PARAMECIUM IN PURE CULTURES OF BACTERIA 435 



fresh added with a sterile pipette. This was repeated five times 

 in the same way. Samples of the five wash fluids were then 

 plated in Petri-dishes and the following number of bacteria found 

 present. 



TABLE 3 

 Showing reduction iti numbers of bacteria by washing Paramecium through 5 wash 



fluids in a centrifuge 



This showed that it was possible to reduce the number of 

 bacteria greatly, but the results were not satisfactory; absolute 

 sterility of Paramecium was desired. The method was not very 

 practical since a great deal of time was consumed, the wash 

 waters had to be drawn off immediately after the centrifuge 

 stopped or the paramecia rose in a body and prevented the 

 removal of the wash water. The method has the advantage of 

 permitting the reduction in number of bacteria to approximate 

 sterilization of large numbers of the protozoa at one time, and 

 would be useful for any experimental work which demanded 

 large numbers of the protozoa. 



Another serious drawback to the centrifuge method is the 

 difficulty of keeping the wash waters free from contamination by 

 bacteria of the air. The air may contain such enormous numbers 

 of bacteria that instruments and media which are sterile to 

 start with will be contaminated unless precautions are taken 

 to prevent the contact of air bacteria. A sterile pipette laid 

 down on the table is no longer sterile, a wash water left unpro- 

 tected is soon contaminated by air bacteria. Any successful 

 sterihzation method must, therefore, not only rid Paramecium 

 of the bacteria sticking to it, but must prevent the possibihty 

 of contamination from any source, air or otherwise. 



In the next method many of these difficulties were overcome. 

 A single Paramecium was isolated with a sterile capillary pipette 

 and washed through a number of sterile wash fluids in watch 

 crystals placed inside of sterile Petri-dishes. Under the pro- 

 tecting cover of the dish danger of outside contamination was 



