Antimicrobial activity of pediocin PA-1 against Oenococcus oeni and other wine bacteria

Pediocin PA-1 is an antimicrobial peptide produced by lactic acid bacteria (LAB) that has been sufficiently well characterised to be used in food industry as a biopreservative. Sulphur dioxide is the traditional antimicrobial agent used during the winemaking process to control bacterial growth and wine spoilage. In this study, we describe the effect of pediocin PA-1 alone and in combination with sulphur dioxide and ethanol on the growth of a collection of 53 oenological LAB, 18 acetic acid bacteria and 16 yeast strains; in addition, production of pediocin PA-1 by Pediococcus acidilactici J347-29 in presence of ethanol and grape must is also reported. Inhibitory concentrations (IC) and minimal bactericide concentrations of pediocin PA-1 were determined against LAB, and revealed a bacteriostatic effect. Oenococcus oeni resulted more sensitive to pediocin PA-1 (IC₅₀ = 19 ng/ml) than the other LAB species (IC₅₀ = 312 ng/ml). Cooperative inhibitory effects of pediocin PA-1 and either sulphur dioxide or ethanol were observed on LAB growth. Moreover, the pediocin PA-1 producing P. acidilactici strain J347-29 was able to grow and produce the bacteriocin in presence of ethanol (up to 4% ethanol in the fermentation broth) and grape must (up to 80%), which indicated that pediocin PA-1 can be considered as a potential biopreservative in winemaking.     

Production of pediocin PA-1, and coproduction of nisin A and pediocin PA-1, by wild Lactococcus lactis strains of dairy origin

Heterologous production of pediocin PA-1 in nisin and non-nisin-producing Lactococcus lactis strains, which had been previously selected because of their technological properties for cheese making, was investigated. Plasmid pFI2160, which contains a hybrid gene (L-pedA) encoding the fusion between the lactococcin A leader and propediocin PA-1, and also the genes lcnC and lcnD, that encode the lactococcin A secretion apparatus, was introduced into L. lactis ESI 153 and L. lactis ESI 515 (Nis⁺). The pediocin production level of their respective transformants, L. lactis CL1 and L. lactis CL2 (Nis⁺), was approximately 600 and 400 ng mL⁻¹, respectively, which represents a 30% and a 20% of the quantity produced by the natural pediocin PA-1 producer Pediococcus acidilactici 347. Transformation of L. lactis ESI 515 with pFI2160 did not affect its ability to produce nisin. Pediocin bioassays showed the stability of pFI2160 in both heterologous hosts under selective and non-selective conditions.     

Antibacterial activity of antimicrobial peptide gcDefb1 against foodborne pathogenic bacteria and its application in pork storage

Food Science and Biotechnology - Pork and its products are preferably contaminated by bacteria; thus, it is essential to develop low-cost, high-efficiency and biologically safe preservatives to...     

Chimeras of mature pediocin PA-1 fused to the signal peptide of enterocin P permits the cloning, production, and expression of pediocin PA-1 in Lactococcus lactis

Chimeras of pediocin PA-1 (PedA-1), a bacteriocin produced by Pediococcus acidilactici PLBH9, fused to the signal peptide of enterocin P (EntP), a sec-dependent bacteriocin produced by Enterococcus faecium P13, permitted the production of PedA-1 in Lactococcus lactis. Chimeric genes encoding the EntP signal peptide (SP(entP)) fused to mature PedA-1 (pedA), with or without its immunity gene (pedB), were cloned into the expression vector pMG36c to generate the recombinant plasmids pMPP9 (SP(entP):pedA) and pMPP14i (SP(entP):pedA + pedB). Transformation of competent L. lactis subsp. lactis IL1403, L. lactis subsp. cremoris NZ9000, and L. lactis subsp. lactis DPC5598 with the recombinant plasmids has permitted the detection and quantitation of PedA-1 and the coproduction of nisin A and PedA-1 in supernatants of producer cells with specific anti-PedA-1 antibodies and a noncompetitive indirect enzyme-linked immunosorbent assay. Recombinant L. lactis hosts carrying pMPP9 or pMPP14i displayed antimicrobial activity, suggesting that mature PedA-1 fused to SP(EntP) is the minimum requirement for the synthesis, processing, and secretion of biologically active PedA-1 in L. lactis. However, the production and antimicrobial activity of the PedA-1 produced by L. lactis was lower than that produced by the P. acidilactici control strains.     

