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Biofilm is a community of microorganisms that adhere to abiotic or biotic surfaces, and is problematic in a wide range of food industries, as well as to human health. Although bacteriocin from lactic acid bacteria is well known as a natural antimicrobial agent in food preservation, it has been poorly investigated that bacteriocin inhibits the biofilm formation of foodborne pathogens. In this study, we demonstrated that bacteriocin produced by Lactobacillus brevis DF01 was partially purified and investigated whether the bacteriocin inhibited the biofilm formation of Escherichia coli and Salmonella Typhimurium. Assessment by the microtiter plate method, as well as by fluorescence and scanning electron microscopy, showed that the biofilm formation of E. coli and S. Typhimurium was reduced when the bacteria was co-incubated with crude bacteriocin of L. brevis DF01 (DF01 bacteriocin). Although pre-treated DF01 bacteriocin also significantly inhibited the biofilm formation of E. coli and S. Typhimurium (P = 0.0035 and P = 0.0003, respectively) post-treatment with DF01 bacteriocin did not significantly inhibit the biofilm formation (P = 0.1314 for E. coli and P = 0.2939 for S. Typhimurium), suggesting that DF01 bacteriocin interfered with biofilm formation, but did not disrupt the established biofilm of E. coli and S. Typhimurium. In addition, biofilms of both E. coli and S. Typhimuriun on the surface of stainless steel coupons were decreased in the presence of DF01 bacteriocin. Taken together, these results suggest that DF01 bacteriocin may be applied to control the biofilm formation of E. coli and S. Typhimurium.

A bacteriocin-producing lactic acid bacterium was isolated from malts. The isolate was identified as Pediococcus acidilactici by morphological, API 50 CHL kit and 16S rRNA analysis, and designated as strain HW01. The antimicrobial activity was maintained over a broad pH range (pH 2–11). Proteolytic enzymes and α-amylase treatments inactivated the bacteriocin activity; while catalase, heat, detergents, and solvents did not. The molecular weight of the crude bacteriocin was approximately 6 kDa by SDS-PAGE. The bacteriocin production phenotype (Bac+) was linked to a 9.8 kb plasmid and was not increased by co-culture with an indicator microorganism. Bacteriocin HW01 inhibited several Gram-positive bacteria, including health-threatening microorganisms (Staphylococcus aureus, Streptococcus mutans, Lactobacillus curvatus, Listeria monocytogenes, and L. innocua). Particularly, Pediococcus damnosus and P. claussenii, which were isolated from spoiled beer, were strongly antagonised by the bacteriocin. P. acidilactici HW01 has tolerance to hop compounds but is unable to grow in a simulated beer environment. The potential of the HW01 strain, as a starter culture in brewing, to control beer spoilage lactic acid bacteria, is discussed.

In a previous study, indigenous bacteriocin-producing Pediococcus acidilactici HW01 was isolated from malt. The bacterium was chosen as it is properly adapted to the malting conditions, and it showed a strong antagonistic effect against beer-spoilage lactic acid bacteria (LAB) in the previous study. Strain HW01 was added to the malting process for bio-acidification and bio-preservation of LAB. Changes in the microorganisms, enzymes, pH, total soluble nitrogen, free amino nitrogen, colour, filtration time and viscosity of wort were compared with the control. The LAB starter culture improved several characteristics of malt modification, such as microbiological stability, viscosity and filtration time. Especially, the starter culture protected the malt from the beer-spoilage bacteria of the genus Pseudomonas, by producing several acids and antimicrobial substances. The potential of using HW01 strains for bio-acidification and the bio-preservation of LAB are discussed.

The effects of two crude bacteriocins (DF01 and K10) on lactic acid bacteria (LAB) communities and pH during kimchi fermentation were analyzed by polymerase chain reactiondenaturing gradient gel electrophoresis (PCR-DGGE). Crude LAB bacteriocins, prepared by ammonium sulfate precipitation, were added at 5 AU/mL, and kimchi was incubated at 20°C for 7 days. The pH and titratable acidity of the kimchies were determined daily, and the amplified 16S rRNA products were analyzed by PCR-DGGE. The common and main LAB were Weissella spp., Leuconostoc spp., and Lactobacillus spp. from both control and bacteriocin-treated samples. Among them, W. koreensis, W. confusa, and Lb. sakei were the predominant microorganisms throughout the fermentation period. Some obligate and facultatively heterofermentative LAB were detected from the bacteriocin-treated samples. The pH of the kimchi samples treated with each bacteriocin was higher (ca. 0.8 unit) than that of the control at day 4 and 5.

