J. R. HADDOW AND J. A.
IDNANI
53
One or two outbreaks of the disease have occurred on farms which possessed vaccinated
fowls and in all cases the vaccinated birds resisted infection.
In view of the isolated result obtained by Iyer [1941], in England in which one chicken
was proved immune two years later, taken together with the fact that even very young
chickens show no diminution in immunity after six months as they do in the case of gel
vaccine, it is hoped that a serviceable immunity will persist for the normal life span
Dried virus
Undiluted virus may be kept at refrigerator temperatures without noticeable deteriora¬
tion for three to four months, after which a progressive reduction in virus content occurs,
but, as the virus in amniotic fluid does not stand exposure to high temperatures for long, it
may be completely inert after three days at 37°C.—and as considerable difficulty was
experienced in transporting the vaccine on ice, a few experiments have been carried out
with virus dried on glucose powder, cotton wool, muslin, and filter paper. Dried yolk,
which has a similar virus content to amniotic fluid was also used. Of these, glucose powder
and cotton wool were the most successful. However, it is not easy to get uniform suspensions
from the cotton wool material, but the glucose is easily handled. The method consisted of
dropping 1 c. c. amniotic fluid on to 1 gm. of glucose (or proportionate smller quantities), and
then drying this in a vacuum desiccator over calcium chloride. More than 200 chicks were
vaccinated with this material, but neither a standard procedure for its preparation nor
the temperature limit at which it may be safely kept have yet been defined.
SUMMARY
1. The results of vaccination with virus adsorbed on alumina gel previously described
have been amplified. It has been shown that vaccination can be carried out successfully and
safely at all ages, but that the immunity conferred is not of long duration and in very young
chicks may not exceed two months. Preparation of the vaccine is tedious and the safe time
limits for use after preparation restrict the practical application of this vaccine to short range
work in areas adjacent to a well-equipped laboratory.
2. By passage of the virus in pigeons a satisfactory vaccine has been evolved.
3. A more rapid reduction in virulence was attained by a few passages of the virus
through one to ten-day old ducklings.
4. Various factors which may influence the loss of virulence of the virus during passage
in hens' eggs have been examined. The virus content of embryos decreases if incubation
exceeds three days. Vaccines could not be obtained from viruses heated at 60°C.—70°C.
The same was true of virus isolated from gel mixtures, virus mixed with immune serum, with
bile, crystal violet, or glycerol.
5. Attenuation to various degrees occurred on passage of the virus in hens' and ducks'
eggs. The practical application of one such strain, in which the attenuation was maximal,
has been described. Over 100,000 birds of all ages over one month have been successfully
vaccinated at casualty-rates varying from 2½ to 6 per cent according to the general standard
of health of the birds vaccinated. The vaccine consists of amniotic fluid taken 48 hours after
the inoculation of nine or ten day embryos. This is ampouled in 1 c. c. quantities and may
be kept at refrigerator temperature for three to four months. It is diluted 1 : 500 with sterile
water immediately prior to use and the dose given is 0.5 c. c. subcutaneously. Immunity
develops after 48 hours and lasts for more than one year. Birds undergoing vaccination are
infective, but the infection that may be so transmitted is 'silent'.
ACKNOWLEDGEMENTS
The authors wish to thank the following gentlemen for their assistance : Dr N. B. Das,
Biochemist, for the supply of the alumina gels, Mr S. R. Nanda, Engineer, for the construc-
tion of incubators, Messrs C. Seetharaman and P. C. Gangulee, Research Assistants, for
assistance with field inoculations, Mr A. J. MacDonald, Officer-in-charge, Poultry Research
Section, for supplying eggs, chickens and fowls, Maj. Gen. C. M. P. Durnford, Col. J. R. Brown
and Lt. Col. E. G. Young of the Production Branch, Central Command, for placing the flocks
of the Command at our disposal and for organizing the army field work, Sir Frank Ware,
for facilities for work in the U. P. and to Dr F. C. Minett, for encouragement and sugges-
tions regarding this article.
REFERENCES
Haddow, J. R. and Idnani, J. A. (1941). Indian J. vet. Sci 11, 113
Iyer, S. Ganapathy and Dobson, N. (1940). Vet. Rec. 52, 889
Iyer, S. Ganapathy (1941). Indian J. vet. Sci. 13, 1
Willstatter, R., Kraut, H., and Erbacher, P. (1925). Ber. 58, 2448, 2454