VIABLE BACTERIA IN YOUNG CULTURES 421 



do not appear till tho socond or third days after inoculation. 

 Later than this no furtlier numerical increase was observed. 



The effect of shaking on broth cultures 



In view of the fact that the discrepancy in the relation of 

 the ^•iable to the total count has frequently been attributed to 

 clumping of the organisms, leading to the development of a 

 single colony from one mass of organisms instead of to several, 

 and in view of the fact that the aim in this work was to eliminate 

 every factor hindering the development of the maximum number 

 of colonies, it was decided to investigate the effect of shaking on 

 the cultures before the counts were put up. Shaken and un- 

 shaken cultures were compared, both total and viable counts 

 being performed on them, the conditions being rendered identical 

 except for the single process of shaking. As a result of several 

 experiments on broth cultures of different ages it appeared that 

 shaking in the case of a broth culture under twelve hours old led 

 to no increa.se in cither count; in a twenty-four hour culture, 

 however, an increase of somewhere in the neighbourhood of 15 to 

 25 per cent could be substantiated in both. In the estimation 

 of the count in young cultures, therefore, shaking is of no value: 

 for the estimation of the count in older cultures, shaking leads 

 to an absolute increase in both total and viable counts, but as the 

 relation between the two remains unaltered, it is of no advantage 

 so long as it is only this ratio wliich is being investigated. 



The optimum number of colonies per tube 



Speaking of the technique of the plate count Park and Williams 

 (1908) remark: 



It is very important to remember that when more than 200-300 

 bacteria start to develop in the agar or gelatin contauied in a plate 

 some develop colonies which fuse together, while others are inhibited 



before they develop visible colonics If possible, dilutions 



should be made so that plates will contain between 40 and 400 colonies. 



This is one of the few references I have been able to find to the 



