Genetic Characterization of Enterococcus Faecalis N1‐33 Bacteriocin and Obtained of the Mutant Strain that Produce Many Bacteriocin by Novobiocin Agent Effect, its Merit as A Food Preservative in Steamed Rice Model

Enterococcus faecalis strain N1‐33 produces a peptide that shows bacteriocin activity, enterocin MR10A (EntMR10A). We cloned and sequenced the structural gene of the EntMR10A locus (entMR10A), and the peripheral region of entMR10A was analyzed. Inverted repeat‐like structure was found for at both ends of entMR10A and so that it suggested entMR10A of strain N1‐33 could be formed from an insertion sequence. Attempting to eliminate the plasmid from strain N1‐33 using the plasmid‐curing agent novobiocin was obtained the mutant strain of N1‐33 which seemed to overproduction of bacteriocin than the original strain, termed N41‐51. The number of originated bacteriocin structural genes detected by real‐time PCR showed that its level of the strain N41‐51 was about twofold higher than that of N1‐33, and correspondingly appeared to produce about twofold more bacteriocin compared with the original strain in these culture broth, respectively. This difference as a food preservative potential was examined whether it proves the same result in the actual food systems, by using steamed rice. The culture supernatant of strain N41‐51 has confirmed to inhibit the growth of Bacillus cereus by significantly less amount added to steamed rice than that of N1‐33. PRACTICAL APPLICATIONS: Besides having it with their usual meals, Japanese often eat steamed rice such as rice balls and in their lunch. The most important point we care at these cases is the food poisoning caused by Bacillus cereus, and thus, growth inhibition of B. cereus in steamed rice has been an important task for food industry. In this study, efficacy of the bacteriocin produced by Enterococcus faecalis N1‐33 against B. cereus in steamed rice had shown. The difference in bacteriocin activity of the wild‐type strain N1‐33 and its mutant strain N41‐51, which showed about twice as much bacteriocin production, was also reshown well in the actual food system. Also that, the high bacteriocin producing strain of N1‐33, termed N41‐51, could be obtained with a simple plasmid curing process.