MEDIA MAKING AND APPARATUS 263 



the best methods are used, the count is always less than the actual 

 number of bacteria present, for the following reasons : 



(a) Many bacteria in process of multiplication are held to- 

 gether by adhesive membranes in pairs, chains or masses. It is for 

 the purpose of separating bacteria thus joined, as well as to obtain 

 an even mixture, that the sample itself and the diluted sample when 

 plating are shaken. This shaking, while it breaks up larger masses 

 and shortens long chains, does not to any great extent break apart 

 the shorter chains, diplococci, etc. Each of these groups of bacteria, 

 when caught in the solid medium, develops as a single colony. 



(b) It is impossible to obtain a medium suited to the food 

 requirements of all species or races of bacteria. It has been found 

 by experiment that a medium consisting chiefly of a watery extract 

 of raw meat and peptone', alkaline to litmus and slightly acid to 

 phenolphthalein, will furnish the best food for the greatest number. 



(c) These varying forms of minute vegetable life require 

 "varying temperatures for their best development. Many forms 

 which will develop at room temperature will not grow at body 

 temperature. Some require a very high temperature for their best 

 growth. 



(d) Some bacteria develop in an atmosphere free from oxy- 

 gen, some only where oxygen is present ; many are facultative, i. c, 

 growing under either condition. Bacteria which require an oxygen- 

 free atmosphere do not develop in plates as generally prepared. 

 Bacteria requiring oxygen, if deep in the medium, develop but 

 slowly, as they then obtain free oxygen only by diffusion. 



(e) Many forms are slow in developing into visible colonies, 

 some requiring three or four days. On the other hand, in plates 

 grown for several days, many small colonies are obscured in the 

 growth of larger ones. 



(f) Each bacterium requires a certain amount of nourish- 

 ment for development. There are also antagonistic forms which 

 will not develop in close proximity to each other. It therefore 



