160 ROYAL SOCIETY OV CANADA 



Reaction was made -f-0.6, and the medium was filtered. 



B. In this medium the same proportion of ingredients was 

 used as in A, but with the addition of 3 grams of commercial bile 

 (salt sodium taurocholate). The final acidity was -|~ 0-6. 



These media were filled into test tubes, about 10 cc. in each tube. 



The water for analysis was added in quantities not exceeding 

 5 cc. per tube. 



The tests with culture medium A. (without bile salt) gave the 

 following results. 



In a series of 77 tests the tubes were black in 24 hours in 40 

 cases. Subcultures in aesculin agar plates shewed black colonies 

 in 48 hours at 37°C. In 25 cases the tubes were black in 24 hours 

 at 37°, but it took 120 hours to develop black colonies on aesculin 

 agar plates. In 12 tests the tubes were unchanged after 48 hours 

 at 37°C. Subcultures in aesculin agar gave no black colonies in 

 168 hours at 37°C. ' 



Ninety tests were carried out with culture medium B. (with 

 bile salt). In 20 tests the tubes were black in 24 hours, and sub- 

 cultures on aesculin plates showed black colonies in 24 hours. In 

 35 tests the tubes were black in 48 hours, and subcultures on aescu- 

 lin plates showed black colonies after 24 hours. 



In 30 tests the tubes were unchanged in colour after 48 hours; 

 subcultures on aesculin agar developed, however, black colonies in 

 120 hours. In 5 tests, tubes and subcultures on plates remained 

 unchanged (108 hrs.). 



In most of the above tests we used both varieties of media for 

 the same sample of water and observed that the culture medium 

 without bile salt (A) blackened quicker than the medium with bile 

 salt (B). Undoubtedly weak individuals of B. coll are checked in 

 their growth by the bile salt. A very good instance of this was ob- 

 tained from some experiments on the duration of life of B. coli in 

 river water. We inoculated river water with a strong B. coli giving 

 pronounced black colonies in aesculin media. 



Every week part of this water was subcultured, and we noticed 

 that the ability of />. coli to produce typical colonies on aesculin 

 media gradually decreased, and at one time the culture would pro- 

 duce black discoloration in aesculin media without bile salt, but was 

 without effect on liquid aesculin media containing bile salt. This 

 differentiation enables us to distinguish between an old and a new 

 infection, or between a strong or weak species. 



In the preparation of liquid media only 0.05% of aesculin and 

 iron citrate are necessary as compared with the 0.1% in the solid 



