Theoretical Base for the Application of the DiVSaL Method to Bacterial Spores to Evaluate Injured Populations Occurring After Exposure to Lethal Stress.

To assess injury in bacterial spore populations exposed to lethal stress, we proposed a theoretical basis for applying the DiVSaL method, which has already been reported for general microorganisms as a double subculture method. We constructed a mathematical model in which both injuries to the germination system and the spore body were taken into the theory. In this theory, we reasonably assumed that the viable and germinable spore count is constant before the subsequent vegetative growth and that the delay of germination and outgrowth can be included in the concept of λ injury previously reported as the growth-independent injury. By introducing these assumptions, the double subculture method can be considered to apply to spores as well. As examples of the application of this theory, the growth delays of Bacillus subtilis spores treated with heat and UV irradiation were analyzed and the numbers of injured spores were evaluated. Based on the results obtained, heat is indicated to have a higher injury generation ability than UV irradiation. The applicability of the DiVSaL method as a tool for food preservation and sanitation designs is presented.

Low Temperature Heating-Induced Death and Vacuole Injury in Cladosporium sphaerospermum Conidia.

The mechanism of thermal death of mold conidia has not been understood in detail. The purpose of this study is to analyze the death kinetics of heated conidia of Cladosporium sphaerospermum and to ascertain the expectant cell injury responsible for the death. The death of the dormant (resting) conidia of Cladosporium sphaerospermum was examined at temperatures of between 43 and 54℃ with the conventional colony count method. The death reaction apparently followed the first order kinetics, but the Arrhenius plot of the death rate constant demonstrated seemingly a break. The linearity at temperatures higher than that at the break was lost at lower temperatures, suggesting the involvement of an unusual mechanism in the latter temperatures. In the cell morphology, we observed with quinacrine staining the vacuole rupture at a lower temperature but not at a high temperature. Interestingly, the vacuole rupture by low-temperature heating was found to correlate with the viability loss. Furthermore, active protease originally locating in vacuoles was detected in the cytoplasm of the conidia after heated at a low temperature. The results obtained suggest the involvement of potent autophagic cell death induced by low temperature heating of C. sphaerospermum conidia.