Antimicrobial activity of lactoferrin against foodborne pathogenic bacteria incorporated into edible chitosan film

The objectives of this research were to develop and characterize edible chitosan film containing lactoferrin as a natural antimicrobial agent, and to investigate the combination effects of lactoferrin with lysozyme in chitosan film against the growth of Escherichia coli O157:H7 and Listeria monocytogenes. Chitosan films containing lactoferrin, lysozyme, or nisin were fabricated, and the antimicrobial concentrations were 0.5, 1, or 2 mg in a circular disc of chitosan film. Three concentrations of lactoferrin or EDTA (0.28, 0.56, or 1.12 mg per disc) were also incorporated into the chitosan film containing lysozyme to investigate the combination effects of lactoferrin. The water barrier properties of the chitosan films containing lactoferrin were characterized. The antimicrobial activities against E. coli O157:H7 and L. monocytogenes were determined using the agar diffusion assay and cell count assay. The chitosan films containing lactoferrin less than 1 mg per disc did not alter the water vapor permeability of the chitosan film. Although the film containing lysozyme exhibited significant antimicrobial activity, the incorporation of lactoferrin alone into chitosan film did not exhibit significant antimicrobial activity against both E. coli O157:H7 and L. monocytogenes. However, the combination of lactoferrin with lysozyme-containing chitosan film significantly decreased the growth of E. coli O157:H7, exhibiting a comparable effect to that of the combination of EDTA with lysozyme (P < 0.05). Furthermore, the combination of lactoferrin with lysozyme in chitosan film exhibited greater reduction in the growth of L. monocytogenes than did the combination EDTA with lysozyme, resulting in an approximate 3-log reduction.     

Screening of antimicrobial activities of disinfectants and cleaning agents against foodborne spoilage microbes

 The antimicrobial activities of six disinfectants and cleaning agents against five food and brewery spoilage microbes were studied using the 5-5-5 suspension and a surface test simulating surface disinfection in actual use. The tests were carried out using the lowest recommended concentrations. The surface test was performed both with and without organic soil for various exposure times. In the suspension test, almost all the disinfectant and cleaning agents had adequate antimicrobial activity, except against Bacillus spores. The peroxide-based disinfectant and the isopropanol-based cleaning agent were both ineffective against Saccharomyces cerevisiae. In the surface test without soil the hypochlorite-based disinfectant was effective after an exposure of 10 min against all the microbes tested. The isopropanol-based cleaning agent was effective against all the vegetative cells tested. In the presence of soil, hypochlorite was effective against Listeria monocytogenes and Pseudomonas aeruginosa. In the test performed the lubricant which contained disinfectant had no antimicrobial activity against the surface-attached contaminants tested. P. aeruginosa was the most sensitive test organism and the Bacillus spores were the most resistant towards the agents tested. Received: 30 June 1998     

Surface application of lysozyme, nisin, and EDTA to inhibit spoilage and pathogenic bacteria on ham and bologna

A study was conducted to determine if the effectiveness of an antimicrobial treatment for cooked ham and bologna would be increased or maintained when applied in a surface coating. Cooked 10-g disks of ham and bologna sausage received one of three treatments: no coating (control), coating with 0.2 g of 7% (wt/vol) gelatin gel (gel-control), or coating with 0.2 g of 7% gelatin gel containing 25.5 g/liter of lysozyme-nisin (1:3) plus 25.5 g/liter of EDTA (gel-treated). The samples were then inoculated with one of six test organisms: Brochothrix thermosphacta, Escherichia coli O157:H7, Lactobacillus sakei, Leuconostoc mesenteroides, Listeria monocytogenes, or Salmonella Typhimurium. Inoculated samples were vacuum packed and stored at 8 degrees C for 4 weeks. The antimicrobial gel treatment had an immediate bactericidal effect up to 4 log CFU/cm2 on the four gram-positive organisms tested (B. thermosphacta, Lactobacillus sakei, Leuconostoc mesenteroides, and Listeria monocytogenes) and inhibited the growth of these organisms during the 4 weeks of storage. The antimicrobial gel treatment also had a bactericidal effect on the growth of Salmonella Typhimurium during storage. The numbers of E. coli O157:H7 on ham were reduced by 2 log CFU/cm2 following treatment with both antimicrobial-containing and non-antimicrobial-containing gels during the 4-week storage period. No effect was observed on the growth of E. coli O157:H7 on bologna.     

