INTKACELLULAR MULTIPLICATION OF BACTERIAL VIRUSES 241 



studying bacteriophage multiplication, consists of infecting a growing 

 suspension of bacteria with a suitable number of phages, incubating this 

 mixture for a few minutes in order to allow adsorption to the bacteria of most 

 of the phage particles, and then diluting this mixture from ten thousand- to a 

 millionfold into nutrient medium. The diluted culture is then incubated 

 further and aliquots plated on sensitive indicator bacteria at regular time 

 intervals in order to assay the instantaneous number of infective centers in 

 the culture. The result of Ellis and Delbriick's one-step growth experiment 

 was that the number of infective centers in the culture remains constant for 

 the first 30 minutes after infection of the bacterial culture. This initial time 

 interval, during which the number of mfective centers shows no increase, is 

 the latent period. After 30 minutes have elapsed, the number of infective 

 centers in the culture begins to augment until a final plateau is attained 50 

 minutes later, when no further increase in infectivity occurs; the time interval 

 during which the number of infective centers increases is the rise period and 

 the ratio of the titer represented by final plateau to the initial titer is the burst 

 size, which in Ellis and Delbriick's case was approximately 60. The latent 

 period thus represents the minimum time which must elapse between the 

 moment at which the bacterial culture is infected with a phage population and 

 the moment at which the first infected cells in the culture lyse to liberate into 

 the medium the progeny phage particle which have grown within them. The 

 rise period represents the interval during which more and more of the infected 

 bacteria lyse, and the final plateau represents the stage at which all of the 

 infected bacteria have lysed and no further phage multiplication occurs, 

 progeny phage and any remaining uninfected bacteria in the culture having 

 been separated from one another in the growth tube by the high dilution made 

 shortly after the outset of the experiment. The burst size, finally, represents 

 the average number of infective progeny phage particles produced per 

 infected bacterium. 



One of the few later improvements in the one-step groAvth technique has 

 been an induction of more synchronous phage growth by adsorbing the 

 infecting phage particles to starved bacteria in a non-nutrient medium and 

 then declenching phage development simultaneously in all bacteria by addition 

 of nutrient (Benzer, 1952). Adsorption of phage to bacteria suspended in 

 growth media contaming cyanide (Benzer and Jacob, 1953) or chloram- 

 phenicol (Tomizawa and Sunakawa, 1956; Hershey and Melechen, 1957) and 

 subsequent initiation of phage development by removal of the drug can 

 achieve similar results. These procedures eliminate that variation in the onset 

 of intracellular phage growth which is due to the spread in times at which 

 different bacteria adsorb their first phage particle. 



The parameters of phage multiplication defined by the one-step growth 

 experiment may differ very much, according to the exact physiological 



VOL. II — 16 



