VACCINATION WITH VIRUS 17D IN THE CONTROL OF
JUNGLE YELLOW FEVER IN BRAZIL

by Fred L. Soper and H. H. Smith, International Health
Division of the Rockefeller Foundation, Rio de Janeiro 3 3

Reprinted from the Transactions of the Third International Congress
of Tropical Medicine and Malaria, 1938, Vol. I, pp. 295-313

PRINTED IN HOLLAND BY C. A. SPIN & ZOON N.V.
VACCINATION WITH VIRUS 17D IN THE CONTROL OF
JUNGLE YELLOW FEVER IN BRAZIL?!
by Fred L. Soper and H. H. Smith, International Health

Division of the Rockefeller Foundation, Rio de Janeiro

Vaccination in Jungle Yellow Fever

The first attempt to protect an exposed population against
jungle yellow fever by. vaccination was made in Parana, Brazil, early
in 1936 (1), using hyperimmune goat and monkey sera and a virus
modified by culture in mouse embryo tissue (17E) (2). The difficulties
encountered were such as to cause the discontinuation of this method
in the field, and during the yellow fever season (January to May)
of 1937, no attempt was made to protect exposed populations.

Work with another modified virus (17D) developed in the
laboratories of the International Health Division of The Rockefeller
Foundation in New York, began in Brazil in February, 1937 (3).
By June, these studies had progressed far enough to justify field
vaccination, and the county of Varginha, Minas Geraes, in a region
where jungle yellow fever had been found a few weeks previously,
was chosen for the first field application of the new vaccine virus.
During the next three months, 2,746 persons were vaccinated in
the field, with satisfactory results, and, in September, routine field
vaccination began, which increased the total field vaccinations for
the year 1937 to 36,104.

The 1938 yellow fever season in South Brazil began early in
January, with an outbreak of jungle yellow fever at Presidente
Wenceslau, Sao Paulo (4), and shortly thereafter the disease was
found at Mathias Barboza, Minas Geraes. Vaccination units were
moved into both these districts, and an attempt was made throughout
the following months to vaccinate threatened populations wherever
yellow fever was found. The 1938 yellow fever season was an active
one, with outbreaks in some of the richest and most heavily populated
agricultural districts of Brazil, in the states of Minas Geraes, Rio de
Janeiro, Sao Paulo and Santa Catharina. The need for vaccine greatly
exceeded the initial production capacity of the laboratory and the

1 This report is based on work of many colleagues of the Cooperative Yellow Fever
Service, jointly maintained by the Ministry of Education and Health of Brazil
and the International Health Division of The Rockefeller Foundation. Special
credit for the rapid expansion of vaccination in 1938 must go to the Brazilian
Government, which furnished the necessary additional funds.

295
FRED L. SOPER AND H. H. SMITH

ability of the field service to apply it. The Brazilian Government
opened a special credit of 2,000 contos, or approximately $ 100,000
UScy., to cover the cost of a program for the vaccination of at least
one million persons during the year 1938.

From January first to July 31st, a total of 557,861 persons were
vaccinated, and the final figures for the year will almost certainly
exceed the preliminary estimate of one million.

Table J gives the number of persons vaccinated per month in
Brazil, from September, 1937, to July, 1938, by states. Table II
gives the distribution of the same persons by population groups.

Origin of Vaccine Virus 17D

In December, 1933, Lloyd transferred the Asibi strain to tissue
culture containing mouse embryo tissue and monkey serum ; after
18 subcultures, a second transfer was made to a medium containing
whole chick embryo tissue, from which, after 56 passages, it was
transplanted to tissue culture containing chick embryo, from which
the central nervous system had been removed. After 39 passages in
this medium, without central nervous system tissue, this strain of
virus, now known as 17D, was tested and found to have lost much
of its viscero- and neurotropism, while still retaining the property
of stimulating the production of antibodies (5). Virus 17D was first
used for human inoculation on November 30, 1936, in New York
(6), with material transferred 227 times in tissue culture since its
last previous passage in an animal host. Subcultures used as source
of vaccine in Brazil have ranged from the 205th to the 317th.

Results Obtained with Virus 17D

The points on which a method of vaccination for general use
as a public health measure should be judged, may be grouped under
the following headings :

a. Ease of manufacture of standard product.

Ease of application under field conditions.
Safety and comfort of persons vaccinated.
Safety of persons not vaccinated.
Antibody production. ,

Sun >

A. Ease of Manufacture of the Standard Product

The titer of virus in tissue culture material is much below that
obtained by growth in the developing chick embryo. The vaccine

296
VACCINATION WITH VIRUS 17D IN JUNGLE YELLOW FEVER

virus is maintained in tissue culture free of central nervous system
tissues, to avoid any possible reversion to type, but for the preparation
of vaccine, tissue culture is inoculated in the allantoic sac of the
six-day old chick embryo. After further incubation for four days,
at a temperature of 37° C., the embryo is removed, triturated, and
suspended (10%) in inactivated human serum diluted with equal
amounts of distilled water. The filtrate of this suspension, which is
the vaccine material, is distributed in ampoules, frozen, dried in
yacuum, sealed and stored at about 2° C. In addition to the usual bac-
teriological controls for sterility, each lot of vaccine is titrated for
virus content by intracerebral inoculation in serial dilutions in
white mice, and is inoculated intracerebrally into a rhesus monkey,
to test for possible increase in either viscero- or neurotropism.
Altough laboratory studies indicate (3) that a much smaller
dose may be sufficient, between 350 and 800 MLD 1 for mice are
now being allowed for each person vaccinated in Brazil. On this
basis the Rio de Janeiro Laboratory is producing some 120,000 doses
of vaccine per month, at a total cost, including overhead, excepting
rent, of less than $ 3,000 UScy., or 2% cents UScy., per dose.