Efficacy of plant essential oils against foodborne pathogens and spoilage bacteria associated with ready-to-eat vegetables: antimicrobial and sensory screening

The objectives of this study were to evaluate the antimicrobial activity of plant essential oils (EOs) against foodborne pathogens and key spoilage bacteria pertinent to ready-to-eat vegetables and to screen the selected EOs for sensory acceptability. The EOs basil, caraway, fennel, lemon balm, marjoram, nutmeg, oregano, parsley, rosemary, sage, and thyme were evaluated. The bacteria evaluated were Listeria spp., Staphylococcus aureus, Lactobacillus spp., Bacillus cereus, Salmonella, Enterobacter spp., Escherichia coli, and Pseudomonas spp. Quantitative antimicrobial analyses were performed using an absorbance-based microplate assay. Efficacy was compared using MIC, the half maximum inhibitory concentration, and the increase in lag phase. Generally, gram-positive bacteria were more sensitive to EOs than were gram-negative bacteria, and Listeria monocytogenes strains were among the most sensitive. Of the spoilage organisms, Pseudomonas spp. were the most resistant. Oregano and thyme EOs had the highest activity against all the tested bacteria. Marjoram and basil EOs had selectively high activity against B. cereus, Enterobacter aerogenes, E. coli, and Salmonella, and lemon balm and sage EOs had adequate activity against L. monocytogenes and S. aureus. Within bacterial species, EO efficacy was dependent on strain and in some cases the origin of the strain. On a carrot model product, basil, lemon balm, marjoram, oregano, and thyme EOs were deemed organoleptically acceptable, but only oregano and marjoram EOs were deemed acceptable for lettuce. Selected EOs may be useful as natural and safe additives for promoting the safety and quality of ready-to-eat vegetables.     

Antimicrobial activities of Asian plant extracts against pathogenic and spoilage bacteria

Food Science and Biotechnology - This study was conducted to investigate the antimicrobial effects of 300 Asian plant extracts (PEs) against pathogenic and spoilage bacteria. The antimicrobial...     

Antimicrobial activity of Microgard against food spoilage and pathogenic microorganisms

Microgard, a commercially available fermented milk product containing antimicrobial metabolites, was a potent inhibitor for Gram-negative bacteria such as Pseudomonas, Salmonella, and Yersinia when 1% concentration was incorporated into agar media. Gram-positive Bacillus cereus, Staphylococcus aureus, and Listeria monocytogenes were insensitive to Microgard. Kluyveromyces marxianus, an unidentified black yeast, and Penicillium expansum were partially suppressed, whereas Aspergillus niger and a yogurt spoilage yeast were tolerant to 5% Microgard. Optimum activity of Microgard was at pH 5.3 and below; the concentration that gave complete inhibition depended upon the number of bacteria present as well as the genus tested. Blood agar base reversed the antagonistic activity of Microgard against Pseudomonas putida compared with plate count agar.     

Composition and antimicrobial properties of Sardinian Juniperus essential oils against foodborne pathogens and spoilage microorganisms

In this work, the chemical compositions and antimicrobial properties of Juniperus essential oils and of their main components were determined. Five berry essential oils obtained from different species of Juniperus growing wild in Sardinia were analyzed. The components of the essential oils were identified by gas chromatography-mass spectrometry (GC-MS) analysis. The antimicrobial activities of the oils and their components against food spoilage and pathogenic microorganisms were determined by a broth microdilution method. The GC-MS analysis showed a certain variability in the concentrations of the main constituents of the oils. Alpha-pinene was largely predominant in the oils of the species J. phoenicea subsp. turbinata and J. oxycedrus. Alpha-pinene and myrcene constituted the bulk (67.56%) of the essential oil of J. communis. Significant quantitative differences were observed for myrcene, delta-3-carene, and D-germacrene. The results of the antimicrobial assay show that the oils of J. communis and J. oxycedrus failed to inhibit any of the microorganisms at the highest concentrations tested (MLC > or = 900 microg/ml), while the oils extracted from J. turbinata specimens were active against fungi, particularly against a strain of Aspergillus flavus (an aflatoxin B1 producer). Of the single compounds tested, delta-3-carene was found to possess the broadest spectrum of activity and appeared to contribute significantly to the antifungal activity observed for J. turbinata oils. This activity may be helpful in the prevention of aflatoxin contamination for many foods.     

