SYLLABUS 115 



wild type forms a turbid plaque and the host range mutant a clear 

 plaque. 



To determine the time between infection and lysis and the average 

 phage yield of singly or multiply infected bacteria, the one step growth 

 technique is used. After mixing bacteria and phages in the desired 

 proportion, and allowing a few minutes for adsorption, unadsorbed or 

 free phage (and only those!) may be inactivated by anti-phage serum. 

 The mixture is thereupon highly diluted so that a sample used for 

 plating contains only a few hundred infected bacteria. Each infected 

 bacterium gives a single plaque if it did not burst before plating. In 

 case the free phage were not inactivated previously by antiserum, there 

 would be two indistinguishable kinds of plaques from two kinds of 

 "infective centers," i.e., the infected and not yet lysed bacteria and the 

 free phages. Plating samples of this diluted mixture at short time inter- 

 vals give constant plaque counts for a certain length of time, then 

 suddenly the count rises from plate to plate until it reaches a new, 

 much higher level. The sudden rise in count is brought about by the 

 first bursts of infected bacteria, which distribute the phages produced 

 by them evenly throughout the dilution tube, increasing thereby the 

 number of infective centers, since each newly released phage particle 

 can now form a plaque of its own. The count levels off after the last 

 infected bacterium has burst. The time interval between infection and 

 the beginning of lysis is called the latent period, the time between be- 

 ginning and end of lysis the rise period, and the ratio of the count at 

 the end to that at the beginning the step size. From the latter it is easy 

 to calculate the average burst size of an infected bacterium. The whole 

 experiment can be plotted on graph paper (count vs. time) in order 

 to obtain a one step growth curve. 



Individual burst sizes are determined by another type of experi- 

 ment where the infected bacterial suspension is diluted so far that a 

 single drop of the suspension contains on the average much less than 

 one bacterium. Distributing drops into a great number of tubes and 

 plating each tube after the latent period is over, gives no plaques at all 

 for most of them, the rest containing mostly the yields of single bursts. 

 Again the Poisson Law serves for detailed analysis of this single burst 

 experiment. 



In many experiments it is necessary to count free phage and in- 

 fected bacteria separately. This can be done either by phage antiserum, 

 which, as indicated above, inactivates only unadsorbed phages, or by 

 short centrifugation, which throws down bacteria and leaves free phage 

 in the supernatant. 



