SEWAGE CONTAMINATION OF OYSTER BEDS. 207 



BACTERIOLOGICAL ANALYSIS OF WATER SAMPLES FEOM NAREAGANSETT 



BAY. 



Jfet/tods employed. — The usual methods for isolation of B. coli from 

 water and sewage were used in this work. Fermentation tubes con- 

 taining a neutral 1 per cent dextrose broth were inoculated with 1 c. c. 

 of the suspected water and incubated for three daj's at 37° C. In some 

 cases a 0.1 per cent phenol broth was also inoculated with 1 c. c. of the 

 water and allowed to develop at 37° C. for twenty-four hours. In a 

 few tests litmus-lactose-agar and agar containing 1 per cent neutral 

 red were sown with varying amounts of water and grown at incubator 

 temperature. 



If no gas was formed in the fermentation tubes in twenty-four hours 

 the test was considered negative without further procedure. If, how- 

 ever, any considerable quantity of gas developed within this time, 

 litmus-lactose-agar plates were inoculated in most cases from these 

 tubes and incubated at 37° C. twenty -four hours longer. When litmus- 

 agaf plates were not used, a gelatin medium was substituted. Any red 

 colonies developing on the litmus medium, and any colonies showing 

 the characteristic growth of B. coll on gelatin, were fished out and 

 transferred to slant agar tubes. From the cultures thus o])tained 

 subcultures were made in neutral dextrose and lactose broth, nitrate 

 solution, milk, sugar-free broth containing 2 per cent of peptone 

 and gelatin. When growth occurred in the phenol broth, although 

 sufficient gas to indicate the presence of B. coll was not developed in 

 the feraientation tubes, litmus-lactose-agar plates were inoculated 

 from the phenol broth and treated as alread}" described. 



Organisms giving the following positive reactions to tests were 

 regarded as members of the colon group of bacteria: 



1. A small more Or less motile bacillus in twenty-four-hour bouillon 

 or agar cultures. Usually not all the Imcilli in one microscopic field 

 are motile — often sluggishh" motile. 



2. Fermenting dextrose broth with the production of gas. The large 

 part, if not all, of the gas is formed during the first twenty-four hours. 

 The liquid in the tube must be distinctly acid to indicate B. coll. The 

 ratio of h3^drogen to carbon dioxide is approximateh' 2 to 1. This 

 ratio is, however, more or less variable in cultures from a single 

 strain. The total amount of gas produced in dextrose usually does not 

 exceed 55 per cent, though there is also more or less variation in this 

 characteristic. 



3. Fermenting lactose with the production of much gas; reaction 

 strongly acid. 



4. Indol produced in sugar-free broth containing 2 per cent of 

 peptone. 



5. Milk coagulated in three days at room temperature; in twenty- 

 four hours at 37.5° C; casein not liquefied; reaction acid. 



