156 THE BACTERIOPHAGE AND ITS BEHAVIOR 



When we filter a fecal suspension, a specimen of river water, or even 

 tap-water, we obtain a filtrate which is a suspension of bacteriophage 

 corpuscles, but there is no reason for affirming that all of the corpuscles 

 present belong to the same race. Indeed, it is certain that quite the 

 contrary is true. Therefore, it is essential, especially for a study of the 

 processes of bacteriophagy, to isolate the corpuscles so that one may 

 work with strictly pure races, the issue of a single corpuscle. 



In order to accomplish such an isolation we have only to imitate bac- 

 teriological methods. For example, assume that we have a material 

 containing, aside from a variety of banal bacteria, a very few organisms 

 pathogenic for a given animal, such as the guinea pig. How would we 

 obtain a pure culture of this pathogen? We would inoculate a guinea 

 pig with some of the material, the pathogenic bacterium would multiply 

 within the body of the animal and at the death of the animal we would 

 find a pure culture of the bacterium in the blood or in some lesion. But 

 it is evident that if the initial material contained two bacteria pathogenic 

 for the particular animal used the two organisms might be recovered 

 in a mixed culture. If we wish to separate all of the bacteriophages 

 which may be found in a stool filtrate virulent for a given bacterial 

 species, B. typhosus for example, eUminating all of the races which do 

 not possess a virulence for this bacterium, we have only to adopt an 

 analogous method. 



Inoculate 0.1 cc. of this filtrate into 10 cc. of a B. typhosus suspension. 

 After 24 hours, bacteriophagy having taken place and the Typho- 

 bacteriophage corpuscles having developed, filter through a candle. 

 Inoculate 0.01 cc. of this filtrate into a fresh suspension of B. typhosus. 

 Incubate, filter, and continue such passages. After a sufficient number 

 of these have been carried out we may be sure that, because of the 

 extreme dilution brought about through the passages, no trace of the 

 original filtrate from the stool will remain. At this time all of the 

 corpuscles present will obviously have reproduced at the expense of the 

 typhoid bacillus; that is to say, there will be present only those resulting 

 from the development of corpuscles present in the initial filtrate virulent 

 for B. typhosus. None of the corpuscles of the original filtrate which 

 were virulent for other bacterial species and non- virulent for B. typhosus 

 will be recoverable after a certain number of transfers. At whose ex- 

 pense could they have multiphed? And if they have not multiplied 

 they have necessarily disappeared when the degree of dilution has 

 reached a certain limit, readily calculable.* 



* Determination of the limiting dilution in which there no longer remains a 



