NationalLibraryOfScotland/medical-history-of-british-india huggingface dataset:
RESEARCH WORK ON FOOT-AND-MOUTH DISEASE
199
He has found that the best conditions for survival of the virus in a given
medium at 37°C. vary within narrow limits, and the most favourable degree of
acidity in a strongly reducing medium may be more acid than that in one which
provides less reducing conditions. In simple solutions of phosphates the
reducing conditions in an anaerobic chamber in the absence of oxygen were found
to be distinctly more hostile to the virus than when it was exposed to the air,
if the liquid medium was kept at the degree of alkalinity usually considered most
favourable to survival. At a lower level of alkalinity, however, survival was
more prolonged in the absence of oxygen than when it was present. A slight increase
in the oxidising properties of the medium in the presence of air was also harmful.
On the other hand, bran infusion, which prolongs life well in the presence of air,
is a bad medium for survival when the oxygen is removed, apparently because
the reducing properties of the bran are then excessive.
Dr. Edwards has compared by the use of dyes the oxidation-reduction poten-
tial of liquids made by mixing different substances whose effect on survival has
been found experimentally. These mixtures were also tested by means of dyes
which indicates the degree of acidity or alkalinity.
The properties of some of these mixtures changed quickly on exposure to air
and in many instances it was, therefore, necessary to keep the test-tubes
in an anaerobic jar with reduced oxygen pressure and to secure sufficiently
oxidising conditions by means of additional ingredients. Another expedient was
the addition of weak agar jelly and a layer of soft paraffin in the upper part of the
test-tube to hinder the penetration of oxygen from the air. As a result of his
work he has found that certain substances, such as extracts of hay or bran, which
were known to favour survival from previous work done for the Committee, and
certain mixtures which he had determined experimentally to have a like effect in
promoting survival, had very similar qualities when the oxidation-reduction and
the alkalinity or acidity were considered together. He also found that when the
indicator-dyes were injected intradermally into guinea-pigs' pads either alone or
mixed with virus, as in an ordinary inoculation experiment the state in the skin
as regards the above-mentioned properties was comparable to that present in some
of the media in which the survival results were the best.
These experiments tend to confirm the hypothesis that one of the most im-
portant desiderata for survival of foot-and-mouth virus in vitro is the maintenance
of suitable oxidation-reduction and acidity alkalinity conditions. Although no
definite evidence of multiplication of the virus in vitro in simple media under these
conditions has so far been obtained, it is hoped that a step in this direction has
been made.
200 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [ II, II
The success which has attended attempts at propagation in vitro in cultures
of living tissues of foot-and-mouth disease and some other viruses, e.g., vaccinia,
suggests that the conditions inside the living animal cell must be imitated in order
to obtain growth in artificial media. The maintenance of a definite steady oxida-
tion-reduction potential is among the conditions known to be characteristic of the
living cell.
D.—SURVIVAL IN SEWAGE AND DISINFECTION BY AMMONIACAL LIQUIDS.
It has been shown by Wagener at Riems that although virus contained in
pieces of skin will survive for a long time, even over 100 days in winter, when kept
in a sewage tank at ordinary temperatures, if the contents of the tank have not
become very acid or alkaline, still the virus is rapidly destroyed in closed tanks
into which the cowsheds drain and which contain the urine and washings from the
stalls. The reason for this is the high content of ammonia and the consequent
alkalinity of the liquid. If the effluent from the stalls is freely exposed to the air
the virus is not so rapidly destroyed since the ammonia escapes as it is formed.
E.—SURVIVAL OF VIRUS OUTSIDE THE ANIMAL BODY.
Continuing the observations described in previous reports, a series of experi-
ments has been made by Dr. Andrews and his colleagues with a view to testing
the continuance of activity of virus under conditions likely to be met with in
practice on farms or in the meat trade. They have tested the times of survival of
virus in small pieces of skin from vesicles when immersed in liquids of weak but
varying alkalinity and have found the virus still active after 71 to 105 days at a
temperature of—1°C. (29°F.), if the alkalinity approached that of the blood in the
living body. They also examined blood from infected animals which had been
allowed to dry at ordinary temperatures, 59° to 68°F. (15° to 20°C. on a variety
of substances. The virus only survived 2 or 3 days in blood dried on glass, iron,
zinc, tile, brick or wood, but on hide, leather from a boot or rubber from a gum-
boot, its life was materially prolonged, up to 80 to 102 days respectively. On the
last three materials the survival was exceptional, though not so long as had been
found in earlier work when watery filtrates of virus were allowed to dry on bran,
in which case the duration of activity was 20 weeks. On hessian or stockinette,
the materials used for wrapping chilled and frozen carcases, blood containing virus
was found infective after 45 days at 56° to 87°F (13.4° to 30.4°C.) or 28.4° to
35.6°F. (—2° to + 2°C.).
