centers may explain
why day-care attendance might facilitate PRSP transmission. The likelihood
that day-care settings may serve as reservoirs for antibiotic-resistant
pneumococci indicates that the efficacy of prophylactic antibiotics for
otitis media should be reassessed, especially when PRSP is present in a
community.

Furthermore, carriage of or infection with PRSP has been associated with
recent use of antibiotics (17). Our study showed that patients with PRSP
infections were more likely to have taken an antibiotic in the 3 months
before their illness than patients with penicillin-sensitive pneumococcal
infections. This finding supports the theory that antibiotic resistance has
developed because of the widespread availability and use of antibiotics.
Since the beginning of the antibiotic era 50 years ago, it has been well
recognized that antibiotics have been and continue to be inappropriately
used (18).

The emergence of drug-resistant S. pneumoniae emphasizes the importance of
following the recommendation of the Immunization Practices Advisory
Committee that all persons 2 years of age and older who are at high risk
for pneumococcal disease receive the 23-valent pneumococcal capsular
polysaccharide vaccine. Because of its lack of immunogenicity and efficacy,
the pneumococcal vaccine has not been licensed forchildren under 2 years of
age (14). The high prevalence of PRSP among young children (3, 17), and the
potential for these children to transmit PRSP to susceptible persons,
underscore the need for an effective pneumococcal vaccine for this age
group.

Antimicrobial resistance contributes to increased morbidity, mortality, and
health care costs (19). The solution lies in changing antibiotic
prescribing patterns, changing patient attitudes about the necessity of
antibiotics, increasing surveillance of drug-resistant organisms, improving
techniques for antibiotic susceptibility testing, and investing in research
and development of newer antimicrobial agents.

Acknowledgments

We thank the Health Statistics Survey Research Unit in the Colorado
Department of Public Health and Environment for assistance with the
telephone interviews, and the supervisors of the microbiology laboratories
and the staff in the medical records departments of participating hospitals
for their cooperation. This publication was supported by Grant No. STC-T2
D33AH18002-09 from the Division of Medicine of the Health Resources and
Services Administration.

Crystal B. Kronenberger, M.D., M.S.P.H.,* Richard E. Hoffman, M.D.,
M.P.H., Dennis C. Lezotte, Ph.D.,* and William M. Marine, M.D., M.P.H.*

*Department of Preventive Medicine and Biometrics, University of Colorado
Health Sciences Center, Denver, Colorado, USA; Colorado Department of
Public Health and Environment, Denver, Colorado, USA

References

  1. Hansman D, Bullen MM. A resistant pneumococcus. Lancet 1967;1:264-5.
  2. Appelbaum PC. Antimicrobial resistance in Streptococcus pneumoniae: an
     overview. Clin Infect Dis 1992;15:77-83.
  3. Hofmann J, Cetron MS, Farley MM, Baughman WS, Facklam RR, Elliott JA,
     et al. The prevalence of drug-resistant Streptococcus pneumoniae in
     Atlanta. N Engl J Med 1995; 333:481-6.
  4. Williams WW, Hickson MA, Kane MA, Kendal AP, Spika JS, Hinman AR.
     Immunization policies and vaccine coverage among adults. Ann Intern
     Med 1988;108:616-25.
  5. Butler JC, Breiman RF, Campbell JF, Lipman HB, Broome CV, Facklam RR.
     Pneumococcal polysaccharide vaccine efficacy: an evaluation of current
     recommendations. JAMA 1993;270:1826-31.
  6. Ward J. Antibiotic-resistant Streptococcus pneumoniae: clinical and
     epidemiologic aspects. Rev Infect Dis 1981;3:254-66.
  7. Radetsky MS, Istre GR, Johansen TL, Parmelee SW, Lauer BA, Wiesenthal
     AM, et al. Multiply resistant pneumococcus causing meningitis: its
     epidemiology within a day-care centre. Lancet 1981;2:771-3.
  8. Lauer BA, Reller LB. Serotypes and penicillin susceptibility of
     pneumococci isolated from blood. J Clin Microbiol 1980;11:242-4.
  9. Istre GR, Humphreys JT, Albrecht KD, Thornsberry C, Swenson JM,
     Hopkins RS. Chloramphenicol and penicillin resistance in pneumococci
     isolated from blood and cerebrospinal fluid: a prevalence study in
     metropolitan Denver. J Clin Microbiol 1983;17:472-5.
 10. Colorado Vital Statistics, 1990. Health Statistics Section, Health
     Statistics and Vital Records Division. Colorado Department of Health,
     1990.
 11. Spika JS, Facklam RR, Plikaytis BD, Oxtoby MJ, Pneumococcal
     Surveillance Working Group. Antimicrobial resistance of Streptococcus
     pneumoniae in the United States, 1979-1987. J Infect Dis
     1991;163:1273-8.
 12. Centers for Disease Control and Prevention. Drug-resistant
     Streptococcus pneumoniaeKentucky and Tennessee, 1993. MMWR
     1994;43:23-5,31.
 13. Hieber JP, Nelson JD. A pharmacologic evaluation of penicillin in
     children with purulent meningitis. N Engl J Med 1977;297:410-3.
 14. Centers for Disease Control and Prevention. Prevalence of
     penicillin-resistant Streptococcus pneumoniaeConnecticut, 1992-1993.
     MMWR 1994; 43:216-7,223.
 15. Reichler MR, Allphin AA, Breiman RF, Schreiber JR, Arnold JE, McDougal
     LK, et al. The spread of multiply resistant Streptococcus pneumoniae
     at a day care center in Ohio. J Infect Dis 1992;166:1346-53.
 16. Committee on Infectious Diseases, American Academy of Pediatrics.
     Antimicrobial prophylaxis. In: Hall PG, Lepow ML, Phillips CF,
     editors. Report of the Committee on Infectious Diseases. 21st ed. Oak
     Grove Village, IL: American Academy of Pediatrics, 1988:465-8.
 17. Nava JM, Bella F, Garau J, Lite J, Morera MA, Mart C, et al.
     Predictive factors for invasive disease due to penicillin-resistant
     Streptococcus pneumoniae: a population-based study. Clin Infect Dis
     1994;19:884-90.
 18. Kunin CM. Problems in antibiotic usage. In: Mandel GL, Douglas RG Jr,
     Bennett JE, editors. Principles and practice in infectious diseases.
     3rd ed. New York: Churchill Livingstone;1990:427-34.
 19. Holmberg SD, Solomon SL, Blake PA. Health and economic impacts of
     antimicrobial resistance. Rev Infect Dis 1987;9:1065-78.

---------------------------------------------------------------------------

Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

---------------------------------------------------------------------------
[Pages 125-129]

Dispatches

Nosocomial Transmission of Multidrug-Resistant Mycobacterium tuberculosis
in Spain

Before 1990, outbreaks of multidrug-resistant tuberculosis (MDRTB) were
uncommon (1); since then, more than 10 outbreaks have been reported, all in
hospitals and prisons in the eastern United States (2-7). Persons
traditionally considered at risk for MDRTB (foreign-born TB patients and
those inadequately treated for TB) have not been associated with these
outbreaks. Instead, the presence of patients with active TB near
immunocompromised patients in HIV-dedicated wards has led to MDRTB-infected
HIV patients whose TB cases often go unrecognized. The patients receive
inadequate treatment in facilities without effective procedures for
isolating acid-fast bacilli; these circumstances favor nosocomial
transmission. Health officials in other geographic areas where HIV and TB
are major public health threats have been alerted to this emerging problem,
and surveillance systems have been designed (8).

Spain has the highest reported incidence rate of AIDS in Europe (143.4
cases per million in 1994) (9). Although in Spain TB is not notifiable at
the national level, reported rates in the autonomous community of Madrid
for 1994 were among the highest in Europe (33.5/100,000) (10). During a
45-month period starting in September 1991, a number of patients and one
health care worker became infected with MDRTB in a 120-bed, infectious
disease reference hospital in urban Madrid. In May 1995, the Field
Epidemiology Training Program of the Spanish Ministry of Health was invited
to assist the Madrid Department of Health and hospital officials in
investigating the outbreak. This report describes the findings of the
epidemiologic and molecular laboratory investigation and analyzes risk
factors associated with the outbreak. The study was designed in three
parts: 1) a description of the reported MDRTB cases, including a laboratory
investigation of isolates; 2) a case-control study comparing HIV-infected
patients who also had MDRTB to HIV-infected patients who did not have
MDRTB; and 3) a study of tuberculin conversion among hospital employees.

We reviewed the medical records and laboratory specimen testing results of
a series of patients with MDRTB in the HIV-dedicated ward of the hospital
for January 1991 through June 1995. Cases were defined as patients with
culture-confirmed TB and drug resistance to at least rifampin and isoniazid
and with no previous history of inadequate treatment. Demographic and
clinical variables were collected to characterize the cluster. Drug
susceptibility testing and DNA subtyping analysis were performed with
resistant strains available at the time of the study. Drug susceptibility
testing was performed by the method of proportions in Middlebrook 7H11
medium distributed in petri plate compartments (reference).

All 48 reported cases of isoniazid and rifampin resistance were among
HIV-infected patients hospitalized in the HIV-dedicated ward from September
15, 1991, to May 1995. One patient was an HIV-infected nurse who worked on
the ward from 1990 to 1994. The mean age of patients was 34.1; 81.3% were
male, and 66.6% were intravenous-drug users (Table 1). Of the 47 (97.9%)
who died, the mean interval from diagnosis to death was 77.6 days.


 Table 1. MDRTB patient characteristics
----------------------------------------------
                   Number
     Variable       (n =       %
                    48)
----------------------------------------------
 Sex

  Male               39       81.3

  Female              9       18.7

 Mean age            34.1     (6.8)a

 HIV status

  Infected           48      100.0

 HIV risk group

  Intravenous
  drug users         32       66.6

  Homosexual         10       20.8

  Others              6       12.6

 Outcome

  Death              47       95.9

  Discharge           1        4.1

 Mean survival
 after                      (107.8)a
 MDRTB diagnosis     77.6
 (in days)
----------------------------------------------
 a Standard deviation.

The epidemiologic curve suggests a propagated transmission pattern of MDRTB
among HIV ward patients, starting in 1991 and continuing until June 1995
(Figure 1). By the first 6 months of 1995, 65% of Mycobacterium
tuberculosis strains seen among HIV ward patients were multi
drug-resistant.

[Image not available in ASCII version]
Figure 1. Hospital outbreak, Madrid, Spain, 1995

At the beginning of the outbreak, no consistent antibiogram pattern was
observed among isolates. However, beginning in 1993, strains were
consistently resistant to isoniazid, streptomycin, ethambutol, and rifampin
(HSER); of the 26 patients with an MDRTB diagnosis since the last trimester
of 1993, 24 (92.3%) had HSER isolates. DNA subtyping analysis was performed
on 12 of these HSER isolates that were available. Eleven of the12 strains
had the same band patterns (Figure 2). Two additional isolates with a
different antibiogram pattern had different banding patterns.

[Image not available in ASCII version]
Figure 2. Evolution of MDRTB strain* over time, hospital outbreak, Madrid,
Spain, 1995.

Subtyping of M. tuberculosis strains was performed (11) by analyzing DNA
located between two copies of repetitive sequence IS6110. A 10-l sample of
the extracted DNA was amplified in a reaction mixture containing 0.5 pM of
each of the four primers (Ris1, Ris2, Pntb1, and Pntb2), 200 UM DNTPs, 50
mM Tris-HCL, 50 mM KCL (pH 8.8), 2.5 mM MgC12, 0.1% Triton x-100, and 0.5%
Taq polymerase. The samples were denatured by incubation at 95C for 10 min
and amplified by 30 cycles of denaturation at 94C for 1 min, primer
alignment at 56C for 2 min, and primer extension at 72C for 1 min. The
amplification products were analyzed by electrophoresis in 2% agarose gel
stained with ethidium bromide and observed with an ultraviolet
transilluminator.

HIV-infected patients hospitalized on the HIV-dedicated ward between
September 15, 1991, and December 31, 1994, who had TB diagnosed in 1994
with a known drug-susceptibility pattern were included in a case-control
study. Case patients isolates were resistant to isoniazid, rifampin,
streptomycin, and ethambutol; control patients isolates were sensitive to
the same antimicrobial drugs.

A time-person line diagram was prepared for each of the case patients and
control patients, including all hospitalization dates, ward and room
number, potentially infectious days, and possible days exposed to infective
patients. Prior admission was defined as all admissions to the
HIV-dedicated ward since the MDRTB patient was diagnosed (9/15/91) until
the end of the period (12/31/94). TB patients were classified as
potentially infective from the 2 week-period before sputum or culture
results were positive for M. tuberculosis until the time sputum results
were negative or the patient died. TB-negative patients were classified as
possibly exposed if they were hospitalized on the HIV-dedicated ward
during the time a potentially infective patient was also present, beginning
on September 15, 1991, until 2 weeks before TB diagnosis.

Thirty-five patients (18 with cases and 17 controls) met the established
study case/control definition. Case patients and control patients were not
significantly different with respect to age, sex, HIV risk factors,
interval between HIV and TB diagnoses or count of CD4 lymphocytes at the
time of TB diagnosis (Table 2).

          Table 2. Population characteristics: case control study
----------------------------------------------------------------------------
                                         Control patients
                     Case patients (%)         (%)
      Variable            n = 18              n = 17         OR (95% CI)a
----------------------------------------------------------------------------
 Mean age (yr)          34.4              35.2                 p = 0.30b

 Male sex               13 (72.2)         14 (82.3)            1.8 (0.3-12.1)

 IVDU                   11 (61.1)         11 (64.7)            1.2 (0.2-6.7)

 Interval HIV to TB                                         p = 0.79b
 diagnosis in days    1403              1293

 CD4 lymphocyte                                             p = 0.30b
 count, median         112.9             199.5
----------------------------------------------------------------------------
 a OR (95% CI) = odds ratio (95% confidence intervals); Fishers Exact Test.
 b p value for analysis of variance (ANOVA). IVDU = intravenous drug user.

Before the hospitalization during which TB was diagnosed, 76.4% of the
case-patients had been hospitalized on the HIV-dedicated ward of the
hospital versus 23.5% of control patients (OR = 7.8 [1.4,50.5]) (Table 3).
Of the five case patients with no prior hospitalization, three were family
members of previously hospitalized HIV-infected patients who had visited
the HIV ward frequently during the outbreak period.

Case patients and controls were compared with respect to possible exposure
to a potentially infective wardmate beginning on 9/15/91 until 2 weeks
prior to the diagnosis of TB. Case patients were more likely to have been
exposed to potentially infective wardmates (72.2% with a median of 26.4
days) than control patients (41.2% with a median of 7.6 days). When we
stratified possible ward exposure by days (0 days vs. 1 to 30 days vs. 30
days), we observed a dose-response effect, and the chi-square for linear
trend was statistically significant (Table 3). Case patients were more
likely to die during their initial hospitalization for MDRTB(94.4%) than
were control patients (29.4%) during their hospitalization for TB (OR =
40.8 [3.6,1842]) (Table 3).

    Table 3. Variables associated with MDRTB
----------------------------------------------------
                Case      Control
              patients   patients
                 (%)        (%)
  Variable     n = 18     n = 17      OR (95%
----------------------------------------------------
 Prior
 admission
 to HIV ward

  Yes        13 (76.4)    5 (29.4)    6.2
                                    (1.2-37.2)

  No          5 (23.6)   12 (70.6)

 Possible
 ward
 exposure
 days

  None        5 (27.8)   10 (58.8)    1.0

  1-30        6 (33.3)    5 (29.4)    2.4

                                      7.0, p =
  30          7 (38.9)    2 (11.8)    0.03b

 Outcome

  Death      17 (94.4)    5 (29.4)   40.8
                                     (3.6-1842)

  Discharge   1 (5.6)    12 (70.6)
----------------------------------------------------
 a OR (95% CI) = odds ratio (95% confidence
 intervals); Fishers Exact Test.
 b Chi-square for linear trend.

A TB screening clinic visit was offered to all hospital employees after the
outbreak was identified. Of the 591 active employees, 565 (95.6%)
participated. Of these, 288 (51%) had not participated in previous hospital
employee screening programs conducted in 1990 and 1994. The overall
prevalence of TB infection among participating employeeswas 450 (80%) of
565; only 115 (20%) of the current employees tested were
tuberculin-negative.

Employees currently working at the hospital, who had a documented negative
(< 6 mm) tuberculin test between January 1993 and June 1995 were eligible
for this skin test conversion study. Many of the employees had received BCG
vaccine. For those who had not received BCG vaccine, conversion was defined
as an induration of 10 mm or greater with a change of at least 6 mm of
induration since the last negative tuberculin test (12). For BCG vaccinees,
conversion was defined as a 15-mm induration change since the last negative
(< 6 mm) skin test.

