RESISTANCE OF BACTERIA 199 



illon. We will thus have on the one hand 1 cc. of bouillon containing 

 10,000 million cocci and on the other hand a flask containing the same 

 number of cocci suspended in 100 cc. of bouillon. Inoculate the two 

 suspensions with the same quantity (1- 10~^) of a bacteriophage of maxi- 

 mum virulence. After incubation it will be found that the hquid in the 

 flask is clear and remains so indefinitely, and the corpuscles present 

 show a maximum virulence. At this same time the cubic centimeter 

 quantity is extremely cloudy and after filtration it will be found that only 

 attenuated corpuscles are present, their virulence being not even 

 moderate. 



It is possible to demonstrate this same fact in a still more conclusive 

 manner. The cubic centimeter of suspension is inoculated with 0.1 

 cc. of a bacteriophage suspension; the hundred cubic centimeters with 

 a hundred-millionth of this amount of the same suspension. After 

 incubation the results are absolutely comparable to those of the preced- 

 ing experiment. Corpuscles of a maximum virulence are found in the 

 100 cc. quantity and corpuscles of a low virulence in the cubic centi- 

 meter. In view of the fact that the number of bacteria was the same in 

 both cases,* the attenuation of virulence can not, then, be explained by 

 assuming that the bacteriophage disseminates its action through too 

 large a number of bacteria. This is, however, a priori certain since 

 we know that the bacteriophage acts as a unit; a single bacteriophage 

 inoculated into 10 cc. of a normal suspension, that is, placed in contact 

 with 2500 million bacteria, effects a complete bacteriophagy. 



What, then, can be the cause of the attenuation in virulence? The 

 single point which differentiates the process of bacteriophagy as it 

 occurs in one suspension from that taking place in the other is that in the 

 100 cc. of suspension a secondary culture does not develop. None of the 

 bacteria there acquire a resistance. In the 1 cc. the bacteria become 

 resistant. All of the other conditions such as the number of corpuscles 

 and the number of bacteria, the state of these bacteria and of the cor- 

 puscles combined with them, the nature of the medium and the tem- 

 perature are alike in the two suspensions. This being the case, it 

 seems necessary to conclude that the single difference observed, that is, 

 the acquisition of a resistance by the bacteria, is responsible for the 

 attenuation of the bacteriophage corpuscles. 



Of interest in this same connection are the observations of Bordet,^'' 

 as well as those of Gratia and de Kruif."^ They have found that if 



* One might even accomplish the experiment by placing three or four times as 

 many bacteria in the 100 cc. as in the cubic centimeter. 



