622 BACTERIOLOGY OF THE SOIL, WATER, AND AIR 



invariably contain more bacteria than ground waters; it may be 

 necessary to dilute a cubic centimeter of the sample with 99 c.c. of 

 sterile water to obtain the requisite distribution of organisms for an 

 accurate estimation, or even higher dilutions may be necessary. 

 Grossly polluted waters are diluted one thousand or even ten thousand 

 times with sterile water before they are plated. In any event, not 

 more than 200 colonies or less than 50 colonies should be present in 

 the final dilution, for experience has shown that greater numbers of 

 organisms materially restrict development, and fewer than fifty 

 colonies upon a plate introduces an error in dilution. 



Technic of Plating. The sample of water, diluted to the required 

 degree if necessary, is shaken vigorously to break up groups and 

 chains of bacteria; a cubic centimeter of water is then removed with 

 a sterile pipette into each of two sterile Petri dishes, being careful to 

 prevent contamination. 



A tube of sterile nutrient gelatin (10 c.c.) previously melted and 

 cooled to 42 C., is then carefully poured over the water in one Petri 

 dish, and melted nutrient agar is similarly poured into the other Petri 

 dish. The. water and culture fluid are intimately mixed by carefully 

 tilting the plates, and then set aside to harden. The agar plate is 

 inverted after it has hardened to prevent condensation of moisture 

 upon the surface of the medium; this procedure reduces the possi- 

 bility of confluence of surface colonies. The gelatin plate is not 

 inverted. 



Incubation at 20 C. for the gelatin plate and 37 C. for the agar 

 follows. The agar plate is counted after forty-eight hours' incubation, 

 the gelatin plate after four days. 



Interpretation of Bacterial Count. At best the quantitative estima- 

 tion of bacteria in water and sewage is inexact and relative only. 

 The many factors of error in sampling, lack of uniformity in media, 

 the difficulties of counting colonies when several hundred have grown 

 in one plate all tend to reduce the accuracy and precision of the 

 method. Again, the normal difference in bacterial content between 

 ground waters, surface waters and polluted waters makes an inter- 

 pretation of the bacterial count somewhat difficult. For example, 

 100 bacteria per cubic centimeter in a deep well water might have 

 greater sanitary significance than 500 bacteria per cubic centimeter 

 in a surface water, where bacterial counts are almost invariably higher. 



Attempts have been made to establish arbitrary bacterial standards; 

 thus, waters containing less than 100 bacteria per cubic centimeter 