Employees were defined as occupationally exposed if they worked in parts of
the hospital where exposure to patients or M. tuberculosis was likely (the
HIV-dedicated ward, HIV outpatient clinic, radiology unit, the
mycobacteriology laboratory, and the internal medicine ward). Employees
were asked to quantify the cumulative number of months spent in these
high-risk areas, regardless of their usual place of work, during the
30-month study period.

According to the Mantoux technique, 2 tuberculin units of purified protein
derivative (PPD) tuberculin RT-23 were administered in the anterior forearm
of the screened employees, and the results were read within 48 to 72 h. Of
the participants, 92 (16.3%) were eligible for the conversion study. The
incidence of conversion during the 30-month period was 24 of 92 (26%).
Employees who had occupational exposure to high-risk areas had higher
conversion rates than employees who did not have occupational exposure to
high-risk areas (RR = 5.0 [2.7,9.6]) (Table 4). The conversion had a
dose-response effect, that is, the more months the person is occupationally
exposed to high risk-areas, the higher the risk for conversion.

     Table 4. Employee tuberculin screening
            conversion study results
----------------------------------------------------
               Convertors  Nonconvertors    RR
                  (%)           (%)        (95%
   Variable      n = 24       n = 68       Cl)
----------------------------------------------------
 Occupational
 exposure to
 high-risk
 areas

                                           5.0
      Yes      14 (70.0)      6 (30.0)    (2.7,
                                           9.6)

       No      10 (13.9)     62 (86.1)

 Months
 exposed                                   ORb
 in high-risk  
 areas

       0        6 (10.7)     50 (89.3)     1.0

      1-6       4 (28.6)     10 (71.4)     3.3

                                          14.6,
      7-36     14 (63.6)      8 (36.4)      p <
                                           0.01
----------------------------------------------------
 a RR (95% CI), relative risk (95% confidence
 intervals).
 b Chi-square for linear trend.

This is the first nosocomially transmitted MDRTB outbreak reported in
Spain, which, with recent outbreaks in the United Kingdom (13) and Italy,
is among the first in Europe. Its characteristics are similar to the other
reported outbreaks on that it occurred in an HIV-dedicated ward among
non-foreign-born patients who had not been treated for TB; it had a high
mortality rate within 3 months of onset during which mycobacteria
laboratory surveillance recognized similar antibiogram resistant patterns;
and identification of MDRTB isolates was followed by DNA subtyping, which
confirmed that the same strain was responsible for the outbreak. Risk
factors identified include admission to the HIV-dedicated ward and more
possible exposure days to potentially infective wardmates; additionally,
skin test conversion rates among employees were directly related to working
in high risk areas of the hospital. The consistency of these findings with
those reported in similar outbreaks and the fact that the same isolate was
cultured over an extensive period among many different patients on a
hospital ward without proper room isolation techniques support the
conclusion that nosocomial transmission was the leading cause of the
outbreak.

CDCs recommended guidelines for hospital TB prevention and control (14)
were not fully implemented in the hospital. Acid-fast bacilli room
isolation techniques were not in place; moreover, no ventilation system was
available to provide negative pressure to prevent bacilli from passing from
the MDRTB patients rooms to the hallway or to provide the six air
interchanges per hour recommended for removing bacilli from room air.
Surgical masks used in the HIV-dedicated ward during the outbreak as
protective masks are not recommended for this purpose because of filtering
and facial sealing problems. Observational visits to the HIV-dedicated ward
revealed that MDRTB patients without masks were walking, talking, and
smoking in the halls and in the lounge next to the unprotected patients,
families, and staff.

Complete follow-up of employees is an essential component of a hospital
control program (14). Because 80% of the employees in this outbreak were
tuberculin-positive,chest radiographs became an important component for
disease screening; yearly surveillance of the tuberculin-negative employees
will help determine if the interventions in place to prevent future
outbreaks have worked in this setting.

Once the case-control study findings were analyzed, a TB control committee
was established. All MDRTB patients were placed in a separate area of the
hospital. Hospital staff were informed about the outbreak and alerted about
future possible cases and the patient management and treatment schemes. The
mycobacteria laboratory expanded the antibiogram service to include all
second-line antibiotics used by the hospital clinicians. Clinicians have
elaborated a treatment flow chart for MDRTB patients. Masks that fulfilled
the sealage and filtering criteria (15) were purchased, and mask use was
aggressively implemented. An employee health clinic was also instituted,
with a prophylaxis and chest X-ray follow-up program for the 410 infected
employees, along with a computerized follow-up surveillance system which
included all employees graded by occupational risk category. Plans were
made to screen tuberculin-negative employees every 3 months to identify
those who had recently seroconverted. Family, community members and
wardmates of all patients whose MDRTB had been diagnosed within the
previous 6 months were notified of their risk and were scheduled for
follow-up evaluations. The HIV-dedicated ward will be transformed into an
acid-fast bacilli isolation zone, with an exclusive ventilation system that
provides 11 individual rooms, each with 12 air interchanges per hour and
negative pressure relative to the hallway.

Health officials in Europe need to be updated about this emerging problem,
especially in areas of high HIV and TB prevalence. Basic universal TB
control and prevention measures (16,17) should be implemented by all
general community hospitals in Spain, especially those with HIV-dedicated
wards in areas where TB is prevalent. TB surveillance of health care
employees is necessary to identify emerging problems as well as to protect
employees, patients, and visitors.

John V. Rulln M.D.,* Dionisio Herrera, M.D.,* Rosa Cano, M.D.,* Victoria
Moreno, M.D., Pere Godoy, M.D.,* Enrique F. Peir, M.D.,* Juan Castell,
M.D.,* Consuelo Ibaez, M.D.,* Arturo Ortega, M.D., Leopoldo Snchez
Agudo, M.D., and Francisco Pozo, M.D.

Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo: *Programa
de Epidemiologa Aplicada de Campo-Centro Nacional de Epidemiologa;
Centro Nacional de Investigacin Clnica y Medicina Preventiva;
Subdireccin General de Salud, Madrid, Spain

Drs. Herrera, Peir, Castell, and Godoy have received a scholarship from
the Fondo de Investigacin Sanitaria del Instituto de Salud Carlos III
during their 2-year epidemiologic training period in the Field Epidemiology
Training Program (Programa de Epidemiologa Aplicada de Campo).

Acknowledgments

We thank the staff of the Center for Clinical Investigation and Preventive
Medicine of the Instituto de Salud Carlos III for their collaboration in
this investigation; in particular, staff from the following departments:
Administration, Respiratory Diseases, Infectious Diseases, Nursing, and the
Mycobacteriology Laboratory. For the employee tuberculin study, we thank
Drs. Rafael Rey, and Dura; Leopoldo Sanchez Agudo; Margarita Muoz Arcaiz;
Consuelo Yubero Higuera; Concepcin Gonzalez de la Calle; Lucia Martin
Gadea; and Pilar Fuertes Rodrigues for their collaboration.We acknowledge
Dr. Jack Crawford and staff from the Division of Bacterial and Mycotic
Diseases, National Center for Infectious Diseases, CDC, for their
collaboration as reference laboratory for DNA subtyping analysis; Dr.
Daniel Fishbein for editing the manuscript; and Dr. Rafael ReyDurn for
providing us with the 1990 and 1994 data set of the hospital employees
tuberculin test screening. We thank Luis Dorado for his graphics
collaboration.

References

  1. Kent JH. The epidemiology of multidrug-resistant tuberculosis in the
     United States. Med Clin North Am 1993;77:1391-1409.
  2. Edlin BR, Tokars JI, Grieco MH, et al. An outbreak of
     multidrug-resistant tuberculosis among hospitalized patients with the
     acquired immunodeficiency syndrome. N Engl J Med 1992;326:1514-21.
  3. Beck-Sague C, Dooley SW, Hutton MD, et al. Hospital outbreak of
     multidrug-resistant Mycobacterium tuberculosisinfections: factors in
     transmission to staff and HIV-infected patients. JAMA 1992;268:1280-6.
  4. Pearson ML, Jereb JA, Frieden TR, et al. Nosocomial transmission of
     multidrug-resistant Mycobacterium tuberculosis: a risk to patients and
     health care workers. Ann Intern Med 1992;117:191-6.
  5. Fischl MA, Uttamchandani RB, Daikos GL, et al. An outbreak of
     tuberculosis caused by multiple-drug resistant tubercle bacilli among
     patients with HIV infection. Ann Intern Med 1992;117:177-83.
  6. Dooley SW, Villarino ME, Lawrence M, et al. Nosocomial transmission of
     tuberculosis in a hospital unit for HlV-infected patients. JAMA
     1992;267:2632-4.
  7. Centers for Disease Control. Nosocomial transmission of
     multidrug-resistant tuberculosis among HIV-infected personsFlorida
     and New York, 1988-1991. MMWR 1991;40:585-91.
  8. Ausina V, Riutort N, Viado B, et al. Prospective study of
     drug-resistant tuberculosis in a Spanish urban population including
     patients at risk for HIV infection. Eur J Microbiol Infect Dis
     1995;14:105-10.
  9. WHO-EC Collaborating Centres on AIDS. AIDS Surveillance in
     Europe-European Centre for the epidemiological monitoring of AIDS.
     Quarterly Report #45; March 1995.
 10. Boletn Epidemiolgico de la Comunidad de Madrid 1995. Mayo #5, Vol 4.
     Informe: Morbilidad por Enfermedades de Declaracin Obligatoria, 1994.
 11. Friedman CR, Stoecle MY, Johnson WD, Riley LW.
     Double-repetitive-element PCR method for subtyping M.
     tuberculosis clinical isolates. J Clin Microbiol 1995;33:1383-4.
 12. Grupo de Trabajo sobre Tuberculosis. Consenso nacional para el control
     de la tuberculosis en Espaa. Med Clin (Barc) 1992;98:24-31.
 13. Communicable Disease Report. Outbreak of hospital acquired multidrug
     resistant tuberculosis. United Kingdom PHLS Communicable Disease
     Surveillance Centre Weekly 1995.
 14. Centers for Disease Control and Prevention. Guidelines for preventing
     the transmission of Mycobacterium tuberculosis in health-care
     facilities, 1994. MMWR 1994;43 (No. RR-13).
 15. Lilienfeld AM, Lilienfeld DE. Foundations of epidemiology. 2nd ed.
     Oxford, UK: Oxford University Press, 1980;292-5.
 16. Maloney SA, Pearson ML, Gordon MT, et al. Efficacy of control measures
     in preventing nosocomial transmission of multidrug-resistant
     tuberculosis to patients and health care workers. Ann Intern Med
     1995;122:90-5.
 17. Wenger PN, Otten J, Breeden A, et al. Control of nosocomial
     transmission of multidrug-resistant Mycobacterium tuberculosis among
     healthcare workers and HIV-infected patients. Lancet 1995;345:235-40.

---------------------------------------------------------------------------

Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

---------------------------------------------------------------------------
[Pages 130-132]

Dispatches

Application of Pulsed-Field Gel Electrophoresis to an International
Outbreak of Salmonella agona

Between 1 December 1994 and 31 January 1995, Salmonella agona infections
increased abruptly in England and Wales; isolates of S. agona from 41
patients with diarrheal illness were referred to the Laboratory of Enteric
Pathogens of the Public Health Laboratory Service, compared with nine cases
in the previous 12 months. Most isolates were from children under 10 years
of age. Many of the cases were in Jewish children in London and several
other parts of England; two children required hospital admission. S. agona
can be subdivided by phage typing and, by using a method developed in the
Laboratory of Enteric Pathogens (L. R. Ward, unpublished manuscript), we
found that the isolates belonged to S. agona phage type (PT) 15. The
outbreak was traced to a kosher savory snack imported into the United
Kingdom from Israel, and all isolates from the contaminated product and
from patients who had a history of consuming this product were found to
belong to S. agona PT 15 (1). (Full details of this outbreak will be
published elsewhere.)

After notification of the outbreak in the United Kingdom, health
authorities in countries where the contaminated snack had also been
distributed were warned of the possibility of an upsurge in S. agona
infections; these countries included Israel, the United States, and Canada.
Countries in the European Union (EU) were also notified through SALM-NET,
the European salmonella surveillance network (2). As a result of such
notification, isolates of S. agona subsequently identified as PT 15 were
received from patients in Israel, the United States, and France and from
food samples in Israel, the United States, and Canada.

For outbreak investigations, the policy of the Laboratory of Enteric
Pathogens is to use serotyping, phage typing, and antibiogram analysis
(R-typing) for the primary identification and subdivision of isolates,
supplemented when appropriate by a range of DNA-based methods, including
plasmid analysis, ribotyping, insertion sequence (IS) 200 fingerprinting,
and pulsed-field gel electrophoresis (PFGE). The use of these techniques
has recently been reviewed (3). Of these methods, PFGE has been used for
the molecular fingerprinting of S. typhi (4, 5) and for subdivision within
epidemic phage types of S. enteritidis (6, 7).

Because a small number of patients in England and Wales had been infected
with S. agona PT 15 in the 11 months before the kosher snack outbreak, we
decided to characterize the outbreak isolates by genotypic methods and, if
possible, to use such methods for subdivision within the phage type. For S.
agona, strains of this serotype do not possess IS200 elements (8), and
studies by the Laboratory of Enteric Pathogens have demonstrated that the
serotype is unlikely to be subdivided by ribotyping (M.D. Hampton, E.J.
Threlfall, unpublished observations). PFGE was, therefore, considered the
method most likely to provide a genotypic fingerprint suitable for
epidemiologic investigations. Isolates from the food product and from all
patients infected both during the outbreak and in the preceding 11 months
were, therefore, examined by PFGE. Similarly, because of the international
distribution of the contaminated food product, S. agona PT 15 organisms
isolated in Israel and Canada from the food product and also from specimens
from patients in Israel, the United States, and France were examined by
PFGE.

Analysis by PFGE of the fragments resulting from Xba I digestion of genomic
DNA from 78 isolates of S. agona PT 15 made in the United Kingdom, Israel,
the United States, Canada, and France between December 1993 and April 1995
showed 11 distinct pulsed-field profiles (PFP) and one variant profile,
with 14 to 17 resolvable chromosomal fragments, ranging from approximately
25 kb to 680 kb (Figure). These profile types have been designated S. agona
PFP (XbaI) 1 through to S. agona PFP (XbaI) 9, and S.agona PFPs (XbaI) 11
and 12; the variant type has been designated S. agona PFP (XbaI) 6a. The
pattern designated S. agona PFP (XbaI) 10 had been assigned to an isolate
of S. agona of an unrelated phage type, which has been used as a control in
PFGE analysis of this serotype.

[Image not available in ASCII version]
Figure. PFGE profiles of XbaI-digested genomic DNA from strains of S. agona
PT 15. Legend: Tracks 1-14 contained: 1 and 14, lambda 48.5-kb ladder
(Sigma);2, S. agona PFP (XbaI) 6; 3, PFP 6a; 4, PFP 4; 5, PFP 10 (= control
PFP type for S. agona); 6, PFP 9; 7, PFP 7; 8, PFP 3; 9, PFP 2; 10, PFP 5;
11, PFP 1; 11, PFP 8; 13, PFP 9. Gels were run at 6.0 V cm-1 for 36 h with
a 25- to 75-s pulse ramp time.

The predominant PFGE profile, S. agona PFP (XbaI) 6, gave at least 15
resolvable fragments, ranging from 25 kbp to 485 kbp; this profile type was
exhibited by 51 of the 78 isolates examined. The PFP 6 variant, designated
PFP 6a and identified in one of the Canadian food isolates, could be
differentiated from PFP 6 by the presence of an additional fragment of
approximately 395 kbp; in all other respects, S. agona PFP 6a was
indistinguishable from S. agona PFP 6.

The distribution of XbaI-generated PFGE types between source group and
country of isolation in isolates of S. agona PT 15 is shown in the Table.
Of the 10 PFGE profiles in 46 isolates from persons in the United Kingdom,
26 isolates (56.5%) belonged to PFP 6. All isolates with this profile
pattern were made between 25 November 1994 and 17 February 1995 and were
from patients associated with the kosher snackrelated outbreak. Two
isolates of S. agona PT 15 from the contaminated food product were also
examined and both belonged to PFP 6. In contrast, PFP 6 was not identified
in any isolates of S. agona PT 15 made in the United Kingdom in the 11
months before the outbreak. In one case, S. agona PT 15 with the PFGE
profile PFP 9 was isolated from a patient just after the outbreak began, in
November 1994. However, the PFGE profile of this isolate was sufficiently
unrelated to that of the outbreak isolates (PFP 6) to warrant exclusion of
the patient from the kosher snackrelated cases. Nine PFGE profiles were
identified in the remaining 19 U.K. isolates, with PFPs 5, 2, and 1
predominating (Table). Four of five patients infected with isolateswith the
PFP 2 pattern lived in northwestern England, and three of five patients
with the PFP 5 profile lived in southwestern England or southern Wales.
None of these nine PFGE profiles were identified in isolates from
case-patients associated with the kosher snack outbreak.

