Isolation and characterisation of the bacteriocin-producing Leuconostoc citreum HW02 from malts

Herein, a lactic acid bacterium which produces bacteriocin was isolated from malted barley. The isolated strain was identified by 16 S rRNA analysis and designated Leuconostoc citreum HW02. Bacteriocin HW02 was shown to have growth inhibition against health-threatening Gram-positive microorganisms such as Staphylococcus aureus, Listeria monocytogenes, and Latilactobacillus curvatus. Bacteriocin HW02 was reported to be incapable of inhibiting the growth of health-threatening microorganisms after treatment with proteolytic enzymes; thus, it was indicated that bacteriocin is proteinaceous in nature. Bacteriocin HW02 showed stability against a broad range of temperatures (60–121 °C), pH (2–10), various detergents, or organic solvents. Its antimicrobial activity against the indicator strain, Lactiplantibacillus plantarum NCDO 955, was maximised (320 AU mL⁻¹) midway through the stationary phase at 12 h. The crude bacteriocin HW02 molecular weight was estimated to be about 7.7 kDa. After the treatment of the indicator strain with bacteriocin, the maintenance of the optical density of the culture, the morphological analysis of cell-structure abnormalities, and cell shrinkage implied that bacteriocin showed bactericidal activity.     

Inactivation of Staphylococcus aureus in Oat and Soya Drinks by Enterocin AS‐48 in Combination with Other Antimicrobials

The presence of toxicogenic Staphylococcus aureus in foods and the dissemination of methicillin‐resistant S. aureus (MRSA) in the food chain are matters of concern. In the present study, the circular bacteriocin enterocin AS‐48, applied singly or in combination with phenolic compounds (carvacrol, eugenol, geraniol, and citral) or with 2‐nitro‐1‐propanol (2NPOH), was investigated in the control of a cocktail made from 1 methicillin‐sensitive and 1 MRSA strains inoculated on commercial oat and soya drinks. Enterocin AS‐48 exhibited low bactericidal activity against staphylococci in the drinks investigated when applied singly. The combinations of sub‐inhibitory concentrations of enterocin AS‐48 (25 μg/mL) and phenolic compounds or 2NPOH caused complete inactivation of staphylococci in the drinks within 24 h of incubation at 22 °C. When tested in oat and soya drinks stored for 7 d at 10 °C, enterocin AS‐48 (25 μg/mL) in combination with 2NPOH (5.5 mM) reduced viable counts rapidly in the case of oat drink (4.2 log cycles after 12 h) or slowly in soya drink (3.8 log cycles after 3 d). The same combined treatment applied on drinks stored at 22 °C achieved a fast inactivation of staphylococci within 12 to 24 h in both drinks, and no viable staphylococci were detected for up to 7 d of storage. Results from the study highlight the potential of enterocin AS‐48 in combination with 2NPOH for inactivation of staphylococci.     

Effect of Extrinsic Parameters on the Production of a Bacteriocin by Lactobacillus buchneri,Isolated from Mexican Raw Sausages

Bacteriocins are antimicrobial peptides acting on certain pathogens and spoilage microorganisms. They have recently been considered as natural alternative to food preservatives. The objective of this work was to study the inhibition spectrum of a bacteriocin produced by Lactobacillus buchneri, isolated from a Mexican sausage, against Listeria monocytogenes and several lactic acid bacteria, and the influence of several extrinsic parameters on bacteriocin activity. Culture media (MRS or APT) had no significant effect on bacteriocin production (p < 0.060) although higher activity was observed in MRS (220 AU/mL). Conversely, temperature significant affected production (p > 0.001). Bacteriocin activity was also increased by 100% nitrogen atmosphere (180 AU/mL) as compared to N₂/CO₂ (50:50) (150 AU/mL) and 100% CO₂ (60 AU/mL). Maximum activity occurred at the end of the exponential growth phase. This bacteriocin inhibited some Listeria monocytogens strains and several lactic acid bacteria, mainly Lactobacillus sp.     