Milk. The use of infected milk is believed to be a means of spreading
disease on the Continent since, if it is derived from infected cows and is not heated,
RESEARCH WORK ON FOOT-AND-MOUTH DISEASE
201
it may infect calves or pigs which are fed on it. It is well known that the milk
given by cows during the acute stage of the disease is infective. Mr. Galloway has
examined in detail how long milk to which virus has been added will retain its
activity under different conditions.
The virus is soon killed when the milk becomes sour, and how soon this
happens varies with the bacterial content at the beginning of the experiment and
the temperature at which the milk is kept.
Virus in milk kept at 18°C. (64.4°F.) remained active for 7 to 9 days.
Virus in milk kept at 4°C. (39°F.) remained active for 15 to 17 days.
Virus added to sterilised milk kept at 18°C. (64.4°F.) was active after 30
days.
Virus added to sterilised milk kept at 4°C, (39°F.) was active for at least 47
days.
Milk contaminated with virus and allowed to dry on pieces of wood was inac-
tive after 2 days at 18°C.
F.—DRIED MILK.
Milk powder is usually made by drying on a heated roller or by spraying
milk into a dry and heated chamber. These commercial processes could not be
imitated exactly with naturally infected milk, but attempts were made to imitate
the conditions. Milk to which virus had been added when dried rapidly and kept
dry was active after 32 to 43 days. It therefore seemed probable that the virus
would not be killed by the spraying method of drying but that drying on hot rollers
was much more likely to inactivate it.
G.—DISINFECTION.
A series of experiments has been made at. Pirbright on the best means of
disinfecting materials which are likely to become contaminated in practice during
an outbreak. These have led to the conclusion that the best disinfectants for
practical purposes are such as raise the alkalinity of the liquid used. This is in
agreement with the results obtained by Minett (Third Progress Report, p. 51),
by Trautwein and Reppin at Riems, and by Olitsky.
The workers at Riems found that a one per cent. solution of caustic soda was
the most efficient disinfectant for general use, and instructions have been issued in
Prussia for farmers and others giving directions for the preparation of such a solu-
tion. This is now preferred at Riems to the preparation of sulphur formerly re-
commended there. Mr. Dobson finds that a solution of common washing soda is
very efficient. This is especially suitable since it is in common use for cleansing
I
202 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [ II, II
purposes, is easily procured and is unlikely to damage materials with which it
comes in contact. When dissolved in hot water at about 60°C. (140°F.) in a
strength of 4 per cent. it can be used very effectively for washing butchers'
utensils and for scrubbing tables and floors in slaughter houses, etc.
Most other disinfectant solutions which were examined, such as acids and
metallic salts, were liable to damage metal surfaces or the hands and clothes of
those who used them. The disinfection of hides has been given special attention,
since these are damaged by alkalis and many other disinfectants. Immersion in a
solution of sodium bisulphate (1 in 10,000) for 5 hours or sodium bifluoride
(1 in 20,000) for 2 hours killed the virus and did not damage the hide.
H.—CHEMOTHERAPY.
Many claims have been put forward at various times that an attack of the
disease can be prevented or rapidly cut short by the injection into infected
animals of chemical substances, but none has so far withstood the test of carefully
controlled experiments. Dr. Andrews records that a thorough trial of iodine both
on. guinea-pigs and cattle, at Pirbright, gave no confirmation of the success
reported from India. Similar negative results in guinea-pigs- were reported by
Maitland in the Third Progress Report, p. 81. Mr. Dobson has tested the value of
a number of colloidal preparations of metals on infected guinea-pigs without
obtaining any encouraging results.
Mr. Galloway has made a very large number of experiments on guinea-pigs and
rabbits with chemical substances in the hope of finding some drug which would
destroy or influence the virus in the living animal. Altogether he has examined 90
different substances belonging to all classes of drugs which have been found or
have been stated to be useful in various infections.
The preparations tested include 36 dyes and 54 other chemical products such
as salts or compounds of arsenic, bismuth, mercury, or others related to
formaldehyde, etc. Especial attention has been given to acriflavine, trypan blue
and bismuth preparations on account of their known value in other diseases.
None of these has definitely modified the course of the disease in doses which
were not seriously poisonous to the animals. The Committee is much indebted to
Dr. King at the National Institute for Medical Research for information about the
substances used and other help in the course of this work.
J.—DISTRIBUTION AND SURVIVAL OF VIRUS IN THE BODIES OF ANIMALS AFTER
DEATH.
