ISOLATION OF THE COLON BACILLUS. 61 



the B. cob', we may assume that some 40 of these organ- 

 isms were present to the cubic centimeter. Irons (Irons, 

 1901), in a comparative study of various methods for the 

 isolation of B. cob, showed that the preliminary enrich- 

 ment frequently gave positive results when the results 

 of the direct use of the agar plate were negative and 

 concluded that " where the waters to be examined are 

 much poUuted, or where the amount of B. cob is small 

 and the colony count large, the lactose plate for plating 

 water direct is inferior to the dextrose fermentation- 

 tube." Gage obtained similar results (Gage, 1902). 

 The method of direct plating in phenol-agar is quicker 

 and there is a certain danger that colon bacilh originally 

 present may be overgrown and killed out in the enrich- 

 ment process. If the incubation is not too prolonged, 

 however, this need not occur. 



The medium ordinarily used for the preliminary enrich- 

 ment is ordinary broth to which 2.5 per cent of dextrose 

 has been added, and the reaction brought to the neutral 

 point. Into each of a number of fermentation-tubes of this 

 medium a measured quantity of the water to be examined 

 is inoculated, and the culture is incubated for twenty-four 

 hours at 37.5 C. At the end of this time the tubes are 

 examined for gas formation. If gas is found, a small 

 amount of the culture should be added, after suitable 

 dilution, to litmus lactose agar and plated. 



With polluted waters it will be found advantageous to 

 plate out on the first appearance of gas (4-8 hours). It 

 has been shown by one of us (Prescott, i.902 b ) that a very 



