BACTERIOLOGICAL ASPECTS OF DEHYDRATION 117 



have invariably been absent. The methods of quantitative de- 

 terminations are at best imperfect because of the character of 

 the materials examined. It was found necessary therefore to 

 make multiple examinations, and derive the "normal" figure 

 from the results obtained. The nature of the materials studied 

 rendered it impossible to extract, plate and count all the organ- 

 isms present, but it was possible to secure a result which showed 

 the general order of magnitude of the microbic population. The 

 method found most satisfactory was as follows : 



Ten grams of the sample, well broken up, with aseptic precau- 

 tions, were weighed out into a tared sterile dish and later trans- 

 ferred to an Erlenmeyer flask containing 200 cc. of sterile tap 

 water. After thorough mixing in order to get all the particles 

 completely wetted, the sample was placed in a 37° incubator for 

 two hours to permit absorption of water, return of tissues to as 

 near normal as possible and easy separation of bacteria. At the 

 end of this time the sample was again thoroughly agitated and ali- 

 quot portions removed, dilutions prepared and plates made as 

 rapidly as possible. The two hour period of infusion was deter- 

 mined upon as a result of preliminary studies. Soaking is neces- 

 sary to make it possible to remove the bacteria from the surfaces 

 of the particles, but too long continued infusion permits the 

 growth of bacteria which have been restored from the plasmo- 

 lysed condition. 



The media which were found most satisfactory for quantitative 

 studies were plain agar and glucose agar or litmus glucose agar 

 for the bacteria, and Czapek's medium for fungi. Plating on 

 beer wort gelatine or agar was carried out for a time for the pur- 

 pose of enumerating yeasts, but the results were so uniformly 

 negative that this process was discontinued. 



Incubation at 37° for forty to forty-eight hours with plates in- 

 verted was found to be most satisfactory. Counts were made 

 with a reading glass, the dish being placed on a black ruled plate. 

 Examination of each plate was made for predominant types of 

 colonies, and these were fished and later identified in order to 

 determine as fully as possible the source and kinds of organisms. 