Antibacterial efficiency of Finnish spice essential oils against pathogenic and spoilage bacteria

The antibacterial properties of 13 essential oils, derived from spices grown in Finland, were examined with an agar diffusion method against 12 bacterial strains. The organisms tested included both spoilage and pathogenic bacteria. The gram-positive bacteria appeared to be more sensitive than the gram-negative organisms, Clostridium botulinum and Clostridium perfringens being the most sensitive. Oregano, savory, and thyme showed the broadest antibacterial activity by distinctly inhibiting the growth of all the organisms tested. By gas chromatography-mass spectrometry analysis, differences were noted in the composition of oregano and thyme oils in comparison to previous reports.     

Antibacterial activity of carvacrol and 2-nitro-1-propanol against single and mixed populations of foodborne pathogenic bacteria in corn flour dough

Cereal doughs are an important part of human diet, but at the same time can act as vehicles for the transmission of human pathogenic bacteria. In the present study, four pathogenic or toxinogenic bacteria (Escherichia coli O157:H7, Salmonella enterica serovar Enteritidis, Bacillus cereus and Staphylococcus aureus) were inoculated in a dough made from corn flour in combination with the single antimicrobial compounds carvacrol and 2-Nitro-1-propanol (2NPOH). Survival of single and mixed populations in the treated doughs incubated at 37 °C was followed by culture-dependent and independent methods (TTGE). All strains were completely inactivated within 24 h by the tested compounds at 5% final concentration, but showed variable inhibition at lower concentrations of 0.5% and 2%. Sensitivity to antimicrobial compounds was in general modified when strains were tested in cocultures compared with single cultures. B. cereus was more sensitive to carvacrol (minimum bactericidal concentration, MBC, 0.5%) in coculture with S. aureus. It was also more sensitive to 2NPOH in cocultures with S. aureus and with S. enterica (MBC, 2% in both cases). S. aureus was more resistant to carvacrol (MBC, 5%) in cocultures with B. cereus, E. coli, as well as S. enterica. However, sensitivity to 2NPOH was not modified in any of the coculture experiments (MBC, 2%). E. coli was more resistant to carvacrol (MBC, 5%) in cocultures with S. aureus and with S. enterica. Resistance of E. coli to 2NPOH also increased in cocultures with B. cereus (MBC, 5%) and with S. aureus (MBC, 2%), but not with S. enterica (MBC, 0.5%). S. enterica was more resistant to carvacrol (MBC, 5%) in cocultures with E. coli, but it was more sensitive to 2NPOH in cocultures with B. cereus as well as with S. aureus (MBC, 2% in both cases). TTGE analysis of survivors from cocultures treated with 2NPOH or carvacrol allowed a good estimation of the identity of survivors according to their DNA band patterns. Results from this study indicate that the efficacy of antimicrobials such as carvacrol and 2NPOH is greatly influenced by the complexity of the microbial populations under target and the relationships between individual populations.     

Characterization and evaluation of antimicrobial activity of actinonin against foodborne pathogens

This study revealed the antimicrobial properties of actinonin against major foodborne pathogens, Escherichia coli O157:H7, Listeria monocytogenes, Salmonella Typhimurium, Staphylococcus aureus, and Vibrio vulnificus. Among them, actinonin caused growth defect in S. Typhimurium and V. vulnificus. Minimal inhibitory concentration (MIC) values of actinonin were determined by broth microdilution methods. The MICs of actinonin were ≤0.768 μg/ml for S. Typhimurium and ≤0.192 μg/ml for V. vulnificus. Susceptibility to actinonin in both pathogens was measured by colony-forming ability and disc diffusion test. The results showed actinonin had antimicrobial activity against S. Typhimurium and V. vulnificus in a dose-dependent manner. The inhibitory effects on swarming motility were determined, and cytotoxicity of each pathogen against HeLa cells was decreased significantly by actinonin treatment. Furthermore, actinonin showed an antimicrobial efficacy in food models infected with these pathogens. These results demonstrate that actinonin is potentially an effective agent for food sanitization or preservation.     

