Feb. 10, 1923 
A Bacterial Leaf spot of Tobacco 
487 
Stained by the Caesar-Gil and modified Pittfield methods for flagella 
from 24-hour-old cultures, the organism shows from one to several 
polar flagella, usually two or three, but as many as seven have been 
counted. The flagella are ordinarily from three to five times as long 
as the organism itself.. The organism has been stained with carbol 
fuchsin, methylene blue, and anilin gentian violet. No spores or involu¬ 
tion forms have been observed. Capsules are formed. Pseudozoogloeae 
are apparently absent. It is Gram-negative and is not acid-fast. 
Potato-dextrose agar. —Most of the cultural work has been done on 
potato-dextrose agar, as this had been found to be most useful for rapid 
comparative purposes on account of the color imparted to the agar. 
Colonies in agar plates are first visible in about 36 hours at about 22 0 to 
26° C., increasing to 3 to 5 mm. in diameter in six days. Colonies are 
at first grayish white, changing on most media on about the third day to 
a distinct yellow, after which a transparent light yellow tinge develops 
in the potato agar. The colonies are round, shining, convex, and yellow 
with opaque centers. The submerged colonies are lenticular. On agar 
slants a distinct growth may appear along the line of inoculation in 24 
hours, and this broadens gradually at the base, becoming echinulate 
(PI. 4 A). The growth becomes fairly heavy in 3 to 5 days but rarely 
covers the slant, usually developing a fairly deep orange color. Certain 
potato-dextrose agar media will color up in a few days, but ordinarily 
more gradual coloration occurs, the media becoming usually a bright 
honey yellow, which may extend to a depth of 2 inches or more from the 
growth down into the tube. Where the pigment is rapidly absorbed by 
the agar, the growth does not take on a deep color. Most of the yellow 
bacterial plant pathogenes known to science have been cultivated 
simultaneously, but with none has this charactristic of yellowing potato- 
dextrose agar been nearly as distinct, and with the majority seemingly 
it does not occur. The pigment on potato-dextrose agar is soluble in 
water, sulphurous acid, ammonia, methyl and ethyl alcohols, and hydro¬ 
gen peroxid. It is apparently destroyed by hydrochloric acid, tolulene, 
xylol, benzol, and carbon disulphid. Its production is slowed up mark¬ 
edly at io° C., but between 20° and 33 0 it is normal. Light apparently 
does not influence its production. Its intensity varies considerably on 
different potato agars, which may possibly be due to differences in 
reaction of the media. 
Gelatin. —Gelatin is rapidly liquefied in thickly sown plates, usually 
within 48 hours. In gelatin stabs liquefaction usually begins in 48 hours 
(assumes a stratiform shape), and may require two to three weeks for 
completion. 
Nutrient broth. —Decided clouding occurs in 24 hours in beef- 
peptone broth (+10 Fuller’s scale) with moderate flaky sediment. On 
slight shaking the sediment readily breaks up into a fine suspension. 
Cloudiness does not appreciably increase after 3 days. A very thin 
surface membrane may be formed, after several days, but this charac¬ 
teristic is not marked. 
Potato cylinders. —Good growth in 48 hours, of brownish yellow 
color. Growth is profuse in 5 days, with increasing yellowing of organism 
along line of streak and darkening of medium. Tests with iodin indicate 
marked conversion of starch to amylodextrin, but diastatic action on the 
whole is feeble as compared with Bacterium campestre. 