Nine isolates of S. agona PT 15 from patients and two isolates from the
contaminated food product that had been made in Israel were also examined
by PFGE and, without exception, all strains belonged to PFP 6. Likewise,
all of 10 isolates from outbreak-associated cases in the United States
belonged to PFP 6 as did two of three strains isolated in Canada from the
contaminated food product; the PFP of the remaining Canadian strain, PFP
6a, was closely related to PFP 6. Three PFPs were identified in the six
isolates of S. agona PT 15 from France, of which two (PFP 2 and PFP 5) had
been observed in isolates of PT 15 made in the United Kingdom before the
outbreak. However, none of the isolates of S. agona PT 15 made in France
were of PFP 6 (Table).
--------------------------------------------------------------------------
      Table. Distribution of Xbal pulsed-field profiles in isolates of
  Salmonella agona phage type 15 made in the United Kingdom, Israel, USA,
                       Canada, and France, 19931995a
----------------------------------------------------------------------------
                                       Pulsed-field profiles (PFP)
   Country   Source   Number
                      studied  1  2  3  4  5   6    6a 7  8  9  10  11   12
----------------------------------------------------------------------------
 United      Human   46        3  5  1  2  5  26b      1  1  1       1
 Kingdom
             Snack    2                        2

 Israel      Human    9                        9

             Snack    2                        2

 United
 States      Human   10                       10

 Canada      Snack    3                        2     1

 France      Human    6           1        4                             1
----------------------------------------------------------------------------
 Total               78        3  6  1  2  9  51     1  1  1  1  0   1    1
----------------------------------------------------------------------------
 a Isolates from Israel, the United States, and Canada were made in 1995;
 of the isolates from France, 3 were obtained in 1994 and 3 in 1995.
 b Isolated 25 November 1994 to 17 February 1995.
--------------------------------------------------------------------------

In conclusion, 11 distinct PFGE profile types and one variant type have
been identified in 78 isolates of S. agona PT 15 made in the United
Kingdom, Israel, the United States, Canada, and France between December
1993 and April 1995. One PFGE profile type, PFP 6, was specifically
associated with isolates from a contaminated savory snack and from persons
who consumed this product in the United Kingdom, Israel, and the United
States. These results demonstrate that one strain of S. agona PT 15, with a
characteristic PFGE profile, was responsible for outbreaks in the United
Kingdom, Israel, and the United States in 1994-95. In contrast, the PFGE
profile types of isolates of S. agona PT 15 identified in the United
Kingdom either in the 11 months before the outbreak (or in one case at the
start of the outbreak) and of isolates of PT 15 obtained from patients in
France were unrelated to PFP 6. We conclude that PFGE fingerprinting
provides a genotypic method for subdivision within phage type 15 of S.
agona and that this method has provided an invaluable supplement to phage
typing for investigation of international outbreaks. It is, however,
important to realize that PFGE typing may not be applicable to all
salmonella serotypes and phage types; thus, the methods results should be
carefullyevaluated by using both epidemiologically-related and unrelated
isolates before it is used for outbreak investigations.

E. John Threlfall, Michael D. Hampton, Linda R. Ward, and Bernard Rowe
Laboratory of Enteric Pathogens, Central Public Health Laboratory, London,
United Kingdom

Acknowledgments

We are grateful to the following microbiologists for sending strains of S.
agona for typing: Dr. N. Andorn, Ministry of Health, Government Central
Laboratories, Jerusalem, Israel; Dr. Mehdi Shayedgani, New York State
Department of Health, New York, NY, USA; Dr. Rasik Khakhria, National
Laboratory of Enteric Pathogens, Laboratory Centres for Disease Control,
Ottawa, Canada; and Professor Patrick Grimont, Unites des Enterobacteries,
Institut Pasteur, Paris, France.

References

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  2. Fisher IST, Rowe B, Bartlett CLR, Gill ON. Salm-Netlaboratory-based
     surveillance of human salmonella infections in Europe. PHLS Microbiol
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  3. Threlfall EJ, Powell NG, Rowe B. Differentiation of salmonellas by
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  5. Nair S, Poh CL, Lim YS, Tay L, Yoh KT. Genome fingerprinting of
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     1995; 114: 403-11.
  8. Gibert I, Barb J, Casadesus J. Distribution of insertion sequence
     IS200 in Salmonella and Shigella. J Gen Microbiol 1995; 136: 2555-60.

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Pages 133-135]

Dispatches

Potential Risk for Dengue Hemorrhagic Fever: The Isolation of Serotype
Dengue-3 in Mexico

The Americas have a long history of dengue epidemics, which present public
health problems because of the potential emergence of dengue hemorrhagic
fever (DHF) (1). Efforts to control Aedes aegyptithe only demonstrated
vector of dengue virus in the Americaswere effectively deployed in the
1950s and 1960s when the Pan American Health Organization launched a
continental eradication campaign against yellow fever (2). Aedes aegypti
was eliminated in Mexico in 1963 (3). However, subsequent social and
economic changes in the Americas have permitted the rapid reinfestation of
the vector throughout the region. In Mexico, population movement from rural
areas to urban centersbrought about by intensive industrializationwere
not matched with adequate housing and sufficient water, sewage, and waste
management systems. The introduction and proliferation of nonrecyclable
products provided numerous and effective breeding sites for urban
mosquitoes. For example, from 1960 to 1990, the annual production of
bottles in Mexico increased from 500,000 to 3.5 million, and the annual
production of tires increased from 2 to 17 million (4). Tourism and travel,
promoted as essential to the national economy, have also become important
mechanisms for transporting dengue viruses. Additionally, surveillance,
prevention, and control programs lack the infrastructure and human
resources needed to tackle this neglected health problem (1,4). Millions of
people living in the tropical and subtropical areas of the region face the
reemergence of dengue and DHF (2).

In Mexico from 1984 to 1993, DHF cases were sporadically reported; only 26
cases were identified, followed by 30 cases in 1994 (4). During 1995,
however, the General Directorate of Epidemiology of the Ministry of Health
in Mexico confirmed 358 DHF cases in 18 states with a case-fatality rate of
7.8% (unpublished data). The widespread distribution of DHF cases and of
the vector and the circulation of different serotypes demonstrate the risk
of serious illness throughout the country.

Dengue fever in endemic-disease areas is often not diagnosed properly
because of its nonspecific clinical manifestations. Furthermore, only
patients with symptoms are treated, and patients rarely demand medical
care; thus, the proportion of infected persons in the population is usually
underestimated (5). On the other hand, DHF is an acute, life-threatening
disease that requires specialized treatment in a medical setting.
Identifying dengue serotypes in the continent is one of the most serious
problems faced by every surveillance system in the region. The serotype,
strain, and sequence of infections by different serotypes are among the
most meaningful risk factors for DHF; thus, creating a strong dengue virus
surveillance system in every country in the Americas should be a high
priority (6, 7).

Serologic evidence of dengue in the Americas can be traced back to 1941 in
Panama (8). DEN-2 was isolated in Trinidad in 1953 (9). DEN-3 was isolated
in the Caribbean and Venezuela in 1963 (2,10), DEN-1 was introduced to the
Americas in 1977, and DEN-4 affected the region 4 years later. In 1981,
Cuba had a major DHF epidemic caused by a new strain of DEN-2 (11). DEN-3
was detected in Nicaragua and Panama in 1994 and in Costa Rica in 1995
(12), after a long absence from the region; a strain similar to one in Sri
Lanka and India in the 1980s caused the DHF epidemics in those countries
(12). The identification of DEN-3 in the region increases the probability
of DHF cases associated with a newly circulating serotype. In Mexico, this
particular situation may have important epidemiologic consequences for
several reasons: 1) DEN-3 has not been identified in the country, and the
population is totally susceptible to infection by this serotype; 2)
infection by DEN-3 would most likely be of the secondary type; 3)
population movements through Mexico and towards other countries, might
disseminate this new serotype to areas where susceptible persons will be
exposed to a new serotype; and 4)intensive transmission of dengue would
naturally increase the risk for DHF epidemics.

Surveillance of dengue virus in Mexico began in 1982 when seven isolates of
DEN-1 and DEN-2 were identified from outbreaks reported in the south and
southeastern regions of the country. From 1982 to 1995, the National
Institute of Epidemiological Diagnosis and Reference (INDRE) identified 681
dengue virus isolates. Serotypes were identified by indirect
immunofluorescence with specific monoclonal antibodies donated by the
Division of Vector-Borne Infectious Diseases, Centers for Disease Control
and Prevention, Fort Collins, Colorado.

DEN-1 was the serotype most frequently isolated from 1982 to 1995 (47% of
all isolates), followed by DEN-4 (30%) and DEN-2 (21%) (Table). In 1995,
DEN-3 was identified in 19 patients with classic dengue fever with no
hemorrhagic manifestations (Table). Beginning in 1995, active surveillance
for dengue cases was begun in areas where transmission had been documented.
Sentinel surveillance centers were implemented to obtain serum samples from
febrile patients with a clinical picture suggestive of dengue and to
isolate and identify the serotype involved. From August to December 1995,
245 isolates of dengue virus were obtained, which represented 36% of
isolates obtained during the 14-year period. The prevalence of serotypes
isolated in 1995 differed from those isolated from 1982 to 1994; DEN-1 was
isolated in only 16% of the samples processed, whereas 40% were DEN-2, 8%
were DEN-3, and 36% were DEN-4 (Figure 1). It is unclear whether the change
in distribution of serotypes is due to more intensive surveillance in
certain areas or in a manifestation of herd immunity to serotype 1. This is
the first report of DEN-3 in Mexico and reflects the strengthening of the
surveillance at INDRE for dengue viruses in areas at risk.
--------------------------------------------------
  Table. Number of isolates of dengue virus
             serotypes in Mexico*
--------------------------------------------------
  Year   DEN-1  DEN-2  DEN-3   DEN-4  Total
--------------------------------------------------
  1982        2      5      0       0       7
  1983        5      6      0       2      13
  1984       89      2      0      38     129
  1985       30      8      0       9      47
  1986       65      0      0      24      89
  1987       13      0      0       0      13
  1988       28      0      0       0      28
  1989       21      0      0       0      21
  1990        6      0      0       0       6
  1991        4      0      0      20      24
  1992        1      5      0      19      25
  1993        0     10      0       0      10
  1994       15      9      0       0      24
  1995       40     98     19      88     245
--------------------------------------------------
 Total      319    143     19     200     681
--------------------------------------------------
 * Serum samples from suspect cases were
 added to C6-36 cells grown in D-MEM with 5%
 fetal calf serum for 7 days at 28C and
 incubated for 1 hour. Cells were washed and
 further incubated in D-MEM with 0.4% bovine
 albumin for identification of cytopathic
 effect.
--------------------------------------------------

The geographic and temporal distribution of DEN-3 isolated in 1995 in
Mexico (Figure 2) shows a pattern similar to the one followed by the first
dengue epidemics in the early 1980s (2) and may be related to population
movements towards the northern border. The role of DEN-3 in increasing DHF
cases is still to be determined; to date none of the DHF cases in which
dengue virus was isolated have been associated with this serotype. Five
cases were associated with DEN-1 and 20 cases with DEN-2. Nevertheless, the
infection of DEN-3 in persons sensitized by previous infections with other
serotypes and the widespread susceptibility of the Mexican population to
this serotype must be considered a potential risk factor for future
outbreaks.

The cost of each DHF case has not been fully determined, but the resources
needed to face a DHF epidemic are certainly not available in countries
where the health sector has financial constraints due to unstable economic
conditions. The development of dengue vaccines is encouraging, but the
widespread dispersion of mosquito breeding sites exceeds the capabilities
of vector control programs. Moreover, the potential role of Aedes
albopictus in the transmission of dengue virus in Mexico must be evaluated
because DHF cases have appeared in areas where A. albopictus has been
identified (14). The role of this vector in dengue transmission could
increase should its geographic distribution expand and its susceptibility
to infection increase (15).

[Image not available in ASCII version]
Figure 1. Frequency of dengue serotypes isloated in 1982 to 1994 and in
1995.

[Image not available in ASCII version]
Figure 2. Geographic and temporal distribution of DEN-3 serotype in Mexico.

The challenge faced by national health services is to improve the early
detection of dengue transmission, prevent DHF, and decrease the
case-fatality rate in DHF patients. This strategy must be supported by a
strong surveillance network for viral diseases, which is now being
implemented on a regional basis according to the risk of dengue
transmission in the country. The detailed knowledge of the serotypes
involved in future epidemics will provide useful information that will
define the role of each serotype in the genesis of DHF cases and target
control measures. The threat of a major epidemic requires a control
strategy that will prevent the emergence of this public health problem.

Baltasar Briseo-Garca,* Hctor Gmez-Dants, Enid Argott-Ramrez,*
RaulMontesano, Ana-Laura Vzquez-Martnez,* Sergio Ibez-Bernal,*
Guillermina Madrigal-Ayala,* Cuauhtmoc Ruz-Matus, Ana Flisser,* and
Roberto Tapia-Conyer

*Instituto Nacional de Diagnstico y Referencia Epidemiolgicos, Direccin
General de Epidemiologa, Secretara de Salud, Mxico D.F.,
Mxico;Direccin General de Epidemiologa, Secretara de Salud, Mxico
D.F., Mxico; Present address: Centro de Investigacin de Enfermedades
Infecciosas, Instituto Nacional de Salud Pblica, Secretara de Salud,
Cuernavaca, Mxico

References

  1. Gubler DJ, Trent DW. Emergence of epidemic dengue/dengue hemorrhagic
     fever as a public health problem in the Americas. Infect Agents Dis
     1993; 2: 383-393.
  2. Pan American Health Organization. Dengue and dengue hemorrhagic fever
     in the Americas: Guidelines for prevention and control. Scient Publ
     1994;548:3-22.
  3. Torres-Muoz A. La fiebre amarilla en Mexico: erradicacin de Aedes
     aegypti. Salud Publica Mex 1995;37:S103-S110.
  4. Narro RJ, Gmez-Dants H. El dengue en Mexico: un problema prioritario
     de salud pblica. Salud Publica Mex 1995; 37:S12-S20.
  5. Dietz VJ, Gubler DJ, Rigau-Prez J, Pinheiro F. Epidemic of dengue 1
     in Brazil, 1986: evaluation of a clinically based dengue surveillance
     system. Am J Epidemiol 1990;131:693-701.
  6. Gubler DJ. Vigilancia activa del dengue y de la fiebre hemorrgica del
     dengue. Bol Of Sanit Panam 1989;107:22-30.
  7. Rico-Hesse R. Molecular evolution and distribution of dengue virus
     type 1 and 2 in nature. Virology 1990;174:479-93.
  8. Rosen L. Observations on the epidemiology of dengue in Panama. Am J
     Trop Med Hyg 1958;68: 45-58.
  9. Anderson CR, Downs WG, Hill AE. Isolation of dengue virus from a human
     being in Trinidad. Science 1956;124:224-5.
 10. Pinheiro F. Dengue in the Americas 1980-1987. PAHO Epidemiol Bull
     1989;10:1-7.
 11. Gubler D. Dengue/dengue hemorrhagic fever in the Americas: prospects
     for the year 2000. In: Halstead SB, Gomez-Dantes H, editors. Dengue: a
     worldwide problem, a common strategy. Proceedings of the International
     Conference on Dengue and Aedes aegypti community-based control,
     Merida, Mexico, July 11-16, 1992; Ministry of Health,
     Mexico,1992:19-27.
 12. Gubler JD, Clark GG. Dengue/dengue hemorrhagic fever: the emergence of
     a global health problem. Emerging Infectious Diseases 1995;1:55-7.
 13. Gubler DJ. Aedes aegypti and Aedes aegypti-borne disease control in
     the 1990s: top down or bottom up. Am J Trop Med Hyg 1989;40: 571-8.
 14. Ibaez-Bernal S, Martnez-Campos C. Aedes albopictus in Mexico. J Am
     Mosq Control Assoc 1994; 10: 231-2.
 15. Shope R. Global climate change and infectious diseases, Environ Health
     Perspect 1991;96:171-4.

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Pages 136-140]

Dispatches

Improved Serodiagnostic Testing for Lyme Disease: Results of a Multicenter
Serologic Evaluation

The diverse clinical manifestations of Lyme disease (1-3) have led to
frequent confusion in clinical diagnosis, a confusion compounded by
problems in the accuracy and precision of diagnostic serologic tests (4-11)
and the difficulty of isolating the causative organism (12-14), Borrelia
burgdorferi. In 1990, more than 20 commercially prepared serologic test
kits for Lyme disease were being sold in the United States, but no
nationally standardized reference test was available. A collaborative
evaluation of a selected sample of the commercial test kits by the Centers
for Disease Control and Prevention (CDC) and the Association of State and
Territorial Public Health Laboratory Directors (ASTPHLD) demonstrated poor
concordance of results among these test kits and among a selected group of
state health department laboratories (11). Because of the lack of a
rigorously defined reference serum panel, conclusions could not be drawn
about the sensitivity and specificity of the test kits evaluated. An
unexpected finding in this study was the low concordance in test results
between CDC and two consulting academic reference center laboratories. A
number of other studies also have demonstrated low concordance of Lyme
disease serologic test results obtained by a variety of laboratories
(4-10).