Antibacterial activity of eicosapentaenoic acid (EPA) against foodborne and food spoilage microorganisms

Eicosapentaenoic acid (EPA), a long-chain polyunsaturated fatty acid of ω-3 type was evaluated for its antimicrobial action against the range of foodborne and food spoilage pathogens, using agar disc diffusion assay in Luria broth (LB) media. The EPA exhibited antimicrobial activity against Bacillus subtilis ATCC 6633, Listeria monocytogenes ATCC 19166, Staphylococcus aureus ATCC 6538, S. aureus KCTC 1916 and Pseudomonas aeruginosa KCTC 2004. The minimum inhibitory concentrations (MICs) of the EPA against the tested bacterial strains were found in the range of 500-1350 μg/ml using broth dilution method. EPA reduced the viability of S. aureus at 250, 125 and 62.5 μg/ml after 15 min exposure and a steep decline in colony forming units (CFUs) was observed at 125 μg/ml after 30 min exposure, while similar reduction in CFU rate was exhibited by EPA when treated with 62.5 μg/ml after 180 min. EPA also reduced the CFU numbers of P. aeruginosa at all the concentrations used in this study after 15 min exposure. On the other hand, scanning electron microscopy (SEM) study of bacterial cells clearly exhibited the antibacterial effect of EPA as evidenced by the damages found in the outer membrane of the cells when treated with EPA. The results demonstrated that EPA exerted significant bactericidal and bacteriostatic effects against both P. aeruginosa and S. aureus.     

Antimicrobial activity of nisin against Oenococcus oeni and other wine bacteria

Nisin is a bacteriocin used against food spoilage bacteria. Sulphur dioxide is a potent antioxidant as well as an antimicrobial agent widely used in the wine industry. In this study we describe the effect of these important antibacterial agents on the growth of a collection of 64 lactic acid bacteria (23 Oenococcus, 29 Lactobacillus, 3 Leuconostoc and 9 Pediococcus), 23 acetic acid bacteria and 20 yeast isolates, most of them recovered from wine. Minimal inhibitory concentrations (MIC) and minimal bactericide concentrations of nisin, potassium metabisulphite and ethanol were determined. Nisin MIC(50) values for the tested isolates were as follows: 0.024, 12.5, 200 and > or micro for oenococci, lactobacilli-pediococci-leuconostoc, acetic acid bacteria and yeasts, respectively. Synergistic effects on bacterial growth inhibition were observed, and potassium metabisulphite MIC(50) values decreased from one to three orders of dilution when it was combined with subinhibitory concentrations of nisin in the growth media. This effect was observed in all lactic acid bacteria species of our study. Significant differences in nisin sensitivity were observed between Gram-positive and Gram-negative bacteria, and between Oenococcus oeni and other species of lactic acid bacteria. It is concluded that appropriate combinations of nisin and metabisulphite could control the growth of spoilage bacteria in wine and therefore allow a decrease in the levels of sulphur dioxide currently used by the wine industry.     

Antibacterial activity of bioconverted eicosapentaenoic (EPA) and docosahexaenoic acid (DHA) against foodborne pathogenic bacteria

The antimicrobial activity of bioconversion extracts of EPA and DHA against a range of foodborne pathogenic bacteria was investigated. The bioconverted EPA and DHA exhibited antibacterial activities against four gram-positive bacteria, Bacillus subtilis, Listeria monocytogenes, Staphylococcus aureus (ATCC 6538) and S. aureus (KCTC 1916) and seven gram-negative bacteria, Enterobacter aerogenes, Escherichia coli, E. coli O157:H7, E. coli O157:H7 (human), Pseudomonas aeruginosa, Salmonella enteritidis and S. typhimurium. The growth inhibition by both bioconverted EPA and DHA was similar against gram-positive bacteria, while the bioconverted extract of DHA was more effective than EPA against gram-negative bacteria as determined by minimum inhibitory concentration.     

Effectiveness of trisodium phosphate treatment against pathogenic and spoilage bacteria on poultry during refrigerated storage

To determine the efficacy of trisodium phosphate (TSP) against pathogenic and spoilage bacteria on poultry and to assess the influence of the bacterial combination of inoculum on TSP effect, chicken legs were coinoculated with similar concentrations of a pathogenic (Salmonella enterica ser. Enteritidis or Listeria monocytogenes) and a spoilage (Pseudomonas fluorescens or Brochothrix thermosphacta) bacteria. Samples were dipped in TSP (12%, 15 min) or were not treated (control). Microbiological analysis and pH determinations were carried out at 0, 1, 3, and 5 days of storage (3 degrees C). Significant bacterial reductions (marked in gram-negative species) were observed on TSP-treated samples throughout refrigerated storage. Inoculum composition scarcely influenced the TSP effect against gram-positive bacteria. However, greater reductions were observed on gram-negative bacteria (Salmonella Enteritidis and P. fluorescens) when samples were coinoculated with B. thermosphacta and L. monocytogenes, respectively. Values of pH were higher in TSP-treated than in control samples at all sampling times. The combination of bacteria in inocula did not have a significant influence on pH values.