INVESTIGATIONS ON A LYSOGENIC BACILLUS MEGATERIUM 



After two hours, there are 82 bacteria: all of the liquid is sampled, no bacterio- 

 phages. 



Discussion. These experiments, as well as those which shall be described in the 

 two following paragraphs, lead to the thought that a lysogenic bacterium 

 multiplies without producing bacteriophages. They demonstrate that B. 

 megaterium 899 can grow and multiply without liberating bacteriophages. Thus, 

 liberation of bacteriophages is not a phenomenon which necessarily accompanies 

 growth of a population of a lysogenic bacteria ; it appears, rather, to be a contin- 

 gent phenomenon. 



IV. Maintenance of Lysogenic Power 

 IN THE Absence of Free Bacteriophage 



A motile diplo-bacillus is sampled from a culture in peptone water (proteose 

 peptone at 1%). This bacillus is successively transferred in six drops of the same 

 medium. It is left in the sixth. During the entire length of this experiment the 

 diplo-bacillus has remained motile. With B. megaterium the separation of the 

 daughter members generally takes place when the daughter bacteria are already 

 partitioned off. Once this separation releases two daughter diplo-bacilli, one 

 injects into the drop approximately \ of its volume of new medium. One assures 

 a mixing of the fluids by repeated aspiration back and forth of approximately 

 half of the fluid and then removes one of the bacteria in a volume essentially equal 

 to the volume of fluid which has been added. This sample is transferred into a 

 suspension of sensitive mutilat in soft agar and this suspension is then spread on 

 an agar surface. This operation was repeated until the 10th division. At this 

 point, the two daughter bacteria were sampled separately, the latter with the 

 total of the residual fluid. The 11 plates that had been inoculated all showed a 

 single plaque centered by a bacterial colony, with the exception of the first and 

 the eighth, which showed no plaque whatsoever. The absence of the lysogenic 

 colony on the two negative "plates" in this experiment, as in the following, 

 probably results from an accidental loss of the bacterium. This experiment was 

 repeated in peptone water to which glucose and a liver extract had been added. 

 The original diplo-bacillus was followed until the 19th division, after which the 

 two final bacteria were plated. The 20 plates, with the exception of the 4th and 

 the 16th which showed no plaque whatsoever, each had, as in the previous series, 

 one plaque centered by a lysogenic colony. One of these centered colonies, after 

 being picked and isolated, proved itself to be lysogenic: the filtrate of its culture 

 contained bacteriophages. 



The absence of any plaque, except for the single plaque centered by a bacterial 

 colony, shows that no free bacteriophages ever appeared in these microcultures. 

 During the experiments, there was never any noticeable decrease in the size of 

 the diplo-bacillus, as would have occurred if one of its members had lysed, nor 

 was lysis ever observed. Incidentally any lysis would have brought some 

 disturbance in the rhythm of the divisions; but the 19 divisions took place in 

 486 minutes at regular intervals of 25 ± 3 minutes. 



Discussion. It was established by Bordet and Renaux (1928) that lysogenic 



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