Sept. 22, 1923 
Bacterial Leafspot 0} Clovers 
477 
of the stroke to the contoured margin (PI. 6). No odor is developed, and 
the agar is unchanged. 
Gelatin plates. —On gelatin plates visible growth was slow, but after 
five days the colonies were 3 to 4 mm. in diameter with a smooth surface 
and entire margin. They were finely granular within, especially toward 
the center. The organism is not capable of liquefying the substratum. 
Gelatin stabs. —Best growth occurs at the surface in stab cultures, 
with neither liquefaction nor discoloration of the medium. 
Potato cylinders. —On steamed potato cylinders growth is first 
manifest by a faintly yellowish white, spreading streak. It becomes 
abundant within six to eight days, but remains flat, grayish, and gelat¬ 
inous in consistency. The cylinder along the line of growth becomes 
smoky gray. No marked dissolution of the potato tissue occurs, and the 
potassium iodid test indicates a weak diastasic activity only in 3-week- 
old cultures. 
Milk. —Plain sterilized milk turns creamy in color after two weeks; af¬ 
ter five weeks a soft curd forms which slowly separates into whey and a 
rather firm curd which was not digested at the end of four months. The 
whey becomes alkaline with litmus as an indicator. 
Litmus milk. —Lavender-colored litmus milk changes rather rapidly 
through dark plumbago blue at the end of 5 days, deep Dutch blue at 
the end of 10 days, to light Tyrian blue at the end of 2 weeks. Curd be¬ 
gins to form soon afterwards and becomes dull tan, whereas the whey is 
dark blue. 
Methylene blue in milk. —Decolorization had been completed in 
two weeks, but slowly returned with the separation of curd .and whey. 
The color reappeared in the whey. 
Blood serum. —Stroke cultures showed in two to three days’ mod¬ 
erate growth, spreading, flat, smooth, and glistening with an echinulate 
margin. No liquefaction occurred and no discoloration of the medium 
was noted. 
Synthetic media. —Cohn’s solution does not appear to support 
growth. 
In Fermi’s solution it soon develops turbidity, and after three days 
sufficient growth has taken place to produce a milky white cloudiness. 
After five days a thin viscid pellicle will have formed. The medium 
gradually takes on a bluish green fluorescence. 
In Uschinsky’s solution two or three days’ growth results in a milky- 
white cloudiness. Delicate, flocculent pellicle-like growths appear at the 
surface. The fluorescence which gradually develops is not so marked as 
in Fermi’s solution. 
Digestion of casein. —Poured plate cultures in casein agar after a 
week’s incubation had developed colonies 5 to 6 mm. in diameter. When 
these cultures were tested by flooding with a 1 per cent solution of hydro¬ 
chloric acid to precipitate the casein, it was found that a narrow zone 
surrounding each colony remained transparent, whereas the remainder of 
the plate was milky white. This indicates that the organism is able to 
digest the casein in the zone immediately surrounding the colonies. 
Ammonia production.— Cultures in peptone broth and beef extract- 
peptone bouillon 7 days old showed ammonia to be present when tested 
with Nessler’s reagent. 
Reduction of nitrates. —Tests were made with Trommsdorf’s 
reagent in tube cultures containing 2 per cent peptone broth to which 2 