B. Ease of Application under Field Conditions

Virus 17D, even when dried and sealed, is susceptible to
ordinary temperatures and to direct sunlight ; the vaccine leaves
the Rio laboratory, packed with ice and salt, in wide-mouthed
thermos flasks, and is thus kept chilled until the moment of rehy-
dration. Even after rehydration with distilled water the ampoule
is kept on ice, and the vaccine is finally diluted in physiological
saline solution in the syringe itself immediately preceding inoculation.
To determine the viability of the virus used, mice are inoculated
intracerebrally with the remaining vaccine after the last person has
been inoculated.

Experience shows that a vaccination unit, consisting of three
persons, a doctor, a technical assistant and a secretary-chauffeur, can,
under optimum conditions, register and inoculate from 1,000 to
2,000 persons a day *, The actual cost of applying vaccine in Brazil

1 The end point of titration in mice is considered as that dilution which, when
injected in 0.03 c.c. amounts intracerebrally in mice, will produce a mortality.
of 50% (7).

2 The use of three Forsbeck needle-racks by each unit is advisable, to avoid unne-
cessary delays in waiting for needles to cool after boiling. It is believed that certain
irregular results of postvaccination protection tests are due to failure to cool needles
after boiling, with consequent inactivation of the vaccine virus.

297
FRED L. SOPER AND H. H. SMITH

in 1938 has not exceeded, including initial cost of automobiles and
equipment, 7 cents UScy., per capita. The actual field operating
expense has dropped from 5% cents UScy., per capita, in January, to
3 cents UScy., in June. However, the per capita cost of application
must increase rapidly in sparsely populated regions and in areas
where transportation is difficult.

C. Safety and Comfort of Persons Vaccinated

Since the beginning of work with virus 17D in February,
1937, a conscientious search has been made among vaccinated
groups for evidence of:

1. Severe reaction at site of inoculation ;
. Sensitization to foreign protein ;
Serum sickness ;

Virus reaction, visceral and neural ;

Delayed jaundice, and

Nw ff Win

Infection with other viruses.

Special attention should be called to the distribution of vacci-
nated persons by population groups (Table II). Employees of the
Yellow Fever Service, of the airlines, the population of large coffee
fazendas, inmates of schools, laborers and highway construction
gangs and members of military units, all form very useful groups for
observation. Even where it has not been possible for physicians of
the Yellow Fever Service to make personal observation, fazenda
owners, military medical officers, school directors and other respon-
sible persons have given information as to the severity of postvacci-
nation reactions. :

The sum total of observations on vaccinated groups may be
stated briefly as follows :

For the eighteen months’ period, during which almost 600,000
persons were vaccinated, there is no evidence of severe reaction
at the site of inoculation, of sensitization to foreign protein 1, of
serum sickness, of delayed jaundice (8), (9), nor of infection with
other viruses.

The type of relatively mild reaction which is observed seems
to be a general, not neural, reaction to the virus itself, after an

1 A number of cases have received second and third inoculations of 17D, without
any evidence of sensitization to chick protein.

298
VACCINATION WITH VIRUS 17D IN JUNGLE YELLOW FEVER

incubation period of generally from five to eight days. 1

The symptoms most frequently noted are : headache, backache,
body pains, weakness and malaise, lasting from a few hours to a
couple of days. The reaction to virus 17D is not severe enough to
have any influence against its general acceptance by the people.
Fazenda owners, and others responsible for large groups, generally
report from 5 to 8% of reactions, with not more than 1 to 2% of
reactions severe enough to cause loss of time from work. A person-
to-person canvas, however, will result in 20, 40 or even 50% of
individuals questioned reporting at least a slight headache, but the
number of severe reactions does not increase correspondingly. The
most severe reactions reported are those related to each other by
members of the foreign colony in the capital city of Rio de Janeiro !

Considering the number vaccinated, it seems truly remarkable
that many more conditions occurring after vaccination have not been
credited to the inoculation. Experience has failed to reveal any
contraindications to the use of virus 17D, early restrictions have been
entirely removed, and children of all ages and women in all stages
of pregnancy are routinely inoculated.

D. Safety of Persons Not Vaccinated

In using a living virus for vaccination, the possibility of such
living virus being picked up from the blood stream by some insect
vector, and sooner or later reverting to its original virulence, must
be considered. Such return to virulence of a yellow fever vaccine
would have to depend upon the following factors :

1. Circulation of virus in the blood stream in quantities sufficient

to infect the insect vector ;

2. Ability of the infected vector to transmit the vaccine virus,

and

3. Ability of the vaccine virus to revert to a virulent state.

Experimental work indicates that sufficient virus does not
circulate to infect the traditional vector, Aédes aegypti, and that even
when this mosquito has been infected by special methods it does