As a result of those findings, the study described here was designed to
fulfill the following objectives: 1) to assemble a serum panel from
patients who had clinically well-defined Lyme disease (preferably confirmed
by isolation of B. burgdorferi); healthy controls, and persons residing in
nonendemic-disease areas whose potentially cross-reactive specimens had
yielded equivocal ELISA results in earlier CDC tests; 2) to test this panel
in a blinded fashion by several recognized Lyme disease reference and
research laboratories; and 3) to compare the accuracy and precision of
tests as a prelude to developing national recommendations for standardized
serologic testing for antibodies to B. burgdorferi.

Tests were performed by five academic centers active in Lyme disease
research (the Marshfield Clinic, Marshfield, Wisconsin; University of
Medicine and Dentistry of New JerseyRobert Wood Johnson Medical School,
New Brunswick, New Jersey; State University of New York at Stony Brook,
Stony Brook, New York; Tufts/New England Medical Center, Boston,
Massachusetts; and the University of Connecticut Health Center, Farmington,
Connecticut) and CDCs Division of Vector-Borne Infectious Diseases,
National Center for Infectious Diseases, based in Ft. Collins, Colorado.

Serum samples from Lyme disease case-patients were obtained from the
participating academic investigators (n = 72) and from the CDC Lyme disease
reference serum collection (n = 37). All case-patient serum samples (total
= 109) were from patients who met the CDC clinical case definition for
surveillance of Lyme disease (15). The clinical manifestations in these
patients ranged from acute erythema migrans (EM) to late neurologic disease
accompanied by Lyme arthritis. B. burgdorferi had been cultured by the
method of Berger et al. from 14 of 34 (41%) acute-phase specimens provided
by CDC (14). Duplicate specimens (n = 85) were randomly selected from the
109 case-patient samples for precision analysis, making a total of 194
case-patient samples in the panel.

Control serum samples were provided by CDC from unpaid healthy blood donors
(n = 113) who resided in areas where Lyme disease is not endemic
(Cincinnati, Ohio, and Atlanta, Georgia; travel histories were not
available from these donors, however. Duplicate specimens (n = 87) also
were randomly selected, resulting in 200 noncase samples in the serum
panel. Additional control samples were obtained from persons who resided in
areas where Lyme disease was not endemic but whose physicians submitted
their serum for Lyme disease testing to CDC through their state health
department (n = 113). These specimens from patients with suspected cases
had borderline (equivocal) seroreactivity in the whole cell sonicate (WCS)
enzyme-linked immunoassay (ELISA)used by CDC before 1992 and are referred
to hereafter as WCS-suspects (16). The addition of duplicate specimens (n
= 87) brought this group to 200 equivocally seroreactive samples.

Serum was separated and frozen by the original collectors and shipped
frozen to CDCs facilities in Ft. Collins, Colorado. The specimens were
divided into aliquots and coded; code labels were applied by CDC staff not
involved in serologic testing of the specimens (n = 594). The panels were
then refrozen and shipped on dry ice for blind testing by participating
investigators. All specimens were received frozen. To calculate test
sensitivity and specificity, only the result of the sample with the lower
random code number of each pair was used.

Each laboratory employed the testing method that it used routinely at the
time this study was undertaken (1992). CDC used an ELISA with a WCS antigen
prepared from highly passaged strain B31 (gift of A. Barbour, University of
Texas Health Sciences Center, San Antonio, Texas) and an ELISA with a
strain B31 flagellar antigen (FLA) then being evaluated (16, 17). The other
five participating investigators used ELISA tests that employed a WCS
antigen of B. burgdorferi. Four used assays developed in their own
laboratories, and one used a commercially available test kit (18-22). Three
investigators also tested all specimens by Western blotting using published
methods (19, 20). Two of these three performed immunoblotting for IgM and
IgG antibodies separately. One laboratory tested for IgM and IgG together.

Each participating laboratory submitted the raw data of its results, along
with a dichotomous interpretation of those results as either positive or
negative. By prior agreement, ELISA results that fell into a range
ordinarily reported as equivocal by that laboratory were treated as
negative for this analysis. Statistical analyses undertaken at CDC included
calculations of sensitivity (true positives correctly identified),
specificity (true negatives correctly identified), precision (frequency of
obtaining the same result on duplicate analysis of a specimen), and a
measure of concordance (agreement among investigators) of results among the
tests using the kappa statistic.

The accuracy and precision of the serologic tests as performed in 1992 by
all six laboratories is summarized in Table 1. The test methods of
investigators 1, 2, and 3 produced essentially equivalent results, with
moderately high sensitivity (73% to 79%) for the aggregate of all
case-patient samples tested and high specificity (98% to 99.5%). Precision
was high in these three laboratories for both blood donor samples (97% to
99%) and the WCS-suspects samples submitted from areas where Lyme disease
is nonendemic (94% to 98%). Precision was somewhat lower for the
case-patient samples (82% to 91%).

 Table 1. Accuracy and precision of serologic tests for Lyme disease
                          performed in 1992
---------------------------------------------------------------------
                    Accuracy (%)               Precision(%)
---------------------------------------------------------------------
                                         Case   Non-case-    WCS
 Investigator Sensitivity Specificity  patients patients  suspects
---------------------------------------------------------------------
 CDC
   (WCS)      93          71           93       77        69

 CDC
   (FLA)      92          82           92       79        62

  1           73          99.5         89       99        98

  2           76          99           82       99        97

  3           79          98           91       97        94

  4           49          91           79       94        93

  5           40          72           63       74        77
---------------------------------------------------------------------
  Specimens from patients with suspected cases that had borderline
 (equivocal) seroreactivity in an enzyme-linked immunosorbent assay
 with whole-cell sonicate antigen (WCS).
---------------------------------------------------------------------

The performance of the other three laboratories, including CDCs, was poor.
Both CDC ELISA tests had high sensitivity (92% to 93%), but low specificity
(71% to 82%). Precision for case-patient specimens was fairly high (92% to
93%), but low for both non-case-patient (77% to 79%) and WCS-suspects
groups (62% to 69%). The method of investigator 4 gave very low sensitivity
(49%), moderately high specificity (91%), poor precision with Lyme disease
case-patient specimens (79%), but good precision with blood donor and
WCS-suspects samples (93% to 94%). Investigator 5, who used a commercial
test, obtained results with low accuracy and precision.

Concordance was high (kappa statistic 0.700) between the results of
investigators 1, 2, and 3. The CDC FLA test showed moderate concordance
(kappa 0.400) with results from investigators 1, 2, and 3. The results of
investigator 4 showed moderate concordance with those of investigators 1
and 2 (kappa 0.400) and low concordance (0.400) with the other results. The
results of investigator 5 had low concordance with all other results. The
CDC WCS test showed moderate concordance with the FLA test, but low
concordance with results of the ELISA tests of the other laboratories.

The three investigators with the best results all used Western blot to
supplement their ELISA. Two of these three investigators submitted their
dichotomous test interpretation with and without using Western blot
results. The sensitivity improved by 20% for one investigator and by 30%
for the other when Western blot results were included. The improvement
resulted from identifying as positive by Western blot those case-patient
specimens from which an equivocal result was obtained by ELISA and which by
study design would have been counted as negative by ELISA results alone.
Specificities were not affected by Western blot analysis in this group of
three investigators, since the serum panel in this study did not contain
cross-reactive sera; and the negative controls and WCS-suspects had
negative results by both ELISA and Western blot.

Test sensitivity from the three laboratories with the best test specificity
(98%) was analyzed according to the clinical manifestations in the
case-patients (Table 2). As expected, the sensitivities of the tests were
lowest in specimens from patients with early disease, 59% to 66% for
erythema migrans and 63% to 75% for early neurologic disease. Sensitivities
were much higher for samples of patients with late disease. Sensitivities
of 89% to 95% were obtained for Lyme arthritis patients and 91% to 100% for
persons with late neurologic disease, primarily encephalopathy or
polyneuropathy.

The emergence of a disease can outstrip the development of reliable methods
for its laboratory diagnosis. The serodiagnosis of Lyme disease has been
fraught with problems of precision and accuracy. This study provided an
opportunity for selected academic research centers and CDC to compare the
performance of their individual tests by using a serum panel from
clinically well-characterized patients and controls from
nonendemic-disease areas. The clinical diagnosis of early Lyme disease was
supported by the isolation of B. burgdorferi from skin biopsy specimens
(14), when possible. The panel, which was coded blind had a sufficiently
large number of samples (n = 335) to provide adequate statistical power for
the comparison.

  Table 2. Test sensitivity of laboratories demonstrating a
                   test specificity of 98%
------------------------------------------------------------
                            Sensitivity, % (positive
                                 samples/total)
------------------------------------------------------------
 Clinical            Laboratory   Laboratory   Laboratory
 Manifestations      1            2            3
------------------------------------------------------------
 Erythema migrans,
 all                 59 (55/94)   60 (56/94)   66 (62/94)

   Acute phasea      65 (11/17)   65 (11/17)   76 (13/17)

   Convalescent
 phaseb              57 (44/77)   58 (45/77)   64 (49/77)

 Carditis            100 (2/2)    100 (2/2)    100 (2/2)

 Lyme arthritisc     89 (58/65)   95 (62/65)   92 (60/65)

 Neurologic, all     85 (28/33)   88 (29/33)   91 (30/33)

   Early             63 (5/8)     63 (5/8)     75 (6/8)

   Late              91 (10/11)   100 (11/11)  91 (10/11)

   Late and
 arthritis           93 (13/14)   93 (13/14)   100 (14/14)
-------------------------------------------------------------
 Total               74           77           79
                     (143/194)    (149/194)    (154/194)
-------------------------------------------------------------
 a <=30 days from onset of erythema migrans to blood
 collection.
 b > 30 days from onset of erythema migrans to blood
 collection.
 c Without neurologic signs or symptoms.

Laboratories that supplemented their primary test, an ELISA, with
immunoblotting achieved greater test accuracy than those that did not. The
use of Western blot as a second test enabled the best performing
laboratories to increase test sensitivity without a concomitant loss of
specificity. This increase in sensitivity occurred as a result of
identifying as true positives by Western blot a number of those specimens
from patients with clinical cases of Lyme disease that were interpreted as
equivocal by ELISA and would have been otherwise considered in this study
as dichotomously negative results. Although the investigators employing
Western blot tested all panel specimens with this method, they did so at
that time to evaluate the potential value of Western blot in Lyme disease
serologic diagnosis.

The observation that Western blotting could be employed to resolve
equivocal ELISA results gave additional impetus for evaluating its
potential adjunctive role in Lyme disease serodiagnosis and eventually led
to the finally recommended two-test approach (23). The potential utility of
Western blotting, however, pointed out the lack of standardized methods for
producing blots and standardized interpretive criteria.

Performance of the CDC WCS and FLA ELISA in this study that did not include
known cross-reactive sera suggested that the positive cut-off value for
these tests was inappropriately low, thereby increasing sensitivity at the
expense of specificity. These results then explained the large number of
borderline WCS ELISA results obtained by CDC when it tested the sera of
patients residing in areas where Lyme disease was not endemic. This group
of WCS suspects was nearly uniformly found to be negative on ELISA by the
three laboratories with the best performance (Table 1) (23).

Specificity in this study was determined by testing specimens from blood
bank donors. With these samples, specificity in the three laboratories that
used immunoblotting was very high (98% to 99.5%). The test panel did not,
however, contain specimens frompatients with conditions known to produce
cross-reacting antibodies (e.g., syphilis) or polyclonal B-cell activation
(e.g., Epstein-Barr virus infection or systemic lupus erythematosus). Thus,
reported specificities in this study are likely higher than they would have
been if cross-reactive specimens were included in the evaluation.
Subsequent studies that included cross-reactive sera demonstrated that
Western blotting correctly identifies many false-positive ELISA reactions
(23, 24).

This study confirmed in the reference and research laboratory setting the
previously documented problems with accuracy and precision of
serodiagnostic tests by using WCS antigens of B. burgdorferi (4-11). The
study confirmed that a serious disparity existed between the test results
obtained by CDC and those obtained by academic reference centers with the
best testing performances. These results guided corrective action and led
to the adoption by CDC and ASTPHLD of a two-test approach to serodiagnosis
(23), which forms the basis for the future national standardization of Lyme
disease serologic testing methods.

Robert B. Craven,* Thomas J. Quan,* Raymond E. Bailey,* Raymond Dattwyler,
Raymond W. Ryan, Leonard H. Sigal, Allen C. Steere, Bradley Sullivan,#
Barbara J.B. Johnson,* David T. Dennis,* and Duane J. Gubler*

*Centers for Disease Control and Prevention, Fort Collins, Colorado, USA;
State University of New York at Stony Brook, Stony Brook, New York, USA;
University of Connecticut Health Center, Farmington, Connecticut, USA;
University of Medicine and Dentistry of New JerseyRobert Wood Johnson
Medical School, New Brunswick, New Jersey, USA; Tufts/New England Medical
Center, Boston, Massachusetts, USA; #Marshfield Clinic, Marshfield,
Wisconsin, USA

A portion of this information was presented at the VIth International
Conference on Lyme Borreliosis, Bologna, Italy, June 1994 and at the Second
National Conference on Serologic Diagnosis of Lyme Disease, Dearborn,
Michigan, October 1994.

References

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  3. Steere AC, Bartenhagen NH, Craft JE, et al. The early clinical
     manifestations of Lyme disease. Ann Intern Med 1989;99:76-82.
  4. Bakken LL, Case KL, Callister SM, Bourdeau NJ, Schell RF. Performance
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  5. Hedberg CW, Osterholm MT, MacDonald KL, White KE. An interlaboratory
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  6. Hedberg CW, Osterholm MT. Serologic tests for antibody to Borrelia
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  7. Jones JM. Serodiagnosis of Lyme disease. Ann Intern Med 1991;114:1064.
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  9. Luger SW, Krauss E. Serologic tests for Lyme disease: interlaboratory
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 10. Schwartz BS, Goldstein MD, Ribeiro JMC, Schulze TL, Shahied SI.
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 12. Steere AC, Grodzicki RL, Kornblatt AN, et al. The spirochetal etiology
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 13. Benach JL, Bosler EM, Hanrahan JP, et al. Spirochetes isolated from
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     1983;308:740-2.
 14. Berger, BW, Johnson RC, Kodner C, Coleman L. Cultivation of Borrelia
     burgdorferi from erythema migrans lesions and perilesional skin. J
     Clin Microbiol 1992;30:359-61.
 15. Centers for Disease Control and Prevention. Case definitions for
     public health surveillance. MMWR 1990;39(RR-13):19-21.
 16. Russell H, Sampson JS, Schmid GP, Wilkinson HW, Plikaytis B.
     Enzyme-linked immunosorbent assay and indirect immun ofluorescence
     assay for Lyme disease. J Infect Dis 1984;149:465-70.
 17. Hansen K, sbrink E. Serodiagnosis of erythema migrans and
     acrodermatitis chronica atrophicans by the Borrelia burgdorferi
     flagellum enzyme-linked immunosorbent assay. J Clin Micrbiol
     1989;27:545-51.
 18. Craft JE, Grodzicki RL, Steere AC. Antibody response in Lyme disease:
     evaluation of tests. J Infect Dis 1984;149:789-95.
 19. Grodzicki RL, Steere AC. Comparison of immunoblotting and indirect
     enzyme-linked immunosorbent assay using different antigen preparations
     for diagnosing early Lyme disease. J Infect Dis 1988;157:790-97.
 20. Dressler F, Whalen JA, Reinhardt BN, Steere AC. Western blotting in
     the serodiagnosis of Lyme disease. J Infect Dis 1993;167:392-400.
 21. Fister RD, Weymouth LA, McLaughlin JC, Ryan RW, Tilton RC. Comparative
     evaluation of three products for the detection of Borrelia burgdorferi
     antibody in human serum. J Clin Microbiol 1989;27:2834-37.
 22. Dattwyler RJ, Volkman DJ, Luft BJ, Halperin JJ, Thomas J, Golightly
     MG. Seronegative Lyme disease: dissociation of specific T- and
     B-lymphocytic response to Borrelia burgdorferi. N Engl J Med
     1988;319:1441-6.
 23. Association of State and Territorial Public Health Laboratory
     Directors and the Centers for Disease Control and Prevention.
     Recommendations. In: Proceedings of the Second National Conference on
     Serologic Diagnosis of Lyme Disease (Dearborn, Michigan). Washington,
     DC: Association of State and Territorial Public Health Laboratory
     Directors 1995:1-5.
 24. Johnson BJB, Robbins KE, Bailey RE, et al. Serodiagnosis of Lyme
     disease: accuracy of a two-step approach using a flagella-based ELISA
     and immunoblotting. J Infect Dis 1995; submitted.