Beer is generally considered to be a beverage that has high microbiological stability. However, some undesirable lactic acid bacteria (LAB) can grow in beer and consequently spoil this beverage. In this study, bacteriocin-producing Pediococcus acidilactici K10 was used as a means of bio-acidifying the mash and reducing the spoilage LAB content of the beer. The K10 strain had antimicrobial activity against two beer spoilage LAB strains in wort and did not grow in a beer environment. The K10 strain was inoculated before the mashing step. The effect of K10 as a starter culture was investigated and compared with a control. As a result, filtration time was shortened by 17 min, alcohol content was increased by 137%, foam stability was increased by 156%, bitterness was increased by two bitterness units and there was a significant difference (p

In this study, antagonistic activities and probiotic potential of lactic acid bacteria (LAB) derived from a plant-based fermented food, kimchi, were demonstrated. The cell free supernatants (CFS) from Lactobacillus curvatus KCCM 43119, Leuconostoc mesenteroides KCCM 43060, Weissella cibaria KCTC 3746, and W. koreensis KCCM 41517 completely inhibited the growth of foodborne pathogenic bacteria, while neutralized CFS (pH 6.5) partially inhibited the growth. The competition, exclusion, and displacement of foodborne pathogenic bacteria by the LAB strains from adhesion to HT-29 cells were investigated. The LAB strains were able to compete with, exclude, and displace the foodborne pathogenic bacteria. However, the degree of inhibition due to the adhesion was found to be a LAB strain-dependent phenomenon. The LAB strains showed high coaggregation with foodborne pathogenic bacteria, and they also exhibited high resistance to acidic condition. Except W. cibaria KCTC 3746, all LAB strains were capable of surviving in the presence of bile salts. Furthermore, while all LAB strains were resistant to chloramphenicol, kanamycin, streptomycin, gentamicin, and erythromycin, only W. cibaria KCTC 3746 and W. koreensis KCCM 41517 displayed resistance to vancomycin. These results suggest that the LAB strains derived from kimchi exerted antagonistic activities against foodborne pathogenic bacteria with probiotic potential.

The effect of low frequency airborne ultrasound energy on microbiological and physicochemical properties of soybean koji during preparation was investigated. Following inoculation of steam-heated soybeans with spores of Aspergillus oryzae, koji samples were treated with airborne sonication at increasing frequencies of 20, 25, 30, and 35 kHz and incubated at 30°C for 5 days. Amylase and protease activities, the number of fungal spores, bacterial growth, the moisture content, pH, and the titratable acidity were analyzed during koji preparation. All frequencies of low ultrasound treatment tested enhanced enzyme activities. After continuous ultrasonication at 30 kHz, amylase, and protease activities were significantly (p

From the previous study, Enterococcus faecium SH01 was isolated from mukeunji, an over-ripened kimchi, and it produced bacteriocin SH01. Bacteriocin SH01 showed an inhibitory effect against Listeria monocytogenes ATCC 19111, a bacterial strain causing human listeriosis. Crude bacteriocin SH01 was purified by ammonium sulfate precipitation and its inhibitory activity at two concentrations (500 and 1,000 AU/g) against Listeria monocytogenes ATCC 19111 was investigated in ground beef at increasing temperatures (5, 10, 15, and 20℃) for 8 d. The number of Listeria monocytogenes ATCC 19111 significantly decreased (p

Bacillus cereus contamination is one of the major problems for food safety in Korean fermented soybean pastes. A one-sample inspection is not sufficient to determine the overall level of contamination. The aims of this study were to propose new statistical sampling plans for B. cereus in Korean fermented soybean pastes. The acceptable probability over 95% is considered suitable for sampling plan. We suggest statistical sampling plans for B. cereus A (n=5, c=2, m=3, and M=4), B (n=5, c=0, and m=4), or C (n=5, c=1, m=4, and M=5) in Korean fermented soybean pastes based on the B. cereus standard in current Korean Food Code and the median and mean value from the 150 pastes estimated in this study. The statistical sampling plans for B. cereus proposed in this study will provide an advanced food regulation and permit harmonization with international food standards.

