Studies on the Heat Resistance of Bacterial Spores

Part 3: The Effects of Sugars in the Subculture Media on the Survival Times of Bacillus natto

Abstract

(1) The influences of subculture media on the survival times of spores were studied. The, spores of Bacillus natto were suspended in neutral phosphate buffer and heated at. 100°C, and. then subcultured by various media. The yeast-extract broth (10% bakers' yeast extracted cted by nutrient both, adjusted to pH 7.0). liver-extract broth (10% pork-liver extracted by nutrient broth), and glucose-broth (1% glucose added to nutrient broth), showed survival time-twice as long as plain nutrient broth. The addition of reducing agents, cysteine and thioglycollate to nutrient broth, have shown no increasing effect on the survival time. (Table 1)
(2) The effects of adding various carbohydrates and organic acids were examined. Fructose, mannose, galactose, sucrose, maltose, and soluble starch were found to have the increasing effect on the survival time of spores of Bacillus natto to the same extent as glucose, but other eight sugars, arabinose, xylose, lactose, trehalose, mannitol, glycerol glycogen, and Cori-ester have had no effects.
The addition of pyruvate or alpha-glycerophosphate to nufttent broth also increased the, survival time, but the addition of organic acids such as lactic, acetic, and succinic had noeffects.
It was found that in a synthetic medium⁽⁹⁾, which is composed of eighteen amino acids, nine vitamins, four purine-pyrimidine bases, and salts, the vegetative cellls of Bac. natto grew well without the addition of glucose, whereas the spores could not grow without glucose (carbohydrates). And it was also found that the kind of carbohydrates which were effective in increasing the survival time of the spores of Bacillus natto was the same with that which served, as the energy source and which enabled the spores to grow in the synthetic medium. (see Table 3 and 5). The minimum effective, concentration of glucose in increasing the survival time of the spores was between 10^-5M and 10^-6M, and the same order of glucose concentration also has limited the growth of spores of the strain in the synthetic medium (see Table 4 and 5).
(4) It should be noted that when subcultured by glucose broth, the survival time at each spore concentration was commonly about twice that of plain nutrient broth, throughout a wide range of spore concentration. The relation between spore concentration (N) and survival time (t) in the case of subculturing by glucose broth, can be expressed by the following general formula as reported in the previous paper⁽⁷⁾. logN=a+blogt
(5) The discussions on the mechanisms of spore survival time increasing activities of glucose were delivered.