DIPHTHERIA BACILLUS. 341 
medium employed to get by plating methods a pure 
culture from the original serum tube. The agar should 
be freshly melted and poured in the Petri dish for this 
purpose. After it has hardened the layers in a number 
of plates are streaked across with bacteria from colonies 
on the serum culture, which appear in size and color 
like the diphtheria bacilli. Other plates are made from 
a general mixture of all the bacteria, selected, as a rule, 
from the drier portion of the serum. The plates are left 
in the incubator for twelve hours at 37° C. In the 
examination of the plates one should first seek for 
typical colonies and then later for any that look nearest 
the characteristic picture. Diphtheria colonies are very 
apt to be found at the edges of the streaks of bacterial 
growth. 
Growth in Bouillon. The diphtheria bacillus usu- 
ally grows readily in broth slightly alkaline to litmus. 
The characteristic growth in neutral bouillon is one 
showing fine grains. These deposit along the siles and 
bottom of the tube, leaving the broth nearly clear. A 
few cultures in neutral bouillon and many in alkaline 
bouillon produce for twenty-four or forty-eight hours 
a more or less diffuse cloudiness, and frequently a film 
forms over the surface of the broth. On shaking the 
tube this film breaks up and slowly sinks to the bottom. 
This film is more apt to develop during the growth of 
cultures which have long been cultivated in bouillon, and 
indeed after a time the entire development may appear 
on the surface in the form of a friable pedicle. The 
diphtheria bacillus in its growth causes a fermentation 
of the meat sugars and the glucose, and thus changes 
the reaction of the bouillon, rendering it distinctly less 
alkaline within forty-eight hours, and then, after a vari- 
