ISOLATION OF ANAEROBES 449 



1 . To segarate non-sporulating anaerobes from aerobes 



1. Heat to 56°-58° for five or ten minutes. This occasionally 

 serves the purpose. 



2. Try to induce sporulation by growing the mixture on al- 

 kaline sugar-free medium, such as alkaline egg, or serum medium 

 (von Hibler, 1908, p. 189). When the anaerobes form spores, 

 heat. This procedure is a sure method of freeing B. WelcMi from 

 ordinary aerobes : incubate for four days. This organism is found 

 in a sporulating condition in soil and in fecal material. 



3. Try a pathogenicity test. If the organism sought is patho- 

 genic it may be recovered in pure culture from the animal tissues. 

 Use this method for B. Welchii, B. egens, B. fallax. 



4. Use selective media. For the Welch bacillus use milk or 1 

 per cent glucose broth. Inoculate it with a pipette, a fresh tube 

 of medium every twelve hours if possible. 



5. Use good anaerobic methods. Cultivate the material on 

 meat medium in strict anaerobiosis, inoculate in agar dilution 

 tubes that have been thoroughly boiled, and fish the colonies. 

 This technique is described on page 461. 



6. Northrup suggests the use of a 25-cc. burette, in which the 

 organisms of an inoculated mixture will, on short incubation, sort 

 themselves out, the aerobes growing above, the anaerobes below, 

 where they may be drawn off through a stopcock. 



IL To separate non-sporulating anaerobes or reluctantly sporu- 

 lating anaerobes from other sporulating anaerobes 



1. Use selective media, milk, with short incubation periods, 

 for B. Welchii. 



2. Use animal inoculation. 



3. Use shake cultures. 



4. Use semi-anaerobiosis: The non-sporulating anaerobes are 

 naturally more resistant to oxygen than the sporulating ones. 



Aside from B. Welchii this sort of organism is rarely sought after 

 or noticed. Few non-sporulating anaerobes are described, and 

 the group has been generally neglected, but careful methods show 

 that non-sporulating anaerobic rods and cocci are not uncommon. 



