489 
Nov. 15,1924 A Bacterial Leaf spot of Martynia 
brown, then the agar became light 
brown in the upper half and finally after 
2 months light brown throughout. A 
small amount of H 2 S was produced. 
Relation to oxygen. —Growth was 
observed in fermentation tubes con¬ 
taining 1 per cent peptone and 2 per 
cent sugars from which the oxygen 
had recently been driven off by heating. 
At the end of a week there was light 
clouding in the closed arms of tubes 
containing saccharose, dextrose, and 
mannit, but not in tubes containing 
maltose, lactose, and glycerin. In 
agar shake cultures containing 2 per 
cent sugars a few small colonies grew 
in the agar for the depth of half a centi¬ 
meter in saccharose and dextrose only. 
The organism is aerobic. 
Diastasic action. — Streaks were 
made on beef peptone agar plates con¬ 
taining 0.2 per cent starch. Tested 
the second day with a saturated solu¬ 
tion of iodine in 50 per cent alcohol 
there was no clear zone around any of 
the colonies. 
Tested after 8 days there was still 
no cleared zone about the streaks. In 
some cases a zone 4 to 5 mm. wide was 
slightly more reddish than the sur¬ 
rounding agar. Twelve plates were 
tested and on one a fungus contamina¬ 
tion had a definite cleared zone about 
it. There is very little diastasic action. 
Milk. —There is evidently a deli¬ 
cate balance between the separation 
or nonseparation of whey and curd. 
When separation occurs the curd is evi¬ 
dent after 2 days but is always soft. 
Litmus milk shows very slight acid re¬ 
action remaining about the same color 
as the checks with a slight reddish 
tinge. A test for rennet gave negative 
results. Cultures were heated for a 
half hour at 55° C. and then a few cubic 
centimeters poured into sterile milk. 
There was no separation of curd and 
whey when all the organisms had been 
killed. The curd is evidently an acid 
curd. Peptonization becomes evident 
in 2 to 3 days, progressing in definite 
bands, the clearest at the top. Pep¬ 
tonization is completed in about 2 
weeks. Reduction of litmus begins in 
about 5 days and is complete in 8 to 14 
days. When cultures containing re¬ 
duced litmus are killed by boiling, the 
color returns a deeper shade than the 
checks but neither red nor blue. Re¬ 
duction of methylene blue in milk is 
completed in 4 to 5 days. The blue 
color returns after the organism has 
been killed by heating the reduced cul¬ 
ture. After several months milk turns 
amber to deep brown and becomes 
gelatinous or has gelatinous lumps sus¬ 
pended in the liquid. 
Nitrate reduction. —No gas is 
produced. Two tests for nitrites were 
made in nitrate broth cultures with 
sulphanilic acid and a-naphthylamine 
in 5 N acetic acid. Cultures showed 
good growth. Results were negative 
during the first few days but showed 
definite reduction after 2 to 3 weeks. 
Two tests for nitrites in the synthetic 
nitrate medium KNO3 1 gm., K2HPO4 
0.5 gm., CaCl 2 0.5 gm., glucose 10 gm., 
distilled water 1,000 c. c. gave positive 
results. Reduction of nitrates is evi¬ 
dent in this synthetic medium in 24 
hours. There was no reduction in the 
checks. 
Fermentation. —Tests were made 
in media containing 1 per cent peptone 
and 2 per cent carbohydrate or one-half 
per cent peptone and 1 per cent carbo¬ 
hydrate. Gas was not formed. Hy¬ 
drogen-ion concentrations were deter¬ 
mined colorimetrically and the results 
are given in Table I. Lactose showed 
no acid reaction, although tests were 
made after 1, 2, 3, 5, 6, 10, and. 16 days, 
nor were acids detected from raffinose, 
maltose, mannit, rhamnose, or glycerin. 
Acid is produced from dextrose, sucrose, 
galactose, and arabinose. 
Litmus sugar agars .—Litmus 
showed reddening in dextrose, galac¬ 
tose, and arabinose after 24 hours; 
in sucrose in 3 days; in levulose a 
brownish red in 4 days. Mannit also 
showed a trace of this browning after 
3 days, and when a month old all tubes 
were slightly redder than the checks. 
Lactose and maltose were bluer than 
the checks at the end of a week, and 
in 11 days the upper parts of cultures 
of arabinose were turning blue. In 
from 11 to 20 days partial reduction 
occurs in sucrose, maltose, levulose, 
galactose, arabinose, and glycerin. 
EXPLANATORY legend for PLATE 3 
A —Culture I, 2-day colonies on beef peptone agar. Photographed Mar. 9, 1923. X 10. 
B. —Culture II, 3-day colony on beef peptone agar. Photographed Mar. 10, 1923. X 10. Showing the 
thin margins of young colonies. Transmitted light. Also buried and bottom colonies. 
C. —Culture II, 8-day colonies on beef peptone agar. Photographed Mar. 15, 1923. X 10. Showing 
depressed centers. Reflected light. 
D. —Culture II, 5-day colonies on beef peptone agar. Photographed Mar. 12,1923. X 10. Transmitted 
light. Showing thin depressed centers and raised heavier borders. 
E. —Culture I, 1-day beef peptone agar. Casares-Gil stain. Dec. 15, 1923. X 1900. 
F. —Culture I, smear from 8-day beef peptone agar slant. Ribbert’s capsule stain. Chains. Photo¬ 
graphed Jan. 22, 1924. X 1400. 
G. —Culture I, 2-day beef peptone agar cultures. Casares-Gil stain. Capsules. Photographed Jan. 
24, 1924. X 1900. 
