71 



A priori, then, we might expect that the addition of gentian violet to our 

 culture media in proper dilution would result in eliminating many saprophytic 

 bacteria, still permitting those forms of sewage origin to flourish. If we 

 used a sugar medium and added litmus we could still further emphasize the 

 colon group, as these are acid-forming organisms. The violet stain partly 

 masks the coloration of the litmus indicator, but not sufficiently to make the 

 picture of acid fermentation uncertain. 



My work to date has not been extensive enough to warrant any definite 

 conclusions, but it is at least suggestive. I have analysed various samples 

 of water taken chiefly from the Wabash River, which is rather highly polluted 

 at Lafayette. Duplicate plates of proper dilutions have been made of litmus- 

 lactose agar and litmus-lactose-violet agar, the latter being the same as the 

 former with the exception of the addition of a standardized loop full of 

 gentian violet solution to the agar tube just before pouring. The plates have 

 been examined after 24 hours incubation at 37° C. The total number of 

 organisms growing, the total number of red colonies — acidifers — and the pre- 

 sumptive coli colonies growing on the two media have been recorded. The 

 suspected coli growths have been "fished" and planted in lactose-peptone-bilc 

 for confirmation and almost without exceptioii the fermentation of this media 

 has checked the presumptive colony growth. 



The colonies on the violet plates appear somewhat smaller and the acid 

 production is less distinct. The stain is picked up by the cells so that the 

 colonies appear, especially the sub-surface colonies, as distinctly pur])le 

 growths. Viewed under the microscope the cells show a light purple color, 

 indicating vital staining. 



So far I have found pretty generally what was expected, viz., that the 

 total count is materially reduced on the violet plates but that the number 

 of red colonies, and especially of coli, are approximately the same on the two 

 media. It has been found possible to plate a larger sample of water and to 

 intensify the picture of presumptive pollution by the use of the violet. A 

 few typical examples of actual tests will illustrate this: 



