142 LABORATORY EXERCISES IN BACTERIOLOGY. 



at from counting the number of colonies developing in the cultures probably represents 

 incompletely the real proportion of bacteria in the sample. Each colony is supposed to 

 represent one original bacterium, but if the diffusion has not been entirely successful, 

 a clump of organisms might well -have developed into a single colony; moreover, it 

 is quite probable that not all of the germs will have developed on the medium selected 

 and at the temperature to which the cultures were exposed. Close watch should 

 be maintained upon the preparations so that all the colonies are included in the enu- 

 meration, a magnifying glass being used to detect the smaller ones. Where greater 

 exactness is demanded it is well that not only gelatine preparations be made, but 

 that agar be also inoculated, plates of which, after being grown for several days in 

 the room temperature, are placed in the incubator in order that organisms growing 

 only in warmth may be afforded opportunity for development. By this measure 

 one may evade the difficulty which is apt to arise from the liquefaction of the gelatine 

 by the bacteria, a source of much confusion in many cases. As a rule the common 

 water bacteria develop best in these cultures during the first few days while exposed 

 to the temperature of the room, the incubator preventing their free development. 



Especially upon these agar preparations at incubator temperature is there proba- 

 bility of appearance of colonies of such pathogenic and parasitic organisms as may 

 have been present in the sample. In ordinary investigations they are very likely 

 to be overlooked and lost. Of course, their recognition must depend largely upon 

 the skill, experience, and watchfulness of the observer; but in the midst of the hundreds 

 of colonies of much the same general appearance on the plate, even should they appear 

 they may be unrecognized by the well-informed and skilled. In order to favor their 

 recognition, it is best to induce their increase in the sample of water in some way 

 before subjecting it to culture, and then make cultures on agar preparations at incu- 

 bator temperature. This modification of the original sample is not to be made, of 

 course, until after the preparation of the cultures above described, which are intended 

 for the exhibition of the common bacteria and for their enumeration. Thereafter 

 there may be added to the remainder of the sample in the collecting bottle either 

 peptone (one per cent.) and sodium chloride (one-half per cent.) or a small amount 

 . of sterile bouillon ; after which the bottle and the contents are placed in the incubator 

 for twenty-four or forty-eight hours for the more satisfactory development of the 

 suspected bacteria. Under such conditions of added nutrition and warmth the typhoid 

 organism, the colon bacillus, and other important forms are apt to increase to a marked 

 extent, while the common water germs develop but poorly or are prevented entirely. 

 Diffusion or smear preparations on solid media are now made from this and grown 

 in the incubator. The final recognition and identification of such organisms must 

 depend thereafter upon the experience and skill of the observer, the separation of the 

 typhoid organism from the other varieties of the colon group being a difficult and 

 often uncertain task, although the recognition of the organisms as members of this 

 group is not difficult and quite convincing as evidence of the fecal contamination 

 of the sample of water from which they were obtained. 



Milk Examination. Milk as obtained directly from the cow is invariably con- 

 taminated with bacteria which have been on the surface of the teats or which have 

 been growing in the milk close to the openings of the milk ducts. Under the most 

 rigid cleanliness the fresh milk is apt to contain from ten to twenty thousand bacteria 

 to the cubic centimeter ; and milk is often supplied to purchasers which contains as 

 many millions to the same volume. The New York Board of Health prescribes as a 

 limit for good milk two hundred thousand to the cubic centimeter. For the most 