Characterisation of anti‐Staphylococcus aureus activity of quercetin

Although many antibiotics are available for the treatment of bacterial infections, the emergence and global spread of antibiotic‐resistant bacteria is a community‐wide problem. To overcome this problem, we must explore alternative antimicrobials. This study investigated the antibacterial properties of quercetin, a flavonoid present in vegetables and fruits. Quercetin was tested against gram‐positive and gram‐negative bacteria and was found to exert selective antibacterial activity against Staphylococcus aureus, including methicillin‐resistant S. aureus (MRSA), and Staphylococcus epidermidis. Some clinical MRSA strains showed remarkable susceptibility to quercetin. In combination with antibiotics, such as oxacillin, ampicillin, vancomycin, gentamicin, and erythromycin, quercetin showed markedly enhanced antibacterial activity against MRSA. We also report quercetin‐induced aggregation of S. aureus cells; the morphological changes in these cells, as assessed by electron microscopy; and the colony‐spreading ability of quercetin‐sensitive MRSA, all of which revealed the unique antibacterial properties of quercetin against S. aureus.     

First report of methicillin‐resistant Staphylococcus aureus in tank cultured dusky kob (Argyrosomus japonicus), and evaluation of three phenotypic methods in the detection of MRSA

This study was conducted to ascertain the occurrence of methicillin‐resistant Staphylococcus aureus (MRSA) in marine finfish (Argyrosomus japonicus) harvested from the wild and two re‐circulatory aquaculture systems, and evaluating the reliability of three phenotypic methods in the detection of methicillin resistance. A total of 120 dusky kob fish were sampled for S. aureus detection using conventional methods and confirmed by polymerase chain reaction (PCR) targeting the nuc gene. Methicillin resistance was determined by molecular detection of the mecA gene. Using mecA as the defining standard, the specificities and sensitivities of cefoxitin disc diffusion, oxacillin screen agar, and growth of S. aureus on Brilliance MRSA II Agar were evaluated. A total of 321 presumptive S. aureus isolates were recovered by culture, out of which 202 (62.9%) were confirmed by PCR. Of these, 33 (16.3%) strains were mecA positive while 169 (83.7%) were mecA negative. The sensitivities and specificities of MRSA detection was 93.9 and 91.7%, 81.8 and 92.3%, and 87.9 and 94.1% for cefoxitin disc, oxacillin screen agar test, and Brilliance MRSA II agar, respectively. This is so far the first report of MRSA in dusky kob aquaculture in South Africa. In the absence of molecular techniques, cefoxitin disc diffusion test is recommended along with any other phenotypic method to improve MRSA detection from samples of veterinary origin.

Practical implications

Methicillin‐resistant Staphylococcus aureus currently presents one of the greatest challenges for medical research worldwide, as well as is one of the most important causes of bacteria gastroenteritis due to preformed toxins in foods. There is a dearth of knowledge on marine foods as carriers/sources of MRSA infection. There are also major discrepancies obtained with MRSA detection methods, making effective detection of this pathogen complicated. The results from this study show that healthy aquaculture fish are reservoirs of MRSA, thus it is necessary to regularly monitor marine foods. Cefoxitin disc diffusion test is recommended as the preferred method for detection of MRSA from fish/food samples where molecular methods are lacking.     

Antibacterial activity of proteins extracted from the pulp of wild edible fruit of Bromelia pinguin L.

Bromelia pinguin L. is a natural source of bioactive compounds. The main purpose of this research was to isolate and characterize bioactive proteins from its fruit. B. pinguin proteins were fractionated by gel filtration chromatography, and analyzed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The antibacterial activity of the proteins was analyzed against Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 25923, and the enzymatic activity was evaluated by protease activity and trypsin inhibitions assays. Protein fraction obtained by gel filtration chromatography exhibited antibacterial activity against E. coli (minimum inhibitory concentration [MIC] 0.3492 mg/mL) and S. aureus (MIC 0.6845 mg/mL). The proteolytic activity of the fraction was 0.985 U_cas/mL. The substrate-sodium dodecyl sulfate-polyacrylamide gel electrophoresis assay detected protease inhibitors with molecular weights of 43 and 74 kDa. Antibacterial studies of E.coli and S. aureus were determined by comparing the protein fraction with different antibiotics. The antibacterial activity of proteins extracted from the pulp of the fruit of Bromelia pinguin L. could be related to the presence of enzymes, protease inhibitors and peptides.     

Production,mode of action and sequencing of the corresponding gene of a bacteriocin produced by Lactococcus lactis 19.3

Lactococcus lactis 19.3 produces a bacteriocin with a wide inhibitory spectrum, including the food pathogen Listeria monocytogenes. The producing strain was able to grow and produce similar amounts of bacteriocin in MRS medium, cow's milk and soya milk, respectively. The mode of action of this bacteriocin was further investigated using two indicator strains, Lactobacillus delbrueckii subsp. bulgaricus LMG 6901^T and Listeria monocytogenes ATCC 1911‐1. The bacteriocin displayed a bactericidal effect, causing a rapid decrease of the cell viability. Bacteriocin treatment of the sensitive strains resulted in major morphological changes, including pore formation, as shown by scanning electron microscopy. Finally, the bacteriocin‐encoding gene was identified by sequencing. The presence of nisin gene was confirmed. Based on all our data gathered so far, L. lactis 19.3 is a good candidate for a starter or protective culture in the manufacturing of both fermented dairy and vegetarian food products.     

