AGf~ICULTURAL RESEARCH, PUSA.

312 PRACTICAL BAC'l'EIUOLOGY
DIAGNOSIS
Films arc prqmred frum the pus, discharge from
sores, etc., or from nodules ill internal organs found at
post-mortem; these arc stained with methylene blue
and by Gram's method. '1'he appearance of beaded
Gram-negative organisms corresponding to B. mallei
:is suggestive. It :::;hould be noted that glunucr:::; bacilli
arc not easily demonstrable in preparations from the
lesions.
Cultures arc also made on glycerol-agar, and if a
mixeu growth results, pure cultures are obtained from
single colonies. 'rho chocolate-coloured growth on
potato is an important criterion in identification.
In all cases the nature of the infection must be
confirmed by animal inoculation. A male guinea-pig
is injected intraperitoneally' with the pathological
matcrial or the culture isolated; in two or three days
an enlargement of the testis results, and the animal
subsequently dies, showing the lesions of acute
glanders (vide 8upm). If pyogenic organisms are
also present in the material inj eeted, and a septic
peritonitis results, the glanders bacilli will be found
more numerous in the tunica vaginaEs. If the inoculum
contains a large number of other organisms it may be
introduced by subcutaneous injection.
Agglutination Test.-In the diagnosis of glanders
in horses, the agglutination test has been used. The
technique followed is that described 011 pp. 10G-199,
with certain minor differences. The bacterial suspension
is prepared from young glycerol-agar slope
cultures which, before emulsification, are placed at
60° C. for two hours to kill the organisms. For personal
safety it is necessary to use dead cultures. SerulU
dilutions ranging from 1 in 100 to 1 in 4000 are tested.
The agglutination tubes are incubated for twenty-four
hours at 37° C. before readings are made. N onnal horse
serum may agglutinate the bacillus ill low dilutions,
and only agglutination in high titres, 1 in 1000 or more.
can be accepted as proof of infection. Agglutillation in