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Pages 141-144]

Dispatches

Emergence of Bartonella quintana Infection among Homeless Persons

Bartonella quintana has episodically emerged as a cause of infection among
distinct and diverse populations during the 20th century. The organism was
first identified as an important human pathogen during World War I when it
caused epidemics of louse-borne trench fever that affected an estimated 1
million troops in Europe (1, 2). Trench fever was characterized by fever,
rash, bone pain, and splenomegaly and ranged in severity from a mild
flulike illness to a more severe, relapsing disease. B. quintana infections
were rarely recognized from the end of World War II until the 1980s when
the organism reemerged as an opportunistic pathogen among HIV-infected
persons. In this population, B. quintana has been identified as a cause of
bacillary angiomatosis, endocarditis, and bacteremia (3-5) and has been
isolated from AIDS patients in France (6) and the United States (3-5).

In the 1990s, B. quintana has emerged among homeless persons in North
America and Europe. In 1993, the organism was isolated from the blood
specimens of 10 patients at a single hospital in Seattle, Washington,
within a 6-month period (7). These patients had illnesses characterized by
fever and persistent bacteremia. Endocarditis developed in two patients,
one of whom required a heart valve replacement. All 10 patients had chronic
alcoholism, eight were homeless, and the six who were tested for HIV
infection were HIV-negative. These six were the first cases of invasive B.
quintana infection among HIV-negative persons reported in the United
States. Results of a case-control study indicated that the patients with
Bartonella bacteremia were more likely than controls (other hospitalized
patients from whom blood specimens were obtained at approximately the same
time) to be homeless (p = 0.001), to have a history of alcohol abuse (p =
0.001), and to be nonwhite (p = 0.007). The isolates from the 10 patients
were identical by polymerase chain reaction restriction-fragment-length
polymorphism testing, which further suggests that the cases were
epidemiologically linked. Patients characteristics were obtained by
retrospective medical record review, and at the time they sought treatment,
three patients reported a recent cat scratch, five had scabies, and one had
lice. More complete information, however, on patients past exposures to
animals and ectoparasites was not available.

In 1995, Drancourt and co-workers reported three cases of B. quintana
endocarditis among HIV-negative, homeless, alcoholic men in France (8). One
of the patients had reported contact with a dog, and one had reported
contact with dogs and cats; however, a current or past history of infection
with lice or scabies was not documented for any of the patients. In 1995,
Stein and Raoult also reported serologic evidence of B. quintana infection
in an HIV-negative, homeless man from Marseilles, who had a relapsing
febrile illness and a history of louse infestation (9).

As a follow-up to the 1993 B. quintana outbreak in Seattle in 1994, we
conducted a seroprevalence study of anti-Bartonella antibodies among
patients at a community clinic in the skid row section of Seattle, which
serves a primarily homeless and indigent population (10). The median age of
the 192 patients included in the study was 45 years, 156 (81%) of the 192
were male, and 126 (66%) were classified as homeless. B. quintana IgG
titers >= 64 were detected by an indirect fluorescence antibody assay (11)
in 39 (20%) of the 192 clinic patients. In contrast, only 4 (2%) of 199
banked blood specimens from an age-matched and sex-matched comparison group
of Seattle volunteer blood donors had titers >=64 (p < 0.001). Among clinic
patients, seropositivity (titer >=64) was associated by univariate analysis
with older age, homelessness (relative risk [RR] 2.0; 95% confidence
interval [CI] 1.0-4.1), alcohol abuse (RR, 2.5; 95% CI 1.4-4.2), smoking
(RR, 2.0; 95% CI 1.2 -3.4), and injection drug use (RR, 2.5; 95% CI
1.3-4.8). By multivariate analysis, only alcohol abuse remained
independently associated with seropositivity (odds ratio 3.3; 95% CI
1.6-6.9), and of 39 seropositive patients, 21 (54%)had a history of chronic
alcoholism. Reliable data on past exposure to animals or ectoparasites were
also not available for patients in this study.

The study was limited by the well-described cross-reactivity of the assay
between Bartonella species (12, 13), and most (62%) clinic patients with B.
quintana >= 64 also had titers >= 64 to B. henselae. It is, therefore,
possible that some of the seropositive patients may have been exposed to
Bartonella species other than B. quintana. These findings do, however, show
that a surprisingly high proportion of clinic patients without a history of
documented Bartonella infection had detectable anti-Bartonella antibodies
and may have been exposed to B. quintana.

Multiple factors, including those related to disease transmission, host
susceptibility, and ability to detect the organism, have likely contributed
to the emergence of B. quintana infection among the homeless. Transmission
of B. quintana from human to human by the body louse, Pediculus humanus,
has been experimentally documented (1) and is believed to have been the
predominant mode of transmission of epidemic trench fever in World Wars I
and II. Lice reside primarily in the seams of clothing and are easily
killed by immersion in water 50C or warmer (14), which explains the
propensity for louse-borne infections among displaced persons or wartime
troops. Although these reports of B. quintana infection among homeless
persons lack sufficient information to conclusively determine the disease
vector, louse-borne infection remains a plausible hypothesis. Lice,
however, have not been associated with bacillary angiomatosis among AIDS
patients, although exposure to cats (and, therefore, possibly to fleas) has
been associated with bacillary angiomatosis and bacillary peliosis caused
by Bartonella species (15) and with cat-scratch disease caused by B.
henselae (16). Thus, it is possible that B. quintana infection is spread
among homeless persons by as yet unidentified vectors or reservoirs.

Homeless persons are also at risk for non-vectorborne infectious diseases.
An increased risk for tuberculosis in this population is well documented
(17, 18), and outbreaks of meningococcal disease (19, 20), pneumococcal
disease (21), and diphtheria (22, 23) have been reported. It is likely that
factors such as crowding, altered immunity due to alcoholism or other
co-existing health problems, and inadequate or infrequent access to medical
care affect the transmission and spread of infectious diseases among the
homeless. Previous studies have shown that the clinical response to a
standard inoculum of B. quintana varies substantially in experimental study
patients (1); this variation indicates that host factors are likely
important determinants of the risk for clinical infection following
exposure to the organism.

Although cases of B. quintana bacteremia among homeless persons have thus
far been reported only from France and Seattle, Washington, the problem is
probably not confined to these locations. B. quintana is a fastidious and
slow-growing bacterium that generally requires special culturing techniques
for isolation (3-5, 24), and many clinical laboratories do not routinely
use blood culturing methods that are sensitive for isolating this organism.
Moreover, B. quintana infection can result in a broad range of often
nonspecific clinical manifestations (1, 3-5); therefore, case-patients
evaluated for suspected bacteremia may represent only a small proportion of
infected persons, as suggested by the results of the Seattle seroprevalence
survey. To better define the geographic distribution and prevalence of B.
quintana infection among homeless populations, a heightened awareness for
this infection on the part of clinicians and the use of appropriate culture
techniques by microbiology laboratories serving this population are needed.
In addition, more specific serologic tests would aid in the diagnosis and
assessment of the epidemiologic characteristics of B. quintana infections.

The optimal treatment regimen for HIV-negative patients with suspected or
confirmed B. quintana infection has not been established. Minimal published
data exist regarding antimicrobial therapy for this infection, and in vitro
susceptibility testing has provenunreliable (25). Nonetheless, on the basis
of limited data, we believe it is reasonable to treat immunocompetent
patients who have uncomplicated B. quintana bacteremia with at least 14
days of oral therapy with erythromycin, azithromycin, doxycycline, or
tetracycline. In the 1993 Seattle outbreak, most patients had a
satisfactory response to treatment with a beta-lactam agent followed by
either erythromycin or azithromycin for 14 days (7). Although the number of
patients identified with B. quintana endocarditis is small, most of these
patients have required cardiac valve replacement despite intravenous
antimicrobial therapy (5, 8, 26). Therefore, we recommend that patients
with B. quintana endocarditis receive a more prolonged course of at least 4
to 6 months of antimicrobial therapy and cardiac valve replacement if
needed. Further study is needed to determine the role of bactericidal
agents, such as third generation cephalosporins or quinolones, as
monotherapy or in combination with a bacteriostatic agent for treating
invasive B. quintana infections.

Many aspects of the acquisition and pathogenesis of B. quintana infections,
and specifically B. quintana infections among the homeless, are not well
defined. Changes in the organism itself that have led to increased
virulence may in part account for its reemergence; however, microbiologic
data that can support or refute this hypothesis are lacking (27). The
absence of recently identified cases in Seattle and in other areas with
laboratories that use culture techniques appropriate for isolating
Bartonella species suggests an episodic pattern of disease, with few or no
cases occurring during interepidemic periods. It seems clear, however, that
this most recent emergence of an old disease is related, at least in part,
to societal factors that have contributed to urban decay and the existence
of large homeless populations in our cities. As with other emerging
infectious diseases, further efforts to identify, evaluate, and control B.
quintana infections among homeless persons are challenges that will require
the coordinated effort of clinicians, microbiologists, and public health
officials.

Lisa A. Jackson, M.D., M.P.H.*, and David H. Spach, M.D.
*School of Public Health and Community Medicine, and the Division of
Infectious Diseases and the Department of Medicine, University of
Washington, Seattle, Washington, USA

References

  1. Vinson JW, Varela G, Molina-Pasquel C. Trench fever. III. Induction of
     clinical disease in volunteers inoculated with Rickettsia quintana
     propagated on blood agar. Am J Trop Med Hyg 1969;18:713-22.
  2. Slater LN, Welch DF. Rochalimaea species (recently renamed
     Bartonella). In: Mandell GL, Bennett JE, Dolin R, editors. Principles
     and practice of infectious diseases, 4th ed. New York: Churchill
     Livingstone, 1995:1741-7.
  3. Welch DF, Pickett DA, Slater LN, Steigerwalt AG, Brenner DJ.
     Rochalimaea henselae sp. nov., a cause of septicemia, bacillary
     angiomatosis, and parenchymal bacillary peliosis. J Clin Microbiol
     1992;30:275-80.
  4. Koehler JE, Quinn FD, Berger TG, LeBoit PE, Tappero JW. Isolation of
     Rochalimaea species from cutaneous and osseous lesions of bacillary
     angiomatosis. N Engl J Med 1992;327:1625-31.
  5. Spach DH, Callis KP, Paauw DS, et al. Endocarditis caused by
     Rochalimaea quintana in a patient infected with human immunodeficiency
     virus. J Clin Microbiol 1993;31:692-4.
  6. Maurin M, Roux V, Stein A, Ferrier F, Viraben R, Raoult D. Isolation
     and characterization by immunofluorescence, sodium dodecyl
     sulfate-polyacrylamide gel electrophoresis, western blot, restriction
     fragment length polymorphism-PCR, 16S rRNAgene sequencing, and
     pulsed-field gel electrophoresis of Rochalimaea quintana from a
     patient with bacillary angiomatosis. J Clin Microbiol 1994;32:1166-71.
  7. Spach DH, Kanter AS, Dougherty MJ, et al.Bartonella (Rochalimaea)
     quintana bacteremia in inner-city patients with chronic alcoholism. N
     Engl J Med 1995;332:424-8.
  8. Drancourt M, Mainardi JL, Brouqui P, et al. Bartonella (Rochalimaea)
     quintana endocarditis in three homeless men. N Engl J Med
     1995;332:419-23.
  9. Stein A, Raoult D. Return of trench fever [letter]. Lancet
     1995;345:450-1.
 10. Jackson LA, Spach DH, Kippen DA, Sugg NK, Regnery RL, Sayers MH, Stamm
     WE. Seroprevalence to Bartonella quintana among patients at a
     community clinic in downtown Seattle. J Infect Dis 1996;173:1023-6.
 11. Regnery RL, Olson JG, Perkins BA, Bibb W. Serologic response to
     Rochalimaea henselae antigen in suspected cat scratch disease.
     Lancet 1992;339:1443-5.
 12. Waldvogel K, Regnery RL, Anderson BA, Caduff R, Caduff J, Nadal D.
     Disseminated cat-scratch disease: detection of Rochalimaea henselae in
     affected tissues. Eur J Pediatr 1994;153:23-7.
 13. Dalton MJ, Robinson LE, Cooper J, Regnery RL, Olson JG, Childs JE. Use
     of Bartonella antigens for serologic diagnosis of cat-scratch disease
     at a national referral center. Arch Intern Med 1995;155:1670-6.
 14. Elgart ML. Pediculosis. Dermat Clin 1990;8:219-28.
 15. Tappero JW, Mohle-Boetani J, Koehler JE, et al. The epidemiology of
     bacillary angiomatosis and bacillary peliosis. JAMA 1993;269:770-5.
 16. Zangwill KM, Hamilton DH, Perkins BA, et al. Cat scratch disease in
     Connecticut: epidemiology, risk factors, and evaluation of a new
     diagnostic test. N Engl J Med 1993;329:8-13.
 17. Barnes PF, El-Hajj H, Preston-Martin S, et al. Transmission of
     tuberculosis among the urban homeless. JAMA 1996;275:305-7.
 18. Nardell E. McInnis B, Thomas B, Weidhaus S. Exogenous reinfection with
     tuberculosis in a shelter for the homeless. N Engl J Med
     1986;315:1570-5.
 19. Filice GA, Englender SJ, Jacobson JA, et al. Group A meningococcal
     disease in skid rows: epidemiology and implications for control. Am J
     Public Health 1984;74:253-4.
 20. Counts GW, Gregory DF, Spearman JG, et al. Group A meningococcal
     disease in the U.S. Pacific Northwest: epidemiology, clinical
     features, and effect of a vaccination control program. Rev Infect Dis
     1984;6:640-8.
 21. DeMaria A Jr, Browne K, Berk SL, et al. An outbreak of type I
     pneumococcal pneumonia in a mens shelter. JAMA 1980;244:1446-9.
 22. Pedersen AHB, Spearman J, Tronca E, et al. Diphtheria on skid road,
     Seattle, Washington, 1972-75. Public Health Rep 1977;92:336-42.
 23. Heath CW Jr, Zusman J. An outbreak of diphtheria among skid row men. N
     Engl J Med 1962;267:809-12.
 24. Larson AM, Dougherty MJ, Nowowiejski DJ, Welch DF, Matar GM,
     Swaminathan B, Coyle MB. Detection of Bartonella (Rochalimaea)
     quintana by routine acridine orange staining of broth blood cultures.
     J Clin Microbiol 1994;32:1492-6.
 25. Myers WF, Grossman DM, Wisseman CL. Antibiotic susceptibility patterns
     in Rochalimaea quintana, the agent of trench fever. Antimicrob Agents
     Chemother 1984;25:690-3.
 26. Spach DH, Kanter AS, Daniels NA, et al. Bartonella (Rochalimaea)
     quintana species as a cause of culture-negative endocarditis. Clin
     Infect Dis 1995;20:1044-7.
 27. Relman DA. Has trench fever returned? N Engl J Med 1995;332:463-4.

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Pages 145-146]

Dispatches

The Reemergence of Visceral Leishmaniasis in Brazil

Because of a complex array of factors, an increasing number of new and
reemerging infectious diseases are being recognized in both industrialized
and developing countries in the Americas (1,2). The expanding population,
living in overcrowded conditions with inadequate housing and sanitary
facilities, has been exposed to new diseases and human pathogens. For
example, the appearance of the South American arenaviruses (Junin, Machupo,
and Guanarito) illustrates how exploitation of new areas for human
settlement and agriculture increases the likelihood that new infectious
diseases will emerge. Cholera, plague, AIDS, dengue hemorrhagic fever, and
urban/periurban visceral leishmaniasis are examples of new and reemerging
diseases in the region.

In tropical America, zoonotic visceral leishmaniasis caused by Leishmania
chagasi, an intracellular protozoon, is a long-lasting infectious disease
characterized by weight loss, cough, fever, diarrhea, hepatosplenomegaly,
and lethargy. Since not all symptoms appear simultaneously, and many other
conditions manifest the same symptoms, the diagnosis is not always
opportune. This disease is different from the anthroponotic form of
visceral leishmaniasis found in Sudan and India. In the New World this
disease is more common among poor, malnourished children (3) under 15 years
of age who live in semiarid regions. The case-fatality rate is low if
pentavalent antimony therapy is introduced promptly (4). The disease has
also been reported in immunodepressed persons, especially those with HIV
infection.

The domestic dog is the principal animal reservoir of L. chagasi and, as
the constant companion of humans throughout the endemic-disease area,
contributes to the dispersal of diseases during human migrations. The
protozoon is transmitted from one mammalian host to another, including
humans, primarily by the bite of a sand fly who has fed on an infected dog.
In the Americas, the principal sand fly vector is Lutzomyia longipalpis
(5). This 2- to 3-mm long fly has peridomestic and intradomiciliary habits
(6) and avidly bites humans at night, primarily during twilight, while the
host is resting.

