Heat‐resistance of spores of non‐proteolytic Clostridium botulinum estimated on medium containing lysozyme

Heat treatment of spores of non‐proteolytic strains of Clostridium botulinum at 75–90°C, and enumeration of survivors on a nutrient medium containing lysozyme gave biphasic survival curves. A majority of spores were inactivated rapidly by heating, and the apparent heat‐resistance of these spores was similar to that observed by enumeration on medium without lysozyme. A minority of spores showed much greater heat‐resistance, due to the fact that the spore coat was permeable to lysozyme, which diffused into the spore from the medium and replaced the heat‐inactivated germination system. The proportion of heated spores permeable to lysozyme was between 0.2 and 1.4% for spores of strains 17B (type B) and Beluga (type E), but was about 20% for spores of strain Foster B96 (type E). After treatment of heated spores with alkaline thioglycolate, all were permeable to lysozyme. D‐values for heated spores that were permeable to lysozyme (naturally and after treatment with thioglycolate) were: for strain 17B, D85°C, 100 min; D90°C, 18.7 min; D95°C, 4.4 min; for strain Beluga, D85°C, 46 min; D90°C, 11.8 min; D95°C, 2.8 min. The z‐values for these spores of strains 17B and Beluga were 7.6°C and 8.3°C.