ACTIVE FROM INACTIVATED BACTERIOPHAGE 97 



The occurrence of reactivation for certain phages accounts for deviations 

 from the logarithmic inactivation rate found for these same phages 

 (see figure 1). As the dose of radiation increases beyond a certain point, the 

 survival, as determined by plaque count, appears to diminish less rapidly. 

 This is due to an unavoidable partial reactivation. In a phage titration, we 

 mix approximately 5X10^ bacteria with an amount of phage suspension such 

 as to give approximately 100 plaques, and pour the mixture on an agar plate. 

 On the one hand, if we are dealing with a fully active sample, the bacteria 

 only come in contact with 100 active particles. On the other hand, in 

 assaying an irradiated suspension containing, for example, one active 

 particle in 10®, we expose the bacteria to 10^ inactive particles plus a corre- 

 spondingly large amount of other lysate constituents, besides the residual 

 100 active particles. Conditions permitting reactivation, therefore, obtain in 

 these plating mixtures, and reactivation disturbs and often completely ob- 

 scures the count of the residual active phage. For this reason, the survival of 

 fully active phage for high doses must be obtained by extrapolation from the 

 logarithmic part of the curve, a necessarily inefficient procedure. If deviations 

 in the survival rate for high doses occurred, this extrapolation would not be 

 justified. One possible cause of error — screening of some phage particles from 

 radiation by components of the lysate itself — was excluded by irradiating 

 concentrated phage T6 mixed with phage Tl, and testing for the inactivation 

 rate of the latter, which is not disturbed by reactivation phenomena. The 

 inactivation rate of Tl remains the same as in the absence of T6 up to doses 

 that correspond to 100 hits per particle of phage T6. 



Identification of the reactivating factor 



What is the "factor" present in irradiated stocks of phages T2, T4, T5, or 

 T6, which, if acting on bacteria that have adsorbed inactive phage particles, 

 allows production of active phage? Since phage stocks are lysates produced 

 by lysis of the common host E. coli B, the factor might be either of phage or of 

 bacterial origin. 



The factor was identified as being phage itself, inactive or active, in that 

 reactivation occurs in bacteria that adsorb either more than one inactive particle of 

 a given phage, or one inactive particle of one phage plus some active or inactive 

 particles of a related phage. The evidence for this conclusion, which is illustrated 

 in part by the data in table 3, can be summarized as follows. 



(a) Addition of an excess of supernatant from a heavy bacterial culture to 

 a mixture of dilute irradiated phage plus bacteria gives no increase in plaque 

 count. The factor in the lysates is not a normal bacterial secretion. 



(b) Concentrated lysates of phages Tl, T5, or T7 added to a mixture of 

 bacteria and dilute irradiated T2 (or T4, or T6) do not cause reactivation. 

 No heterologous lysate causes reactivation of T5. The test for cross-reactivation 

 is done by plating the mixtures, before lysis, with bacterial indicator strains 

 sensitive to the phage whose reactivation is tested, but not to the others. 

 Since all phage stocks are lysates of common host cells, the factor is not an 

 unspecific bacterial product liberated upon lysis. 



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