Visceral leishmaniasis is usually diagnosed by identifying the parasite in
spleen aspirates; microscope examination of bone marrow aspirates offers a
satisfactory alternative. Specific antibodies may be detected by
immune-enzymatic reactions and immunofluorescence, but these methods lack
specificity and are too complex to be carried out in the field. TRALd, a
recently developed diagnostic test, consists of a 60-second dipstick, based
on a recombinant protein rK39 of a sequence of 298 amino acids and an
improved serologic procedure (7). The procedure is undergoing field testing
and appears to be a promising tool for control programs.

Although visceral leishmaniasis was previously known as a rural disease,
large outbreaks and epidemics of visceral leishmaniasis have been reported
recently in large cities in Brazil because of the favorable epidemiologic
conditions associated with the reduction of the natural ecologic space of
this zoonosis. Waves of droughts, lack of available farm land, and famine
have led to a large migration of the population, to the peripheral suburbs
of large cities, creating densely populated settlements of shanties
(favelas) with minimal infrastructure and sanitation. Most families that
migrate are young, less-established farmers of peak child-bearing age;
children under 15 years of age account for a large percentage of the entire
population. In these communities, the newly introduced disease (parasite)
encounters a vast number of nonimmune hosts who, because of poor living
conditions, are malnourished; malnutrition is one of the prime risk factors
for L. chagasi infection and visceral leishmaniasis (3). The habit of
keeping domestic animals such as dogs, chickens, and horses in the back
yard provides an abundance of blood meals for sand fly vectors and raises
vector population densities dramatically.

Two recent examples of reemerging visceral leishmaniasis in Brazil occurred
in the citiesof Teresina (population = 678,000), the capital of Piau
State, and So Lus (population = 918,000), the capital of Maranho State.
In Teresina, which has the best-documented records, an epidemic of visceral
leishmaniasis occurred from 1981 to 1985; it was initially limited to rural
settings, but later spread to peripheral areas of the city (9) (Figure 1).
Spraying with residual insecticide helped bring the disease under control
during the subsequent 3 years. Since 1989, visceral leishmaniasis has
gradually reemerged; it reached epidemic levels by the end of 1992 and
peaked in 1994. In the State of Maranho, the disease reemerged in 1993,
(Figure 2), 10 years after a previous epidemic (1982 to 1986). In both
Teresina and So Lus, the epidemics were preceded by prolonged, severe
drought. The disease was found predominantly among young people who came
from rural areas and lived in periurban, substandard housing and kept
domestic animals in their back yards. The two states accounted for
approximately 40% to 50% of the number of cases reported in the country
during 1993 to 1994 (3,000/year).

[Image not available in ASCII version]
Figure 1. Number of cases of visceral leishmaniasis from the state of
Pianu, Brazil, 1971 to 1994.

[Image not available in ASCII version]
Figure 2. Number of cases of visceral leishmaniasis from the state of
Maranho, Brazil, 1979 to 1994.

Emergency control plans based on rapid diagnosis, complete treatment of
human patients, spraying of residual insecticide, ultra-low volume
spraying, elimination of infected (seropositive) dogs, and health education
were initiated in these two cities in July 1994 (10). The incidence rate
dropped markedly from January 1995, as expected, since the incubation
period of visceral leishmaniasis is 4 to 6 months (11, 12). The remaining
reported cases include those with longer incubation periods, as well as
others, because of the lack of complete coverage of the control
interventions and reduced sensitivity of the current diagnostic tools.

Technical expertise and effective mechanisms to prevent visceral
leishmaniasis epidemics and urban transmission exist, but they require
social and political commitment as well as the allocation of funds to
provide adequate sanitary and satisfactory housing conditions to the
population at risk. Visceral leishmaniasis in the Americas must be
addressed before it becomes a serious public health problem. Even though it
is a zoonosis, control interventions are available that, when properly
used, can eliminate urban transmission and keep disease incidence in rural
areas at a very low level.

Jorge R. Arias, Ph.D.,* Pedro S. Monteiro, and Fabio Zicker, M.D., Ph.D.

*Pan American Health Organization, Brasilia, Brazil; National Visceral
Leishmaniasis Control Program, Ministry of Health, Brasilia, Brazil; Pan
American Health Organization, Washington, D.C.

References

1. Pan American Health Organization/World Health Organization. Regional
   plan for action for combating new, emerging, and reemerging infectious
   diseases. Resolution CD38/17, August 1995. Pan American Health
   Organization, Washington, D.C., USA.

2. World Health Organization. Report of the Second WHO Meeting on Emerging
   Infectious Diseases. Document WHO/CDS/BVI/95.2. Geneva, Switzerland: World
   Health Organization, January 1995.

3. Cerf BJ, Jones TC, Badaro R, Sampaio D, Carvalho EM, Rocha H, Texeira R,
   Johnson Jr WD. Malnutrition as a risk factor for severe visceral
   leishmaniasis. J Infect Dis 1987;156:1030-2.

4. World Health Organization. Control of the leishmaniases: report of a WHO
   expert committee. Technical Report Series 793. Geneva, Switzerland: World
   Health Organization, 1990.

5. Lainson R, Shaw JJ. Epidemiology and ecology of leishmaniasis in Latin
   America. Nature 1978;273 (Parasit Suppl):595-600.

6. Young DG, Arias JR. Flebotomos: vectores de leishmaniasis en Las
   Americas. Pan American Health Organization, 1992: Cuaderno Tcnico No. 33.
   Pan American Health Organization, Washington, D.C., USA.

7. Reed SG, Sheffler WG, Burns Jr JM, Scott JM, Orge MG, Ghalib HW, et al.
   An improved serodiagnostic procedure for visceral leishmaniasis. Am J Trop
   Med Hyg 1990;43:632-9.

8. Wilson ME. Travel and emergence of infectious diseases. Emerging
   Infectious Diseases 1995;1:39-49.

9. Costa CHN, Pereira HF, Arajo MV. Epidemia de leishmaniose visceral no
   estado do Piau, Brasil, 1980-1986. Rev Sade Pbl S Paulo 1990;24:361-72.

10. Ministerio da Sade. Controle, diagnstico e tratamento da leishmaniose
   visceral (calazar). normas tcnicas. Normas Tcnicas. 1a. edio, Brasilia,
   Brazil: Fundao Nacional de Sade, Brasil, 1994.

11. Manson-Bahr PEC, Southgate BA, Harvey AEC. Development of kala-azar in
   man after inoculation with a Leishmania from a Kenya sandfly. Br Med J
   1963;I:1208-10.

12. Kirk R. Studies in leishmaniasis in Anglo-Egyptian Sudan. Trans R Soc
   Trop Med Hyg 1942;35:257-70.

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Pages 147-150]

Commentary

Molecular Epidemiology of Pneumocystis carinii Pneumonia

Pneumocystis carinii pneumonia (PCP) was first recognized as a distinct
clinical entity in orphanages in Europe during World War II (1). Today it
is the most frequent serious opportunistic infection in AIDS patients.
Despite advances in research, numerous questions remain regarding the basic
biology and epidemiology of P. carinii.

Transmission and Patient Care

Although reactivation of latent infections has long been considered the
primary explanation for PCP in immunosuppressed patients, over the years a
steady flow of reports has described clusters of PCP cases (2). In
addition, recent studies have suggested that the duration of latency is
very limited, i.e., usually less than 1 year (3,4). Still other studies
have demonstrated genetic variation in PCR-amplified P. carinii DNA from
the lungs of patients during subsequent PCP episodes (5). Together, these
observations provide strong circumstantial evidence of person-to-person
transmission of P. carinii. Consequently, establishing the role that
person-to-person transmission plays in the epidemiology of PCP is urgent.

Another important area of PCP epidemiology is determining the predisposing
factors for disease. The most frequently discussed predictor of disease is
CD4+ cell count, specifically as it relates to care and management of AIDS
patients (6); however, it has long been known that malnutrition can be an
important contributor (7). The degree to which other factors such as viral
infections or pneumonitis of other causes, may come into play, is yet to be
shown.

Much can also be learned regarding the epidemiology of PCP in HIV-infected
infants. Recent studies report that primary infections in these infants
often develop when the child is 3 to 6 months old (8,9). The source of
these patients P. carinii infections (i.e., the hospital setting, their
mothers, other children, or an environmental source) is not known.

Clinicians working with AIDS patients need a sensitive, reliable, and
noninvasive tool for early detection and diagnosis of PCP infections
(10,11). Besides the standard procedures of bronchoalveolar lavage (BAL)
and induced sputum (IS) sampling, recent studies indicate that it is
possible to amplify P. carinii DNA sequences by polymerase chain reaction
(PCR) directly from blood or serum samples and from nasopharyngeal
aspirates of PCP patients (11,12). Further studies are needed to confirm
that a PCR-based diagnostic tool superior to microscopy can be adapted for
use in clinical settings. A serologic tool that will distinguish recent PCP
infections from those past is also needed.

Prophylaxis failures have been reported for both
trimethoprim-sulfamethoxazole (TMP-SMX) and pentamidine (13-16). Studies
evaluating these cases, however, are frequently complicated by the
difficulties in assessing and confirming patient compliance with the
prophylaxis regimen. The only factor that has a significant correlation
with failure in most cases, however, is the patients CD4+ T lymphocyte
count (14). Although this correlation would be expected because of the
general increased risk for PCP associated with CD4+ cell depletion (6) and
the increase in prophylaxis complications in HIV-infected patients (17),
these drugs may not eliminate all organisms, and some degree of patient
immunity may be required to clear or control the infection. What role, if
any, specific antimicrobial resistance mechanisms play in the reported
treatment failures has not been shown; however, the emergence of resistance
is always a threat. Likewise, long-term TMP-SMX prophylaxis increases the
possibility for the selection of antimicrobial resistance in bacterial
pathogens, some of which are important potential causes of pneumonia in
HIV-infected patients (18). Identifying potential antimicrobial resistance
mechanisms in P. carinii is difficult because of the lack of an established
culture system for human P. carinii that would allow traditional
antimicrobial sensitivity testing.

At least three separate lines of data suggest that P. carinii is a commonly
encounteredorganism: the high seroprevalence rates reported in normal
populations (19), the rapid rate at which infants acquire primary
infections (8) and AIDS patients become reinfected after successful
treatment (20), and the amplification of P. carinii-specific DNA from
ambient air sampled from the environment (e.g., an apple orchard) (21) and
from rooms of animals and patients with PCP (22). Airborne transmission has
been demonstrated for PCP in rats (23-26) and is by far the most likely
mechanism proposed for natural exposure to P. carinii in humans (2,22).
Given the similarities between P. carinii and various fungal agents and the
enigma surrounding the issue of environmental sources for P. carinii, it
has been suggested that P. carinii may in fact be a dimorphic fungus,
ubiquitous in the environment and disseminated by airborne spores (27).
Identifying the specific environmental source or sources of P. carinii is
critical to understanding the epidemiology of PCP and establishing
guidelines for preventing its transmission.

It is generally accepted that P. carinii strains from rats do not infect
humans and that human strains do not infect rats; however, we do not know
the host boundaries for a given P. carinii strain or if all isolates from a
given host display the same degree of host restrictions (28,29). In fact, a
careful evaluation of the available data concerning P. carinii of numerous
hosts suggests that P. carinii may represent a collection of diverse fungal
species (30). Like drug resistance research, studies aimed at
strain/species characterization are generally hindered by the difficulties
in culturing human P. carinii and the lack of refined molecular biological
methods that allow strain identification and characterization.

Molecular Biologic Techniques and Specific Epidemiologic Issues

One of the essential reasons for cultivating any particular pathogen is for
strain identification and characterization that would elucidate such
specific phenotypic characteristics as virulence factors, antimicrobial
sensitivity levels, and factors associated with transmissibility. The
isolation and cultivation of individual strains, and ultimately of clones,
would provide a homogeneous population of organisms from which the desired
information can be obtained and a pure source of genetic material for
constructing DNA libraries and identifying relevant genes.

In the absence of cultivation, investigators have been able to begin
addressing some of the basic epidemiologically important issues by applying
PCR-based technology. In these studies, the DNA sequence of specific
genetic loci from P. carinii is usually amplified from BAL, IS, or serum
samples from PCP patients, using highly specific oligonucleotide primers.
Inherent problems exist in this approach (which are discussed below);
however, the approach has allowed the identification of genotypic
differences in P. carinii populations sampled from the lungs of different
patients and even from the lungs of the same patient during different PCP
episodes. Great potential exists in applying this technology to develop
molecular profiles of P. carinii isolates that could ultimately allow the
particular genotypes to be linked to specific epidemiologically relevant
phenotypes.

Molecular Typing

Five to ten different genetic loci have been identified as potentially
informative for molecular characterization and typing (30-33). Concerning
the typing that has actually been performed on human samples, the primary
loci evaluated include: 1) a 346-bp region of the mitochondrial large
subunit rRNA gene (mt lsrRNA) (10) and 2) a 550-bp fragment containing the
nuclear ribosomal internal transcribed spacer regions 1 and 2 (ITS1 and
ITS2) (34). When these loci are considered collectively, nucleotide
variation can be detected at approximately 37 different positions. Work is
in progress in several laboratories, both to type patient isolates
according to the available loci and to identify additional genetic loci to
more thoroughly define a given genotype.

The primary obstacles to the development of a molecular typing scheme based
on PCR-amplified DNA sequence data obtained from PCP patients include the
following: 1) multiple strains may infect a single patient at a given time;
2) a diploid organism of a single strain may be heterozygous with respect
to a particular polymorphic locus; 3) presumed single genes could have
multiple copies in a single genome, which could give the appearance of
genetic polymorphism; 4) amplified DNA sequence data might be confounded
because of other fungal agents such as Cryptococcus or Candida; and 5)
inferences that can be drawn from restricted sequence data (i.e., gene
typing versus strain typing) are limited. Although these problems are not
insurmountable, they must be considered when evaluating data obtained by
this approach. We propose the following recommendations.

Recommendations

In September 1995, the U.S. Centers for Disease Control and Prevention
hosted a meeting to discuss issues relating to molecular biologic
approaches for studying the epidemiology of P. carinii pneumonia. The
meeting was attended by representatives of research laboratories and by CDC
personnel involved in the epidemiology and molecular biology of P. carinii.
These recommendations regarding research priorities emerged from this
meeting.

Molecular Epidemiology

1. Recent advances in molecular-based typing should be combined with
epidemiologic studies to investigate the transmission of P. carinii and new
strategies for control.

2. Additional genomic regions must be identified for use in typing, along
with the genetic loci that are available. These new loci must be shown to
represent single-copy genes. Also, new molecular approaches should be
developed that will increase the current capacity to resolve genotypic
variation among P. carinii strains.

3. Genetic variation should be investigated among P. carinii strains that
could be linked to variations in factors such as strain virulence, drug
resistance, or transmissibility.

4. The critical issue regarding person-to-person transmission is not so
much whether it occurs, as whether it contributes to infection
significantly more than airborne sources in the environment. Thus, it must
be determined whether there is any benefit to establishing complex
protocols that ensure that patients are carefully protected from each other
if they can become infected from other sources in the environment.
Consequently, the importance of person-to-person transmission in the
epidemiology of PCP should be defined.

5. The role of latent P. carinii infection as a source of PCP in
immunocompromised persons should be clarified.

Diagnosis, Treatment, and Prevention

1. New tools for noninvasive early diagnosis of PCP, including culture
systems, molecular approaches, and serologic tests that can distinguish
recent and past PCP infections are needed.

2. In the United States, clinician compliance with recently published U.S.
Public Health Service/Infectious Diseases Society of America guidelines on
the treatment and prophylaxis of PCP should be evaluated.

3. Studies should be initiated to develop additional drugs for PCP
treatment and prophylaxis.

4. New approaches for improving patient compliance with prescribed PCP
prophylaxis must be devised and evaluated.

5. Methods for detecting the possible emergence of drug resistance to P.
carinii should bestandardized.

6. Standard decontamination procedures for respiratory therapy equipment
and pulmonary diagnostic instruments should be evaluated to confirm that
they effectively eliminate all viable P. carinii.

Environmental Reservoirs and General Biology

1. Environmental sources and the coinciding infective stage(s) of P.
carinii should be detected and evaluated.

2. The host range of P. carinii from various sources (i.e., to what degree
are humans susceptible to P. carinii from nonhuman sources?) should be
determined.

Acknowledgments

We thank the following colleagues for their contributions: C. William
Angus, Marilyn S. Bartlett, Jeanne M. Bertolli, Suzanne Binder, Daniel G.
Colley, Frank H. Collins, Melanie T. Cushion, Alexandre J. DaSilva, Jeffrey
S. Duchin, Brian R. Edlin, Walter T. Hughes, John A. Jernigan, Dennis D.
Juranek, Jonathan R. Kaplan, Scott P. Keely, Chao-Hung Lee, Joseph E.
McDade, Stephen A. Moser, Norman J. Pieniazek, Frank F. Richards, James W.
Smith, Eleanor K. Spicer, James R. Stringer, and Craig M. Wilson. Mary E.
Bartlett provided editorial assistance.

