Heat Inactivation of Intracellular Phage Precursor. ∗

Abstract

Krueger and Mundell 1 recently reported a method for demonstrating intracellular phage precursor. The essential step in the method is the preparation of “activated” suspensions of staphylococci; this is accomplished by growing the organisms in a heavily oxygenated medium. The activated cells are separated from the broth, resuspended in Locke's solution and are then maintained at 5°C for 2 hours before they are used. To demonstrate intracellular precursor 4 ml of activated cell suspension containing 5 × 10⁸ bacteria/ml is added to 1 ml of phage diluted with Locke's solution to contain 1 × 10⁹ activity units/ml. The mixture is kept for 5 minutes at 5°C and is promptly titrated for total phage content. The end titer is 2 × 10⁹ activity units/ml, an increase of 500% in phage concentration.

Krueger and Scribner 2 supplied additional evidence for the existence of the intracellular precursor. They made use of the fact that staphylococci activated in the presence of Mn⁺⁺ have a relatively low lytic threshold and require only small amounts of phage to induce lysis. Starting with a small amount of phage it was possible to transform the phage precursor in successive lots of staphylococci into phage and to obtain the newly formed phage free in solution by lysing the precursor-containing cells. The original phage added at the start of the experiment has been diluted at least 1 to one million without any reduction in plaque count or activity titer.

It seemed likely from data already available that the phage precursor is more thermolabile than the bacterial cell which produces it. In the experiments of Krueger and Fong 3 the relationships shown in Table I between bacterial reproduction and phage formation at various temperatures were observed.