Genetic Diversity and Antibiotic Resistance Patterns of Staphylococcus Aureus Isolated from Leaf Vegetables in Korea

Staphylococcus aureus is an important foodborne pathogen on global basis. The current study investigated the genetic patterns in S. aureus isolates from leaf vegetables (n = 53). Additional isolates from livestock (n = 31) and humans (n = 27) were compared with the leaf vegetable isolates. Genes associated with toxins, antibiotic resistance, and pulsed‐field gel electrophoresis (PFGE) patterns were analyzed. At least 1 enterotoxin‐encoding gene (sea, seb, sec, sed, and see) was detected in 11 of 53 (20.75%) leaf vegetable isolates. When the agr (accessory gene regulator) grouping was analyzed, agr II was the major group, whereas agr IV was not present in leaf vegetable isolates. All S. aureus isolates from leaf vegetables were resistant to more than one of the antibiotics tested. Nineteen of 53 (35.85%) isolates from leaf vegetables exhibited multidrug‐resistance, and 11 of these were MRSA (methicillin‐resistant S. aureus). A dendrogram displaying the composite types of S. aureus isolates from 3 origins was generated based on the combination of the toxin genes, agr genes, antibiotic resistance, and PFGE patterns. The isolates could be clustered into 8 major composite types. The genetic patterns of S. aureus isolates from leaf vegetables and humans were similar, whereas those from livestock had unique patterns. This suggests some S. aureus isolates from leaf vegetables to be of human origin.     

Comparison of the performances of different fermentation strategies on cell growth and bacteriocin production by Lactobacillus curvatus CWBI‐B28

The dynamics of cell growth and bacteriocin production by Lactobacillus curvatus CWBI‐B28 in modified De Man/Rogosa/Sharp (mMRS) broth with various concentrations of glucose and complex nitrogen source (CNS; peptone, yeast extract and meat extract) was investigated in flask fermentations and in a laboratory fermentor using batch and fed‐batch cultivations. In fed‐batch fermentation the rate of feeding of the reactor with the substrates was either maintained constant (0.12 L h^?1) or varied exponentially as a function of time. The results showed that both cell growth and bacteriocin activity were influenced by changes in the concentrations of glucose and CNS. Optimal growth and bacteriocin activity were obtained in mMRS broth containing 40 g L^?1 glucose and 40 g L^?1 CNS (mMRS_40/40). A bacteriocin titre of 4266 AU mL^?1 and a cell count of 8.7 log colony‐forming units (cfu) mL^?1 were recorded when this medium was used for cultivation. In batch fermentation using the same medium, a higher cell count (9.5 log cfu mL^?1) and twice as much bacteriocin as in flask fermentation were produced. The highest bacteriocin titre (8533 AU mL^?1) was obtained with fed‐batch fermentation at an exponentially varying rate of feeding. Bacteriocin activity and cell dry mass did not always correlate. Copyright © 2007 Society of Chemical Industry     

A Multiplex PCR Assay for the Rapid and Sensitive Detection of Methicillin‐Resistant Staphylococcus aureus and Simultaneous Discrimination of Staphylococcus aureus from Coagulase‐Negative Staphylococci

Abstract: Methicillin‐resistant Staphylococcus aureus (MRSA) is a global health concern, which had been detected in food and food production animals. Conventional testing for detection of MRSA takes 3 to 5 d to yield complete information of the organism and its antibiotic sensitivity pattern. So, a rapid method is needed to diagnose and treat the MRSA infections. The present study focused on the development of a multiplex PCR assay for the rapid and sensitive detection of MRSA. The assay simultaneously detected 4 genes, namely, 16S rRNA of the Staphylococcus genus, femA of S. aureus, mecA that encodes methicillin resistance, and one internal control. It was rapid and yielded results within 4 h. The analytical sensitivity and specificity of the multiplex PCR assay was evaluated by comparing it with the conventional method. The analytical sensitivity of the multiplex PCR assay at the DNA level was 10 ng DNA. The analytical specificity was evaluated with 10 reference staphylococci strains and was 100%. The diagnostic evaluation of MRSA was carried out using 360 foodborne staphylococci isolates, and showed 99.1% of specificity, 96.4% of sensitivity, 97.5% of positive predictive value, and 97.3% of negative predictive value compared to the conventional method. The inclusion of an internal control in the multiplex PCR assay is important to exclude false‐negative cases. This test can be used as an effective diagnostic and surveillance tool to investigate the spread and emergence of MRSA.     

