102 Journal of Agricultural Research voi. vm,N0.3 
growth and more constant results were obtained than with that made 
from lean beef and freed from fermentable, carbohydrates by use of B. coli . 
All chemicals employed in making media or testing reactions were 
either chemically pure or were of the highest purity obtainable. In 
addition to Liebig’s extract, Witte’s peptone, “Gold label” gelatin or 
Nelson’s photographic gelatin No. 1 were used, the latter entirely during 
the last of the work. For bouillon 5 gm. of Liebig’s meat extract and 
10 gm. of Witte’s peptone were used for each liter of distilled water. To 
this were added 15 gm. of agar shreds or flour, or 100 gm. of gelatin if a 
solid medium was desired. No sodium chlorid other than that contained 
in the meat extract was. added. All media unless otherwise specified 
were made neutral to phenolphthalein. Distilled water was used for all 
culture media and Griibler’s dry stains formed the basis for all staining 
liquids. 
Tubes of ordinary bouillon and agar were sterilized by heating once 
in an autoclave for 15 minutes at from 3 to 6 or 7 pounds’ pressure. 
All vegetable media or media made from vegetables or containing sugars 
were sterilized by flowing steam, for tubes, 15 to 20 minutes at from 
99 0 to ioo° C. on each of three consecutive days. In the earlier part of the 
work gelatin was also sterilized by the discontinuous steaming method. 
Later it was found that it was less likely to become liquid at ordinary 
room temperatures if the tSubes were sterilized by one exposure in the 
autoclave to steam under 5 to 7 pounds’ pressure for 15 minutes. 
At the beginning of the comparative work with the assembled cultures 
each was transferred to a fresh potato-broth culture every 24 hours 
for several successive days and then gelatin plates were poured from 
the last potato-broth culture. From these plates transfers were made 
from single colonies to tubes of sterile bouillon to furnish fresh 
subcultures with which to work, after proving that the subcultures 
thus obtained were pathogenic. This was to insure that the cultures 
were in as near a uniform condition of vigor as possible before beginning 
to work. Attempts were made to revive in this way the pathogenicity 
of the two strains which were received as B . phytophthorus , but without 
success. A separate series of stock cultures of each strain which had not 
been put through this revivifying process were kept in reserve. 
All transfers to media to test cultural features were made from young 
broth cultures about 48 hours old, when the cloudiness had reached its 
maximum density. For stab cultures to agar and gelatin a straight 
platinum needle of approximately 0.5 mm. in diameter was u3ed. All 
transfers to liquid media were made with a 2-mm. loop of the same 
sized wire. Unless otherwise specified the cultures were incubated at 
20 0 C. In making records of the detailed features of the morphology, 
cultural, physical, and biochemical features, etc., of the various organ¬ 
isms the descriptive chart or card adopted by the Society of American 
Bacteriologists in December, 1907, was used as a model and guide. 