1 So far, only one case has been reported, in which symptoms of involvement of the
central nervous system were attributed by the attending physicians to inoculation
with virus 17D. Case E. R. C., observed by Drs. Raul Azevedo and Deolindo
Couto, Rio de Janeiro, to whom we owe thanks for details of this case, developed
signs of meningeal involvement one month after vaccination with Lot 136 of
virus 17D, the estimated virus used being 220 MLD for mice. Complete recovery
occurred, and studies are now in progress to determine, if possible, the nature of
the infection.’ :

299
FRED L. SOPER AND H. H. SMITH .

not readily transmit the 17D virus, even after prolonged incubation
(10). Attempts to infect Aédes aegypti by postvaccination feeding on
humans and on rhesus monkeys, which have been shown to circulate
more virus than do humans, were failures, no virus being demon-
strated in the mosquito by either feeding on monkeys or inoculation
into mice. The immersion of Aédes aegypti larvae in high concentration
of virus did result in the production of infected mosquitoes, as
demonstrated by mouse inoculation; such infected aegypti failed
completely to transmit virus to susceptible animals, even after
prolonged incubation periods.

The difficulty of getting virus 17D to circulate in appreciable
quantities with regularity, has, so far, prevented conclusive experi-
ments with the jungle vectors of yellow fever, only a few of which
have very recently been definitely incriminated (11). The same
difficulty has prevented the carrying out of a large series of animal
passages, to determine the ability of virus 17D to revert to its
original type; the relative stability of the virus in tissue culture,
embryo passage and in mouse brain passage, suggests that such
reversion to virulence, if it did occur at all, would be slow in appear-
ing. This opinion is strengthened by the results of other workers,
who have not been able to transmit a tissue culture virus with Aédes
aegypti (12), nor to reconvert it to virulence by direct liver-to-liver

passage (13).
E, Antibody Production

The rhesus monkey, which is more highly susceptible to yellow
fever than is man, becomes fully resistant to virulent strains, such
as Asibi, following inoculation with virus 17D. Similar tests on
humans have not been made, but the wide use of virus 17D this
year, among exposed populations, during active outbreaks of jungle
yellow fever, has resulted in a mass of field observation almost as
conclusive as laboratory experiments. Local physicians and other
observers report a sudden reduction in observed cases in infected
districts shortly after mass vaccination, and cite instances in which
individuals, who failed to be inoculated, later contracted the disease
in infected forests, while vaccinated members of the same labor
gangs escaped. Field experience suggests that the protective effect
of vaccination begins not later than a week after inoculation, although
laboratory tests fail to show demonstrable antibodies at this time (3).
While it is probable that a much larger number of cases of yellow

300
VACCINATION WITH VIRUS 17D IN JUNGLE YELLOW FEVER

fever must have occurred among persons infected before vaccination,
only eight of these have been reported, four in Minas Geraes, three
in Santa Catharina and one in Sao Paulo. Onset in two was on the
same day as vaccination, in the other four, between the first and
fourth days following. Two of the three fatal cases in this group were
confirmed by viscerotomy, and a virus, quite different from the
vaccine virus, was isolated from one of the non-fatal cases.

Two additional cases of postvaccination yellow fever have been
found, one mild case with onset thirty days, and one fatal case with
onset six weeks after vaccination. These cases had received virus
from lots 95 and 117, both of which gave irregular results, as measured
by the protection test (Table IV). It is possible that neither received
active virus.

The mouse protection test (14) has been used since 1931,
for determining the presence of yellow fever antibodies in the blood
serum of persons and animals. It is customary to inoculate six mice
with highly neurotropic virus and with the serum to be tested.
Results are read as a fraction showing the proportion of mice living
on the fourth day (denominator), which survive to the tenth day
(numerator) after inoculation.

Seven readings are possible, of which only two, 6/6 and 5/6,
are, in analyzing critical immunity surveys, considered definite
evidence of previous infection with yellow fever; 4/6 and 3/6
results are considered inconclusive, and 2/6, 1/6 and 0/6 as negatives.
It has been noted in immunity surveys that bloods from regions
where yellow fever has never been present give remarkably clearcut
negative readings, whereas bloods from endemic regions give an
appreciable number of inconclusives, as well as positives and nega-
tives. The majority of these inconclusives are probably from indivi-
duals who have at some time been exposed to yellow fever infection,
and are, almost certainly, not apt to ever again develop clinical
yellow fever. It seems’ reasonable at the present time to read mouse
protection test results as indicating full protection, partial protection
and no protection, without attempting to interpret too rigidly these
readings in terms of reaction to yellow fever infection, further than
to assume that those showing full protection are, at the moment
tested, adequately protected against fully virulent virus. Postvacci-
nation results, when compared with prevaccination results (Table III),
suggest that virus 17D does produce some measurable antibody
formation in almost 100% of persons receiving so MLD or more

301
FRED L. SOPER AND H. H. SMITH...

of living virus. It has been noted that in many postvaccination pro-
tection tests, in which the final reading is ; 2/6, 1/6, or even o/6,
the average length of survival of inoculated animals is from one to
two days longer than for similar negative tests in vaccinated groups.
This suggests that sufficient antibody is present to definitely delay
the action of virus inoculated in animals,

Table III gives the results of pre- and postvaccination tests on
the same individuals, including both laboratory and field groups },
during the preliminary phase of observation, before routine field

_ vaccination began, Attention must be called to the fact that on one
occasion, the virus was apparently inactive before inoculation began,
since all of the persons tested failed to give evidence of antibody
development, and the inoculation of the remaining vaccine into mice
failed to cause any deaths.

