40 FREDERICK EBERSON 



tional" picture. Repeating Mellon's technic I have not succeeded in demon- 

 strating this and I find that his paper contains no evidence that such a 

 postulate has been fulfilled. Subcultures were made on agar, after 12, 24, 30, 

 48, 72, and 96 hours, from individual flasks which had been seeded with the 

 sediment obtained from tubes incubated at 25-28 C. for 3 days. In none of 

 these transplants, incubated at 37 C. for 24 hours, was a culture of cocci 

 obtained. There was, however, an abundance of extremely short forms 

 coccoids and some diplococcoids, but nothing to indicate that the bacillary type 

 had undergone any definite change. When cocci are present, the picture is 

 unmistakable and there can be no confusion of this type with the very short 

 forms which look like diplococci (Fig. 26). 



It is questionable whether the refined technic of Mellon is needed to demon- 

 strate what is erroneously called a transformation. That all diphtheroid cul- 

 tures show two distinct forms of organisms is beyond question and that these 

 types may be seen under usual laboratory conditions, is also a matter of 

 common experience. Great caution is necessary in interpreting a morphologic 

 appearance as the actual process responsible for the occurrence of different 

 forms in bacterial culture. Repeated subcultures gave negative results and at 

 no time was it possible to show that complete disintegration had taken place. 

 The "mucus-like debris" and "concentration of chromatin" described by Mellon 

 was seen after 24 hours, not only in broth cultures made according to the 

 special technic described, but also in agar slants grown under conditions to 

 be given. Transfers from broth containing "disintegrated bacillary forms" 

 naturally failed to give mucus-like shreds which are found in smears, but 

 the explanation for this is very simple. Detritus can hardly be expected to 

 grow in Transplants. As for the "chromatin masses approaching each other in 

 transplants until the figure cannot be told from a diplococcus," the fact remains 

 that these forms only resemble diplococci. In order to study the effect of lower 

 temperatures on the possible suppression of bacillary forms which might occur 

 during the interval- when the broth cultures are incubated at 25-30 C., a few 

 experiments were performed with agar cultures. 



Kinyoun's method for the staining of diphtheria was used. This stain gives 

 most marked granular appearance and is best adapted to differentiating cocci 

 from bacillary bodies with chromatin substance so disposed as to mislead one 

 into taking these for true cocci. Agar slants were prepared from a bacillary 

 culture of C. enzymicus and incubated at 37 C. for 24 hours. As usual, some 

 coccus forms could be discerned in smears. The picture, however, was bacillary. 

 Such cultures when incubated further at room temperatures, 21-23 C. for 72 

 hours, showed in smears numerous diplococci and a marked overgrowth of 

 very short forms. Subcultures maintained at these temperatures always yielded 

 the same results. When transfers were made to agar and incubated at 37 C, 

 however, the diplococci became rare and finally could not be found at all. 

 At 28 C. the effect on morphology was evident also. After 48 hours at this 

 temperature, diplococci became numerous, and after 5 days they were present 

 in practically pure culture. Transfers to agar incubated at 28 and 37 C. gave, 

 respectively, numerous diplococci and short forms and few diplococci. Broth 

 cultures, kept at 28 C. for 1 week, were found to contain numerous diplococci 

 and extremely short forms which could not be distinguished readily from these. 



Coccus Type. A pure culture of cocci was obtained from the original 

 diphtheroid strain, as described previously, and attempts were made to alter 

 its morphology. The results of the experiments showed conclusively that 

 when working with such a pure strain of cocci it is impossible to transform 



