134 
. Journal of Agricultural Research 
Vo!. XXI, No. > 
and in 20 days the liquid was semitransparent with a yellow precipitate 
in the bottom of the tube. The consistency of the liquid was unchanged. 
Litmus milk. —Lavender-colored litmus milk was completely decolor¬ 
ized in 10 days, but no pink color was produced. Therefore no aciditv 
was produced. In 20 days the blue color reappeared in the bottom of 
the tube, but the upper part of the liquid remained amber-colored. 
Clearing caused by digestion took place as described above, and a yellow 
surface film and precipitate were formed. 
Brom cresol purple in milk. —In accordance with modern bacterio¬ 
logical technic (2), the litmus milk tests were supplemented by the use 
of milk containing brom cresol purple in a concentration of 0.0016 per 
cent, which gives a distinct bluish color. This indicator changes from 
its alkaline purple color at a hydrogen-ion concentration of P„ 6.8 to its 
acid yellow color at P H 5.2 and affords a more reliable index of the changes 
in the true acidity of milk than does litmus. Twelve days after inocula¬ 
tion there was no change in color, but in 16 days the inoculated tubes 
were a deeper blue than the controls. At no stage did the color become 
lighter. The hydrogen-ion concentration in milk was therefore lowered 
by the growth of this organism. 
Methylene blue in milk.— Reduction of methylene blue ocurred. 
Decolorization began in 2 days, and after 5 days all the liquid except a 
surface layer 5 mm. deep was completely decolorized. At the end of 20 
days the upper layer of 20 mm. in depth was a light grass-green. 
Digestion of the milk occurred as previously noted, and a yellow preci¬ 
pitate was formed. 
Reduction of nitrates— In fermentation tubes containing 1 per 
cent potassium nitrate bouillon, a light growth occurred in the open arm, 
but there was no growth in the closed arm. No gas was formed. Tests 
with Trommsdorf’s reagent indicated that there was no nitrite formed. 
Additional tests were made with test tube cultures in 2 per cent Difco 
peptone water containing 1 per cent potassium nitrate. Good growth 
occurred, but no test for nitrites nor ammonia was secured at 4, 6, and 11 
day intervals. Therefore nitrates were not reduced. 
Carbon metabolism. —For fermentation tubes, a 2 per cent peptone 
solution was used as the base for six solutions made by adding 2 per cent 
of the following carbon compounds: dextrose, saccharose, maltose, 
lactose, glycerin, and mannit. Cultures were run in duplicate and in¬ 
cubated at 22 0 C. In 3 days there was heavy clouding in the open arm 
of each tube and no growth whatever in the closed arm. There was a 
sharp plane of separation between the clouded and the clear medium. 
In 10 days a yellow surface pellicle and yellow precipitate had formed in 
the open arm in each case. The plane of separation mentioned above 
was not quite so sharp, and in some cases where the connecting tube was 
of greater diameter a slight clouding was noted in the closed arm. No 
gas was produced in any of these tubes, whereas in the Bacillus colt 