Table IV covers a special investigation to determine the results
obtained with different dosages of virus, and to evaluate the viability
test as an indication of efficiency of the preceding vaccination. The
groups bled for this special study were selected as probably represen-
tative of the poorest work of the season, and included groups receiving
the lowest doses of virus used during the height of the yellow fever
outbreak, working with newly trained personnel, far from head-
quarters. The results indicate that doses as low as 50, 85 and 100
MLD per person are adequate to give satisfactory results. They also
indicate that the viability test, in and of itself, is not a safe indication
of the efficiency of the vaccination. For example, lot 117 of virus
17D was used and tested in five groups, of which only one gave
satisfactory results, the viability tests for which, o/s, 1/5 and 1/4,
were poor. Postvaccination mouse protection tests on a number
of persons from vaccinated groups are proving a better method of
checking the work of field units than is the test for viability of the
remaining vaccine. .

Table V gives a general summary of all postvaccination protection
test results for work with virus 17D in Brazil.

A study of Tables III, IV and V and other available information
suggest that the differences in the results of vaccination depend in
great part upon the delivery of relatively small amounts of active
virus below the skin of the individual vaccinated, The results show

 

1 Vaccination of these groups was carried out under the direct supervision of
Dr. H. H. Smith, who, with Drs. Henrique Penna and Adhemar Paoliello (3)
has published a report covering observations on the first 60,000 vaccinations in Brazil.

302
VACCINATION WITH VIRUS 17D IN JUNGLE YELLOW FEVER

that with standardized methods of vaccine production and with
adequate supervision of the administration of virus in the field,
highly satisfactory results can be obtained.

Anticipated Epidemiological Results of Vaccination

Admitting that the individual can be protected by vaccination,
the epidemiological results of vaccination must vary with the con-
ditions under which infection, occurs. Where man is an essential
element in the cycle of infection, responsible for maintaining the
virus, as in urban aegypti-transmitted yellow fever, artificial immuni-
gation of the bulk of the population should effectively protect the
remaining non-immunes. It is probable that occasional mass vacci-
nation will be found more economical and practicable in certain
regions, for breaking the cycle of infection, man-aegypti-man, than
is the traditional maintenance of antimosquito services for the
prevention of aegypti breeding.

In considering jungle yellow fever, however, in which man is,
apparently, not an important factor in maintaining the virus, vacci-
nation should alter the epidemiological picture, mostly by preventing
the infection of vaccinated persons, and, only in a very minor
degree, by reduction of the source of virus for forest vectors.

Vaccination promises to be a great aid in preventing the transfer
of yellow fever infection from one place to another by the human
host ; the long-distance transfer of virus, by modern methods of
rapid transportation, can be prevented by vaccination, as can also
the introduction of virus from jungle to urban areas. Since the jungle
infection, apparently, exists independent of the human population,
and spreads from place to place by other than human carriers,
vaccination cannot be expected to completely eradicate yellow fever.

Summary.

During the period, September 1937 to July 1938, over half a
million persons were inoculated with the modified yellow fever
virus 17D. Vaccination with this virus was widely used throughout
the 1938 epidemic of jungle yellow fever in South Brazil. Field obser-
vations indicate that vaccination becomes effective within a week
after inoculation. Reaction to vaccination is relatively mild, and no
contraindications have been found. The results of approximately
3,000 mouse protection tests are presented, showing that a high
percentage of persons vaccinated develop demonstrable antibodies.

393
FRED L. SOPER AND H. H. SMITH

Bibliography
Soper, F. L. Vacinagao contra a febre amarella no Brasil, de
1930 4 1937. Arch. de Hig. Rio de Janeiro. 1937. 7: 379-390.
Lloyd, Wray, Theiler, M., and Ricci, N. I. Modification of the
virulence of yellow fever virus by cultivation in tissue in vitro.
Tran. Roy. Soc. Trop. Med. Hyg. 1936. 29: 481-529.

. Smith, H. H., Penna, H. A., and Paoliello, A. Yellow fever

vaccination with cultured virus (17D) without immune serum.
Am, jl. Trop. Med. 1938. In Press.

Aragao, H. de B. Observacies a respeito de um foco limitado de
febre amarella sylvestre no Estado de Sao Paulo. Brasil Medico.
Rio de Janeiro. 1938. 52: 401-412.

Theiler, M., and Smith, H. H. The effect of prolonged culti-
vation in vitro upon the pathogenicity of yellow fever virus.
jl. Exp. Med. 1937. 65: 767-786.

Sawyer, W. A. Experience in vaccinating against yellow fever

~ with immune human serum and virus fixed for mice. Am. ji.

To,

Il.

12,

13.

14.

304

Hyg. 1937. 25: 221-231.
Reed, L. J., and Muench, H. A simple method of estimating
fifty per cent endpoints. Am. Jl. Hyg. 1938. 27: 493-497.

. Findlay, G. M., and MacCallum, F. O. Note on acute hepatitis

and yellow fever immunization. Trans. Roy. Soc. Trop. Med. Hyg.
1937. 31: 297-308.

Soper, F. L., and Smith, H. H. Yellow fever vaccination with
cultivated virus and immune and hyperimmune serum. Am. Jl.
Trop. Med. 1938. 18: 111-134.

