21 



The test for acetyl-methyl-carbinol is but little affected by the period 

 or temperature of incubation of the culture. Positive reactions have been 

 obtained after one, three, or five days at 30 to 37 degrees C. and Chen and 

 Rettger obtained positive results in 10 to 14 hours at 30 degrees C. The 

 kind of peptone does not influence the test, but brighter reactions are ob- 

 tained in peptone than in synthetic media. As neither the character of 

 the medium nor the period of incubation of culture interferes seriously 

 with the test, the Voges Proskauer reaction should serve as a convenient 

 and valuable index for the differentiation of aerogenes from the more 

 objectionable coli section. 



Gas Production and Gas Ratio. When the presumptive test for the 

 colon group was first suggested, it was pointed out that a volume of 25 to 75 

 percent gas was particularly likely to be due to colon bacilli. Escherich 

 in 1885 determined gas ratios for Bact. aerogenes, but Theobcjd Smith 

 in 1895 first called attention to the significance of the ratio of the gases 

 evolved in the decomposition of glucose, pointing out that, whereas Bact. 

 coli produced twice as much hydrogen as carbon dioxide, the Bact. aerogenes 

 differed in that it produced equal volumes of these two gases. These ratios 

 were obtained with the Smith fermentation tube and may be referred to 

 as the crude gas ratio. The determination of the composition of the gases 

 in the Smith tube is very inaccurate and unreliable due in part to the ab- 

 sorption and solution of carbon dioxide and to neutralization by amphoteric 

 substances in the culture medium which would tend to reduce the amount 

 of carbon dioxide observed. Referring to Table VII., it will be noted 

 that in the careful quantitative studies of glucose fermentation by Harden 

 and Walpole, Bact. coli evolved carbon dioxide and hydrogen in ap- 

 proximately equal volumes and not in the ratio of one to two as had 

 been observed by Smith. On the other hand, Bact. aerogenes forms twice 

 as much carbon dioxide as hydrogen instead of equal volumes observed 

 with the Smith tube. These differences are easily accounted for, as has 

 been stated above, by the loss of carbon dioxide in the Smith tube. 



Keyes and Gillespie carried out a series of very careful experiments 

 on the gas ratio and their work has since been confirmed and amplified by 

 Rogers and his associates. They conclude that the accurately determined 

 gas ratio, obtained by growing the test organism in glucose medium in 

 a vacuum and measuring all gas formed, is of fundamental significance 

 and importance in studies on the colon group. 



The real significance of this accurately determined gas ratio was not 

 fully appreciated until 1914 when Rogers called attention to the striking 

 correlation between this ratio and the source of the organisms. In 

 three papers by Rogers, Clark, and Davis (1914) and Rogers, Clark, and 

 Evans (1914 and 1915), it is demonstrated very conclusively that the 

 colon strains obtained from bovine feces decompose glucose with the 

 liberation of carbon dioxide and hydrogen in about equal volume, while 

 strains isolated from grains formed two 'or more times as much carbon 

 dioxide as hydrogen. The former group (C0 2 /H 2 =1:1) seemed in 

 their other characteristics to resemble Bact. coli; the latter (C0 2y /H 2 =2:l) 



