SPIRILLUM CHOLERZ ASIATICZ. 587 
Plan of Procedure for the Biological Diagnosis of the 
Cholera Vibrio. A. Dejecta (fluid) or intestinal con- 
tents of a cholera patient or cholera suspect. 
1. Use one drop (one platinum loop) for gelatin plate- 
cultures, making two dilutions. Do this in duplicate 
or triplicate. Cultivate at 22° C. 
2. Inoculate a couple of bouillon tubes and a couple 
of Dunham’s 1 per cent. peptone solution with one drop 
each, and place them in the incubator (37° to 38° C.) 
for six to eight hours. 
3. Examine a drop of the dejecta in the hanging drop. 
4, Examine a drop of the dejecta in stained cover- 
glass preparation.’ 
5. Make gelatin plates from one drop taken from 
the surface of each of the bouillon and peptone solution 
tubes and cultivate at 22° C. 
6. As soon as the plates (see 1 and 5) are sufficiently 
developed (thirty-six to forty-eight hours) fish the 
suspected cholera colonies and use the material for the 
following procedures : 
7. Inoculate six or eight peptone tubes (1 per cent. 
peptone, 0.5 per cent. NaCl in distilled water) and 
place them at once in the incubator. Note the time. 
8. Examine hanging drop for form, size, and motility 
(and arrangement). 
9. Make stained cover-glass preparations and exam- 
ine. 
1 These direct microscopical examinations of the intestinal contents are, as 
a rule, very unsatisfactory, at least in those in which the symptoms are not 
marked. Ina few the spirals will make up from 50 to 100 per cent. of the 
bacteria present. In most of the cases during the last epidemic in New York 
Dunham found abundance of columnar epithelium from the intestinal mucous 
membrane, numerous straight, thick bacilli, and only a few curved bacilli or 
segments of spirals—too few to identify. Plate cultures from these showed 
from 20 to 80 per cent. of all the colonies developing to be cholera spirilla, 