A bacteriocinogenic strain identified as Enterococcus faecium SH01 was isolated from mukeunji, a Korean traditional over-ripened kimchi with antimicrobial activities against Listeria monocytogenes KCTC 3569 and Lactobacillus curvatus KFRI 166. The maximum bacteriocin titer (1,280 AU/mL) was detected at the early stationary phase and was maintained for 28 h with no activity loss. The bacteriocin activity, which disappeared after treatment with the proteolytic enzymes α-chymotrypsin, pronase E, proteinase K, and trypsin, was partially inactivated using α-amylase. The activity of bacteriocin SH01 remained after heat treatment (121°C, 15 min) and exposure to pH values from 2–12. The molecular weight of crude bacteriocin SH01 was 3 kDa. The bacteriocin production phenotype (Bac+) was linked to a 6 kb plasmid. Bacteriocin SH1 production was not induced by the co-presence of viable cells, heat treatment, or a cell-free indicator supernatant. The mode of action of bacteriocin SH1 was bactericidal.

The inhibitory effect of Lactobacillus brevis DF01 and Pediococcus acidilactici K10, which had been isolated from kimchi, on enteropathogenic bacterial adhesion to intestinal epithelial cells as well as their probiotic potential was studied. After the incubation of Lb. brevis DF01 and P. acidilactici K10 in simulated gastrointestinal juice, they showed higher tolerance to acidic juice (pH 3.0) and porcine bile extract (0.3%) than Lb. rhamnosus GG. Both Lb. brevis DF01 and P. acidilactici K10 also showed high percentages of autoaggregation and coaggregation with the enteropathogenic bacteria Salmonella Typhimurium and Escherichia coli O157:H7 at 37 °C after 24 h-incubation. It was shown that P. acidilactici K10 showed the highest adhesion to a human intestinal epithelial cell line, HT-29, followed by Lb. brevis DF01 and Lb. rhamnosus GG. In addition, the ability of Lb. brevis DF01 and P. acidilactici K10 to inhibit the adhesion of S. Typhimurium and E. coli O157:H7 to HT-29 cells was also evaluated. P. acidilactici K10 showed the strongest inhibitory effect on the adhesion of S. Typhimurium and E. coli O157:H7 on the basis of competition, displacement and exclusion assays. Lb. brevis DF01 was also shown to effectively inhibit the adhesion of S. Typhimurium and E. coli O157:H7, in particular, inhibition by exclusion. The results suggested that Lb. brevis DF01 and P. acidilactici K10 can be considered as promising probiotic candidates with antimicrobial activities against enteropathogenic bacteria.

To enhance the production of bacteriocin DF01, produced by Lactobacillus brevis DF01, cultivation conditions and medium composition were optimized by using response surface methodology (RSM). The selected 5 factors based on MRS medium were glucose, yeast extract, MgSO4, temperature, and initial pH. Fractional factorial design (FFD) was effective in searching for the main factors. By a 25−1 FFD, glucose, yeast extract, and initial pH were found to be significant factors and had positive effects on bacteriocin production. The effects of the 3 main factors on bacteriocin production were further investigated by a central composite design (CCD). RSM revealed that the maximum bacteriocin production was achieved at yeast extract concentration of 14.56 g/L, glucose concentration of 28.95 g/L, and initial pH of 6.8. After RSM, the titer of bacteriocin was increased by 4-fold.

The bacteriocin-producing lactic acid bacteria Enterococcus faecium DB1 was isolated from Korean traditional gajami sikhae. Culture conditions were optimized by response surface methodology (RSM) to maximize bacteriocin DB1 production. E. faecium DB1 displayed the highest bacteriocin activity when grown in modified MRS medium containing sucrose, rather than glucose, as a carbon source. The effects of temperature, initial pH, and sucrose concentration were tested to determine the optimum conditions for maximum bacteriocin production by E. faecium DB1. A central composite design was used to control the three variables in the experiment. RSM revealed that the optimum values for bacteriocin production were 27.66 g/L sucrose, temperature of 34.37 C, and an initial pH of 6.54. A 2.08-fold increase in bacteriocin production was obtained with sucrose-containing MRS medium compared to production in standard MRS medium.

