260 THE BACTERIOPHAGE AND ITS BEHAVIOR 



alcohol and through sterile distilled water. Fragments of these nodules, 

 cut up with sterile instruments, were then distributed over solid media 

 which had previously been seeded with a culture of B. radicicolum, 

 derived either from clover, lupin, or sarrandella. 



They found, after incubation, a zone of clarijfication attaining a 

 diameter of from 1 to 2 cm. around the fragments of nodule. This 

 activity was specific, that is to say, the nodule fragments from lupin 

 were the center of a plaque with the culture of B. radicicolum derived 

 from lupin, while the culture was normal when the bacterium was 

 derived from clover; mutatis mutandis. 



They also ground up these nodules with sterile saline in a mortar and 

 filtered the material through a candle. By this procedure they isolated 

 very virulent bacteriophages, usually specific. The filtrates brought 

 about a dissolution of the bacteria, but rather slowly, requiring about 

 10 days at ordinary temperature. Nevertheless, it was complete, while 

 after this same interval the controls were very turbid. When drops 

 of the filtrate were placed on agar no colonies developed within the area 

 covered by the drop. When suspensions, to which a small quantity 

 of the filtrate had been added, were spread over an agar surface plaques 

 appeared after incubation. It may be added that the paper is accom- 

 panied by photographs which show beyond question that the reaction 

 was a typical bacteriophage phenomenon. 



Among the races isolated they found some whose action extended to 

 strains of B. radicicolum other than those isolated from nodules of the 

 same plant. A race of bacteriophage isolated from clover nodules 

 possessed a weak but nevertheless definite virulence, for a B. radicicolum 

 strain isolated from the bean. A bacteriophage isolated from lupin was 

 weakly virulent for strains of B. radicicolum isolated from the clover 

 and from the bean. On the contrary B. coli, Azotohacter chromaceum, 

 Radiobacter, B. fluorescens liquefaciens and B. violaceus were absolutely 

 insusceptible to any of the races of the bacteriophage isolated. 



In three attempts out of seven they isolated bacteriophages from the 

 stalks of the clover virulent for the B. radicicolum present in the nodules 

 of the roots. All attempts to isolate them from the leaves were un- 

 successful. Working with 100 gram samples of garden soil and of cul- 

 tivated earth they obtained virulent races of this bacteriophage but 

 with forest earth and with uncultivated soils their results were uniformly 

 negative. 