It was known that in the bodies of animals which were killed during the
height of the disease, the virus was present in much greater concentration in the
RESEARCH WORK ON FOOT-AND-MOUTH DISEASE
203
vesicle fluid and epithelial tissues of the skin of the feet and of the mucous
membrane of the mouth than elsewhere. It has been found, too, that whereas the
vesicle fluid can often be diluted five million times and still be infective, the blood
cannot be diluted more than five or ten thousand times without losing its activity.
If the whole of a guinea-pig's body, excluding the skin, bones and intestine, is
ground up with phosphate salt solution so as to make 400 cubic centimetres of
emulsion, this cannot be diluted more than 1,000 to 5,000 times and still prove
infective. In order to obtain a supply of virus there is, therefore, no obviously
great advantage in using the whole carcase, although the blood serum contains a
definite amount of virus in addition to the affected parts of the skin and mucous
membrane. It is not likely, however, that the actual amount of virus in the
whole body would be twice as much as that present at the sites of vesicles on the
feet and mouth.
The amount of virus which can be obtained is at its highest when the primary
vesicles are well developed and the generalised eruption has begun, i.e., in the
guinea-pig usually between the 24th and 48th hours after inoculation, but on the
third day it has fallen to one-tenth or one-fiftieth of that on the second day. The
virus content of the body then falls rapidly, and is usually nil at the end of 7 to
10 days in guinea-pigs or 12 days in cattle.
K.—RESISTANCE TO PICKLING OF THE CARCASE.
It was shown by Minett (Second Progress Report, Appendix I, p. 41) that the
virus in the feet of infected guinea-pigs remained active in different pickling
solutions of salt, nitre, sugar, etc., at a mean temperature of 17°C. (62. 5°F.) for
periods up to 35 and 49 days. He also found that the virus in crushed bones was
infective for pigs by feeding after 42 days storage at a chilling temperature of
about —4°C. (24. 8°F.), whether the bones were derived from a bacon carcase or the
carcase of an ox.
L.—EFFECT ON THE INFECTIVITY OF CARCASES OF STORAGE AT DIFFERENT
TEMPERATURES.
On account of the importance of the persistence of infectivity in stored
carcases several series of experiments have been made on the carcases of cattle and
guinea-pigs.
For this purpose guinea-pigs killed at the height of the disease, about 48 hrs.
after inoculation, have been used for the most part in previous experiments.
From three carcases which were kept at 18°C. (64.4°F.) for 13 days and
had become partly putrid, the virus was recovered from the bone-marrow
204 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [II, II
(Appendix III, p. 284). That the virus usually remains active for very much
longer periods when carcases are kept at " chilling " or freezing temperatures has
been shown by numerous experiments.
It is known that repeated freezing and thawing has the effect of killing some
bacteria, and it has been suggested that the same result would follow in the case
of foot-and-mouth virus. Mr. Galloway has made many observations on the
virus in tap water, in blood, in the skin of guinea-pigs' pads, in bone-marrow, and
in the whole carcase, and has not found that the repeated freezing and thawing has
any effect in lessening the activity of the virus.
Carcases of guinea-pigs with primary or secondary lesions, which were frozen
for 26 days and then kept at 18°C. (64.4°F.) for 5 days, contained active virus
in the bone-marrow ; virus was also recovered from another carcase after chilling
at —4°C. (24. 8°F.) for 25 days, and from a third which had been 25 days at 4°C.
(39.2°F.) and then for 3 days at 18°C. (64. 4°F.).
In another experiment the virus in bone-marrow from an infected carcase,
which had been frozen and thawed 6 times in the course of 30 days, was still active.
In many of these experiments, however, there was reason to suppose that the
virus was deteriorating somewhat during the period of storage.
Dr. Andrews and his colleagues, working with guinea-pigs, made observations
on the survival of virus in the different organs and tissues which were removed
after death and stored separately at a chilling temperature of—2°C. (28.4°F.).
He found virus still active in the tongue, lungs, heart, liver, kidney, fat, bone-marrow
and tendons, at periods ranging from 62 to 145 days after the death of the animal.
In order to test more exactly the conditions under which carcases are kept
when stored for trade purposes, cattle were allowed to acquire the infection by
contact with other animals and were slaughtered by a butcher at the stage of the
disease which was believed to be the most infective.
The carcase was dressed, quartered and cooled at about 30°F (—1°C.). After
3½ days it was wrapped in stockinette and hessian and stored at 29°F. (—1. 6°C).
The offal, liver, tongue, cheeks, heart, skirt and gall-bladder, and also pieces of
stockinette and hessian soaked in blood were kept separate and stored at 18°F.
(—7. 8°C.).
