254 



Bacteriology of Water 



to make a number of cultures with different quantities of the 

 water, using, for example, o.oi, o.i, 0.5, and i.o c.c., re- 

 spectively, to a tube of liquefied gelatin, agar-agar, or gly- 

 cerin agar-agar. 



The details of the method depend upon the quality of the 

 water to be examined. If the number of bacteria per cubic 

 centimeter be small, large quantities may be used; but if 

 there be millions of bacteria in every cubic centimeter, it 

 may be necessary to dilute the water to be examined in the 

 proportion of i : 10 or i : 100 with sterile water, mixing well, 

 and making the plate cultures from the dilutions. 



It is best to count all the colonies developed upon the 



culture, if possible; but when 

 hundreds of thousands are scat- 

 tered over it, an estimate made 

 by counting and averaging the 

 number in each of the small 

 squares of some counting appa- 

 ratus, such as those devised by 

 Wolfhiigel (Fig. 72), Esmarch 

 (Fig. 73), or Frost (Fig. 74). 

 In counting the colonies a lens 

 is indispensable. 



In some cases, as in the study 

 of sewage, badly contaminated 

 waters, inflammatory exudates, 

 and in the preparation of bac- 

 terial vaccines, it is expedient 

 to directly enumerate the bac- 

 teria without resorting to the cultivation method, where all 

 of the organisms may not grow. 



Excellent methods for achieving this have been devised. 

 That of Winslow and Willcomb* being as follows: 



"The cover-slips should be boiled in a 10 per cent, solution 

 of potassium bichromate in 50 per cent, sulphuric acid and 

 allowed to lie in this cleansing mixture. Just before using 

 they may be rinsed in 50 per cent, alcohol and dried on a 

 silk cloth, not in the flame. One-twentieth of a cubic centi- 

 meter of water placed on such a cover-slip spreads evenly 

 and should be allowed to dry in the air without sudden 

 heating. After drying it is fixed by passing through the 

 flame, covered with Ziehl-Neelson's carbol-fuchsin, warmed 

 * "Jour, of Infectious Diseases," Supplement No. i, May, 1905, p. 273. 



Fig. 73. Esmarch's instru- 

 ment for counting colonies of 

 bacteria in Esmarch tubes. 



