768 EXPERIMENT STATION RECORD. 



from highly seeded plates are uniformly lower than the counts of the same milk 

 from low-seeded plates. The best results are obtained only when the plates 

 contain somewhere between 40 and 200 colonies. (3) Methods of counting. 

 This has seemed to be the cause of the widest amount of irregularity. The 

 greatest difference was associated with the use or nonuse of a counting lens, 

 or with differences in magnifying power of the lens used. Further results 

 show that the personal equation in counting is still a factor of very large im- 

 portance. When the same plate is counted by two different laboratories the 

 results are not infrequently 100 per cent apart, and occasionally more. Another 

 factor modifying the counting is the method adopted by each laboratory of esti- 

 mating numbers rather than actual counting. (4) Irregularities in samples 

 from the same bottle of milk. Two samples taken from the same bottle of milk, 

 even after thorough shaking, are by no means identical. This is due to the 

 cluiuplng of bacteria and to the fact that inasmuch as bacteria are not in solu- 

 tion, but are solid objects, they can not be expected to be uniformly distributed 

 through the liquid. (5) In low counts variation between duplicate samples is 

 sometimes considerable, due to the irregularity of the distribution of bacteria. 

 In high counts variation in reports is also sometimes very great, due to exces- 

 sive crowding and to methods adopted in estimating the number of colonies. 



The extent of variation in the results obtained from the analysis of dupli- 

 cate samples varies widely with the care that is taken in the laboratory tech- 

 nique. At first the causes of irregularity were sufficient to give results disagree- 

 ing as much as tenfold in the number of bacteria that would be reported from 

 any sample of milk. After the laboratories had adopted methods of bringing 

 about uniformity in technique so far as possible, the variations were very 

 greatly reduced, the last tests showing that when sufficient care is given the 

 variations need not be more than twofold. It is not possible to rely upon a 

 greater accuracy than 100 per cent even when an average of more than one 

 analysis is obtained, although most of the results fall considerably below this 

 limit. 



There is no essential difference in the results whether the milk dilution is 

 directly inoculated into the Petri dish and the agar poured upon it or whether 

 the milk dilution is inoculated into the melted tube of agar and subsequently 

 poured into the Petri dish. When examinations of cream are made the plate 

 inoculation is unreliable, and the inoculation must be made in the agar tube 

 followed by thorough agitation. 



Five days' incubation (48 hours at 37° C. and 72 hours at 20°) gives a very 

 slightly greater count than a two-day incubation. A 24-hour count gives on the 

 average about one-half as high numbers as a 48-hour count. 



The results of the tests seem to warrant three broad grades, essentially the 

 three grades that have been adopted by the commission on milk standards, but 

 they are not as yet accurate enough to warrant a closer grading than the com- 

 mission's grades. 



The series of tests has proved that if a sample of milk can be put into iced 

 water, containing floating ice, it may be kept for 20 hours with very little change 

 in bacteria count. This makes it possible to keep samples sent to a laboratory 

 for analysis for a number of hours without any fear of change in bacterial 

 content, provided the samples are immersed in water containing floating ice. 



It is said that in making a comparison of the bacteriological analysis by the 

 plate count and the microscopic count, the latter should be a count of groups 

 rather than individuals, plate colonies representing groups only. Considerable 

 experience by the person making the count is needed to distinguish between 

 bacteria and dirt particles, particularly when the milk contains minute micro- 

 cocci. When the microscopic count is made by one who has had sufl5cient 



