PREPARATION OF PURE CULTURE. 47 



is facilitated. Thus the treatment described with tartaric 

 acid or hydrofluoric acid gradually converts the mixture into 

 a growth of wild yeasts. If a mass of yeast is strongly con- 

 taminated with bacteria, a cultivation at very low temperatures 

 may possibly suppress the bacteria if the yeast are able to 

 develop under these conditions. If it is desired to obtain a 

 pure culture of the lactic acid bacteria from a mash, the material 

 may be prepared by keeping it a short time at 50 to 55 C. 

 At this high temperature many bacteria cannot thrive, whilst 

 certain species of lactic acid bacteria can stand a high degree 

 of heat, and thus spread throughout the material. In the 

 same way in the cultivation of film-forming bacteria, such as 

 acetic acid bacteria, the growth may undergo a preliminary 

 purification by repeated inoculation of the film in fresh liquids. 

 This process was used by Pasteur in his researches on acetic 

 acid bacteria. To make an approximate separation of a large 

 and small species of yeast in a mixture we may resort to 

 decantation or filtration through a medium which will allow 

 the small cells to pass. 



It is common to all these methods that with more or less 

 luck it is possible to bring about the preponderance of one or 

 more groups of micro-organisms in a mixture, but it is obviously 

 impossible to obtain in this way the exclusive presence of one 

 particular species. 



(6) Dilution Methods. The second group of methods 

 employed for physiological purposes embraces the dilution 

 methods, or " fractional cultivation," the principle of which 

 is to dilute the material to such a degree that it is ultimately 

 possible to isolate a single cell. Brefeld used the dilution 

 process for his botanical investigations of moulds, where 

 he was able, owing to the size of the cells, to insure that only 

 a single cell was present in a small drop of water in the moist 

 chamber. He added sterile nutritive fluid, and observed the 

 growth of the cell. Pasteur utilised air (Etudes aur la biere, 

 1876) as a diluting medium for preparing pure cultures. He 

 started from the fact that if nutritive liquids are exposed 

 to the action of air, fermentation takes place, excited by the 

 germs which fall on to the surface. To isolate single germs 

 from the yeast mass, he proceeded as follows : A small 



