14 



The alcohols have been very frequenlty utilized in investigational 

 studies. Thus dulcitol was employed by MacConkey and Jackson in their 

 classifications, but it is now being generally supplanted by other sugars. 

 Mannitol is fermented by practically all members of the colon group 

 except a small group observed by Rogers in his grain series. The al- 

 cohol adonitol was recently suggested as a means of differentiating the 

 fecal from the non-fecal Bact. aerongenes and this was adopted by the 

 Committee of Standard Methods of Water Analysis in the 1917 report. 



Glycerol has been found by Kligler and later by Levine to correlate 

 well with gelatin liquefaction and they both distinguish Bact. aerogenes 

 from Bact. cloacae on the basis of fermentation of this material. The alco- 

 hols are thus an important group of carbohydrates for studies of the colon 

 group of bacteria. 



The glucoside salicin has been recently suggested to supplant dulcitol 

 for classification purposes by Kligler and by Levine and Castellani and 

 Chalmers employ it for primary subdivision of their sucrose negative strains. 

 Salicin fermentation is an important differential test. 



Voges Proskauer reaction. (Acetyl methyl carbinol test). By 

 the Voges Proskauer reaction is meant the production of an eosin-like col- 

 oration in dextrose broth cultures by some members of the colon group, 

 if made strongly alkaline with potassium hydroxide. It takes its name 

 from the fact that it was first observed by Voges and Proskauer in 1898. 

 The coloration develops slowly from the surface of the medium gradually 

 extending throughout the culture. The test is ordinarily carried out by 

 adding two or three c. c. of 10 percent potassium hydroxide to an equal 

 volume of a 48 to 96 hour dextrose broth culture and after thoro shaking 

 the mixture is allowed to stand exposed to the air. The characteristic 

 eosin-like color will develop in a few hours but it is well to record after 

 24 hours exposure if negative in a shorter time. 



It is suggested that the term Voges Proskauer reaction be restricted 

 to designate the formation of acetyl methyl carbinol from glucose but when 

 referring to its production from other carbohydrates or alcohols, the 

 term acetyl-methyl-carbinol test be applied. The nature of the sub- 

 stance being tested for is thus indicated just as is the case with indol. The 

 Voges Proskauer reaction has been found very valuable and many investi- 

 gators have observed that it is characteristic of the colon-like organisms 

 of the soil and grains while it is very rare to encounter intestinal members 

 of the colon group which give this test. 



Methyl Red Test. Clark and Lubs in 1915 devised the socalled 

 methyl red test which serves to split the colon group into a methyl-red- 

 positive subgroup, found to be characteristic of the organisms obtained 

 from cow feces and other intestinal sources, and the methyl-red-negative 

 subgroup, which is the predominating type in the soil and on grains. 



The test is made by adding a few drops of methyl red indicator to a 

 0.5 percent dextrose-peptone- (Witte)-dipotassium phosphate culture and 

 noting the reaction; a yellow coloration indicates alkalinity or a negative 

 test and a red coloration denotes acidity or a positive test. The reaction 



