Inhibition of Listeria monocytogenes by using bacteriocin PA-1 produced by Pediococcus acidilactici PAC 1.0.

The bacteriocin produced by Pediococcus acidilactici PAC 1.0, previously designated PA-1 bacteriocin, was found to be inhibitory and bactericidal for Listeria monocytogenes. A dried powder prepared from PAC 1.0 culture supernatant fortified with 10% milk powder was found to contain bacteriocin activity. An MIC against L. monocytogenes and lytic effects in broth cultures were determined. Inhibition by PA-1 powder occurred over the pH range 5.5 to 7.0 and at both 4 and 32 degrees C. In addition, inhibition of L. monocytogenes was demonstrated in several food systems including dressed cottage cheese, half-and-half cream, and cheese sauce.     

Inhibition of Listeria monocytogenes by using bacteriocin PA-1 produced by Pediococcus acidilactici PAC 1.0

The bacteriocin produced by Pediococcus acidilactici PAC 1.0, previously designated PA-1 bacteriocin, was found to be inhibitory and bactericidal for Listeria monocytogenes. A dried powder prepared from PAC 1.0 culture supernatant fortified with 10% milk powder was found to contain bacteriocin activity. An MIC against L. monocytogenes and lytic effects in broth cultures were determined. Inhibition by PA-1 powder occurred over the pH range 5.5 to 7.0 and at both 4 and 32 degrees C. In addition, inhibition of L. monocytogenes was demonstrated in several food systems including dressed cottage cheese, half-and-half cream, and cheese sauce.     

Collapse of the proton motive force in Listeria monocytogenes caused by a bacteriocin produced by Pediococcus acidilactici.

The effect of pediocin JD, a bacteriocin produced by Pediococcus acidilactici JD1-23, on the proton motive force and proton permeability of resting whole cells of Listeria monocytogenes Scott A was determined. Control cells, treated with trypsin-inactivated bacteriocin at a pH of 5.3 to 6.1, maintained a pH gradient and a membrane potential of approximately 0.65 pH unit and 75 mV, respectively. However, these gradients were rapidly dissipated in cells after exposure to pediocin JD, even though no cell lysis had occurred. The pH gradient and membrane potential of the producer cells were also unaffected by the bacteriocin. Whole cells treated with bacteriocin were twice as permeable to protons as control cells were. The results suggest that the inhibitory action of pediocin JD against L. monocytogenes is directed at the cytoplasmic membrane and that inhibition of L. monocytogenes may be caused by the collapse of one or both of the individual components of the proton motive force.     

Collapse of the proton motive force in Listeria monocytogenes caused by a bacteriocin produced by Pediococcus acidilactici.

The effect of pediocin JD, a bacteriocin produced by Pediococcus acidilactici JD1-23, on the proton motive force and proton permeability of resting whole cells of Listeria monocytogenes Scott A was determined. Control cells, treated with trypsin-inactivated bacteriocin at a pH of 5.3 to 6.1, maintained a pH gradient and a membrane potential of approximately 0.65 pH unit and 75 mV, respectively. However, these gradients were rapidly dissipated in cells after exposure to pediocin JD, even though no cell lysis had occurred. The pH gradient and membrane potential of the producer cells were also unaffected by the bacteriocin. Whole cells treated with bacteriocin were twice as permeable to protons as control cells were. The results suggest that the inhibitory action of pediocin JD against L. monocytogenes is directed at the cytoplasmic membrane and that inhibition of L. monocytogenes may be caused by the collapse of one or both of the individual components of the proton motive force.     

Morphological changes induced in listeria monocytogenes V7 by a bacteriocin produced by Pediococcus acidilactici

Pediococcus acidilactici produces bacteriocin, which kills Listeria monocytogenes. The bactericidal mode of action of the bacteriocin against L. monocytogenes V7 was investigated by transmission electron microscopy. The bacteriocin was purified partially from the cell-free extract using Micro-Cel and cation-exchange chromatography, and the specific activity was increased 1,791 fold. The bacteriocin (6,400 AU/ ml) was inoculated with L. monocytogenes V7 and incubated for 0.5 h, 1 h, 3 h, and 6 h. The bacteriocin was found to destroy most of the cell wall and released most of the inclusions in the cells after 6 h of incubation. These results suggest that the bactericidal effect of the bacteriocin was due to bacterial lysis.     

Purification and amino acid sequence of a bacteriocin produced by Pediococcus acidilactici.

