4 Journal of Agricultural Research voi. ix, n 0 . i 
MORPHOLOGY AND PHYSIOLOGY OF BACTERIUM CITRAREFACIENS 
MORPHOLOGICAL CHARACTERS 
When the organism is taken from 24-hour-old agar stroke cultures it 
is rod-shaped, with rounded ends, usually single, but occasionally in 
pairs. The limits of size ate 1.2 to 3.0 by 0.3 to 0.9 ix, the most common 
size being 1.8 by 0.6 fx. No spores or capsules have been observed; 
zoogloea are not found; motility has been observed in affefcted tissue 
and from young cultures, one to four flagella at one or both poles having 
been shown from 24-hour agar cultures. Involution forms, long filament¬ 
like bodies in which there are thickened masses of protoplasm that take 
stains more heavily, are found in old cultures and especially in saccharose 
bouillon (PI. i, 2 ?). 
The bacterial rods may be readily stained by watery-fuchsin, gentian- 
violet, carbol-fuchsin, and Loffler’s alkaline methylene-blue. When 
stained lightly, a bipolar effect is brought out in the organism, the ends 
of the rods taking the stain more densely, leaving a lighter area in the 
middle. The organism is Gram-negative, and is not acid-fast. 
cultural characters 
Agar poured plates. —On +10 peptone-beef agar at 20° to 22 0 C., surface colonies 
are apparent on the third day; on the fourth day colonies are 2 to 3 mm. in diameter, 
white, round, smooth, glistening, convex, finely granular (under the compound 
miscroscope), with entire edges sometimes becoming lacerate, due to the semifluid, 
consistency of the bacterial mass. When 5 days old, the agar beneath the colonies 
becomes greenish and fluorescent, and in 7 days the colonies lose their convex glisten¬ 
ing surface and dry down, frequently becoming concentrically ringed and the edges 
undulate to lobate. Buried colonies are biconvex. 
Agar stabs. —Stabs in +10 peptone-beef agar when 13 days old show growth along 
the line of inoculation to four-fifths of the total depth of the stab. Growth is best 
toward the surface, the line of puncture beaded; there is no liquefaction; surface 
growth is restricted. The agar medium is uniformly colored a javal-green 1 when seen 
against a black background. 
Agar slants. —On slant agar, stroke cultures make an abundant growth in two 
days; filiform becoming somewhat echinulate, convex, glistening, smooth, trans¬ 
lucent, of slimy consistency, and with a slight odor of putrefaction. A cream-white 
sediment forms in the V; the medium is colored uniformly a fluorescent-green. 
Nutrient-gelatin plates. —Growth rapid, at first punctiform, then round, crateri- 
form; liquefaction spreading. 
Gelatin stabs. —At 18 0 to 19 0 C., in +15 peptone gelatin, liquefaction starts on 
the first day, is crateriform until on the fourth day the pit of liquefaction reaches the 
walls of the tubes when it becomes stratiform. The liquefaction gradually slows up 
and takes 60 days or more to reach completion. 
BEEP bouillon. —In -f 10 peptone-beef bouillon a moderate uniform clouding 
appears within 24 hours; there is no turbidity. At first there is formed a very thin 
pellicle which disappears within a few days. Old cultures contain a sediment which 
is very slightly viscid. There is a slight odor of putrefaction. 
1 All references to color are expressed according to Ridgway (Ridgway, Robert, Color Standards and 
Color Nomenclature. 43 p. ( 53 pi. Washington, D. C., 191a). 
