EXAMINATION OF AIR, SOIL, AND WATER 225 



water, for instance, or a mountain stream, the better way is to 

 inoculate two or three lactose or dextrose bouillon fermentation 

 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. If 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 coli should produce between 30 and 

 70 per cent, of gas, of which about one-third is CO 2 and 

 is absorbed by the alkali; the remainder is hydrogen. The 

 other broth culture can be tested for the presence or absence of 

 unfermented sugar, for the color reaction of Rivas. 



Quantitative Tests. The number of acid colonies in i c.c. 

 and in 5 c.c. of water is taken as a measure of pollution, together 

 with the total number of colonies of all bacteria present. Thus 

 in i c.c. on the gelatin plate at 20 C. there may be fifty colonies; 

 on the agar plate at 37 C. ten colonies, five of which were 

 acid-formers, or presumably Bacillus coli. 



To count the colonies which develop upon the plates, a 

 special apparatus has been designed, known as 



Wolfhiigel's Apparatus. A glass plate divided into 

 squares, each a centimeter large, and some of these subdivided. 

 This plate is placed above the gelatin plate with the colonies, 

 and the number in several quadrants taken, a lens being used 

 to see the smaller ones. 



The Petri saucers can be used instead of plates, and an 

 apparatus on the Wolfhugel plan can be obtained to count 

 the colonies. It is best to count all the colonies on the plate 

 or dish. 



Diagnostic Points of Colon Bacillus. Microscopic. 

 Non-spore-bearing motile bacillus. 



Gelatin. Non-liquefactive. 



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

 CO 2 ; two-thirds, hydrogen. 

 15 



