Dec. 15,1924 Differentiation of Bact. melitensis from Bad. abortus 587 
colonies, one from heart sterile. The 
organisms seem to have multiplied 
somewhat more rapidly in all cases 
under normal atmospheric conditions 
than in air partially displaced by C0 2 
gas. 
Guinea pig No. 77013. Inoculated 
both intra-abdominally and intra- 
testicularly The inoculated testicle 
showed a slightly abnormal appearance, 
probably at site of injection. An area 
about one-fourth inch in diameter was 
darker in color than the surrounding 
tissue. No other lesions were noted. 
The blood serum of this pig showed a 
titer of 1 to 50 with a Bacterium meli¬ 
tensis antigen and below 1 to 25 with 
a Bact. abortus antigen. Spleen and 
affected testicle cultured. Cultures 
were subjected to the same conditions 
as those of guinea pig No. 77012. On 
January 2, two cultures incubated 
under CO 2 conditions from spleen 
showed from 100 to 200 colonies; two 
tubes from testicle were sterile. The 
colonies developing under normal at¬ 
mospheric conditions were slightly 
larger than those incubated in a 10 per 
cent CO 2 atmosphere. 
Guinea pig No. 77014, inoculated 
into both testicle and abdominal 
cavity, and guinea pigs Nos. 77016 and 
77107, which received the material 
intra-abdominally only, showed no le¬ 
sions. Their blood serum did not 
agglutinate suspensions of either Bact. 
abortus or Bact. melitensis in dilutions 
of 1 to 25 or higher. Cultural results 
were also negative. 
The colonies obtained in cultures 
from the two positive guinea pigs Nos. 
77012 and 77013 closely resembled 
those of Bacterium abortus, but failed 
to attain, in four days, the size com¬ 
monly reached by such colonies on 
serum-agar slants when incubated in 
moist jars containing 10 per cent C0 2 gas. 
A few of the larger colonies were ap¬ 
proximately one-sixteenth inch in diam¬ 
eter. In most cases they were consider¬ 
ably smaller. On January 16 subcul¬ 
tures were made from the original 
spleen and testicle cultures on 3 per 
cent glycerin-agar slants. Slight growth 
was visible on the day after incubation 
under normal atmospheric conditions. 
On the second day the growth was 
reasonably heavy. Antigens prepared 
from subcultures of the organisms 
isolated from guinea pigs Nos. 77012 
and 77013 were agglutinated in a char¬ 
acteristic manner by Bact. abortus and 
Bact. melitensis antiserums. 
Efforts failed to isolate Bacterium 
melitensi from the blood of the 14-year- 
old boy by cultural methods. The 
serum-agar slants sown with different 
dilutions of the blood were heavily 
overgrown with contaminating organ¬ 
isms within a few hours. 
On December 14, through the kind¬ 
ness of Doctor Heck, blood specimens 
were obtained from four of the goats, 
Nos. 96, 26, 61, and 19, exposed to the 
original reacting flock. Milk samples 
were also secured from three of these. 
The agglutination titer of the blood 
serums of goats No. 96, 26, and 61 
with a Bacterium melitensis antigen was 
1 to 1,000; goat 19, 1 to 500. The milk 
whey of goat No. 96 was negative; 
goat No. 26, titer 1 to 1,000; goat No. 
19, titer 1 to 100. 
Different dilutions of the blood and 
milk were sown on serum-agar slants 
and incubated under CO 2 and normal 
atmospheric conditions. On some of 
the tubes there developed colonies 
bearing rather close resemblance to 
those of Bacterium abortus and Bact. 
melitensis; but subcultures of these 
failed to produce antigens that were 
agglutinated by Bact. abortus and Bad. 
melitensis antiserums. 
Four guinea pigs received 2 c. c. 
of milk each from three of the goats 
and the same number received sus¬ 
pensions of the blood of each blood 
sample. The milk was somewhat pu¬ 
trid on arrival. All the guinea pigs 
receiving the milk of goats Nos. 19 and 
96 were dead in less than 48 hours. 
Guinea pig No. 76595, inoculated 
with the blood of goat No. 61, died 
December 22. Cultures from spleen, 
liver, and kidneys remained sterile. 
Guinea pig No. 77411, inoculated • 
with the blood of goat No. 26, died 
December 30. Cultures from the liver 
and spleen remained sterile. 
Guinea pig No. 77496, inoculated 
with milk from goat No. 26, died 
December 31. Cultures from liver 
and spleen developed colonies of 
Bacillus coli; no Bacterium aborts-like 
colonies were detected. 
Guinea pig No. 77497, inoculated 
with the milk of goat No. 26, was 
destroyed January 29. Spleen was 
three times normal size. Liver showed 
evidence of fatty degeneration. Titer 
of blood serum to Bacterium melitensis 
antigen was 1 to 200; to Bact. abortus 
antigen, 1 to 200. 
Guinea pig No. 77498, inoculated 
with the milk of goat No. 26, was de¬ 
stroyed January 29. Spleen was 
slightly enlarged. Right epididymis 
showed evidence of necrosis. Titer of 
blood serum to both Bacterium melitensis 
and Bact. abortus antigens was 1 to 
500. Serum-agar slants, 3 per cent 
glycerin-agar slants, and infusion bouil¬ 
lon were used for culturing the spleens 
and testicles of guinea pigs Nos. 
77497 and 774S8. 