A bacteriocin produced by Pediococcus acidilactici has been purified to homogeneity by a rapid and simple four-step purification procedure which includes ammonium sulphate precipitation, chromatography with a cation-exchanger and Octyl Sepharose, and reverse-phase chromatography. The purification resulted in an approximately 80,000-fold increase in the specific activity and about a 6-fold increase in the total activity. The amino acid composition and sequencing data indicated that the bacteriocin contained 43-44 amino acid residues. The predicted M(r) and isolectric point of the bacteriocin are about 4600 and 8.6, respectively. Comparing the amino acid sequence of this bacteriocin with the sequences of leucocin A-UAL 187, sakacin P and curvacin A (bacteriocins produced by Leuconostoc gelidum, Lactobacillus sake and Lactobacillus curvatus, respectively) revealed that all four bacteriocins had in their N-terminal region the sequence Tyr-Gly-Asn-Gly-Val-Xaa-Cys, indicating that this concensus sequence is of fundamental importance for this group of bacteriocins. The bacteriocin from P. acidilactici and sakacin P were very similar, having at least 25 common amino acid residues. The sequence similarity was greatest in the N-terminal half of the molecules--17 of the first 19 residues were common--indicating the fundamental importance of this region. Leucocin A-UAL 187 and curvacin A had, respectively, at least 16 and 13 amino acid residues in common with the bacteriocin from P. acidilactici.     

Purification, characterization and antimicrobial spectrum of a bacteriocin produced by Pediococcus acidilactici

An antimicrobial peptide designated pediocin AcH was isolated from Pediococcus acidilactici strain H. The pediocin AcH was purified by ion exchange chromatography. The molecular weight of pediocin AcH was determined by SDS-PAGE to be about 2700 daltons. Pediocin AcH was sensitive to proteolytic enzymes resistant to heat and organic solvents, and active over a wide range of pH. Pediocin AcH exhibited inhibition against several food spoilage bacteria and foodborne pathogens including Staphylococcus aureus, Clostridium perfringens and Listeria monocytogenes. It was bactericidal to sensitive cells and acted very rapidly. The bactericidal effect was not produced by either cell lysis or apparent loss of membrane permeability.     

Purification, characterization and antimicrobial spectrum of a bacteriocin produced by Pediococcus acidilactici

An antimicrobial peptide designated pediocin AcH was isolated from Pediococcus acidilactici strain H. The pediocin AcH was purified by ion exchange chromatography. The molecular weight of pediocin AcH was determined by SDS-PAGE to be about 2700 daltons. Pediocin AcH was sensitive to proteolytic enzymes, resistant to heat and organic solvents, and active over a wide range of pH. Pediocin AcH exhibited inhibition against several food spoilage bacteria and foodborne pathogens including Staphylococcus aureus, Clostridium perfringens and Listeria monocytogenes. It was bactericidal to sensitive cells and acted very rapidly. The bactericidal effect was not produced by either cell lysis or apparent loss of membrane permeability.     

Simple method of purification and sequencing of a bacteriocin produced by Pediococcus acidilactici UL5

H. DABA, C. LACROIX, J. HUANG, R.E. SIMARD AND L. LEMIEUX. 1994. A bacteriocin produced by a strain of Pediococcus acidilactici was successfully purified sequentially by acid extraction (at pH 2) and reverse-phase high-performance liquid chromatography (HPLC). Cell extracts of derivative strains deficient in bacteriocin production exhibited a similar HPLC elution profile to the active extracts except for the two peaks containing bacteriocin activity. The sequence of the antibacterial peptide consisted of 44 amino acid residues of which 42 were identified, and its molecular weight was 4624 Da, as determined by mass spectrometry. Moreover, according to the molecular weight of the peptide, the unidentified residues in the bacteriocin sequence must correspond to two tryptophan residues, confirming that the peptide isolated from Ped. acidilactici UL5 is pediocin PA-1. However, oxidized forms of the bacteriocin produced during storage also showed bacteriocin activity and resulted in a second peak with activity in the chromatograms. HPLC chromatograms of cell surface preparations from the active and from the deficient strains were confirmed by capillary electrophoresis. The purification method used is simple and effective in comparison with traditional methods, permitting a selective recovery of cell-associated bacteriocin at low pH, and its isolation in pure form for sequencing.     

Interaction of the bacteriocin produced by Pediococcus acidilactici SJ-1 with the cell envelope of Lactobacillus spp.

In spite of differences in producing strains and their plasmid profiles, amino acid sequence analysis indicates that the bacteriocin produced by Pediococcus acidilactici SJ-1 is identical to that produced by PAC 1.0 and H. Protoplasts prepared from cells of pediocin-resistant strains of Lactobacillus plantarum and Lact. fermentum were lysed by exposure to the pediocin. The interaction of the pediocin with sensitive Lact. plantarum cells did not alter the fluidity of the cell membrane.     