ANTIBACTERIAL ACTIVITY OF EXTRACTS FROM FAMILY ZINGIBERACEAE AGAINST FOODBORNE PATHOGENS

Chloroformic extracts of selected Thai medicinal plants commonly employed to treat infections were investigated for their antibacterial activity against important foodborne pathogenic bacteria. These included Bacillus cereus, Staphylococcus aureus, methicillin‐resistant S. aureus (MRSA), Escherichia coli O157:H7, Salmonella Typhi and Shigella sp. Among 33 extracts tested, only chloroformic extracts of five plant species exhibited antibacterial properties. Alpinia galanga, Boesenbergia rotunda, Zingiber zerumbet and Piper betel were active against S. aureus. Barleria lupulina was active against B. cereus. Only the extract from P. betel leaves possessed activity against gram‐negative bacteria. As extracts from the three plant species belonging to family Zingiberaceae displayed strong activity against S. aureus, they were further tested against 17 clinical isolates. Minimum inhibitory concentration (MIC) values of B. rotunda, A. galanga and Z. zerumbet extracts against most clinical S. aureus isolates were 0.01, 0.19 and 0.79 mg/mL and the minimum bactericidal concentration (MBC) values were 0.19, 1.57 and >12.5 mg/mL, respectively. Significant growth inhibition of MRSA was observed in the cultures incubated in the presence of the B. rotunda extract, A. galanga and Z. zerumbet. B. rotunda exhibited the greatest activity among the three plant species against S. aureus at MIC, 2MIC and MBC within 2 h.     

Characterization of subtilin KU43 Produced by Bacillus subtilis KU43 isolated from traditional Korean fermented foods

An antimicrobial peptide produced by Bacillus subtilis KU25, KU43, and KU44 was isolated from traditional Korean fermented foods and characterized. It was named as subtilin KU25, KU43, and KU44. Subtilin KU25, KU43, and KU44 were sensitive against α-chymotrypsin, protease XIII, and various proteinase enzymes, respectively. B. subtilis KU43 was selected as the producer with the broadest antimicrobial spectrum. Subtilin KU43 was stable at a pH range of 3 to 9 for 4 h, and withstood exposure to temperatures of 50–90°C for 30 min. The mode of inhibition against Listeria monocytogenes ATCC 15313 involved a bactericidal effect by a reduction in the cell numbers and breakage of the indicator cell membranes. The molecular mass of subtilin KU43 was measured at approximately 3.5 kDa. These results demonstrate the development of novel strains from traditional Korean fermented foods, and illustrate the possibility that some of these strains might generate a natural preservative compound.     

Characterisation of an antiviral pediocin-like bacteriocin produced by Enterococcus faecium

The bacteriocin-producing strain Enterococcus faecium ST5Ha was isolated from smoked salmon and identified by biomolecular techniques. Ent. faecium ST5Ha produces a pediocin-like bacteriocin with activity against several lactic acid bacteria, Listeria spp. and some other human and food pathogens, and remarkably against HSV-1 virus. Bacteriocin ST5Ha was produced at high levels in MRS broth at 30 °C and 37 °C, reaching a maximum production of 1.0 × 10⁹ AU/ml, checked against Listeria ivanovii ATCC19119 as target strain and surrogate of pathogenic strain Listeria monocytogenes. The molecular weight of bacteriocin ST5Ha was estimated to be 4.5 kDa according to tricine-SDS-PAGE data. Ent. faecium ST5Ha harbors a 1.044 kb chromosomal DNA fragment fitting in size to that of pediocin PA-1/AcH. In addition, the sequencing of bacteriocin ST5Ha gene indicated 99% of DNA homology to pediocin PA-1/AcH. The combined application of low levels (below MIC) of ciprofloxacin and bacteriocin ST5Ha resulted in a synergetic effect in the inhibition of target strain L. ivanovii ATCC19119. Bacteriocin ST5Ha displayed antiviral activity against HSV-1, an important human pathogen, with a selectivity index of 173. To the best of our knowledge, this is the first report on Ent. faecium as a potential producer of pediocin-like bacteriocin with antiviral activity.     

