246            STUDIES ON SALMONELLA ABORTUS      [Vol. XXV, Part III,

generally disappeared in three or four days. The temperature-rise with the brews
of 1949 was in no way more severe than that in the previously recorded ones, nor,
was the swelling more pronounced.

These symptoms were observed in experimental ponies following an injection
of 20 c.c. of the vaccine, a flat dose employed for conducting safety test in the labo-
ratory before issuing the product for use in the field. The previous brews had also
been tested at this high dose. However, when the dose was reduced to the one
employed in the field and adult healthy ponies in good condition were tested, very
mild reaction was produced, which could hardly be noticed without very careful
observation. Nevertheless, this was considered a serious disqualification for the
vaccine and work was taken up to modify it so as to render it absolutely safe.

Formol-killed vaccine: In the first instance, additon of formalin was tried with
a view to detoxicating it, and three brews were prepared by emulsifying the growth
of three strains of Salmonella abortus equi, S.128, S.129, S.130, in normal saline
solution to an opacity equal to B.W. 4 and treating with formol in the concen-
tration of 0.25 per cent for seven days at 37°C. This, however, did not bring about
the desired effect for, when injected into a mule which had never received a dose of
Salmonella abortus equi vaccine earlier, all the previously observed symptoms were
noticed. But when tested on a very large number of ponies, it was found that
formalin-treated vaccine produced comparatively mild symptoms.

Formol-killed and alum precipated vaccine: Since even formol-killed vaccine
had not given very satisfactory results, it was considered advisable to add to the
formol-treated vaccine potash alum in the concentration of 1 per cent in order to
precipitate the toxins, so that absorption from the site of inoculation would be
slowed down. Three brews were thus prepared and tested for safety on nine ponies,
each brew being given subcutaneously to three ponies at the rate of 20 c.c. per animal.
The experimental ponies were divided into three groups such that each lot contained
two ponies, which had received the Salmonella abortus equi vaccine previously, and
one pony, which had not been vaccinated at all or at least not during the last
10 years or so. Not a single pony showed any of the untoward symptoms of
toxaemia observed earlier but all excepting one developed abscesses at the site of
inoculation, which even after surgical treatment took a very long time to heal.
This was considered even a more serious disqualification for the vaccine than to have
produced toxaemic symptoms in a few ponies, as the horse-owners would never
allow the use of a product, which would produce unsightly abscesses at the site of
inoculation. In order to overcome this serious drawback of the newly-evolved
vaccine, a change in the route of administration was tried and the same brews in
dosage of 20 c.c. were given intramuscularly at two or three places in the same
ponies. Neither any toxaemia nor abscess formation occurred in the ponies.

Four more brews were prepared by treatment with 0.25 per cent formalin at
37°C for seven days and by the addition of potash alum in 1 per cent concentration
and each brew was tested on two ponies (one, which had received the vaccine pre-
viously and the second that had not been vaccinated during the past 10 years or so)
by the intramuscular route and in not a single animal treated in this manner any
untoward effect was noticed. The vaccine was also given field trials in a number of
stud-farms and from nowhere any adverse report was received. Since then only
formol-killed, alum precipitated vaccine is being issued for field use from the Institute
with very satisfactory results.