INDUCED RESISTANCE TO GAMMA IRRADIATION IN ESCHERICHIA COLI

The ability of microorganisms to become adapted to environmental conditions which have been shown previously to be fatal to the parent strain is a problem with practical as well as fundamental scientific implications. Just as thermophilic organisms present a major problem to the food processor, and antibiotic resistant strains cause difficulties in treatment of infections, so radiation resistant strains well may be a factor in radiation sterilization processes. While the lethal and mutagenic nature of the interaction of radiations with microorganisms is well established, the mechanisms of this action are still a matter of speculation. For many years a "target theory", based on the "one hit-one effect" interpretation of Dessauer (1923), was accepted commonly, but recent experiments have shaken the basic tenets of this theory. In its place an explanation of radiation effects is being evolved based on physical and chemical changes taking place in the environment of the cells during irradiation. Following this view, it is not improbable that certain cells in any mass population may become adjusted to these previously fatal surroundings. Strains of radio resistant organisms then can be developed and selected. Cosmic, short wave, visible, and ultraviolet radiation, to which most organisms naturally are subjected, however, may have maximized alrady the radio resistance of most strains. An organism such as Escherichia coli which, by virtue of its normal habitats and mode of existence, is not subject much to radiation is therefore an ideal subject for this kind of study. The development of strains of E. coli resistant to viruses and to penicillin has been reported by Demerec (1945) and Luria and Delbrtick (1943). These investigators applied the statistical methods of variance analysis evolved by Luria and Delbrlick (1943) to resolve the fundamental question: do these agents induce mutants, resistant to the inducing agent, in an initially homogenous population, or do they select spontaneously produced mutants by eliminating nonresistant organisms? In both the cases of penicillin and virus resistance the close approximation of the variance to the mean of the number of resistant organisms, when the organisms all come from a single culture, is taken as sufficient evidence to indicate that mutations are spontaneous and the agent selective. More recently, Witkin (1946), by a double irradiation technique, isolated a strain of radiation resistant E. coli from strain B which was designated as B/r. The method used is contingent upon the observation of Lea et al. (1937) that E. coli, on incubation for three hours at 37 C, produces a characteristic "long" form that then may be distinguished from the radio resistant B/r which develops normal microcolonies. In these experiments, 50 ergs/mm2 of ultraviolet radiation were used before incubation, and a second dose of 700 ergs/mm2 was used three hours later to kill the long forms. A comparison of the survival curves shows that one erg/mm2 is, in effect, equivalent approximately to 40 roentgens of X-rays. An analysis of the data by the statistical method of Delbriuck indicated that the radiation acted as a selective agent for mutations which were occurring spontaneously at the rate of 10-5 per organism per generation. Further experiments by, Witlin (1947), in which strain B was exposed to 3,800 ergs/mm' of ultraviolet, showed an increase in the "spontaneous" frequency of radio resistant mutants from 0.003 per cent (average) in the original culture to 4.1 per cent (average) in the survivors from high dose irradiations. These later experiments then can be interpreted only to mean that the selecting agent is also the inducing agent and that spontaneous mutations, when they occur, are negligible in comparison to the number of induced mutations. Witkin's experiments are unique in the successful isolation of a radio resistant strain. Other observers report instead increased susceptibility to radiation injury of vegetative bacterial cells (Huber, 1952), Triticum monococcum (linkorn