854 
Journal of Agricultural Research 
Vol. XXXI, No. 9 
In order to determine more accurately the changes in true acidity in these media, 
three series of six test tubes each were made up, one series containing brom 
cresol purple, one brom thymol blue, and the other phenol red in a concentra¬ 
tion of 0.0016 per cent. These media were adjusted to about Ph 7.3 as indi¬ 
cated by the blue-green color of brom thymol blue and the red of the phenol 
red. The dextrose and saccharose cultures with brom thymol blue and phenol 
red became yellow (more acid) seven days after inoculation. All the other 
cultures slowly became less acid than the controls. There was, therefore, acid 
production from dextrose and saccharose, but -none from the other carbon 
sources. Probably the decrease in true acidity was due to ammonia produced 
from the peptone. 
Agar with sugars. —In litmus-dextrose, litmus-maltose, and litmus-lactose 
agar slant cultures there was no evidence of acid production. In two days the 
maltose and lactose cultures were blue under the stroke, and in seven days all 
cultures were bluer throughout than the controls. However, in a later repeti¬ 
tion of this series, in which saccharose, mannite, and glycerin were also tested, 
a pink color developed in the dextrose and saccharose tubes, indicating acid 
production. 
To obtain more accurate information on this point, the three sulphone 
phthalein indicators, brom cresol purple, brom thymol blue, and phenol red, 
were used in a triplicate series of slant cultures with dextrose, maltose, and 
lactose. These media were adjusted to a P H of about 7.0 as evidenced by the 
grass-green color of brom thymol blue. In the dextrose cultures containing 
brom cresol purple and brom thymol blue a yellow color developed, indicating 
acid production. No increase of hydrogen-ion concentration was indicated at 
any time with the other two sugars and most of the cultures became slightly 
more alkaline. In a later repetition of this test, saccharose, mannite, and glycerin 
were also included and distinct acid production occurred only in the dextrose 
and saccharose cultures. Parallel series of cultures of Bacillus coli produced the 
yellow-acid color with all of the indicators. These tests show that there is acid 
production from dextrose and saccharose. 
Action on starch. —There were no signs of diastatic action on either potato 
or corn starch. No halos or cleared zones appeared about the colonies in plates 
of beef agar to which starch was added. 
Tests for indol, skatol, and ammonia. —Cultures in beef-peptone bouillon 
gave no test for indol at 7, 26, and 33 day intervals when tested with potassium 
nitrite and sulphuric acid and no test for skatol when tested with nitric acid and 
potassium nitrite. A positive test for ammonia was obtained after 7 and 33 
days with Nessler’s reagent. 
Fermi’s solution. —Growth occurred in Fermi’s solution, accompanied by 
the formation of a greenish-yellow pigment throughout the medium. 
Uschinsky’s solution. —Good growth occurred in Uschinsky’s solution in the 
case of two of the three strains tested, and a greenish-yellow pigment was 
produced. 
Cohn’s solution. —No growth occurred in Cohn’s solution. 
Blood serum. —The growth in stroke cultures on plain solidified blood serum 
in 10 days was abundant, spreading, flat, smooth, and glistening, and showed a 
brownish tinge. A slight liquefaction of the medium along the stroke was noted 
after 17 days, and at the end of 40 days there was a general liquefaction of the 
upper part of the slant, and a brownish-yellow discoloration of the medium. 
On slants of Loeffler’s blood serum, growth was more rapid than on plain 
blood serum, and in two days was abundant, spreading, flat, and rugose. In 10 
days the medium showed a slight brownish color and slight liquefaction under the 
stroke. At the end of 40 days the medium was almost completely liquefied, and 
was clay color according to Ridgway’s chart (16). 
Toleration of sodium chloride. —In tubes of beef-peptone bouillon neutral 
to brom thymol blue, 4 per cent of sodium chloride was tolerated and 5 per cent 
inhibited growth. In a series neutral to phenolphthalein, 5 per cent of sodium 
chloride was tolerated. Evidence was obtained that the use of hydrochloric 
acid in adjusting the reaction increased the inhibitory properties of sodium 
chloride. 
Toleration of acids and alkalies. —Tubes of beef-peptone bouillon were 
adjusted to +30, +25, +20, +18, +15, +14, +12, +10, +5, 0, -5, -10, -15, 
— 20, —30, and —40 Fuller’s scale by the use of hydrochloric acid and sodium 
hydroxide. The three strains grew in the +10, +5, 0, —10, and —15 tubes, and 
two strains grew in the +12 tubes. In a later series growth occurred in the +15 
and —25 tubes. Growth seemed most vigorous in the +5 media. A greenish- 
yellow pigment was formed in the alkaline cultures. 