Whitman, L. Failure of Aédes aegypti to transmit yellow fever
cultured virus (17D). 1938. In Press,

Shannon, R. C., Whitman, L., and Franca, M. Yellow fever
virus in jungle mosquitoes. Science. 1938. 88: (No. 2274)
110-111,

Roubaud, E., Stefanopoulo,.G. J., and Findlay, G. M. Essais
de transmission par les stégomyies du virus amaril de cultures
en tissu embryonnaire.. Bull, Soc. Path. Exot. 1937. 30: 681-583.
Findlay, G. M., and MacCallum, F. O. Vaccination contre la
fiévre jaune au moyen du virus pantrope atténué employé seul.
Bull. Off. Internat. d’Hyg. Publ. 1937. 29: II4§-1149.
Sawyer, W. A., and Lloyd, Wray. The use of mice in tests of
immunity against yellow fever. JI. Exp. Med. 1931. 54: 533-555.
VACCINATION WITH VIRUS 17D IN JUNGLE YELLOW FEVER

TABLE I

Persons in Brazil vaccinated with virus 17D from September 1937

to July 31st 1938, away from the laboratory

 

 

 

 

 

 

 

 

 

 

Federal | Rio de S20 Minas | Santa | Mato
Months district | Janeiro | Paulo | Geraes | Catarina} Grosso | Total

State State State State State
September ...... 3.759 3.759
October ........ 10.580 10.580
November ...... 7-473 7-473

December ...... 11.540 6 11.546 —

January ........ 46 8.103 | 12.701 20.850
February ....... 3.337 1.861 31.557 It 36.766
March.......... 13.455 | 18.234 64.558 96.247
April. ........5. 10.313 | 17.238 45.084 | 22.363 94.998
May .....ceeere 6.181 | 12.894 66.340 | 12.005 11 97-431
June.........00. 5.224 | 16.760 5.406 | 72.393 99.783
July ........... 6.444 | 10.726 | 11.259 | 83.357 111.786
Total ...... 45.000 | 77-713 | 24.768 |409.342 | 34.368 28 591-219

 

395
FRED L. SOPER AND H. H. SMITH

TABLE II

Distribution by population groups of persons
vaccinated in Brazil

September 1937 to July 31st 1938

 

 

 

: September to January to

Population group December 1937 July 1938 Total
Farms and hamlets ........ 16.530 397.809 414.339
Military units ............, 1.105 23.730 24.835
Schools ....... 0... cece eae 994 34.348 35.342
Labor gangs ............. 368 39.183 39.551
Cities and towns .......... 14.361 53-337 67.698
Miscellaneous ............. _— 9.454 9.454
Total... 2. cee eee eee 33.358 557-861 591.219

 

 

 

 

 

306
Loft

TABLE It

Immunity to yellow fever following vaccination with Virus 17D measured by mouse protection test

 

 

 

 

we >
Lot | Dilu- Dose Viability Per Num- Mouse protection test results? - of tec
, M.L.D! Where used test : ber 0/4 1/4 | i 3/4 4/4) mice | tion
No, | ton 2 | ‘lated. a} o/5 | Us a 2/4) BI) ais | 5/5 | sur- | in-
(Mouse) result’ ated | tested | | 7¢ 1/6 2/ 3/6 4/6 5/6 6/6 \viving| dex*
39 «| 1:1) 850 to | Laboratory 5/6,6/6 | 20 18
1:2) 7-500 Pre-vaccination 14 2 ° ° I I ° 10 0.6
Post-vaccination ° ° o ° 3 4 i 89 5-4
40 | 1:1) | 85.000 | Laboratory 26/26 | 52
1:2) Pre-vaccination 43 6 ° ° ° 2 8 0.4.
Post-vaccination I ° ° 5 7 ir 28 82 5.1
41 [| 1:2 | 25.000 | Field 18/18 | 77) 66
Pre-vaccination 54 9 2 ° ° ° I 5 0.3
Post-vaccination ° ° 3 3 18 22 20 78 4.8
41 | 1:2) | 25.000 | Laboratory 23/23 | 10
und) Pre-vaccination 4 4 ° ° ° I I 26 1.§
Post-vaccination ° ° ° ° o 2 8 96 5.8
42 |1:2 | 11.000 | Field 23/23 | 14x | 132 ,
Pre-vaccination 109 16 3 ° ° 2 2 5 0.3
Post-vaccination ° ° ° 7 25 42 58 85 5.1
52 | rit 25.000 | Field 23/23 | 79 69
Pre-vaccination 54 13 ° I ° ° I 5 0.3
Post-vaccination ° ° ° I 15 29 24 84 5.1
52 | r:10| 2.500 | Field 27/27,1/2 | $589 | 159
Pre-vaccination 122 26 8 2 ° ° I 4 0.3
Post-vaccination 4 I 2 18 46 54 34 76 45
52 | 1:10] 2.500 | Field o/10,0o/7 | 172 | 21
Pre-vaccination ~ 17 2 ° ° ° ° 2 iI 0.7
Post-vaccination 15 4 ° ° ° ° 2 13 0.8
TOTAL 1149 | 527
Pre-vaccination 417 78 13 3 I 5 10 6 0.4
Post-vaccination 20 5 5 34 114 164 185 78 4-7

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

w

The endpoint for titration of virus in mice is calculated on the basis of 50 % mortality.
The fraction indicates the number of mice dying of specific encephalitis (numerator)

(denominator).

The fraction indicates the number of mice surviving to the tenth day (numerator) in comparison with number alive four days after inoculation

(denominator).

