466 CONTROL OF CULTURES 



it was hoped that prolonged desiccation, which we knew would not 

 harm the cysts, would cause the death of the bacteria. Tests were made 

 after one, two, three, and six months (duplicate preparations being 

 placed by desiccators) by placing the plungers into tubes of sterile ex- 

 cysting fluid (yeast extract). While the length of time required to show 

 turbidity increased with the time of drying, viable Aerobacter were pres- 

 ent in every case. Heating the dry sponge to 80° C. for three hours did 

 not kill the cysts, nor did it kill the Aerobacter. The heat resistance is 

 very different in the dry and the wet states, as evidenced by the fact 

 that one hour of heating at 50° C. is enough to kill a suspension of this 

 bacterium. 



It was found, however, that a simple manipulation could be employed 

 with eight-months'-old sponge-glass plunger preparation to obtain large 

 numbers of sterile ciliates. U-shaped tubes were partly filled with yeast 

 extract and sterilized, and the sponge-glass plunger was inserted asepti- 

 cally into one arm. The tube was placed in an upright position and the 

 minimum time was allowed for the ciliates to excyst. Then, without 

 disturbing the fluid, the plug from the other arm was lifted and some 

 of the fluid containing the freshly excysted ciliates was withdrawn. These 

 ciliates were sterile, provided not too long a time had elapsed since the 

 sponge was introduced and provided the fluid was not unduly disturbed. 

 What appears to happen is that the dry bacteria take a considerable time 

 to become active and to disperse through the medium, while a few sec- 

 onds after the ciliates leave their cyst walls they migrate to the top of 

 the fluid. It should be remembered that this method has been used only 

 when the associated bacteria have been reduced to a single species. By 

 careful manipulation and by having the conditions just right, it might be 

 possible to use this method on wild, cyst-forming material, but this is 

 only a conjecture as it has not been done to date. 



The possibility of using radiation for sterilization was investigated by 

 Brown, et al. (1933). They found that E. taylori, trophic or cysts, with- 

 stood a much longer exposure to X-rays (2,110 Roentgen units per 

 second) than did the associated bacteria (in this case Pseudomonas 

 fluorescens and B. coli) . They were able to obtain many sterile ciliates 

 by this method. Here again, the success of the method appears to be due 

 to the specific type of bacteria present, and it is extremely doubtful if. 