Charles B. Beard, Ph.D., and Thomas R. Navin, M.D.
Centers for Disease Control and Prevention
Atlanta, Georgia, USA

References

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  3. Chen W, Gigliotti F, Harmsen AG. Latency is not an inevitable outcome
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  4. Vargas SL, Hughes WT, Wakefield AE, Oz HS. Limited persistence in and
     subsequent elimination of Pneumocystis carinii from the lungs after P.
     carinii pneumonia. J Infect Dis 1995;172:506-10.
  5. Keely SP, Stringer JR, Baughman RP, Linka MJ, Walzer PD, Smulian AG.
     Genetic variation among Pneumocystis carinii hominis isolates in
     recurrent pneumocystosis. J Infect Dis 1995;172:595-8.
  6. Phair J, Muoz A, Detels R, Kaslow R, Rinaldo C, Saah A. The risk of
     Pneumocystis carinii pneumonia among men infected with human
     immunodeficiency virus type 1. N Engl J Med 1990;322:161-5.
  7. Hughes WT, Price RA, Havron SF, Sisko F, Havron SF, Kafatos AG,
     Schonland M, et al. Protein-calorie malnutrition: a host determinant
     for Pneumocystis carinii infection. Am J Dis Child 1974;128:44-52.
  8. Hughes WT. 1994. Clinical manifestations in children. In: Walzer PD,
     editor. Pneumocystis carinii pneumonia. New York: Marcel Dekker,
     1994:319-29.
  9. Simonds RJ, Lindegren ML, Thomas P, Hanson D, Caldwell B, Scott G, et
     al. Prophylaxis against Pneumocystis carinii pneumonia among children
     with perinatally acquired human immunodeficiency virus infection in
     the United States. N Engl J Med 1995;332:786-90.
 10. Wakefield AE, Pixley FJ, Banerji S, Sinclair K, Miller RF, Moxon ER,
     et al. Detection of Pneumocystis carinii with DNA amplification.
     Lancet 1990; 336:451-3.
 11. Atzori C, Lu J-J, Jiang B, Bartlett MS, Orlando G, Queener SF, Smith
     JW, et al. Diagnosis of Pneumocystis carinii pneumonia in AIDS
     patients by using polymerase chain reactions on serum specimens. J
     Infect Dis 1995;172:1623-6.
 12. Richards CGM, Wakefield AE, Mitchell CD. Detection of pneumocystis DNA
     in nasopharyngeal aspirates of leukaemic infants with pneumonia. Arch
     Dis Child 1994;71:254-5.
 13. Montgomery AB, Feigal DW, Sattler F. Pentamidine aerosol versus
     trimethoprim-sulfamethoxazole for Pneumocystis carinii in acquired
     immune deficiency syndrome. Am J Respir Crit Care Med
     1995;151:1068-74.
 14. Saah AJ, Hoover DR, Peng Y, Phair JP, Visscher B, Kingsley LA, et al.
     Predictors for failure of Pneumocystis carinii pneumonia prophylaxis.
     JAMA 1995;273:1197-1202.
 15. Lecuit M, Livartowski J, Vons C, Goujard C, Lamaigre G, Delfraissy
     J-F, et al. Resistance to trimethoprim-sulfamethoxazole and
     sensitivity to pentamidine therapy in an AIDS patient with
     hepatosplenic pneumocystosis. AIDS 1994;8:1506-7.
 16. Torres RA, Barr M, Thorn M, Gregory G, Keily S, Chanin E, et al.
     Randomized trial of dapsone and aerosolized pentamidine for the
     prophylaxis of Pneumocystis carinii pneumonia and toxoplasmic
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 18. Schwartz RH, Khan WN, Akram S. Penicillin and
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     1991;10:782-3.
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 20. Dohn MN, Frame PT. Clinical manifestations in adults. In: Walzer PD,
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 21. Wakefield AE. Detection of DNA sequences identical to Pneumocystis
     carinii in samples of ambient air. J Euk Microbiol 1994;41:116S.
 22. .Bartlett MS, Lee C-H, Lu J-J, Bauer NL, Betts JF, McLaughlin GL, et
     al. Pneumocystis carinii detected in air. J Euk Microbiol 1994;41:75S.
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     infection with Pneumocystis. Proc Soc Exp Biol Med 1971;137:1401-4.
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     experimental model for Pneumocystis carinii infection. Science
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     et al. Is Pneumocystis carinii a deep mycosis-like agent? Eur J
     Epidemiol 1992; 8:460-70.
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     Lab Med 1991;11:957-75.
 29. Armstrong MYK, Cushion MT. In vitro cultivation. In: Walzer PD,
     editor. Pneumocystis carinii pneumonia. New York: Marcel Dekker,
     1994:3-24.
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     protozoan, but a diverse group of exotic fungi. Infect Agents Dis
     1993;2:109-17.
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     Walzer PD, editor, Pneumocystis carinii Pneumonia. New York: Marcel
     Dekker, Inc., 1994:91-105.
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     carinii. J Euk Microbiol 1994;41:121S-22S.
 33. Lu J-J, Chen C-H, Bartlett MS, Smith JW, Lee C-H. Comparison of six
     different PCR methods for detection of Pneumocystis carinii. J Clin
     Microbiol 1995;33:2785-8.
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     al. Typing of Pneumocystis carinii strains that infect humans based on
     nucleotide sequence variations of internal transcribed spacers of rRNA
     genes. J Clin Microbiol 1994;32:2904-12.

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Page 151]

Commentary

Needed: Comprehensive Response to the Spread of Infectious Diseases

In his article Globalization, International Law, and Emerging Infectious
Diseases, Fidler recognizes that biological agents travel by themselves or
with their hosts without any recognition of, or regard for, political
borders. He notes that with the continued expansion of economic commerce
across continents and more rapid transport and travel, persons infected
with infectious diseases of very short incubation periods can act as
vectors across several nations before they even become symptomatic. The
protective effect of clipper ship travel is long gone.

Fidler examines the need for international treaties, agreements, and
policies to manage the spread of new or reemerging infections diseases. His
concern is that the current international climate requires more enforceable
treaties with adequate resources to identify, track, interfere with, and
contain the spread of infectious diseases perceived as an international or
global threat.

International cooperation within the existing legislative mechanisms has,
on occasion, been very successful. International eradication of smallpox
was successful because a specific, cost-effective, efficient vaccine was
developed; the disease attacked persons regardless of their economic,
political, racial, religious, or social affiliations; the amount of funding
was adequate; and all nations recognized the benefits of the eradication
program. A similar effort currently in progress to eradicate poliomyelitis
will also be successful because of international cooperation.

In contrast, international control of other infectious diseases, such as
malaria and tuberculosis, has been attempted for decades with considerably
less success. Notwithstanding the lack of efficacious vaccines, the reality
is that only very limited resources are being committed to prevent and
treat all infectious diseases. Outbreaks of Ebola virus infection and
plague are routinely reported in the local, national, and international
press. However, the continued increased incidence and prevalence of
tuberculosis, AIDS, and other sexually transmitted diseases are accepted by
many as problems of the poor, the immoral, and the expendable portion of
society. Local, national, and international awareness and continued
interest are significant problems.

International cooperation must extend beyond merely restricting the natural
spread of specific diseases. One also has to recognize the need for
effective international treaties to prevent the use of biological agents in
either tactical or strategic circumstances. Fear of combatants using
biological agents on military and civilian targets intensified during and
since the Gulf War. The possibility of biological terrorism is no longer
limited to the imagination of fiction writers. Fidler does not stress the
issue of nonnatural outbreaks of diseases; a global need for an improved
non-ad hoc response to emerging infectious disease agents should be
completely considered by civilian and military planners. The threat of
infectious diseases as weapons provides an additional incentive for
cooperation among governments.

Harold M. Ginzburg, M.D., J.D., M.P.H.
Department of Psychiatry and Neurology, Tulane University Medical Center,
New Orleans, Louisiana, USA

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Page 152]

Letters

Transfusion-Associated Malaria

To the Editor: A recent article by Zucker (1) described two cases of
malaria that were probably transfusion associated. A case of
transfusion-associated malaria in which the source was identified was
reported in San Francisco in 1991. The case was in an elderly man in whom
malaria infection developed after coronary bypass surgery.

The patient was born in China and immigrated to the United States in 1940.
His only travel outside the United States was a trip to Hong Kong in 1951
for 6 months. The patients wife was born in China and had malaria in 1941
during World War II. She received no treatment at that time or at any other
time. She came to the United States in 1960 and has not left the country
since.

The patient had six donors, five of whom had no history of malaria, and had
negative serologic test results for all four malaria species. Both the
patient and his wife had blood smears positive for P. malariae. The
patients wife had positive serologic test results for P. vivax and P.
ovale (1:64), for P. falciparum (1:258), and for P. malariae (1:1024).

Frances Taylor, M.D., M.P.H.
Director of Communicable Disease Control
City and County of San Francisco
Department of Public Health
Bureau of Epidemiology
Disease Control, and AIDS
San Francisco, California, USA

Reference

1. Zucker J. Changing patterns of autochthonous malaria transmission in the
United States: A review of current outbreaks. Emerging Infectious Diseases
1996: 2:37-43.

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[Page 152]

Letters

Reply to F. Taylor:

Dr. Taylors letter calls attention to the small but important number of
induced malaria cases that occur in the United States. From 1957 to 1994,
101 such cases were reported to the Centers for Disease Control and
Prevention (CDC); these (including the 1990 case described by Dr. Taylor
[1]) are reviewed annually and reported by CDC (2). The occasional
occurrence of induced malaria further emphasizes the importance of
including malaria in the differential diagnosis of fevers of unknown
origin, even in patients who have not traveled to countries where malaria
is endemic. Preventing induced malaria requires screening potential blood,
tissue, and organ donors and deferring those with a history of malaria or
travel to malarious areas. Furthermore, timely surveillance must be
maintained to detect induced cases promptly, identify infected blood
donors, and prevent additional cases.

The case described by Dr. Taylor was not included in Changing patterns of
autochthonous malaria transmission in the United States: a review of recent
outbreaks (3) because it was a case of induced rather than autochthonous
malaria. Each reported malaria case is classified according to standardized
terminology (4). Imported malaria (which accounts for most cases in this
country) is acquired outside the United States and its territories. Malaria
acquired within the United States is rare and occurs by one of three
mechanisms: Autochthonous malaria is acquired through the bite of an
infective mosquito. Congenital malaria is acquired when a child is infected
in utero. Induced malaria is transmitted by mechanical means such as
transfusion of blood or blood products, organ transplant, deliberate
infection for malariotherapy, or contaminated needles or injection
equipment. Congenital and induced cases were not included in this review.

When an investigation fails to identify the source of transmission and a
case cannot be epidemiologically linked to another case of malaria, the
case is classified as cryptic. Most cryptic cases are believed to be
autochthonous, and there is often evidence to suggest mosquito-borne
transmission, even when the source of infection remains unidentified. For
this reason, most cryptic cases were included in this review of
autochthonous malaria. The two exceptions noted in the article were
excluded because both patients had recent histories of blood transfusion,
suggesting that their infections were induced.

Jane R. Zucker, M.D., M.Sc., and S. Patrick Kachur, M.D., M.P.H.
Centers for Disease Control and Prevention, Atlanta, Georgia, USA

References

1. Zucker JR, Barber AM, Paxton LA, Schultz LJ, Lobel HO, Roberts JM, et
   al. Malaria SurveillanceUnited States, 1992. In: CDC Surveillance
   Summaries, October 20, 1995. MMWR 1995:44(SS-5):1-17.

2. Centers for Disease Control. Malaria Surveillance Annual Summary, 1990.
   Atlanta: Centers for Disease Control, 1991.

3. Zucker JR. Changing patterns of autochthonous malaria transmission in
   the United States: a review of recent outbreaks. Emerging Infectious
   Diseases 1996;2:37-43.

4. World Health Organization. Terminology of malaria and malaria
   eradication. Geneva: World Health Organization, 1963:32.

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[Pages 153-154]

Letters 

An Outbreak of Hemolytic Uremic Syndrome due to Escherichia coli O157:H-:
Or Was It?

To the Editor: Since the first reported outbreaks of hemolytic uremic
syndrome (HUS) and related conditions more than 10 years ago (1), outbreaks
of HUS due to Escherichia coli O157 have been reported from many parts of
the world, particularly North America and Europe. While most of these
reports have incriminated the motile strains of serotype O157:H7, nonmotile
serotypes (e.g., O157:H-) have also been associated with HUS; these two
serotypes are most commonly associated with both outbreaks and sporadic
cases of HUS and related conditions. Over the last decade, a number of
techniques for the rapid identification of these organisms have been
developed. Of these, the use of sorbitol-MacConkey agar (2) has perhaps
been the most valuable. This technique is based on the fact that these
organisms rarely ferment sorbitol on primary isolation, while most other E.
coli usually ferment this substrate. We believe that outbreaks due to other
enterohemorrhagic E. coli may have been attributed to serogroup O157
because of the limited technology used in investigating these outbreaks.

No outbreaks of HUS due to serogroup O157 have occurred in Australia
despite sporadic cases of HUS caused by such strains. Other serogroups
(particularly serotype O111:H-) have been associated with most cases of HUS
and related conditions in Australia (3). No outbreak of HUS had been
reported in Australia until January 1995, when an outbreak associated with
the consumption of contaminated mettwurst (fermented sausage) was reported
from South Australia (4). Twenty-three children with HUS were hospitalized.
Most required hemodialysis; one died. Verocytotoxigenic strains of E. coli
O111 producing Shiga-like toxin (SLT) I and II were isolated from 19
patients and from samples of mettwurst. In addition, strains of E. coli
O157:H- that produced SLT-I and SLT-II were isolated from three of the
patients and the mettwurst. These strains did not ferment sorbitol on the
sorbitol-MacConkey agar, which facilitated their isolation. The predominant
O111 strains were sorbitol-positive, unlike the O111 strains, recently
described as being sorbitol-negative (5). Symptoms of the patients from
whom theO157:H- strains were isolated, in addition to E. coli O111:H-, were
not significantly different from those of the patients whose specimens
yielded only E. coli O111:H-. In addition to O111 (and O157), other
serotypes of enterohemorrhagic E. coli, including strains of serogroup O23,
O26, and O91, were isolated from the patients. However, antibodies to O111
were detected in nearly all patients, which indicates the serogroups
leading role in the outbreak. The isolation of serogroup O157 is
comparatively easy; therefore, it is less likely that these strains would
have been missed, than it is that O111 and other serotypes would have been.
Even though a negative finding can never be considered conclusive, we
consider the inability to isolate serogroup O157 more conclusive than the
same result for other serotypes. It has frequently been suggested that the
O157 serogroup is cleared from the patient relatively rapidly, which makes
its isolation difficult or impossible. We found a similar situation with
other enterohemorrhagic E. coli serotypes. The fact that most patients
elicited an O111 antibody response (and no anti-O157) almost certainly
proves this serotypes causal role in this outbreak.

The laboratory in South Australia was particularly well disposed to deal
with such an outbreak because some of its ongoing research programs
included studies on aspects of enterohemorrhagic E. coli and related
organisms. The most sophisticated molecular biologic techniques were
immediately available to investigate the outbreak accurately and confirm
epidemiologic leads regarding a common source. Polymerase chain reaction
(PCR) played a major role not only in identifying SLT-I, SLT-II, and SLT-I
and SLT-II producing bacteria in the stool of patients, but also in
identifying the suspected source (mettwurst). In addition, PCR, utilizing
sequences specific for the O111 serogroup, enabled this serogroup to be
rapidly identified in patients feces samples and suspected source
material. Without this technology, the outbreak would not have been
contained so rapidly. On the other hand, if the laboratory had to rely on
conventional microbiologic culture procedures, including sorbitol-MacConkey
agar, strains of serogroup O157 would have been identified from three
patients, as well as from the epidemiologically incriminated mettwurst. The
laboratory would not have found the O111 strains because they all fermented
sorbitol readily and would have been discarded as normal flora as would the
other enterohemorrhagic E. coli serotypes. The outbreak would have been
reported as another O157 outbreak, from which only about 15% of the
patients yielded the incriminating strains. This outbreak could be
recognized as one caused by a number of different enterohemorrhagic E. coli
serotypes, of which serotypes O111:H- and O157:H- were the most prominent.
Other serotypes, however, such as O23, O26, and O91, were also present.
With the widespread nature of verocytotoxigenic strain of different
serotypes as has been reported from many environmental studies, it is not
surprising that a product, such as mettwurst, which is made from meats from
various sources, would contain a number of these potential pathogens.

