APPENDIX 447 



cultures. With an energy source such as lactic acid, aeration 

 is essential for growth. A convenient device for aeration is the 

 Marco air pump.* 



A factor of great importance in the design of experiments is 

 the medium in which the host cells have been grown. E. coli 

 grown in nutrient broth is from the metabolic point of view 

 very different from E. coli grown on the ammonium lactate 

 medium. E. coli grown in broth, centrifuged and resuspended 

 in ammonium lactate medium has a long lag period before 

 growth starts, presumably because the organism has to synthe- 

 size enzymes necessary for synthesis of amino acids and growth 

 factors furnished in the broth medium. This property of broth- 

 grown E. coli was used by Fowler and Cohen (1948) in an inves- 

 tigation of the various constituents of the medium which may 

 play a role in phage synthesis in the organism. The organisms 

 were grown in broth, centrifuged and resuspended in an am- 

 monium lactate medium. The latent period of phage growth 

 and the burst size (p. 473) were then determined on such a cul- 

 ture infected with phage T2. The effect of addition of various 

 amino acids and growth factors on latent period and burst 

 size could then be readily determined. 



Isolation of Bacterial Viruses 



To isolate a virus lysing E. coli, 30 ml. of broth is inoculated with 

 1 ml. of a visibly turbid culture of the host organism. To this 

 is added with thorough mixing 1 ml. of supernatant obtained 

 by centrifuging pooled sewage for a few minutes. After incuba- 

 tion overnight at 37 ° C, an aliquot of the culture is centrifuged 

 to remove bacteria and the supernatant is filtered through a 

 Corning ultrafine sintered glass filfer. This filtrate is tested 

 for presence of phage. 



a. By plating. Dilute the filtrate serially in a broth culture of 

 E. coli and spread 0.1 ml. samples of each dilution on agar 



* Made by the J. B. Maris Company, Bloomfield, N. J. This company also 

 makes valves and other fittings suitable for low pressure air lines. 



