Staining Isolation Cultivation 585 



septicemia, but only half its length (0.2 by 0.5 p). They 

 are usually solitary, but may be united in chains of three or 

 four. 



They are non-motile, and, so far as is known, do not 

 form spores. 



Staining. They stain rather poorly, except with such 

 concentrated and penetrating stains as carbol-fuchsin and 

 Loffler's alkaline methylene-blue, and even with these more 

 deeply at the ends than in the middle, so that they appear 

 not a little like diplococci. They do not stain by Gram's 

 method. 



Canon * recommends a rather complicated method for the 

 demonstration of the bacilli in the blood. The blood is 

 spread upon clean cover-glasses in the usual way, thoroughly 

 dried, and then fixed by immersion in absolute alcohol for 

 five minutes. The best stain is Czenzynke's: 



Concentrated aqueous solution of methylene-blue. . 40 

 0.5 per cent, solution of eosin in 70 per cent, alcohol, 20 

 Distilled water 40 



The cover-glasses are immersed in the solution, and kept 

 in the incubator for three to six hours, after which they are 

 washed in water, dried, and mounted in Canada balsam. 

 By this method the erythrocytes are stained red, the leuko- 

 cytes blue; and the bacilli, also blue, appear as short rods 

 or as dumb-bells. 



Large numbers of bacilli may be present, though some- 

 times only a few can be found after prolonged search. They 

 are commonly inclosed within the leukocytes. It is scarcely 

 necessary to pursue so tedious a staining method for demon- 

 strating the bacilli, for they stain well enough for recognition 

 by ordinary methods. 



Isolation. The influenza bacillus grows poorly upon 

 artificial culture media, and is not easy to isolate, because 

 the associated bacteria tend to outgrow it. When isolated 

 it' is difficult to keep, as it soon dies in unnatural environ- 

 ment. 



Cultivation. The bacillus does not grow in gelatin or 

 upon ordinary agar-agar. Upon glycerin agar-agar, after 

 twenty-four hours in the incubator, minute colorless, trans- 

 parent, dewdrop-like colonies may be seen along the line of 

 inoculation. They look like condensed moisture, and Kita- 

 *"Centralbl. f. Bakt.," etc., Bd. xiv, p. 860. 