Antibacterial activity and mechanism of a novel bacteriocin produced by Lactiplantibacillus plantarum against Escherichia coli and Staphylococcus aureus

The purpose of the study was to explore the antibacterial effect and mechanism of a novel bacteriocin of Lactiplantibacillus plantarum (L. plantarum) against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). The antibacterial activity was determined by the Oxford cup method, and the dynamic growth curves were conducted with continuous shaking incubation. The result showed that the bacteriocin was a protein or protein-like antibacterial substance susceptible to pepsin and trypsin. And it had good antibacterial activity, pH stability, thermostability and enzyme treatment stability against E. coli and S. aureus. The SEM, flow cytometry and nucleic acid leakage showed that the bacteriocin disrupted the cell structures of the two bacteria by damaging cell walls and cell membranes. An agarose gel electrophoresis and SDS-PAGE analysis showed that the bacteriocin inhibited DNA replication and interfered with the protein formation, which resulted in the inhibition of the two bacteria's growth. Therefore, the use of the L. plantarum bacteriocin might be a promising biocontrol strategy to inhibit the pollution of E. coli and S. aureus simultaneously in foods.     

Comparison of the enterotoxigenic types,toxic shock syndrome toxin I (TSST-1) strains and antibiotic susceptibilities for enterotoxigenicStaphylococcus aureusstrains isolated from food and clinical samples

The distribution of enterotoxin types and toxic shock syndrome toxin I (TSST-1) strains for 176Staphylococcus aureusstrains obtained from food samples and 62S. aureusstrains isolated from clinical samples were compared. It was found that for both the food and clinical isolates, staphylococcal enterotoxin A (SEA) strains accounted for the major part (75% and 45% of the total enterotoxigenic strains for food and clinical isolates, respectively) followed by SEB or SEAB, SEC and SED strains. For food isolates, none of theS. aureusstrains was TSST-1 strain while for clinical isolates, three strains (1 SEC, 1 SED and 1 SEAB strain) were found to be TSST-1 strains. When susceptibilities for these enterotoxigenicS. aureusstrains to antibiotics, such as penicillin, oxacillin, vancomycin, methicillin, streptomycin, tetracycline, gentamycin and kanamycin were compared, results showed that 51.6% of the food isolates were resistant to penicillin G only but sensitive to the other antibiotics tested. Also, 8 of the 64 enterotoxigenic strains isolated from food samples were all sensitive to antibiotics while none of the enterotoxigenic strains from clinical samples showed this antibiotic susceptibility pattern. No methicillin resistantS. aureus(MRSA) strains could be found among the food isolates. On the other hand, the majority (42.4%) of the enterotoxigenicS. aureusstrains from clinical samples were penicillin and/or other antibiotics resistant MRSA strains. Since MRSA strains often posed the therapeutic problem, the MRSA strains were further confirmed by PCR assay using the primers specific formecA gene. It was found that results obtained from the disc agar diffusion method and the PCR method were the same.     

Biochemical,antimicrobial and molecular characterization of a noncytotoxic bacteriocin produced by Lactobacillus plantarum ST71KS

Lactobacillus (Lb.) plantarum ST71KS was isolated from homemade goat feta cheese and identified using biochemical and molecular biology techniques. As shown by Tricine-SDS-PAGE, this lactic acid bacterium produces a bacteriocin (ST71KS) with an estimated molecular weight of 5.0 kDa. Bacteriocin ST71KS was not affected by the presence of α-amylase, catalase and remained stable in a wide range of pH and after treatment with Triton X-100, Triton X-114, Tween 20, Tween 80, NaCl, SDS, urea and EDTA. This bacteriocin also remained active after being heated at 100 °C for 2 h and even after 20 min at 121 °C; however, it was inactivated by proteolitic enzymes. Production of bacteriocin ST71KS reached 6400 AU/mL during stationary growth phase of Lb. plantarum cultivated in MRS at 30 °C and 37 °C. Bacteriocin ST71KS displayed a bactericidal effect against Listeria monocytogenes strains 603 and 607 and did not adsorb to the producer cells. Lb. plantarum ST71KS harbors two bacteriocin genes with homology to plantaricin S and pediocin PA-1. These characteristics indicate that bacteriocin ST71KS is a class IIa bacteriocin. The peptide presented no toxic effect when tested in vitro with kidney Vero cells, indicating safe technological application to control L. monocytogenes in foods.     