Three such experiments were made with 10 cattle in all, and are recorded in
Appendix I, p. 40. It was intended that the carcases should be highly infective
at the time of slaughter. The degree of infectivity varies, however, very much
in different animals, and also with different strains of virus. It is not easy to tell
whether an animal is at the most infectious stage of the illness, and whether the
RESEARCH WORK ON
FOOT-AND-MOUTH
DISEASE
205
degree of infectivity, i.e., the concentration of virus in the body, is high without
testing the blood at the time of death.
The first three cattle had so little virus in the blood that the experiment was
abandoned. In the second experiment with another strain of virus, the blood
proved infective for swine at the time of slaughter. From each of the three
carcases, C737, C741, C742, virus was recovered later.
From C737 the bone-marrow proved infective when inoculated into pigs on the
13th and 33rd days after slaughter.
On the 14th day after slaughter inoculation of the tongue and cheeks of
C741 infected one pig each, and on the 33rd day the cheeks infected two pigs by
inoculation and two by feeding. The wrapping materials soaked in blood of C741
and of C742 also each infected two pigs by inoculation on the 40th day after
slaughter.
The remainder of the offal including the bone-marrow of these three animals
did not prove infective 13 days after slaughter nor did the salted hides stored at
out-of-door temperatures.
In the third experiment six cattle were exposed to infection and four of them
reacted—D7, D2, D6 and D4. Intramuscular inoculation of small quantities of the
organs and bone-marrow of these four animals after 33, 24, 7 and 22 days respec-
tively at about 18°F. did not convey infection to pigs. Nevertheless, inoculation
of a larger quantity of bone-marrow taken from D7 after 40 days caused infection
and when large amounts of bone-marrow with crushed bones taken from the
carcase of D4 after 80 days at 29°F. were fed to six pigs and these animals were
at the same time inoculated intramuscularly with small amounts of the bone-
marrow all of them became infected and showed characteristic signs of foot-and-
mouth disease.
From all these experiments it is evident that if animals at a highly infective
stage of foot-and-mouth disease are killed, dressed and stored at a " chilling " or
freezing temperature they may remain infective for some weeks.
In the guinea-pigs used in these experiments the degree of infectivity was
higher and much more uniform than in the cattle. There is, however, no doubt
that a more concentrated and active virus in cattle is sometimes present.
M.—TYPES OF VIRUS, VIRULENCE AND ADAPTATION OF VIRUS.
(i) Virulence. It is a matter of common experience in infectious diseases,
whether due to bacteria or a virus, that the infective agent varies at different times
and in different conditions and epidemics. The most obvious change is one of
206 THE INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [ II, II
virulence, by which is meant that a larger dose of the material or culture is needed
to infect an animal on one occasion than on another, and also that the disease
produced is much more severe and that sometimes the incubation is much shorter
with some samples of virus. The decline in virulence is often a serious obstacle
in experimental work. The reasons for this change are for the most part obscure,
but in some cases explanations of the difference in bacterial cultures are beginning
to emerge. In work with invisible viruses the problem is still for the most part
quite unsolved.
It is, however, known that a strain of bacteria or of a virus of low virulence
is often restored to full activity by infection of one animal after another by direct
transmission or passage.
(ii) Adaptation. Not only are the changes in virulence just mentioned of
common occurrence, but in attempted transmission from one kind of animal to
another the agent may completely fail to infect the new species, although highly
virulent for fresh members of the species from which it was obtained. In the same
way, as in the case of simple loss of virulence, bacteria or a virus may be adapted
to the new kind of host by inoculation of two or more individuals in series.
Like pathogenic bacteria, strains of virus which are adapted to one species of
animal by having infected a number of individuals of the same species in series
may have acquired increased virulence for this kind of animal and still only have
a very feeble power of infecting other species of animal. It has been observed
in the field that outbreaks of foot-and-mouth disease among swine in some cases
do not readily spread to cattle and vice versa. A corresponding course of events
has been found when artificial inoculation is practised. Certain strains of virus
from cattle have only with great difficulty been adapted to guinea-pigs by
Mr. Dobson, and with some such strains attempted infection of guinea-pigs has
never been successful in spite of repeated trials. A similar delay in adapting a
virus was sometimes experienced when a strain which had long been maintained
by passage from guinea-pig to guinea-pig was again inoculated into a bovine.
Dr. Edwards has encountered the same phenomenon when producing the disease
in hedgehogs, for a guinea-pig strain as a rule only gives slight infection in the
hedgehog until it has been transmitted through two or three of these animals.
The infection of rats with a strain from cattle or the guinea-pig may show the
same tendencies. On the other hand some strains are found to infect two or three
species with equal readiness, at least as regards artificial inoculation. These
differences between strains in respect of " adaptation " to particular species do not
appear to be related to the differences of " type " to be discussed later. Observa-
tions on adaptation show how dangerous it is to conclude without prolonged trials