A DF01 strain that inhibits tyramine-producing Lactobacillus curvatus KFRI 166 was isolated from Dongchimi, a tradi-tional Korean fermented vegetable, and identified as Lactobacillus brevis by biochemical analysis and reverse transcriptasesequencing of 16S rRNA. The antimicrobial compound produced by L. brevis DF01 was secreted at a maximum level of640 AU/mL in late exponential phase in MRS broth, and its activity remained constant during stationary phase. The activityof bacteriocin DF01 was totally inactivated by α-chymotrypsin, pronase E, proteinase K, trypsin, and α-amylase, but not bycatalase, which indicates the compound was glycoprotein in nature. The activity was not affected by pH changes rangingfrom 2 to 12 or heat treatment (60, 80, and 100°C for 30 min), but was reduced after autoclaving. Bacteriocin DF01 hadbacteriolytic activity and a molecular weight of approximately 8.2 kDa, as shown by tricine-SDS-PAGE analysis. There-fore, bacteriocin DF01 can be used in the manufacture of fermented meat products due to its inhibition of tyramine-produc-ing L. curvatus and non-inhibition of L. sake, which is used as a starter culture for meat fermentation.Key words: amylase-sensitive bacteriocin, Lactobacillus brevis, dongchimi

Numerous strains of lactic acid bacteria used in the fermentation of foods are known to produce bacteriocins. In general, bacteriocins are a group of proteinaceous antimicrobial substances that inhibit the growth of closely related bacteria. However, some bacteriocins produced by lactic acid bacteria (LAB) exhibit a relatively broad antimicrobial spectrum and are active against several food‐spoilage and health‐threatening microorganisms. Many investigators have reported on the use of bacteriocins as food preservative to extend the shelflife of various foods. This review decribes the research that has been conducted on bacteriocinogenic lactic acid bacteria— isolated from a wide variety of foods and in some instances of animal origin—and the characteristics of bacteriocins. Special emphasis is placed on their potentials for use as food preservative and on their physicochemical nature, antibacterial spectrum, and genetic behavior.

A recombinant DNA, encoding the chimeric protein of the signal sequence for bifidobacterial α-amylase mature pediocin PA-1, was introduced into Bifidobacterium longum MG1. Biologically active pediocin PA-1 was successfully secreted from the strain and showed bactericidal activity against Listeria monocytogenes and the same molecular mass as native pediocin PA-1.

For rapid detection of Escherichia coli 0157:H7 and Listeria monocytogenes, simple methods for sample preparation and PCR were established and applied to a field test. To improve specificity, primer sets LP43-LP44 and C(+)-D(-) were selected for E. coli O157:H7 and L. monocytogenes, respectively. Through centrifugation and partial heat treatment after enrichment, E. coli 0157:H7 and L. monocytogenes were detected at 1 initial CFU without genomic DNA extraction in the culture and with artificially inoculated food samples including milk, chicken, ham, and pork. Based on the optimized PCR method, a feasibility test was carried out using randomly collected field samples. To remove false positives and false negatives, a PCR method using several primer sets, including the optimized primer set, and a standard culture method were used. With the PCR detection and standard culture methods, two pork samples were positive for L. monocytogenes after enrichment, indications that the PCR assay could be effectively used for rapid, sensitive, and species-specific detection of foodborne pathogens.

The bacteriocin from Pediococcus acidilactici M, designated as Pediocin AcM, was rapidly purified to homogeneity by the pH mediated cell adsorption-desorption method and semi-preparative reversed-phase HPLC. The purification yield was 40.4% and the specific activity was increased by 2450-fold. It gave a single band and a single peak on SDS-PAGE and HPLC analysis, respectively. When subjected to electrospray LC-MS analysis, the protein was found to be highly pure and the molecular weight was determined as 4,618 Da. High concentration of the bacteriocin (> 50 μg ml ) showed good resistance to extremes of pH (1–12) and temperature (121 °C). Pediocin AcM was deduced to be a monomer with an intra-peptide disulfide bond from the results of reversed-phase analytical HPLC analyses after reduction, oxidation and trypsin digestion. P. acidilactici M inhibited a large number of bacteria, including Staphylococcus aureus, Listeria monocytogenes, Clostridium perfringens, Bacillus coagulans, B. cereus , and Aeromonas hydrophila .