Average number of mice surviving calculated on basis of six mouse groups.

in comparison with number alive four days after inoculation

WAATAT MOTIAA FTONN NI G41 SNUIA HLIM NOLLYNIOOVA
TABLE IV

Immunity to yellow fever following vaccination with Virus 17D
measured by mouse protection test

 

 

 

Dose Viability Per- Num- Mouse protection test results? % Pro-
Lot | Dilu- sons of | tec-
. M.L.D! Where used test : _| ber 0/4 1/4 3/4 4/4] mice | tion

No. tion inocu 2/ 5 2/. 4 3/ 5 . in-

(Mouse) result? lated | tested o/s 1/5 2/6 3/6 4/6 4/5 5/5 | sur in
ate 0/6 1/6 5/6 |6/6 |viving| dex4

69 Juss 1goo | Farms & Hamlets 0/6 47 10 ° I ° ° I I 7 88 5.2
79 | rss 85 | Cit. & Towns 1/6 338 17 2 ° ° I 3 3 8 81 4.6
Ing 85 | Milit. Units 6/6 143 20 ° ° ° ° ° 4 16 96 5.8

8o J 1:5 130 | Farms & Hamlets 3/5 329 | 21 I ° I ° 1 7 II 84 5.1
1:5 | 130 | Farms & Hamlets 1/5 350 | 23 I ° 1 I 2 7 11 82 5.0

85 Frss so | Cit. & Towns 2/6 364 21 3 ° o ° 2 6 10 77 4.7
Iig go | Cit, & Towns 6/6 245 20 6 ° 0 1 ° 5 8 63 3.8

95 | 1:20 850 | Farms & Hamlets 1/6 237 ar 10 3 2 ° I ° 5 32 2.0
1:20 850 | Farms & Hamlets 4/4 476 | 20 I ° ° 1 I 3 14 87 5.3

Or | 1:20 85 | Schools 2/5 72 12 ° ° ° ° I ° II 97 5.8
1:20 85 | Farms & Hamlets 6/6 108 10 ° ° ° a I 4 5 89 5-4

1:10 170 | Farms & Hamlets 18/18 195 21 ° ° ° ° 5 3 13 89 5.4

102 | 1:20 270 «| Cit. & Towns 3/6 94 22 ° ° ° ° 2 4 16 96 5.6
1:20 270 | Farms & Hamlets 6/6 353 20 ° ° oO 1 3 5 i 88 5:3

1:10 540 | Farms & Hamlets 16/16 Ti 20 I ° ° I 3 I 14 87 5.2

1:20 270 «| Cit. & Towns 6/6 Ir II ° ° I Q ° 3 7 90 5.4

103 | 1:10 too =| Farms & Hamlets 4/6 399 20 ° ° ° ° 4 5 1 89 5.4
I:10 10oo =| Farms & Hamlets 6/6 414 20 ° ° ° ° I 6 13 93 5.6

106 J rsg 8s; | Farms & Hamlets 4/6 269 20 3 ° ° ° 1 7 9 77 47
I:g§ 85 | Cit. & Towns 4/5 353 21 10 I I I ° 4 4 41 2.4

11g | r:10 110 | Farms & Hamlets of4 4l7 21 I ° ° 3 I 6 10 88 4.9
1:10 110 | Schools 0/4 634 21 ° ° ° ° 3 7 II 89 5.4

Ii10 110 | Farms & Hamlets 5/5 119 19 2 ° ° I I 7 8 78 4.7

I:10 110 | Farms & Hamlets 5/5 109 21 I I I I ° 5 12 82 4.9

17 | 1:20 85 | Farms & Hamlets o/s 405 5 ° ° ° o ° I 4 96 5.8
1:20 85 | Farms & Hamlets 1/5 192 II ° I ° ° ° 5 5 84 5.1

1:20 85 | Cit. & Towns 6/6 356 21 I ° o ° 3 5 12 86 5.2

1:20 85 | Cit. & Towns 6/6 243 20 5 I ° ° 2 5 7 64 3.8

I:10 170 | Farms & Hamlets 5/5 6o 20 3 2 ° ° 2 6 7 68 4.1

1:20 85 | Farms & Hamlets 1/6 384 21 6 I ° 1 ° 7 6 bo 3.6

1:20 85 | Farms & Hamlets |4/6,10/12, 5/5 | 1201 54 3 3 5 7 6 12 18 Jo 4.2

126 | 1:20 200 | Farms & Hamlets 3/4 42 7 I 3 ° I 12 18 JI 4.3
1:20 200 | Farms & Hamlets of§ 1307 36 6 I I ° 3 7 18 73 4.4

§
1:20 200 ©| Farms & Hamlets 1/5 42 11 ° 2 ° 5 10 14 63 3.8
137 [1:20 280 | Cit. & Towns 4/4 954 39 I I ° I 3 17 15 83 5.0

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

The endpoint for titration of virus in mice is calculated on the basis of 50 % mortality.

The fraction indicates the number of mice dying

(denominator).

(denominator).

Average number of mice surviving calculated on basis of six mouse groups.

