Influence of food matrix on inactivation of Bacillus cereus by combinations of nisin, pulsed electric field treatment, and carvacrol.

Carvacrol was used as a third preservative factor to enhance further the synergy between nisin and pulsed electric field (PEF) treatment against vegetative cells of Bacillus cereus. When applied simultaneously with nisin (0.04 microg/ml), carvacrol (0.5 mM) enhanced the synergy found between nisin and PEF treatment (16.7 kV/cm, 30 pulses) in potassium-N-2-hydroxyethylpiperazine-N-ethanesulfonic acid (HEPES) buffer. The influence of food ingredients on bactericidal activity was tested using skimmed milk that was diluted to 20% with sterile demineralized water. The efficacy of PEF treatment was not affected by the presence of proteins, and results found in HEPES buffer correlated well with results in milk (20%). Nisin showed less activity against B. cereus in milk. Carvacrol was not able to enhance the synergy between nisin and PEF treatment in milk, unless used in high concentrations (1.2 mM). This concentration in itself did not influence the viable count. Carvacrol did act synergistically with PEF treatment in milk, however not in HEPES buffer. This synergy was not influenced by proteins in milk, as 5% milk still allows synergy between carvacrol and PEF treatment to the same extent as 20% milk.

Sensitivities of germinating spores and carvacrol-adapted vegetative cells and spores of Bacillus cereus to nisin and pulsed-electric-field treatment.

Treatment of Bacillus cereus spores with nisin and/or pulsed-electric-field (PEF) treatment did not lead to direct inactivation of the spores or increased heat sensitivity as a result of sublethal damage. In contrast, germinating spores were found to be sensitive to PEF treatment. Nisin treatment was more efficient than PEF treatment for inactivating germinating spores. PEF resistance was lost after 50 min of germination, and not all germinated spores could be inactivated. Nisin, however, was able to inactivate the germinating spores to the same extent as heat treatment. Resistance to nisin was lost immediately when the germination process started. A decrease in the membrane fluidity of vegetative cells caused by incubation in the presence of carvacrol resulted in a dramatic increase in the sensitivity to nisin. On the other hand, inactivation by PEF treatment or by a combination of nisin and PEF treatments did not change after adaptation to carvacrol. Spores grown in the presence of carvacrol were not susceptible to nisin and/or PEF treatment in any way.

Pulsed-electric field treatment enhances the bactericidal action of nisin against Bacillus cereus.

Vegetative cells of Bacillus cereus were subjected to low doses of nisin (0.06 microg/ml) and mild pulsed-electric field treatment (16.7 kV/cm, 50 pulses each of 2-micros duration). Combining both treatments resulted in a reduction of 1.8 log units more than the sum of the reductions obtained with the single treatments, indicating synergy.

Combined action of nisin and carvacrol on Bacillus cereus and Listeria monocytogenes.

Nisin, a small antimicrobial protein, was tested for its bactericidal action against Listeria monocytogenes and Bacillus cereus and a typical biphasic reduction of the viable count was observed. The reduction was most fast during the first 10 min of exposure, while the viable count remained stable in the last part of the exposure period. Bacillus cereus was more sensitive towards nisin than L. monocytogenes and the inhibitory effect of nisin was stronger towards cells cultivated and exposed at 8 degrees C than towards cells cultivated and exposed at 20 degrees C. Combining nisin with sublethal doses of carvacrol resulted in an increased reduction in the viable count of both organisms, indicating synergy between nisin and carvacrol. Addition of lysozyme as a third preservative factor increased the synergistic effect between nisin and carvone, especially in the last part of the exposure period.

Effect of oxygen concentration and redox potential on recovery of sublethally heat-damaged cells of Escherichia coli O157:H7, Salmonella enteritidis and Listeria monocytogenes.

The measured heat resistance of cells of Escherichia coli O157:H7, Salmonella enteritidis and Listeria monocytogenes was up to eightfold greater when they were grown, heated and recovered anaerobically rather than aerobically. Measured heat resistance was highest when anaerobic gas mixtures were used (time at 59 degrees C for a 6-decimal (6-D) reduction of E. coli O157:H7, 19-24 min); moderate when low concentrations of oxygen (0.5-1%) were included (time for a 6-D reduction, 5-17 min); and lowest when higher concentrations of oxygen (2-40%) were used (time for a 6-D reduction, 3 min). This effect was principally attributed to the recovery conditions, and a greater effect was noted at lower heating temperatures. The use of reduced oxygen concentration (< 2% O2), e.g. packing under an anaerobic gas mixture or a vacuum, might therefore increase the risk of these pathogens surviving heat treatments applied to food. It is also possible that foods that are packed in air but with a low redox potential might allow the survival of heated cells, and thus the anticipated level of safety might not be achieved.