EXAMINATION OF AIR, SOIL, AND WATER 241 



tation tubes and place in an incubator at 40 C. Note the 

 presence of gas, if any, at the end of twelve, twenty-four, 

 thirty-six, and forty-eight hours. // no gas forms, sewage 

 bacteria are absent. 



If gas forms, plate at once a portion of the sediment as 

 above on lactose litmus-agar. Test the other fermentation 

 tubes for acidity, and the nature of the gas, whether any, and 

 how much is absorbed by a 2 per cent, solution of sodium 

 hydroxid. Bacillus coll should produce between jo and 70 

 per cent, of gas, of which about one-third is CO^ and is ab- 

 sorbed by the alkali; the remainder is hydrogen. The other 

 broth culture can be tested for the presence or absence of 

 unfermented sugar by Fehling's solution. 



Diagnostic Points of Colon Bacillus. Microscopic. 

 Non-spore-bearing motile bacillus. 



Gelatin. Non-liquefactive. 



Dextrose Broth. Fifty per cent, gas; one- third absorbed, 

 COz', two-thirds, hydrogen. 



Milk (litmus) coagulated in forty-eight hours and rendered 

 acid; litmus colored red. 



Peptone Solution. Production of Indol. (A peptone solu- 

 tion tube is inoculated with the culture and kept together with 

 a control four days at 37 C. Then 2 drops of concentrated 

 sulphuric acid and i centimeter of a o.oi per cent, solution of 

 sodium nitrate are added. The appearance of a pink color 

 at the end of thrity minutes denotes the presence of indol.) 



Presumptive Test. If a water from a well or spring pro- 

 duces gas in the sugar broth and forms acid colonies on litmus- 

 lactose agar, the presumption is strong that there is sewage 

 contamination. If gas-production continues in a series of 

 samples carefully collected for several days or weeks, there 

 can be no doubt of a contamination, and especially if the well 

 or spring is protected from surface water. Algae which grow 

 in service pipes, reservoirs, and deep wells may give rise to 

 non-acid gas fermentation, but all well-water that, without 

 further testing, forms acid colonies on litmus-agar lactose 

 plates and ferments sugar broth, is open to suspicion, and if 

 16 



