CHAPTER XVIII. 



ISOLATION OF BACTERIA IN PURE CULTURE. 



As has been stated, the thorough study of a bacterium 

 depends on first getting it in pure culture. In the early 

 days of bacteriology supposedly pure cultures were obtained 

 by (1) dilution in liquid media. A series of tubes or flasks 

 containing sterile liquid media was prepared. Number one 

 was inoculated with the material to be examined and thor- 

 oughly mixed. A small portion of the mixture was trans- 

 ferred to number two, and mixed; from this to number 

 three, and so on until a sufficient number were inoculated, 

 the last three or four in the series receiving the same amounts 

 of a very high dilution of the original material. If one or 

 two of these latter showed a growth and the others not, it 

 was assmned that the dilution had been carried so far that 

 only a single organism was transferred and therefore the 

 culture obtained was "pure." The method in this crude 

 form is too uncertain to be of value today and recourse is 

 had to more exact means. The procedure most widely used 

 is that of (2) "plating out" by means of gelatin or agar 

 plates. The material to be plated out is diluted by trans- 

 ferring to three or more tubes of melted gelatin or agar as 

 in the first method and then all the tubes are poured into 

 Petri dishes and grown under suitable conditions. By 

 proper mixing in the tubes the bacteria are well scattered 

 through the medium which holds the individual organisms 

 separate when it solidifies. On some of the plates a suffi- 

 cient dilution will be reached so that the colonies develop- 

 ing from the bacteria will be so few that they are separate 

 and pure cultures may be obtained by inoculating from one 

 of these a tube of the appropriate medium (Figs. 129 

 to 132). The chief uncertainty with this method is that 

 occasionally two kinds of bacteria stick together so closely 