Purification and partial characterization of antilisterial bacteriocin produced by Pediococcus pentosaceus KJBC11 from Idli batter fermented with Piper betle leaves

Bacteriocins are natural antimicrobial agents mainly act against closely related bacteria and at times unrelated organisms including various food spoilage and pathogenic organisms. Bacteriocins from Lactic Acid Bacteria (LAB) have been widely used for food preservation because of their safety nature. In this study, bacteriocin from Pediococcus pentosaceus KJBC11 was purified with the recovery of 15%, using cell adsorption‐desorption technique, gel permeation chromatography, hydrophobic interaction chromatography, and reversed phase HPLC successively. Molecular weight of bacteriocin KJBC11 was determined to be 4.5 kDa using Tricine SDS‐PAGE. The bacteriocin KJBC11 showed strong inhibitory activity against various Gram‐positive and Gram‐negative pathogens. Bacteriocin KJBC11 activity was not altered when treated with amylase, lipase, and trypsin but inactivated by protease and proteinase enzyme treatment. It was heat stable (100°C, 1 hr) and exhibited strong antimicrobial activity within the pH 3–7. Its mode of action was bactericidal in nature as revealed against Listeria monocytogenes showing leakage of internal contents out of the cell. Bacteriocin KJBC11 could belong to Class IIa‐pediocin like bacteriocins based on its characteristics as well as the presence of pediocin gene in the genome of the isolate KJBC11.

Practical applications

Studies on Lactic Acid Bacteria and bacteriocin have attracted increasing attention for a long time, due to their established role in food fermentation and preservation. The antimicrobial properties of bacteriocin became a trademark approach to achieve food safety and to counter the menace of antibiotic resistant pathogens. This study revealed a potent bacteriocin with wide range of antibacterial activity against various foodborne and clinically important pathogens to have potential application in food preservation and biomedicine.     

Biocontrol of Listeria monocytogenes in a model cultured milk (lben) by in situ bacteriocin production from Lactococcus lactis ssp. lactis

Bacteriocin‐producing (Bac⁺) Lactococcus lactis ssp. lactis CCMM/IAV/BK1 isolated from traditional lben was used in the preparation of lben from pasteurized milk to assess its potential inhibitory activity against Listeria monocytogenes ATCC 7644. Production of bacteriocin (arbitrary units, AU) in MRS broth fortified with yeast extract (MRSY) in a fermentor under controlled and uncontrolled pH conditions was also investigated. This Bac⁺ strain yielded about 35 times more bacteriocin when the pH was maintained constant at 6.5 than under varying pH conditions. To test the effect of in situ bacteriocin production against L. monocytogenes, lben was made from cow's milk artificially contaminated with approximately 10⁷ cfu/mL and fermented with a mixed mesophilic starter culture consisting of the lactococcal Bac⁺ organism and Lc. lactis ssp. lactis biovar diacetylactis 66, a diacetyl‐producing strain, in a ratio of 1 : 1. Numbers of L. monocytogenes were monitored during fermentation and storage of lben at refrigeration temperature (c. 7°C) for up to 6 days. Performances of the Bac⁺ starter were compared to those of an isogenic Bac^? derivative strain obtained from the Bac⁺ starter by curing with ethidium bromide. The results showed that the amount of L. monocytogenes decreased to below the detectable level in a 1‐mL sample within 24 h of storage at 7°C in lben fermented with the Bac⁺ starter culture. On the contrary, L. monocytogenes survived for 6 days of storage at 7°C in lben made with the Bac^? starter. The Bac⁺ wild strain of the starter studied could be adequately used to produce lben or similar indigenous fermented milks of improved hygienic quality on an industrial scale. Alternatively, it could be used as an adjunct in minimally processed products or in products obtained from raw milk to add a safety factor.     

Optimisation of bacteriocin production of Lactococcus lactis subsp. lactis MA23, a strain isolated from Boza

Lactococcus lactis subsp. lactis MA23 produces a bacteriocin (6400 AU/mL) that inhibits the growth of many Gram‐positive bacteria but is not active against Gram‐negative bacteria. This bacteriocin inhibits growth of lactococcal strains that are producing nisin, lacticin or lactococcin suggesting it to be different from these bacteriocins. The nutritional requirements and optimal growth conditions for MA23 bacteriocin production were studied with fed‐batch fermentations. The optimal pH, carbon source and nitrogen source for bacteriocin production were pH 6.5, sucrose (0.5%) and yeast extract (1%), respectively.     

Antilisterial and amylase-sensitive bacteriocin producing Enterococcus faecium SH01 from Mukeunji,a Korean over-ripened kimchi

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.