446 HILDA HEMPL HELLER 



molds to grow through them; where a mold can grow a bacillus 

 can follow. (4) During prolonged incubation water of con- 

 densation may even run into the tubes from the top of the jar. 

 (5) If stored in closed cans molds may grow through the plugs. 

 Workers should take these points into consideration in planning 

 their work. Anaerobic jars are exceedingly convenient and prac- 

 tical for periods of incubation under four or five days, and for 

 much anaerobic study twenty-four to forty-eight hours incubation 

 is sufficient. Prolonged incubation should be made under vase- 

 line or in the case of sugar-free media in exhausted sealed tubes. 

 Sealing of tubes is inadvisable where carbon-dioxide may be so 

 confined that it produces an acid end-point. Re-incubation of 

 cultures in exhaust jars should be cautiously undertaken so that 

 the medium may not boil up to the cotton plugs. Anaerobic jars 

 which do not require exhaustion are preferable for re-incubation 

 of cultures. 



The commonest contaminators of my cultures have been cocci 

 and molds, not anaerobes. The reason that anaerobic contami- 

 nation of anaerobe cultures is so very common probably lies 

 principally in the uncritical handling of such cultures. If a coc- 

 cus or mold contaminates a culture the worker immediately kills 

 such an organism, but if an anaerobe enters the tube it proceeds to 

 multiply unmolested. Daily watchful observation of the cul- 

 tures studied is absolutely necessary for successful anaerobic work. 

 I have not found indications of any so-called symbiotic tendency 

 that makes anaerobes more difficult to isolate than aerobes. 



Anaerobes vary greatly in their behavior and requirements, and 

 the method of isolation must be adapted to the problem in hand as 

 it turns up. Each combination of two or more species of organ- 

 isms presents different elements for consideration and for adap- 

 tation of technique. There is no one method that is always best, 

 and it is only after a worker knows something about the nature of 

 the particular organisms that he is dealing with, their cultural 

 behavior, and their morphology in the medium in which he reg- 

 ularly grows them, that he is able quickly and surely to isolate 

 numbers of strains. 



