go DETERMINATION OF SPECIES. 
in the hanging-drop, preferably in a fresh medium 
inoculated with a very small amount of the culture 
to be examined. It is desirable that the form and 
grouping be determined in bouillon, gelatin, and 
on agar, and that any variation found upon the 
examination of the growth on other media be accu- 
rately noted. 
(b) Test for Motility.—For the study of motility 
the hanging-drop preparations should be made from 
young cultures grown at or near the optimum 
temperature for only a few (six to eighteen) hours. 
(c) Tests for Spores.—The tests for the presence 
of spores are : 
' (a) Do colonies develop from cultures which have 
been subjected to a temperature of 80° C. for ten 
minutes ? 
(6) Are there highly refracting bodies within the 
bacteria in unstained preparations, and can they be 
demonstrated by the spore-staining methods?. ~ 
Cultures to be tested should be grown for forty- 
eight hours in bouillon, and when possible at 
36°-38° C. Three loopfuls of this culture’ after 
agitation are transferred to tubes containing 10 c.c. 
of bouillon. This is exposed to a temperature of 
80° C. for ten minutes, and then placed under con- 
ditions favorable for the development of any of the 
organisms which may have survived. . 
(d) Pleomorphism.—In regard to pleomorphism, 
attention is called to the variations in size and shape 
brought about by the following conditions of 
growth : 
(a) At: different temperatures, 
