94 THE BACTERIOPHAGE AND ITS BEHAVIOR 



their diameter remains indefinitely what it was at first. They are never 

 covered or encroached upon by the bacterial culture. At no time does 

 there exist within the extent of the plaque, whatever its size may be, 

 microscopically visible bacterial cells. The plaque is always rigorously 

 sterile. 



As soon as the culture is well developed, as after 18 to 24 hours of 

 incubation, if the centre of such a plaque is touched with a platinum 

 wire and this is immersed in a culture of Shiga bacilli the bacteriophage 

 develops in this suspension and the latter is dissolved after a few hours. 

 The plaque, although sterile, is not ultrasterile; it is in fact a colony of 

 the bacteriophage corpuscles. 



Furthermore, if a trace of the bacillary growth at the periphery of a 

 plaque is taken with a platinum wire and seeded on agar it remains 

 sterile and inoculation into a bacterial culture shows that the bacterio- 

 phage is present there also. But when the bacillary layer is taken, 

 not at the immediate edge of the area, but at a distance of two milli- 

 meters from it, for example, and planted, the tubes show the growth of a 

 normal culture. The bacteriophage is not found. 



If the culture showing the plaques is returned to the incubator and 

 the tests are repeated three or four days later, that is, culturing the 

 bacillary growth at a distance of two millimeters from a plaque onto 

 agar and into a suspension it will be found that the bacteriophage is 

 there present at that time. The bacteriophage has, therefore, gradually 

 invaded the bacillary layer. This invasion is always slow — ^proceeding 

 more and more slowly as time progresses — so that the ring invaded, 

 even after several months, amounts to a zone but a few milUmeters wide. 

 Beyond the hmits of this zone the Shiga organisms remain cultivable 

 just as long as they do in a normal control culture without the bacterio- 

 phage. 



The question immediately arises as to why the bacteriophage does not 

 invade the entire layer of bacterial growth. For this there are two 

 reasons. The bacteriophage attacks the bacterial cell most readily 

 when the bacterium is young. When placed upon agar the bacterio- 

 phagous corpuscles find themselves located in the immediate vicinity 

 of bacilli which reproduce actively as soon as they are deposited upon 

 a nutrient medium. They find then, within their range, very young 

 bacilh distributed in a very thin layer over the agar. Dissolution is 

 thus possible and the apparent sterihty of the plaque results. But 

 beyond this zone invaded by the bacteriophage during the first few 

 hours the bacilh develop freely forming a layer of increasing thickness 