308

of specific encephalitis (numerator) in comparison with number alive four days after inoculation

The fraction indicates the number of mice surviving to the tenth day (numerator) in comparison with number alive four days after inoculation
TABLE V

Immunity to yellow fever following vaccination with Virus 17D
measured by mouse protection test

 

 

 

a _ Mouse protection test results 3 % Pro-
Lot | Dilu- | Dose \ h 4 Viability Fer: Nom- - of tec-
No tion Mouse) Winer wee nek? inocu- te: ted o/4 1/4 2/5 2/4 3/5 3/4 4/4] mice tion
. (Mouse su lated | St€¢] 0/5 | 1/5 | 2j6 | 3/6 {4/6 | 4/5 | 5/5 | sut- | in-
0/6 1/6 5/6 6/6 viving | dex
39 | 1:1) | 850 to | Laboratory 63/65 20 18 Q ° o ° 3 4 Ir 89 5-4
1:2) 7-500
40 1:1) | 85.000 | Laboratory 201/202 71 52 I ° ° 5 7 It 28 82 5.1
1:2)
41 [1:2 | 25.000 | Field and 18/18 77), 66 ° o 3 3 18 22 20 78 4.8
41 I:2 25.000 | Laboratory 27/29 )} 10 ° ° ° ° ° 2 8 96 5.8
42 1:2 11.000 | Field 34/34 141} 132 ° ° ° 7 25 42 58 85 5.1
52 Juit 25.000 | Field 23/23 y 69 ° o fs) I 1§ 29 24 84 5.1
52 | 1:10] 2.500 | Field 28/29 840); 159 4 I 2 18 46 54 34 76 45
52 | t:10| 2.500 | Field 1/18 >) 15 4 ° ° ° ° 2§ 13 0.8
54 | 1:10 ? Field and 79/86 1245)/ 143 3 4 4 20 33 44 35 74 44
54 Laboratory 198/203 76 ° ° ° 3 8 28 37 88 5.3
55 1:10] 6.800 | Field 121/125 Isso} 108 ° 2 2 10 28 32 34 78 47
56 | 1:10 540 | Field 134/138 1868 80 2 ° ° 2 5° 22 49 89 5.4
57 |1t:to0 ? Field 49/88 1204 23 Ir 2 ° ° I 4 5 41 2.4
58 Ju:t 230 | Laboratory 207/240 126 68 I ° ° 6 12 20 29 83 5.0
59 | I:10 2 School & Field 68/84 1680 55 4 ° ° 2 6 17 26 82 4.9
60 |] 1:10 1.700 | Field 100/107 1943 78 I 1 I 7 13 31 24 79 4.8
61 1:10 760 | School & Field 196/207 3389 99 3 3 I 12 16 32 32 76 4.6
62 1:10 1.700 | Army & Field 205/218 2866 | 129 6 I I 5 14 36 66 83 5.0
63 1:10 ? City & Field 134/144 2284 51 I ° 2 2 6 21 19 81 5.0
64 1:10 540 | Field 78/78 1866 56 3 I ° 3 II 16 22 78 4.8
65 1:10 340 | City & Field 81/84 1743 73 I I 2 4 10 29 26 81 4.9
67 Tit 540 | Laboratory 175/188 73 |. 24 ° ° ° ° 7 7 10 85 5.1
68 I:1Io 170 | Field 45/71 1293 18 4 ° ° I 5 3 5 64 3.
69 rig 1.500 | Field 57/70 714 10 I ° ° ° I 1 7 88 5.1
FI I:10 170 | Army & Field 50/105 2152 47 6 4 ° I 4 9 23 3 4.4
76 [1:20 850 | Field 184/190 4414} 93 6 1 I 1 21 24 39 79 4.8
77. J t:i20 850 | City & Field 300/317 5933 | 128 2 I 5 11 22 30 57 81 4-9
78 1:10 400 | Field 139/149 2807 84 3 ° I 3 10 29 38 85 5.0
79 [iss 85 | Army & Field 40/51 1486 37 3 ° ° I 3 7 23 85 5.1
Bo Fis 130 | Field 22/37 1785 44 2 ° 2 I 3 14 22 83 5.0
83 JF 1:10 230 | City 270/279 3191 | 100 2 4 2 5 7 31 49 82 5.0
84 | 1:20 850 | Field 37/47 4514 31 ° ° ° o 3 12 16 go 5-4
85 iif 50 | City 47/54 2182 43 9 ° ° I 2 II 20 71 4.3
88 1:20 850 | Field & Lab. 186/202 4752 62 5 I 2 5 6 19 24 82 4.6
95 | 1:20 850 | Field 145/163 6233 | 41 TI 3 20 I 2 3 19 59 3.6
Io 1:10 170 | School & Field 212/232 6822} 46 ° ° I ° 7 7 31 gt 555
102 1:20 270 | City & Field 264/282 7995 jo I ° I 2 7 II 48 92 5-4
103 1:10 too | Field 101/106 3867 40 ° ° ° ° 5 Ir 24 gt 5-5
106 I:§ 85 | City 20/23 624 41 13 I I I I It 13 58 3.5
wm2 [rit 230 | Laboratory 85/94 21 14 ° ° o I 2 5 6 85 5.1
115 1:10 110 | School & Field 27/42 3614 |- 80 4 I I 5 5 25 39 83 5.0
WI7_ | r:1o0 170 | City & Field 150/208 10454 | 153 18 8 5 9 13 41 59 72 4.2
123 1:20 200 | Field 118/176 = }.10259 5 ° ° ° ° ° I 4 96 5.8
126 | 1:20 200 | City & Field 59/85 6166 | 120 23 3 6 1 9 30 48 69 4.1
136 | 1:20 220 | Laboratory 240/253 9233 9 ° ° ° 1 2 3 3 81 4-9
137 | 1:20 280 | Field 87/89 7400 38 I I ° 1 3 17 15 83 5.0
TOTAL 130897] 2944 | 170 48 48 162 427 858 |1231 80 4.8

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

The endpoint for titration of virus in mice is calculated on the basis of 50 % mortality.

