158 BACTERIOLIGY OF WATER, AIR, MILK, ETC. 



times a minute for ten to twenty minutes. Then remove supernatant milk and add 

 0.5 c.c. of Toisson's solution to the sediment. Instead of Toisson's solution I use 

 Gram's iodine solution which brings out the leukocytes equally well and gives a cleaner 

 preparation. You thus have the leukocytes of 10 c.c. contained in 0.5 c.c. (Con- 

 centrated 20 times.) Make a haematocytometer preparation as for blood and 

 find the average number of cells for each square millimeter. Then multiply this by 

 10 to get the number of cells in a cubic millimeter. As a cubic millimeter is 1000 

 times smaller than a cubic centimeter, you multiply the number per cubic milli- 

 meter by 1000. Then, as the milk was concentrated 20 times, you divide by 20. 

 (If it were diluted 20 times, you would multiply by 20.) 



Example. Found an average of 50 cells per square millimeter. This would make 

 500 per cubic millimeter, and 500,000 per c.c.; then 500,000 divided by 20 would 

 give 25,000. 



There is no agreement as to a standard for allowable leukocytes. Even in ap- 

 parently healthy animals they may exceed 100,000 per c.c. Doane has suggested 

 500,000 per c.c. as a preferable limit. 



The smear methods for determining the number of leukocytes present do not 

 compare in accuracy with the volumetric ones. It is important, however, from a 

 standpoint of examining for tubercle bacilli, etc., as well as for recognition of leuko- 

 cytes, to deposit the sediment from a centrifuge tube, taken up with a capillary bulb 

 pipette, on a glass slide. Smear out slightly and then when dry fix with a mixture of 

 ether and absolute alcohol. Flood with ether to get rid of remaining fat and stain by 

 Gram's method or by acid-fast staining. 



To summarize, we may state that the bacterial count is an indicator 

 of the care used in handling the milk while the presence of harmful 

 bacteria (qualitative examination) or numerous pus cells indicates 

 disease in the cow. During 1912 severe epidemics of sore throat due 

 to a streptococcus, S. epidemicus, were traced to milk of cows having 

 probably suffered from mastitis. In Baltimore the milk had been 

 pasteurized by the flash method which indicates the unreliability of 

 this process. 



Pasteurization of Milk. The objections to this method of preserving milk 

 have been (i) that the lactic acid bacteria which have been by some credited with 

 antagonism to harmful bacteria, would be destroyed by pasteurization, (2) the more 

 rapid development of bacteria in milk that has been pasteurized (3) interference with 

 nutritive qualities and (4) pasteurized milk does not show its deterioration as does 

 unpasteurized milk, thus failing to give a clue as to the age of the milk. 



The United States Bureau of Animal Industry in studying this important phase of 

 the milk question has grouped the milk bacteria into three classes (a) acid-forming, 

 (b) putrefactive (liquefying) and (c) inert bacteria. In their investigations it was 

 found that many acid-forming bacteria withstood temperature as high as i68F., 

 so that pasteurized milk was soured just as is raw milk, but more slowly. They found 

 that pasteurized milk showed fewer putrefactive bacteria than raw milk, so that 

 even should it be a fact that injurious toxins were produced by spore-bearing putre- 





