Heat resistance of an outbreak strain of Listeria monocytogenes in hot dog batter.

The heat resistance of a strain of Listeria monocytogenes responsible for a listeriosis outbreak in hot dogs was not higher than the heat resistance of other L. monocytogenes strains when tested in tryptic soy broth and in laboratory-prepared hot dog batter. For the thermal death time experiments, the cells were grown to stationary phase or were starved in phosphate-buffered saline, pH 7, for 6 h at 30 degrees C. Starvation increased the heat resistance of L. monocytogenes in broth but not in hot dog batter. D-values in hot dog batter were higher than in broth. For the hot dog formulation used in this study, cooking the hot dog batter for 30 s at 71.1 degrees C (160 degrees F), or its equivalent using a z-value of 6 degrees C (11 degrees F), would inactivate 5 logs of L. monocytogenes.

Purification and properties of spore-lytic enzymes from Clostridium perfringens type A spores.

Spores of Clostridium perfringens contain at least two spore-lytic enzymes active in hydrolysing cortical peptidoglycan. One enzyme has been purified 1800-fold and has a molecular weight of 17 400 determined from chromatography on Sephadex G-75. Two protein bands were apparent after SDS-PAGE. The isolated enzyme was investigated for response to temperature, pH, ionic strength and enzyme inhibitors, and for mode of action. A second enzyme activity, differing from the first in apparent molecular weight (29 800) as determined by gel exclusion chromatography, and also in its pH optimum and activity on cortical substrate, was also isolated, although not purified to the same extent.

Extraction of spore-lytic enzyme from Clostridium perfringens spores.

Various chemical reagents known to extract spore coat protein were used to extract spore-lytic enzyme (SLE) from intact and germinated spores of Clostridium perfringens. Of the reagents tested, 7.2 M-urea plus 10% (v/v) mercaptoethanol, pH 2.85, solubilized the most SLE activity per mg spores. The quantity of SLE extracted was dependent on the initial pH of the reagent, with a maximum between pH 2.7 and 3.0. Germinated spores yielded more SLE than non-germinated spores upon urea/mercaptoethanol extraction. SLE release during spore germination probably utilizes a trigger mechanism not satisfied by germination alone. Significant amounts of SLE were released during germination when spores were suspended in potassium chloride or a complex germinant mixture containing brain-heart infusion, yeast extract and chloramphenicol, but not during germination with sodium nitrite, which non-enzymically lysed the cortical peptidoglycan. Greater solubilization of SLE activity was obtained by urea/mercaptoethanol extraction of spores germinated with nitrite than of spores germinated with either potassium chloride or the complex germinant.

Reversal of radiation-dependent heat sensitization of Clostridium perfringens spores.

The effect of solute concentration on the sensitization of Clostridium perfringens spores to heat by ionizing radiation was investigated. As we have shown previously, spores of C. perfringens treated with gamma radiation are now sensitive to subsequent heat treatments than are spores that receive no radiation treatment. When gamma-irradiated spores were heated in the presence of increasing concentrations of glycerol or sucrose, the heat sensitivity induced by irradiation was progressively decreased. The magnitude of the increase in heat resistance induced by extracellular solutes was greater in gamma-irradiated spores than in nonirradiated spores. Based on these observations, it is proposed that the induction of heat sensitivity in spores by radiation is related to the loss of osmoregulatory or dehydrating mechanisms in irradiated spores.

Sensitization of Clostridium perfringens spores to heat by gamma radiation.

Spores of Clostridium perfringens, type A, were given separate or sequential treatments of gamma radiation (0 to 0.7 Mrad) and/or high temperature (93 to 103 degrees C). Prior heating, sufficient to inactivate 40 to 99% of the viable spores, had no effect on the subsequent radiation inactivation rate. Prior irradiation had a sensitizing effect on subsequently heated spores. The degree of sensitization to heat, as measured by thermal inactivation rate, increased with increased radiation pretreatment dose.