64 THE BACTERIOPHAGE 



that the number of granules consequent upon the rupture of a 

 cell amounts to between 15 and 25. There is, therefore, a great 

 probability that the granules are actually the ultramicroscopic 

 bacteriophagous organisms. 



2. We may consider the second case, that of a minimal inocula- 

 tion. In this case the medium becomes more and more turbid 

 before lysis actually commences. 



A suspension of Shiga bacilli, containing 250,000,000 per cubic centimeter 

 is inoculated with 0. 0001 cc. of a culture of the bacteriophage, a very active 

 strain being selected. 



After 30 minutes the medium has its original turbidity; essentially that 

 of a normal culture of the Shiga bacillus. 



After one hour the original turbidity is still maintained . When smeared 

 and stained all the bacilli are of normal shape, but an occasional form 

 stains poorly. 



After two hours the culture is about twice as turbid as at first. There 

 is amorphous debris in the bottom of the tube. All of the bacilli appear 

 to stain as normally. Many of the bacilli (about two in every three) are 

 about four times the normal length, that is, of the bacilli used to seed the 

 culture, and there are all intermediary forms. Oval and spherical forms 

 are relatively numerous, but they are always fewer than would be expected 

 from a comparative ultramicroscopic examination. These forms are 

 indeed very fragile and are particularly liable to destruction during fixation 

 upon the slide so that their demonstration in stained preparations requires 

 great care. 



After three hours the suspension is slightly cloudy. The bottom of the 

 tube is covered with fine debris without definite form, with, from place to 

 place, great amorphous masses and numerous granules resembling those 

 encountered in very old cultures of normally grown Shiga bacilli. Only 

 a single spherical form can be detected in a ten-minute search. Each field 

 may contain a dozen large bacilli, well stained. 



After four hours the turbidity is very slight. There is somewhat less 

 material in the bottom of the tube, and this shows only a single poorly 

 stained bacillus to a field. 



After six hours the medium is limpid. There is still less deposit in the 

 bottom of the tube and it is with difficulty that a single poorly stained 

 bacillus may be found in searching 25 fields. 



After eighteen hours nothing at all can be seen in the preparation. 



As is evident, the aspect of this preparation differs but little 

 from that seen in the former case, the only departure being that 

 the bacilli which have grown immediately after inoculation, be- 

 fore the action of the bacteriophage becomes operative, present 

 abnormally large forms. 



