Felix L. Haas

Mutations and Mutagenic Agents in Bacteria

As previously reported, mutation to penicillin or streptomycin resistance can be induced in Micrococcus aureus by growing the organism in nutrient broth and in certain synthetic media which have been irradiated with ultra-violet light (Stone, Wyss, Haas, 1947) or treated with hydrogen peroxide (Wyss, Stone, Clark, 1947; Wyss, Clark, Haas, and Stone, 1948). Stone, Haas, Clark, and Wyss (1948) showed that these mutations result from an increase in mutation rate and are not due to selective action in favor of pre-existing mutants in the culture. The penicillin and streptomycin resistant mutants grow no faster than the original strains in either irradiated or unirradiated media. Mixtures of normal and mutant Micrococcus aureus exhibited the sam-e ratio after growth in irradiated and unirradiated media that they had before growth. Penicillin resistant mutants had no more resistance to streptomycin than did their original parent strain. When these penicillin resistant mutants were grown in irradiated broth or hydrogen-peroxide-treated broth the treatment induced mutation to streptomycin resistance in these at the same rate as that induced in the normal strain. The same characteristics hold for the streptomycin resistant mutants. In attempts to characterize the mutating principle, and to determine the method of its production by ultra-violet irradiation or hydrogen peroxide treatment, we have investigated some of the physical properties of treatment of the substrate, and of the treated substrate itself.

Quantitative studies on the expression of a transformed character in Bacillus subtilis.

Abstract The effect of media supplementation upon the expression of a transformed genetic character has been studied in Bacillus subtilis . Localization of cells on the surface of a membrane filter disc eliminated bias due to population changes, and also permitted manual relocation of cells from one medium to another. Replicate samples of competent recipient cells were banked in liquid nitrogen so that comparative experiments could be carried out on successive days. Transient supplementation of solid medium with a nutritional factor (tryptophan) required by B. subtilis during incubation periods up to 5 h has no effect on subsequent transformation frequency. Control cells manipulated in the same manner, but incubated throughout the experiment on the unsupplemented medium selective for the transformant phenotype, gave similar results. Identical experiments using a complete medium prior to transferring the filter disc to minimal selective medium yielded quite different results. In this case, transformation frequency increased as a linear function of the incubation time on complete medium. Furthermore, the ordinate value of the transformation frequency was directly proportional to the time the cells were exposed to transforming DNA, although the rate of transformation frequency increase was identical at all times. The data suggest that these techniques effectively separate the transformation event per se (altered genome) from subsequent phenotypic expression of the new potentiality.

Gasometric determination of hydrogen peroxide.

Summary Determinations of the peroxide content of irradiated broth using catalase in a Warburg respirometer were found to be inaccurate. In the presence of excess catalase the oxygen evolved approaches a value of twice that indicated by the accepted general equation for enzymatic decomposition of H2O2; if too little enzyme is used the reaction is slow and incomplete. The method is unsatisfactory for the quantitative determination of peroxide.