ORIGINAL ARTICLES
VACCINATION AGAINST RANIKHET DISEASE
By J. R. HADDOW AND J. A. IDNANI, Imperial Veterinary Research Institute, Mukteswar
(Received for publication on 2 April 1946)
D URING the last three years we have been exploring different methods of preparing
Ranikhet disease vaccine, and the results are surveyed in the present paper. As will be
seen, more than one of these efforts quickly gave very encouraging results. These have not
yet been investigated exhaustively, because at the time other methods appeared to be still
more promising and because the urgent need of the moment was a vaccine which would
help to stem severe losses from Ranikhet disease and so assist the country's war effort. In
due course, it will be profitable to re-examine some of the procedures, in order to come to a
more firm decision as to the vaccine that should be finally recommended.
Strains of virus used. R 1, a virulent laboratory strain kept in fowls, R 10, a freshly
isolated field strain kept in egg cultures, and R2, attenuated egg-passaged virus, were
used.
I. GEL-ADSORBED VACCINE
In a preliminary report [Haddow and Idnani, 1941] an account was given of the pre-
paration of a vaccine by adsorbing virus from a spleen pulp emulsion on an aluminium gel.
We have since made modifications which have given better results. In the first place, since
different brews of gel, prepared by the method previously described, did not give constant
results, we adopted the following technique, mainly according to Willstatter, Kraut and
Erbacher [1925]:
One hundred c. c. of 10 per cent ammonia solution (accurately prepared) were poured
into 600 c. c. of water at 63°C. containing 22 gm. of ammonium sulphate and the tempera-
ture was quickly brought to 58°C. The solution was stirred vigorously, and 150 c. c. of
water at 58°C. containing 76.7 gm. of ammonium alum (Baker Chemical Co.) were poured
in rapidly as one lot. The mixture was shaken in a stoppered bottle for 15 minutes at
60°C. , care being taken to keep the temperature constant during the process. It was then
transferred to a tall jar and was washed each time with about five litres of distilled water
by decantation and not by certrifugation. At the fourth washing, 80 c. c. of 20 per cent
ammonia solution were added to the wash water, before it was used. The washings were
repeated about 20 times, until the supernatant fluid no longer became clear. The last
t wo or three washings occupied several days. The final suspension of 1000 c. c. was left at
room temperature for at least two months before use.
Vaccines prepared from such gels have not varied greatly. It appeared, however, that
the spleen emulsion introduced a very variable factor into the manufacturing process, and
it was, therefore, decided to use instead of spleen allanto-amniotic fluid from hens' eggs, the
fluid being collected 48 hours after the inoculation of 9 or 10 day embryo. This fluid has a
more constant virus content and is much cleaner to work with. Using this method, the
following experiments were tried : (1) diluting the virus before adsorption ; dilutions even at
1/ 10, however, greatly reduced the immunizing capacity; (2) using eluates of prepared vaccine,
-these also were unsatisfactory ; (3) reducing the concentration of formalin,—but concentra-
tions below 1/2000 were not safe ; (4) reducing the incubation period but using formalin at
1/2000 periods below 24 hours were not always safe and ; (5) reducing the dose of vaccine,
—0. 5 c. c. to 1.0 c.c. is satisfactory for adult birds, young chickens can be immunized
satisfactorily with 0.2 c. c. Table I is an example of some experiments on these lines.
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