The fraction indicates the number of mice dying of specific encephalitis (numerator) in comparison with number alive four days after inoculation
(denominator).

The fraction indicates the number of mice surviving to the tenth day (numerator) in comparison with number alive four days after inoculation
(denominator).

Average number of mice surviving calculated on basis of six-mouse groups.

Prevaccination immunes.

 
 

 

 

 

 

 

 

 

 

 

 

 

 

Oe ee ee ee

0 @auS21 SNOSEId

 

VIRUS USED, DOSAGE, MOUSE PROTECTION TEST RESULTS

 

 

 

 

 

 

 

 

 

 

 

 

 

IMMUNITY TO YELLOW FEVER BEFORE AND AFTER VACCINATION WITH VIRUS i7D.

 

 

VIABLITY TEST
RESULT.

  

 

 

 

 

 

 

 

VIRUS USED, DOSAGE, | MOUSE PROTECTION TEST RESULTS

WHERE GIVEN AND | ow

 

FRED L. SOPER AND H. H. SMITH

 

310
VACCINATION WITH VIRUS 17D IN JUNGLE YELLOW FEVER

 

PERSONS VACCINATED

 

MONTHLY PROGRESS OF ROUTINE FIELD VACCINATION
IN BRAZIL, SEPTEMBER 1937 TO JULY 1938

110.000 —
100.000 —
90.000 —
80.000 +

70.000 —

60.000 —
50.000 —
40000 —
30.000 —

20000

10.000 —

i)

pra

 

 

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uo ff wb WwW & & zZ2 <9 2a %
5 5 a, Wo an ao 28 ag A ES Se Sf 5
we 2+ 25 68 35 235,20 St <0 ¢zX ~“H 29 5
MOO We Wi Oo Gr Tes ws SO a a a
90 sHe™ Fo é = Q 6
avTN 5 qa% O= 2 oF IN OM
zio@” & & FF Ayr #
eum wig & Zz
& aS
ne a<
z on
> ere (937 1938
on .
Su
&
>

111786

 

 

311
FRED L. SOPER AND H. H. SMITH

 

 

POST VACCINATION MOUSE PROTECTION TEST RESULTS
ON 2944 PERSONS INOCULATED WITH VIRUS 170

1.300 7

1.200 —
1100 —4
1.000 —
900 -—
800 —
700 —
600 —

§00 —

PERSONS TESTED

400 =
300 —
200 =

100 =

170 | 4g |. 4g | i62.[ 427 | 858 | 1231

 

 

 

oat 1/4 4 gia] 4/4
o/5 v5 oe 2} . ‘es 4/5 5/5
oe =| 176 6 | 3/6 % lose | 66

 

 

 

 

 

 

 

MOUSE PROTECTION TEST RESULTS

 

 

 

 

312

 
VACCINATION WITH VIRUS 17D IN JUNGLE YELLOW FEVER

Disputatio.

W. A. P. Schiiffner (Holland): Mit besonderer Genugtuung
hérte ich die Vorschlage Sopers, die er beziigl. des Ablesens des
mouse-protection-tests machte. Sie stimmen mit den von uns in
Amsterdam gegebenen erfreulich iiberein. Die Anspriiche, die man
an den Mause-Versuch stellen muss, haben sich mit der Zeit ge-
yndert. Urspriinglich von Theiler und von Sawyer ausgearbeitet,
wurden die Methoden von franzésischer und portugiesischer Seite
(im Office international d’hygiéne) als nicht spezifisch angegriffen.
Um diesen Vorwurf zu entkraften, vermehrte Sawyer die Menge
des Virus; statt einer 10% Emulsion nahm er eine 20% ; damit
konnten unspezifische Reaktionen (die iibrigens kaum vorkommen)
mit noch grésserer Sicherheit ausgeschaltet werden. Aber natiirlich
gingen damit schwache spezifische Reaktionen verloren, Heute aber,
wo an der Spezifitét des Mause-Versuchs nicht mehr gezweifelt
werden kann, verlangt die Erforschung der Epidemiologie des Gelb-
fiebers auch das Erfassen einer schwachen Immunitét. Wir haben
daher einmal die schwaichere Emulsion (10%, und davon 0,2 cc. mit
0,4 zu priifendem Serum intraperitoneal gegeben) beibehalten und
zweitens, ebenso wie heute nun auch Soper, vorgeschlagen, die Resul-
tate, die jetzt noch als zweifelhaft oder gar als negativ gelten, mit zu
beriicksichtigen. Ich stimme Soper vollkommen bei, wenn er daran
erinnert, dass selbst ein volkommen negativ abgelaufener Mausever-
such (6/6) noch nicht eine Rest-Immunitat ausschliesst. Zu dieser
Auffassung wurden Snijders und ich friiher bereits bei unsern Dengue-
Untersuchungen gedrungen, spater hatten wir sie fiir das Verstandnis
der Verhiltnisse in Suriname, wo der Eingeborene auffallend resistent
bei Gelbfieberepidemien war, nétig.

313