552 REUBEN L. KAHN 



cally, but not during the aerobic growth on Endo agar, does not 

 appear to hold. Organisms growing on the surface of agar 

 must indeed have the abihty to grow in the presence of air in 

 order to get a start, so to speak. But once the start is gained 

 the surface organisms alone grow aerobically, while these organ- 

 isms below the surface of the colony are probably growing in 

 no less an anaerobic environment than those growing below the 

 surface in carbohydrate broth. 



Observations of the gradual disappearance of the metallic 

 film of colon colonies on Endo agar corroborates this view. 

 It is reasonable to assume that the organisms below the metallic 

 film of a colon colony, are growing in an anaerobic environment. 

 These organisms, after twenty-four to forty-eight hours incuba- 

 tion, proceed to break down the nitrogenous bodies of the nu- 

 trient media into ammonia and amines (Robinson and Rettger), 

 creating a condition favorable for the re-solution of the metallic 

 fuchsin and final decolorization. In other words, we have here a 

 chemical change produced by B. colt growing anaerohically in 

 apparently aerobic colonies. 



As further evidence that the metallic sheen of colon colonies 

 on Endo medium results from the evaporation of volatile sub- 

 stances produced during the metabolism of these organisms, 

 the following two observations are cited. 



1. If colon bacilli are grown anaerohically in Endo agar, the 

 colonies are red but not metallic. Under anaerobic conditions, 

 B. coli breaks down carbohydrate into various acids, aldehyd, 

 and possibly acetone and alcohol (Grey and Mendel). These 

 substances bring out the color of decolorized fuchsin and the 

 colonies become red. The fuchsin, however, remains in solution, 

 because the volatile substances can not evaporate. The same 

 colonies exposed to air become metallic, because surface evapo- 

 ration permits the disappearance of the substances which hold 

 the fuchsin in solution. 



2. The metallic sheen of a given colon colony is markedly 

 pronounced when grown in such a manner that evaporation can 

 take place on a large scale. Thus, if B. coli be inoculated on 

 Endo plates and on Endo agar slants, the metallic film, after a 



