BACTERICIDAL ACTION OF CR INDICATOR 85 



test tubes and autoclaved. 2 After incubating over night to test 

 for sterility, each tube was inoculated by means of a sterile cap- 

 illary with two drops of a well-grown peptone water culture of 

 the organism to be tested. Incubation was carried on at 37.5° 

 for a maximum of four days. A control tube of the same medium 

 without dye was also inoculated at the same time, and served as 

 a standard by means of which comparative amounts of growth 

 could be estimated. 



The composition and method of preparation of this medium 

 is exactly the same as that of the stock media used in this labora- 

 tory for the determination of carbohydrate fermentation, with 

 the exception that in this case carbohydrate was not added. 



The media containing China blue were perfectly clear. The 

 media with 0.125 per cent and 0.05 per cent concentrations of 

 Rosolic acid were turbid at times, and a slight precipitate was 

 often present. In these cases the percentage of dye actually 

 dissolved in the medium is of course not definitely known. 



RESULTS 



China blue, in the highest concentration used, 0.0625 per cent, 

 had no inhibiting action on the majority of the organisms which 

 we tested. Bacillus acidi-lactici was, however, completely in- 

 hibited by 0.025 per cent, and the Micrococcus flavus by a 0.0625 

 per cent concentration of this dye. A few of the other organisms 

 were slightly inhibited by a concentration of 0.0625 per cent. 



s The media with the greatest concentration of the dye (0.0625 per cent) con- 

 tained about 3 per cent of alcohol by volume, and it was suspected that the alco- 

 hol might have some effect upon bacterial growth. However, practically all the 

 organisms which we tested grew as well on media containing China blue and 3 per 

 cent of alcohol as on media without the dye or the alcohol. There was, therefore, 

 practically no inhibition from the alcohol. According to Kligler (1918) 4 per cent 

 alcohol is required to inhibit the growth of B. dysenteriae, a greater concen- 

 tration being required to inhibit the other bacteria which he tested. In our 

 work, the media containing the second highest concentration of dye (0.025 per 

 cent) contained a little more than 1 per cent of alcohol. It is probable that the 

 effect of the alcohol present may be neglected for our purposes. 



