196
Bovine on Pleuropneumonia
nutrition of the part, thus inhibiting any potential fibroblastic activity. As we could not correlate
these lesions with any known pathological condition of the spleen of cattle, and as the lesion is strictly
comparable with the classic pleuropneumonia lesions elsewhere described, there is apparently consider-
able justification for describing the lesion as one not hitherto recorded in studies in pathogenesis.
That the spleen lesion contains the virus was shown by the fact that two hill cattle subcutaneously
inoculated with a saline extract of spleen lesion reacted as if inoculated with fresh lymph virus and
succumbed. Two hill cattle each inoculated with a large dose (100 c.c.) of a Berkefeld V filtrate of
the same extract failed to react, but both proved immune to a later immunity test in which the control
reacted typically and died.
In thirty four of forty eight Kumaoni cattle, i.e. 71 per cent of the series of cattle subcutaneously
inoculated with Assam 1 virus, one of the main lesions has been a profound oedema and occasionally
ulceration of the mucous membrance of the abomasum, the folds being grossly swollen with a clear,
amber-coloured exudate clotting rapidly on removal. In a few cases, other parts of gastro-
intestinal tract, especially the large intestine, have been likewise affected. When the rectum is
involved, there is usually a tendency for ulceration and haemorrhage. The lesions are frequently
accompanied by a more extensive sero-fibrinous infiltration of the adventitious coat. At Izatnagar,
these lesions were found in four of twenty i.e. 20 per cent cattle inoculated subcutaneously with
the Assam 1 virus. The presence of the virus in the œdema fluid is suggested by the observation
that in three instances, subcutaneous inoculation of hill cattle with œdema fluid resulted in typical
swellings, fever and death in all of six cattle tested, three cattle presenting abomasal lesions at
post-mortem. Two animals of the series showed, at post-mortem, pharyngeal diphtheresis with
ulceration, the lesions resembling those seen in rinderpest. A small fragment of the diphtheritic
membrane of one case was emulsified in saline, and 10 c.c. of the extract inoculated subcutaneously
in a hill bull; the reaction that followed was as severe as that resulting from the inoculation of pure
lymph virus.
In the series of twenty-five hill cattle inoculated subcutaneously in the maintenance of the
Australian 264 strain virus, splenic lesions were found in three animals, i.e. 13 per cent of animals
used, but were not so well defined as in cattle of the Assam 1 virus series. Abomasal œdema was a
fairly frequent lesion, being seen in eight cases, i.e. 34 per cent of animals used. No animal of the
Australian 264 series developed lesions of pleuropneumonia.
ISOLATION AND STUDY OF THE PLEUROPNEUMONIA ORGANISM
(A) Filtration
Lymph virus, whether pleural exudate or local lesion exudate, maintains a constant pH-7.4,
even when stored for some months at +5ºC. It was , at first, thought advisable to dilute liberally
and to buffer M/15 phosphate) all samples of exudate to be filtered. It, however, became evident
later that working with porcelain candles buffering was unnecessary, and that dilution of the exudate,
while convenient in facilitating filtration was not essential; provided a preliminary coarse filtration
through gauze was done to remove particulate matter, undiluted lymph virus could be filtered
through coarse candles, such as Berkefeld V, with comparative ease. In some exudate samples that
were rather firmly clotted, it was necessary to break down the clot with saline. Usually, however,
sufficient fluid exudate could be obtained by agitation and removal of the clot. To facilitate filtra-
tion and to obtain adequate quantities of filtrate, the exudate was subsequently diluted 1/5 saline.
The richness of a Berkefeld V filtrate, as shown by seeding a drop of the filtrate over the surface of a
tube of Bennett's serum agar, indicated that there was little tendency for organisms to concentrate
in the clot as compared with the more fluid portion of the exudate. Ten c.c. lymph virus diluted
1/5 saline passed a Berkefeld V filter in just under 10 minutes at a negative pressure of 40 mm. mercury.
Six cattle, each subcutaneously inoculated with 16 c.c. of the filtrate, tested for freedom from bacterial
contamination before inoculation, reacted strongly and died. Spleen lesions were seen in two of
these animals at postmortem. An equally severe reaction was produced in two cattle subcutaneously
inoculated each with 225 c.c. Berkefeld V filtrate of lymph virus diluted 1/15. Fresh lymph virus
was diluted 1/5 saline, centrifuged and the supernatant filtered through a Berkefeld N filter at a