A large number of E. coli serotypes can be verocytotoxigenic and, in a few
cases, outbreaks due to such strains have been reported. Most notable have
been reports from Italy of outbreaks due to enterohemorrhagic E. coli O111
(6); however, the impression is that these are the exception and that the
most prominent serotype is O157:H7. Some of the reported outbreaks due to
O157 strains may in fact have been due to other serotypes and the O157
strains were only present in comparatively small numbers; however, because
of the ease with which these strains can be identified using
sorbitol-MacConkey agar, they were believed responsible for the outbreaks.
For example, in Argentina, E. coli O157:H7 was found in only one (2%) of 51
children with HUS (7) and in the Netherlands, only 5 (19%) of 26 HUS
patients yielded E. coli O157:H7 (8). In a 10-year, retrospective,
population-based study of HUS, this serotype was isolated in 13 (46%) of 28
patients (9), and in their review, Su and Brandt (10) put an overall figure
of 46% to58% as the incidence range of E. coli O157:H7 infection in cases
of HUS. Finding SLT sequences in a fecal specimen by PCR, or free fecal
toxins in many patients of an outbreak while isolating strains of O157 from
only a few, does not exclude the presence of other serotypes, but culture
methods now available would rarely pick these up. Thus there is ample room
to speculate that approximately half the cases of HUS may be caused by
serogroups other than O157 and, by inference, at least half the outbreaks
may be wrongly attributed to this serogroup. We recognize that
enterohemorrhagic E. coli O157 have become extraordinarily widespread
throughout the world since their first description (1); this does not mean
that other serotypes are not also causing infections, either alone, in
conjunction with O157, or even with other known or unknown enteric
infections. It is important to be aware of the existence of these other
serotypes and be vigilant for them. The isolation and characterization of
strains of serogroup O157 from patients with HUS is certainly noteworthy,
but so is the finding of O111 or any other serogroup. Serogroup O111 has
amply demonstrated the ability to cause extensive outbreaks (6). Even
though many laboratories are becoming aware of the importance of testing
for serogroup O157:H7, we think that testing for this serotype only is a
disservice; simple culture techniques can identify this serogroup, but
always at the risk of missing other serogroups. The development of simple
methods to detect all enterohemorrhagic E. coli is now required.

P.N. Goldwater, F.R.A.C.P., F.R.C.P.A.,* and K.A. Bettelheim, Ph.D.
*Womens and Childrens Hospital, Adelaide, South Australia; Biomedical
Reference Laboratory, Victorian Infectious Diseases Reference Laboratory,
Fairfield Hospital, Victoria, Australia

References

  1. Riley LW, Remis RS, Helgerson SD, McGee HB, et al. Hemorrhagic colitis
     associated with a rare Escherichia coli serotype. N Engl J Med
     1983;308:681-5.

  2. March SB, Ratnam S. Sorbitol-MacConkey medium for detection of
     Escherichia coli O157.H7 associated with hemorrhagic colitis. J Clin
     Microbiol 1986;23:869-72.

  3. Goldwater PN, Bettelheim KA. The role of enterohemorrhagic Escherichia
     coli serotypes other than O157:H7 as causes of disease. Communicable
     Disease Intelligence 1995;19:2-4.

  4. Cameron S, Walker C, Beers M, Rose N, and Anear E, et al.
     Enterohemorrhagic Escherichia coli outbreak in South Australia
     associated with consumption of mettwurst. Communicable Disease
     Intelligence 1995;19:70-1.

  5. Ojeda A, Prado, V, Martinez, J, et al. Sorbitol-negative phenotype
     among enterohemorrhagic Escherichia coli strains of different
     serotypes and from different sources. J Clin Microbiol
     1995;33:2199-201.

  6. Caprioli A, Luzzi I, Rosmini F, Resti C, et al. Communitywide outbreak
     of hemolytic-uremic syndrome associated with non-O157
     verocytotoxin-producing Escherichia coli. J Infect Dis
     1994;169:208-11.

  7. Lopez EL, Diaz M, Grinstein S, Devoto S, et al. Hemolytic uremic
     syndrome and diarrhea in Argentine children: the role of Shiga-like
     toxins. J Infect Dis 1989;160:469-75.

  8. van der Kar NCAJ, Roelofs HGR, Muytjens HL, et al.
     Verocytotoxin-producing Escherichia coli infection in patients with
     hemolytic uremic syndrome and their family members in the Netherlands.
     In: Kaarmali MA, Goglio AG, editors. Recent advances in
     verocytotoxinproducing Escherichia coli infections. Amsterdam:
     Elsevier, 1994.

  9. Martin DL, MacDonald KL, White KE, Soler JT, Osterholm MT. The
     epidemiology and clinical aspects of the hemolytic uremic syndrome in
     Minnesota. N Engl J Med 1990;323:1161-7.

 10. 10. Su C, Brandt LJ. Escherichia coli O157:H7 infections in humans.
     Ann Intern Med 1995;123:698-714.


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[Page 155]

Letters

The Dilemma of Xenotransplantation

To the Editor:

I read with considerable interest Robert E. Michlers commentary on
xenotransplantation (1).

From my point of view, that of a basic virologist, the dilemma is not to
know in what foreseeable future, clinical xenotransplantation may achieve
its targeted goal of extended graft survival, but what deadly emerging
infectious disease, most probably viral in nature, would arise in a
recipient of a baboon or chimpanzee heart. While we face the terrific
threat of AIDS, which clearly emerged from Africa and non-human primates 40
to 50 years ago, we are preparing a new infectious Chernobyl.

Monkeys and apes harbor approximately 50 simian viruses; some of them pose
a serious threat to humans, especially the heavily immunosuppressed.
Recently, an outbreak of encephalitis related to a new type of reovirus (2)
occurred among baboons from a colony used in human organ transplants.
Moreover, once unknown or unrecognized simian viruses, like HIV, may be
efficient invaders of the entire earths population.

Xenotransplantation does not simply pose an ethical problem; it concerns
the survival of the human species, an endangered species if transplant
practitioners continue their course. Ronald Montalero, a virologist, was
right when he said unknown viruses were always a major concern in
xenotransplants (2). A moratorium on these procedures seems the best
solution until all simian pathogens are identified and the risks they pose
to humans are clearly established.

Claude E. Chastel
Virus Laboratory, Faculty of Medicine
29285 Brest Cedex, France

References

1. Michler RE. Xenotransplantation: risks, clinical potential, and future
prospects. Emerging Infectious Diseases 1996; 2; 64-70.

2. Mystery virus fells donor baboons. Science 1994; 264; 1523.



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[Pages 155-156]

Letters

The Thucydides Syndrome: Ebola Dj Vu? (or Ebola Reemergent?)

To the Editor:

The plague of Athens (430-427/425 B.C.) persists as one of the great
medical mysteries of antiquity (1-5). Sometimes termed the Thucydides
syndrome for the evocative narrative provided by that contemporary
observer (6, 7), the plague of Athens has been the subject of conjecture
for centuries. In an unprecedented, devastating 3-year appearance, the
disease marked the end of the Age of Pericles in Athens and, as much as the
war with Sparta, it may have hastened the end of the Golden Age of Greece
(3). Understood by Thucydides to have its origin in Ethiopia beyond Egypt,
it next descended into Egypt and Libya and then suddenly fell upon
Athens walled port Piraeus and then the city itself; there it ravaged the
densely packed wartime populace of citizens, allies, and refugees.
Thucydides, himself a surviving victim, notes that the year had been
especially free of disease and describes the following major findings:
After its abrupt onset, persons in good health were seized first with
strong fevers, redness and burning of the eyes, and the inside of the
mouth, both the throat and tongue, immediatelywas bloody-looking and
expelled an unusually foul breath. Following these came sneezing,
hoarseness . . . a powerful cough . . . and every kind of bilious vomiting
. . . and in most cases an empty heaving ensued that produced a strong
spasm that ended quickly or lasted quite a while. The flesh, although
neither especially hot nor pale, was reddish, livid, and budding out in
small blisters and ulcers. Subject to unquenchable thirst, victims
suffered such high temperatures as to reject even the lightest coverings.
Most perished on the ninth or seventh day . . . with some strength still
left or many later died of weakness once the sickness passed down into the
bowels, where the ulceration became violent and extreme diarrhea
simultaneously laid hold (2.49). Those who survived became immune, but
those who vainly attended or even visited the sick fell victim (2.51).

By comparison, a modern case definition of Ebola virus infection notes
sudden onset, fever, headache, and pharyngitis, followed by cough,
vomiting, diarrhea, maculopapular rash, and hemorrhagic diathesis, with a
case-fatality rate of 50% to 90%, death typically occurring in the second
week of the disease. Disease among health-care providers and care givers
has been a prominent feature (8, 9). In a review of the 1995 Ebola outbreak
in Zaire, the Centers for Disease Control and Prevention reports that the
most frequent initial symptoms were fever (94%), diarrhea (80%), and severe
weakness (74%), with dysphagia and clinical signs of bleeding also
frequently present. Symptomatic hiccups was also reported in 15% of
patients (10).

During the plague of Athens, Thucydides may have made the same unusual
clinical observation. The phrase lugx kene, which we have translated as
empty heaving, lacks an exact parallel in the ancient Greek corpus (5).
Alone, lugx, means either hiccups or retching and is infrequently used,
even by the medical writers. Although contexts usually dictate retching,
we note unambiguous hiccups in Platos Symposium (185C). In his thorough
commentary on the Thucydides passage, the classicist D. L. Page remarks:
Hiccoughs is misleading, unless it is enlarged to include retching.
Regarding empty, unproductive retching [he] has noted no exact parallel
 . . . in the [writings of the] doctors, but . . . tenesmus comes very close
to it (5). A CD-ROM search of Mandell, Bennett, and Dolin discloses no
reference to either hiccups or singultus in the description of any
disease entity (6).

The profile of the ancient disease is remarkably similar to that of the
recent outbreaks in Sudan and Zaire and offers another solution to
Thucydides ancient puzzle. A Nilotic source for a pathogen in the Piraeus,
the busy maritime hub of the Delian League (Athens de facto Aegean
empire), is clearly plausible. PCR examination of contemporaneous skeletal
and archaeozoological remains might test this hypothesis against the 29 or
more prior theories.

*P. E. Olson, M.D., *C. S. Hames, M.D., A. S. Benenson, M.D., and E. N.
Genovese, Ph.D.
U. S. Navy Balboa Hospital, San Diego, California, USA; San Diego State
University, San Diego, California, USA

References

  1. Langmuir AD, Worthen TD, Solomon J, Ray CG, Petersen E. The Thucydides
     syndrome: a new hypothesis for the cause of the plague of Athens. N
     Engl J Med 1985;313:1027-30.

  2. Morens DM, Littman RJ. Epidemiology of the plague of Athens. Trans Am
     Philological Assn 1972;122:271-304.

  3. Morens DM, Littman RJ. The Thucydides syndrome reconsidered: new
     thoughts on the plague of Athens. Am J Epidemiol 1994;140:621-7.

  4. Grmek MD. History of AIDS: emergence and origin of a modern
     pandemic.Princeton, NJ: Princeton University Press, 1990.

  5. Page DL. Thucydides description of the great plague. Classical Quart
     1953;47 n.s. 3:97-119.

  6. Thucydides. Peloponnesian War. Bk. 2, chs. 47-52.

  7. Major RH. Classical descriptions of disease. 3rd ed. Springfield, IL:
     Charles C Thomas, 1945.

  8. Benenson AS, editor. Control of communicable diseases manual. 16th ed.
     Washington, DC: American Public Health Association, 1995.

  9. Mandell GL, Bennett JE, Dolin R. Mandell, Douglas and Bennetts
     principles and practice of infectious diseases. 4th ed. New York:
     Churchill Livingston, 1995.

 10. Centers for Disease Control and Prevention. MMWR 1995;44:25:468-75.

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Emerging Infectious Diseases
Volume 2, Number 2, April-June 1996

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[Page 157]

News and Notes

BSE Meeting at CDC

The recent report of a new variant of Creutzfeldt-Jakob disease (V-CJD) in
Great Britain and the possible link between the disease and bovine
spongiform encephalopathy (BSE) has raised a number of health and safety
concerns (1,2). On April 8, 1996, CDC organized a meeting of U.S. agency
representatives to review information about the report of U.K. cases and
about efforts to identify the existence of BSE and V-CJD in the United
States. The meeting covered the scientific evidence for the report of
V-CJD; recommendations from a meeting of international experts organized by
the World Health Organization on April 2-3; and the current and proposed
activities of U.S. agencies with regard to BSE and V-CJD.

Among the observations made during the meeting were the following:

   * There is no evidence from U.S. surveillance activities or from
     scientific studies to indicate that BSE exists in the United States.

   * Active surveillance for BSE is conducted by the U.S. Department of
     Agriculture (USDA). All cattle presented for slaughter in the United
     States are observed for signs of central nervous system (CNS)
     disorders. Livestock showing CNS signs are condemned and not allowed
     to enter the slaughter plant or to become part of the human food
     supply. Since 1990, laboratory testing of nearly 2,800 brain specimens
     from cattle with CNS signs has shown no evidence of BSE.

   * No U.K. cattle or ruminant-based feed has been imported into the
     United States since July 1989, when USDA banned the importation of
     cattle and cattle products from the United Kingdom. U.K. cattle
     imported before the ban will be destroyed as a precaution to ensure
     that these animals do not enter the food chain (human or animal).

   * The Food and Drug Administration plans to issue a ban on
     ruminant-to-ruminant feeding in the United States.

   * Additional research is needed on the characterization of the causative
     agent of BSE and on the epidemiology, rapid laboratory diagnosis, and
     pathogenesis of BSE and CJD.

   * The Centers for Disease Control and Prevention (CDC) monitors the
     occurrence of CJD in the United States through surveillance and
     special epidemiologic studies (3). On the basis of mortality
     surveillance from 1979 to 1993, the annual incidence of CJD remained
     stable at approximately one case per million persons. In the United
     Kingdom, five of eight patients who died of V-CJD since May 1995 were
     younger than 30 years of age; by comparison, in the United States, CJD
     deaths among persons younger than 30 years are extremely rare (fewer
     than 5 per billion per year). CDCs efforts will be expanded to
     include active surveillance studies at four Emerging Infections
     Program sites (Connecticut, Minnesota, Oregon, and the San Francisco
     area) and in Atlanta to provide more up-to-date information on the
     occurrence of CJD and to verify the absence or presence of V-CJD.

     Future cooperative efforts among U.S. agencies, industry, and other
     interested parties in response to the report of V-CJD are planned. The
     report of the April 8 meeting at CDC can be accessed on the CDC NCID
     Web site (connect to http://www.cdc. gov/ncidod/ncid.htm; the report
     is under New, Reemerging, and Drug-Resistant Infections).

     References
       1. Will RG, Ironside JW, Zeibler M, et al. A new variant of
          Creutzfeldt-Jakob disease in the UK. Lancet 1996;347:921-5.
       2. CDC. World Health Organization consultation on public health
          issues related to bovine spongiform encephalopathy and the
          emergence of a new variant of Creutzfeldt-Jakob disease.MMWR
          1996;45:295-6, 303.
       3. Holman RC, Khan AS, Kent J, Strine TW, Schonberger LB.
          Epidemiology of Creutzfeldt-Jakob disease in the United States,
          1979-1990: analysis of national mortality data. Neuroepidemiology
          1995;14:174-81.

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[Page 157]

News and Notes

CDC Foundation Supports Emerging Infectious Disease Projects

The National Foundation for the Centers for Disease Control and Prevention,
Inc. (NFCDC), a not-for-profit corporation established by Congress to
support CDCs mission, announced in August 1995 that one of its initial
funding efforts would be in the area of antibiotic-resistant diseases.

Also in the area of infectious diseases, NFCDC has recently received a gift
from a Pennsylvania foundation to establish fellowships for two
representatives from nongovernmental organizations to spend 1 month at CDC
learning about the agencys AIDS/HIV support services and activities and
advising CDC on the type of support nongovernmental agencies need in this
area.

The foundation creates new and enhanced financial and program partnerships
between CDC and American and international businesses and industries,
philanthropic organizations, foreign governments, international
organizations, and concerned individuals. An initial grant of $1 million
from the Robert W. Woodruff Foundation was awarded to help establish NFCDC
in Atlanta.

The NFCDC board of directors plans to first fund projects that reflect the
following themes: 1) strengthening global health capacity; 2) promoting
healthy lifestyle choices with initial emphasis on adolescent health; 3)
improving the quality and communication of public health information, with
initial emphasis on development of information technology and networks; and
4) promoting prevention in health care delivery systems, in particular,
managed-care systems.

According to foundation executive director Charlie Stokes, tax-deductible
gifts to NFCDC represent an investment in CDCs efforts to address current
or emerging health problems that threaten U.S. citizens and citizens of the
world. Gifts may be designated for a specific purpose or be given without
restriction to be used for CDCs greatest needs. Federal employees can make
gifts to the foundation through the annual Combined Federal Campaign.

For further information about NFCDC, contact

Charlie Stokes, Executive Director
NFCDC
The Hurt Building
50 Hurt Plaza, Suite 765
Atlanta, GA 30305
phone: 404-653-0790; fax: 404-653-0330
e-mail: seg4@nfcdc.em.cdc.gov

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