MUKTESAR, FOR THE TWO YEARS ENDING THE 31ST MARCH, 1924 31
the paraffin seal on cultures of this organism lay in its power to
prevent the escape of carbon dioxide gas from the culture fluid
and that oxygen was necessary for the growth of the organism.
Samples of rinderpest blood were then suspended at body temperature
in atmospheres containing varying proportions of carbon dioxide,
and the finding was made that the virus survived as long when
the tension of this gas approximated that of the circulating blood
as it did under the paraffin seal. The beneficial effects of the gas were
manifested throughout a wide range (from 1 to 10 per cent. of CO 2 in the
tests). Dilution had long been known to render the virus rapidly inert,
and in early tests at the laboratory it seemed that diluting the blood with
five times its bulk brought about destruction of virulence in about a
day at body temperature even under a paraffin seal. It was subse-
quently found, however, that the apparent rapid destruction was a
manifestation of accelerated action of the originally present
destructive agencies in the more highly diluted fluid, and that the
most serious destructive agency was an unfavourable hydrogen-ion
concentration of the diluting fluid. When the blood was suspended
in high dilution in a slightly diluted serum or in peptone solution
adjusted to near the hydrogen-ion concentration of the circulating
blood the period of survival was relatively long. Again, numerous tests
showed that there was no need to invoke the action of a hypothetical
lysin to account for the apparent rapid destruction of the virus in
blood exposed to air: the factor of greatest importance appeared
to be the transformation that took place in the blood owing to the escape
of carbon dioxide gas, and destruction of virus was rapid during this
transformation. In old stored serum, stabilized to the reaction of the
normal blood by addition of acid, the period of survival of the virus in
small amounts of added virulent blood was long. In work published by
Frosch and Dahmen upon the cultivation of the foot-and-mouth disease
virus long after these experiments were undertaken at Muktesar the
authors state that the source of difficulty in obtaining primary cultures
is the presence of a lysin in the vesicular lymph used as seed material.
Some work done by de Blieck and Winkel (1922) indicates also that the
chief deterrent action resides in the rapid change to an unfavourable
alkaline reaction that takes place in the serous fluid. Likewise, the virus
was very sensitive to an unfavourable reaction in the acid direction, such
as takes place during the autolysis of animal tissues. The addition of a
small fragment of sterile rabbit kidney, for example, to stored virulent
blood rapidly rendered it inert. Bacterial contamination usually exer-
cised little effect.
Differential high-speed centrifugation of hæmolysed virulent blood
suspended in bland fluids of various specific gravities, according to the
method published by MacCallum and Oppenheimer (1922) for the de-
monstration of the particulate nature of the small-pox virus, brought
about only with great difficulty any signs of sedimentation or flotation
of the virus ; nevertheless, the results were suggestive that the virus
was organismal and capable of separation, and not fluid.
c