54 MICRO-ORGANISMS AND FERMENTATION. 



after sterilisation infects with the bacteria, and then dilutes to 

 such an extent that small drops of 0-1 to 0-2 mm. diameter 

 contain on an average a single germ. Such drops are placed at 

 suitable distances with a mapping pen on the surface of a layer 

 of nutrient gelatine, where they immediately evaporate, and are 

 then protected by flamed cover-glasses. Much smaller drops 

 can be deposited on gelatine than on a cover-glass. The con- 

 tents of these specks can then be controlled under a high 

 power, and those noted which contain a single germ, a process 

 that is rendered easier by the fact that the bacteria appear 

 clear on a greyish-brown ground. If they can grow in gelatine 

 the development is allowed to continue, but if they require 

 high temperatures they may be conveyed to an agar plate 

 by cautiously raising the cover-glass, and as the speck of ink 

 is more firmly fastened to the glass than to the gelatine, the 

 germ is carried with the glass, so that it can be conveyed along 

 with this to the agar plate. After removing the germ from 

 the gelatine plate a drop of nutritive liquid can be placed 

 on the ink fleck of the cover-glass, and so the germ may be 

 developed in a liquid. The process can also be used for 

 the cultivation of anaerobic bacteria. 



Anaerobic bacteria demand special methods* of cultivation, 

 in which the atmospheric oxygen must be removed both from 

 the substratum and from the space in which the bacteria are 

 growing. Pure cultivations may be carried out in nutrient 

 gelatine or agar, in tubes filled almost to the top, the bacteria 

 growing in the bottom layers. 



A still better process consists in removing the air from 

 the test tubes by means of an air piimp. whilst the glass is 

 immersed in water at 30 to 35 C., after which it is hermetically 

 sealed. Another method is to remove the air with a current 

 of hydrogen. This is conveniently carried out in the following 

 way (Frdnkel) : A wide test-tube is fitted with a stopper with 

 two holes carrying two glass tubes, one of which reaches to 

 the bottom of the test-tube ; the other terminates just below 

 the stopper. When the vessel has been covered with nutrient 

 gelatine or the like, and sterilised, it is inoculated with the 

 growth, and a stream of hydrogen is passed through the long 

 tube. The tube is sealed up as soon as the air is completely 



