56 ISOLATION AND CULTIVATION OF ANTAGONISTS 



pathogenic organisms. Although it had been previously established that 

 many spore-forming bacteria are capable of producing substances that 

 have antibacterial properties, as shown by the work of Pringsheim 

 (705), Much (621), and others, Dubos was the first to succeed in iso- 

 lating in crystalline form the active substances involved and in demon- 

 strating their chemical nature. He utilized for the isolation of the or- 

 ganisms the enrichment culture method. This consisted in adding 

 repeatedly various pathogenic bacteria to a soil which, as a result, be- 

 came enriched with antagonistic organisms capable of destroying the 

 bacteria added j these organisms were then isolated by appropriate pro- 

 cedures. The isolation of the specific substances will be described later 

 (page 156). 



These investigations, as well as the work of Fleming (265) and 

 other British investigators (3, 7, 8, 113) on the antibacterial properties 

 of molds belonging to the PenicilUum notatum group, served as the di- 

 rect stimulus to numerous studies that followed. The entire series of 

 studies led to the development of simple methods for the systematic iso- 

 lation of microorganisms capable of inhibiting the growth of fungi and 

 bacteria, both pathogenic and saprophytic (857, 934), and for separat- 

 ing many of the antibiotic substances produced by these organisms. 



METHODS OF ISOLATING ANTAGONISTIC 

 MICROORGANISMS 



Four, and possibly five, methods are now available for the isolation 

 of antagonistic microorganisms from natural substrates such as soil, 

 stable manure, composts, sewage, water, and food products. These 

 methods are different in nature, but they are all based on the same prin- 

 ciple, that of bringing a living culture of a bacterium or fungus into 

 close contact with a mixed natural population, thereby allowing certain 

 members of this population to develop at the expense of the added 

 culture. 



Soil Enrichment "Method 



By this method a soil Is enriched with known living pathogenic bac- 

 teria. Fresh garden or field soil is placed in glass beakers or pots, and 



