VIRULENCE OF THE BACTERIOPHAGE 163 



ceding titration. If this time it is found that the 10 suspensions inocu- 

 lated are bacteriophaged, that one where the process occurred most 

 vigorously is selected and is filtered. For the next passage, as is obvious, 

 the next higher dilution is selected. Simple observation of the 10 sus- 

 pensions here will show the dilution to be used for the following passage. 

 If there are only one, two, or three of the 10 suspensions bacteriophaged, 

 it may be well to use the same limiting dilution as that previously 

 used. If there are 8, 9, or 10 suspensions bacteriophaged it is desirable 

 to take the next higher dilution for the next passage. 



At first sight this method appears rather complicated but it gives 

 excellent results. In this way I have been able to obtain bacteriophage 

 races causing regularly a complete and invariably permanent dissolution 

 of normal bacterial suspensions. Those who have worked with the 

 bacteriophage realize how difficult it is to attain a virulence sufficiently 

 high to preclude the development of secondary cultures. 



A number of authors, among whom may be mentioned Izar,^" and 

 Janzen and Wolff"^ have observed that all races of the bacteriophage 

 do not acquire virulence with equal facihty. This is quite true. And 

 in this same connection I have found^^^ that certain races of very low 

 virulence may lose their activity with the first passage, and for this we 

 will see the reason later. But it is likewise true that the mode of pro- 

 cedure has a very great influence upon the result. Just to indicate the 

 importance of the matter of the bacterial strain selected I may state 

 that upon several occasions, when working with a single race of bacterio- 

 phage, I have made two parallel series of passages for the purpose of 

 increasing the virulence. One series was made at the expense of one 

 strain of the bacterium, the second at the expense of another strain of 

 the same species. It was found frequently that virulence increased 

 rapidly with one strain and but very slowly with the other. Further 

 study showed that this difference in result was due to the fact that one 

 of the two bacterial strains developed a resistance far more readily than 

 the other. 



With certain strains which possess the faculty of developing resist- 

 ance in a high degree, the result of serial passages is not an increase in 

 virulence but the bacteriophage is overcome and sometimes it disappears 

 even in the first passage. We will study the phenomenon of bacterial 

 resistance in the next chapter so that we will not enter into further detail 

 here, resting content with the statement that the choice of the bacterial 

 strain at the expense of which the passages are to be made is of extreme 

 importance. It might be well to add, however, that once the increase in 



