156 C.-E. A. WiNSLOw AND Anne F. Rogers 



constant difference in size was apparent in comparing solid and li- 

 quid cultures. One series of organisms examined in dextrose broth 

 and on agar, at periods ranging from one day to two weeks, showed 

 the same average size in both media and at all ages. Finally we 

 attempted to see whether prolonged cultivation under special con- 

 ditions would affect the size of the cell. Cultures were grov^n for 

 10 days, in broth at 37°, on nutrient gelatin, and on acid and 

 anaerobic gelatin with daily transfers. The size of each culture 

 was recorded on the loth day, after which time each was transferred 

 to gelatin and examined after one day. The results showed prac- 

 tically no significant differences. 



In a comparison of the size as determined by examination of living 

 organisms and of stained preparations, the cells appeared generally 

 somewhat smaller after staining. This is no doubt partly due to some 

 shrinkage in drying, and partly to the imperfect definition which 

 makes the unstained specimens appear larger than they really are. 

 Occasionally, when the staining was too heavy, the stained cells 

 appeared larger. In any case the differences are unimportant, 

 and we have used the size of the mcthylene-blue-stained preparation 

 throughout our work. 



Staining reactions. — Since the cocci, as far as we have examined 

 them, all stain easily with methylene blue, we have made no special 

 tests with anilin-gentian-violet. The Gram stain has, however, been 

 used on all our cultures, since, in the genus Diplococcus and in many 

 other groups, it has been thought to have such special importance. 



The value of this staining method has been studied with consid- 

 erable care by Mr. A. T. Brant, working in the laboratories of the 

 Institute. Mr. Brant found, as other observers have done, that while 

 certain bacteria are constantly Gram-negative or Gram-positive, 

 others exhibit an intermediate condition, retaining the stain under 

 some conditions and giving it up under others. In his, as yet unpub- 

 lished, paper he notes, for example, that all cultures of B. coli are 

 decolorized by one minute's treatment with alcohol, while B. mega- 

 therium constantly fails to decolorize after three hours. On the 

 other hand, with B. fluorescens, M. pyogenes, M. aureus, and B. 

 diphtheriae the result is affected by the time of decolorization, as well 

 as by the age of the cultures. Between the fixed points at the 



