VIRULENCE OF THE BACTERIOPHAGE 165 



6. ATTENUATION OF VIRULENCE 



Just as it is possible to produce experimentally an increase in the 

 virulence of the bacteriophage, so also is it possible to cause an attenua- 

 tion of virulence. 



Exposure to high temperatures is one of the methods of causing such 

 an attenuation (d'Herelle and Pozerski^'^^) . The effect of heat is clearly 

 indicated by the data which follow. 



In the following experiments the suspension of bacteriophage under 

 test, previously filtered through a bougie, is taken up in capillary 

 pipettes, sealed at both ends, and completely submerged in a water- 

 bath maintained at the temperatures indicated in each experiment. 

 In each series of experiments 8 tubes with the suspension are maintained 

 for thirty minutes at temperatures of 60, 62, 64, 66, 68, 70, 72, and 75°C. 



Anti-Shiga bacteriophage 



Two drops of the suspension from tubes maintained at 60, 62, 64, 

 and 66°C., when introduced into suspensions of Shiga bacilH, cause 

 complete dissolution in less than fourteen hours. The tests repeated 

 with a second strain of Shiga bacilli give identical results. The bacterio- 

 phage heated to 68 and 70°C. causes dissolution with one strain of Shiga 

 baciUi but not with the other. When heated to 72 and 75°C. the bac- 

 teriophage fails to dissolve the organisms. 



One drop of each of these suspensions, which had received the bac- 

 teriophage previously maintained at 68, 70, 72 and 75°C., and which 

 had not been submitted to bacteriophagy is planted on slant agar. After 

 incubation, all of the cultures, except the last, which is normal, show 

 plaques characteristic of the presence of the bacteriophage. 



Serial passages may be effected, thus permitting the enhancement in 

 virulence of the bacteriophage attenuated by the action of temperature. 

 After two such passages, with the corpuscles heated to 68 and 70°C., 

 and after three passages with those heated to 72°C., dissolution in 

 liquid media is obtained. 



Comparable experiments have demonstrated that the bacteriophagous 

 corpuscles active for B. dysenteriae Flexner, B. dysenteriae Hiss, B. coli, 

 and B. paratyphosiis B, act in a quite similar manner. With the bac- 

 teriophage active for B. paratyphosus A attenuation begins at about 

 64°C. (at least with the strain tested). With that virulent for B. 

 typhosus attenuation is already apparent at about 62°C. In all cases, 

 when heated to 75°C. the bacteriophage is completely inactive, either 



