27 
A Stem Bught of Field and Garden Peas 
\ 
6. HYDROGEN SULPHIDE PRODUCTION. —No hydrogen sul¬ 
phide is produced. This was determined by means of both iron 
gelatin and nutrient broth. 
In the foiTner case, stab cultures were made in +io nutrient 
gelatin which contained 0.5% of iron potassiumi tartrate. No 
blackening took place along the line of the stab as the result of 
the formation of iron sulphide. 
In the second method of determination, tubes of nutrient 
broth were inoculated and held at 28° C. After 24 hours, strips 
of filter paper, moistened with lead acetate were suspended in the 
tubes above the fluid; these were remoistened each day for five 
days, but after fifteen days no blackening of the paper had taken 
place, due to the formation of lead sulphide. 
7. TOLERATION OF ACIDS. —The toleration of acids is slight. 
We have determined this point with relation to citric, malic, 
oxalic and tartaric acids by adding different amounts of these to 
neutral nutrient bouillon in quantity sufficient tO' give the broth 
the acidities +15, +30 and +45. Three tubes of each acidity 
were prepared from each acid. They were inoculated with one 
loop of a 24-hour broth culture and subsequently kept at 28° C. 
In 24 hours, the +15 tubes of all the acids were uniformly turbid, 
but none of the others gave any growth. After twenty days the 
results remained unchanged. 
Hydrochloric acid added to nutrient broth sufficient to give 
it a reaction of +27 inhibits growth. 
8 . TOLERATION OF ALKALIES. —So far as our studies go, the 
toleration of alkalies is even less than that of acids, altho this 
point has been determined for only one alkali, NaOH. Sodium 
hydroxide added to nutrient broth sufficient to give it a reaction 
—II inhibits growth. 
9. OPTIMUM REACTION. —The Optimum reaction for growth 
in nutrient broth appears to be about +8. This was determined 
by adding to 20 c.c. portions of sterile broth, with aseptic pre¬ 
cautions, sufficient normal NaOH or HCl as the case required, to 
give a series of broths ranging from +41° to —17° and differing 
from each other by 2 degrees. The contents of each flask were 
distributed into four sterile tubes by means of sterile pipettes, and 
without further sterilization, three of each set were inoculated with 
one loop of a 24-hour broth culture. The fourth tube of each 
set was kept uninoculated as a check against possible contamina¬ 
tion, but in no case did any of these develop growth. The inocu¬ 
lated tubes were kept at 28° C. 
After 24 hours, growth was present from -f-21 to — 5, in 48 
hours, it appeared in —7, and in 72 hours in +23, in 7 days in —9. 
