48 MICRO-ORGANISMS AND FERMENTATION. 



quantity of yeast was dried and ground with powdered gypsum. 

 The fine dust was thrown into the air at as great a height as 

 possible, and whilst the particles were floating down, a series 

 of vacuum flasks were opened. Thus some of the yeast cells 

 which were finely distributed throughout the dust cloud might 

 penetrate singly into some of the flasks. 



The first application of the dilution method to bacteria 

 was made by Lister (1878). To prepare pure cultures of lactic 

 acid bacteria he first determined microscopically the number 

 of bacteria in a minute drop of sour milk, counting them in 

 several fields of the preparation, and thus calculating their 

 whole number. He then estimated the amount of sterilised 

 water it was necessary to add so that after dilution there 

 would be on an average less than one bacterium in each drop. 

 With five such drops he inoculated in one case five glasses 

 containing boiled milk. The result was that the milk in one 

 of these coagulated, showing that it contained Bacterium 

 lactis, whilst the four other glasses remained unaltered, and 

 did not show the presence of bacteria. The same method 

 was subsequently employed by Nageli and Fitz (1882). 



In comparison with the physiological methods the dilution 

 method now described is a distinct advance ; indeed we have 

 thus approached much nearer to the goal. On the other 

 hand, it is clear that, even if the dilution is carried as far 

 as in the case mentioned, in which only one of several flasks 

 shows development, it is not yet proved that this one flask 

 has received only one germ. Thus, there is still great un- 

 certainty, even in cases where the individuals with which we 

 are working can be counted. Moreover, it is extremely 

 difficult to count individual bacteria, and often, indeed, quite 

 impossible. In all cases the accuracy of such calculations is 

 very questionable. Thus, the problem remains to be solved : 

 How are we to distinguish the flasks which have only received 

 one cell from those which, notwithstanding calculation, have 

 been infected with several cells ? For the bacteria, no means 

 have as yet been found of solving this difficulty. 



In the case of yeast the process was further developed 

 by Hansen (1881). He employed dilution with water, in the 

 following manner : The yeast developed in the flask is diluted 



