Thermal Activation and Dry-heat Inactivation of Spores of Bacillus subtilis MD2 and Bacillus subtilis var. niger (original) (raw)
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Comparison of the properties of Bacillus subtilis spores made in liquid or on agar plates
Journal of Applied Microbiology, 2007
Aims: To compare the properties of the spores of Bacillus subtilis prepared in liquid and on plates.Methods and Results: The spores of B. subtilis were prepared at 37°C using a nutrient exhaustion medium either in liquid or on agar plates. The levels of core water, dipicolinic acid (DPA) and small, acid-soluble spore proteins (SASP) were essentially identical in spores made in liquid or on plates. Spores prepared in liquid were killed ∼threefold more rapidly at 90°C in water than the spores prepared on plates, and the spores prepared in liquid were more sensitive to nitrous acid and a diluted stable superoxidized water. Spores prepared in liquid also germinated more rapidly with several agents than those prepared on plates. Pellets of spores prepared on plates were darker than spores prepared in liquid, and spores prepared in liquid had more readily extracted coat protein. However, there were no major differences in the relative levels of individual coat proteins or the cross-linking of the coat protein GerQ in the two types of spores, although the inner membrane of spores prepared on plates had a higher ratio of anteiso- to iso-fatty acids.Conclusions: The preparation in liquid yielded spores with some different properties than those made on agar plates. Spores made in liquid had lower resistance to heat and several chemicals, and germinated more readily with several agents. There were also differences in the composition of the inner membrane of spores prepared under these two conditions. However, there were no major differences in the levels of DPA, core water, SASP and individual coat proteins or the cross-linking of a coat protein in spores made in liquid and on plates.Significance and Impact of the Study: This work demonstrates that the preparation method can affect the resistance and germination properties of bacterial spores, even if an identical medium and temperature are used. Evidence was also obtained consistent with the role of the inner membrane in spore resistance and germination, and that some factor in addition to core water, DPA and SASP content plays a role in spore resistance to wet heat.
Applied Microbiology and Biotechnology, 2011
Bacillus subtilis(B. subtilis) cells were placed in various environmental conditions to study the effects of aeration, water activity of the medium, temperature, pH, and calcium content on spore formation and the resulting properties. Modification of the sporulation conditions lengthened the growth period of B. subtilis and its sporulation. In some cases, it reduced the final spore concentration. The sporulation conditions significantly affected the spore properties, including germination capacity and resistance to heat treatment in water (30 min at 97°C) or to high pressure (60 min at 350 MPa and 40°C). The relationship between the modifications of these spore properties and the change in the spore structure induced by different sporulation conditions is also considered. According to this study, sporulation conditions must be carefully taken into account during settling sterilization processes applied in the food industry.
International Journal of Food Microbiology, 2019
Specific treatments combining high temperatures of up to 150°C and moderate pressure of up to 0.6 MPa have been applied to Bacillus subtilis 168 spores conditioned at different a w levels (between 0.10 and 0.70) corresponding to different residual water contents within the spore core. The spores were treated as a dry powder in a pressurized nitrogen environment or in water/glycerol solutions. These thermodynamic conditions were intended to prevent any water evaporation from the spore core during time/temperature treatments. Our results clearly show that retaining liquid water in the core by applying pressure during the treatment resulted in greater spore destruction (between 2.4 and 4.9 log at 150°C, 120 s and a w 0.5 in powder) than the destruction observed after the treatment at atmospheric pressure (0.7 log), during which the water rapidly evaporated because its boiling point was reached. Moreover, we found that the water activity level of the spore had a significant impact on spore destruction: the higher the a w level, the greater the spore inactivation. We obtained similar results from spores heat-treated in powder and in water/glycerol solution at the same a w , confirming the strong influence of this parameter. We hypothesized that the increased spore inactivation was related to the well-known thermal sensitivity of vital organic molecules such as proteins, enzymes, and ribosomes in the presence of water.
FEMS Microbiology Letters, 2004
A Bacillus subtilis strain with a base substitution in the ribosome-binding site of spoVAC was temperature sensitive (ts) in sporulation and spores prepared at the permissive temperature were ts in L L-alanine-triggered germination, but not in germination with Ca 2+-dipicolinic acid (DPA) or dodecylamine. Spores of a ts spo mutant with a missense mutation in the spoVAC coding region were not ts for germination with L L-alanine, dodecylamine or Ca 2+-DPA. These findings are discussed in light of the proposal that SpoVA proteins are involved not only in DPA uptake during sporulation, but also in DPA release during nutrient-mediated spore germination.
Applied and environmental microbiology, 2015
High heat resistance of spores of Bacillus thermoamylovorans poses challenges to the food industry as industrial sterilization processes may not inactivate such spores, resulting in food spoilage upon germination and outgrowth. In this study, the germination and heat resistance properties of spores of four food-spoiling isolates were determined. Flow cytometry counts of spores were much higher than their counts on rich medium (maximum 5%). Microscopic analysis revealed inefficient nutrient-induced germination of spores of all four isolates despite the presence of most known germination-related genes, including two operons encoding nutrient germinant receptors (GRs), in their genomes. In contrast, exposure to non-nutrient germinant calcium-dipicolinic acid (Ca-DPA) resulted in efficient 50 to 98% spore germination. All four strains harbored cwlJ and gerQ genes, which are known to be essential for Ca-DPA-induced germination in Bacillus subtilis. When determining spore survival upon he...
PLoS ONE, 2013
Spore-forming bacteria are a special problem for the food industry as some of them are able to survive preservation processes. Bacillus spp. spores can remain in a dormant, stress resistant state for a long period of time. Vegetative cells are formed by germination of spores followed by a more extended outgrowth phase. Spore germination and outgrowth progression are often very heterogeneous and therefore, predictions of microbial stability of food products are exceedingly difficult. Mechanistic details of the cause of this heterogeneity are necessary. In order to examine spore heterogeneity we made a novel closed air-containing chamber for live imaging. This chamber was used to analyze Bacillus subtilis spore germination, outgrowth, as well as subsequent vegetative growth. Typically, we examined around 90 starting spores/cells for $4 hours per experiment. Image analysis with the purposely built program ''SporeTracker'' allows for automated data processing from germination to outgrowth and vegetative doubling. In order to check the efficiency of the chamber, growth and division of B. subtilis vegetative cells were monitored. The observed generation times of vegetative cells were comparable to those obtained in well-aerated shake flask cultures. The influence of a heat stress of 85uC for 10 min on germination, outgrowth, and subsequent vegetative growth was investigated in detail. Compared to control samples fewer spores germinated (41.1% less) and fewer grew out (48.4% less) after the treatment. The heat treatment had a significant influence on the average time to the start of germination (increased) and the distribution and average of the duration of germination itself (increased). However, the distribution and the mean outgrowth time and the generation time of vegetative cells, emerging from untreated and thermally injured spores, were similar.