Purification, partial characterization and plasmid-linkage of pediocin SJ-1, a bacteriocin produced by Pediococcus acidilactici

Pediococcus acidilactici SJ-1, isolated from a naturally-fermented meat product, produced an antibacterial agent active against selected strains of Lactobacillus spp., Clostridium perfringens and Listeria monocytogenes. The agent was bactericidal against sensitive indicators, and sensitive to proteolytic enzymes; it was identified as a bacteriocin, and was designated as pediocin SJ-1. It was stable over a wide pH range (3–9), and apparently most stable in the lower part of that range. At pH 3.6, pediocin SJ-1 was stable at heat-processing temperatures within the range 65–121°C; its activity decreased significantly, however, when it was heated at pH 7.0. The activity of pediocin SJ-1 on sensitive indicator cells was lost in the presence of α-amylase, suggesting that it contains a glyco moiety, necessary for its antibacterial action.
Native pediocin SJ-1 exists in the form of monomers and aggregates (with molecular weights in the range 80–150 kDa). Pediocin SJ-1 was purified 262-fold by direct application of cell-free supernatant fluids to a cation-exchange chromatography column, and was resolved by SDS-PAGE as a single peptide band with a MW of ca 4 kDa. The original pediocin SJ-1-producing strain (bac⁺) harbours three plasmids of 4.6, 23.5, and 45.7 MDa. Production of pediocin SJ-1, but not immunity to SJ-1, is associated with the 4.6 MDa plasmid.     

Inhibition of Listeria monocytogenes by bavaricin A, a bacteriocin produced by Lactobacillus bavaricus MI401

Two hundred and forty-five strains of Listeria monocytogenes belonging to 33 different electrophoretic types, grouped by multilocus enzyme electrophoresis, were screened for sensitivity to bavaricin A produced by Lactobacillus bavaricus MI401. Three strains were resistant. Resistance was found to be neither restricted to particular electrophoretic types nor to certain sources of origin. The resistant strains belonged to three different electrophoretic types (ET 1, ET 5 and ET 14) and were isolated from a human case of listeriosis, smoked salmon and pig faeces, respectively.     

Antigenic property of pediocin AcH produced by Pediococcus acidilactici H

Pediocin AcH, a bacteriocin of Pediococcus acidilactici H, inhibits the growth of several food spoilage and pathogenic bacteria. The antigenic property of partially purified pediocin AcH was tested by immunizing mice and a rabbit. Pediocin AcH was not immunogenic in these animals as determined by immunoblotting even after conjugation to bovine serum albumin. The non-immunogenic nature of pediocin AcH, its non-toxicity to laboratory animals and its hydrolysis by gastric proteolytic enzymes may be considered favourably in its possible use as a food preservative.     

Antigenic property of pediocin AcH produced by Pediococcus acidilactici H

Pediocin AcH, a bacteriocin of Pediococcus acidilactici H, inhibits the growth of several food spoilage and pathogenic bacteria. The antigenic property of partially purified pediocin AcH was tested by immunizing mice and a rabbit. Pediocin AcH was not immunogenic in these animals as determined by immunoblotting even after conjugation to bovine serum albumin. The non-immunogenic nature of pediocin AcH, its non-toxicity to laboratory animals and its hydrolysis by gastric proteolytic enzymes may be considered favourably in its possible use as a food preservative.     

A modified method to directly detect in SDS-PAGE the bacteriocin of Pediococcus acidilactici

A modified direct detection system was developed to identify the antimicrobial peptide, pediocin AcH in SDS-PAGE. The gel containing pediocin AcH was stained for 30 min, destained for 1.5 h, and then overlaid with indicator bacteria. The pediocin AcH band was identified by a zone of inhibition of bacterial growth surrounding the stained protein band.     

Characterization of bacteriocin produced by Pediococcus pentosaceus from wine

A.M. STRASSER DE SAAD AND M.C. MANCA DE NADRA. 1993. Twenty strains of Pediococcus pentosaceus isolated from wine were examined for production of bacteriocins. Only two of them showed inhibitory activity, Ped. pentosaceus N4p against the indicator strains of the same species and N5p against 19 strains of the three genera of lactic acid bacteria from wine. The antimicrobial substance from N5p strains was removed by membrane (0.2 μm) filtration, destroyed by organic solvents and proteolytic enzymes. It was stable for 60 min at 100C. The bacteriocin was produced early in the growth cycle and its production was maximum after 48 h of culture in tomato juice medium at an initial pH of 6.5. The bactericidal effect was observed.     

Enterocin 226NWC, a bacteriocin produced by Enterococcus faecalis 226, active against Listeria monocytogenes

F. VILLANI, G. SALZANO, E. SORRENTINO, O. PEPE, P. MARINO AND S. COPPOLA. 1993. Enterococcus faecalis 226, isolated from natural whey cultures utilized as starters in the manufacture of mozzarella cheese from water-buffalo milk, produces a bacteriocin designated enterocin 226NWC. The bacteriocin was isolated from culture supernatant fluids of the producer strain and was active against strains of the same species and Listeria monocytogenes, but not against useful lactic acid bacteria. Enterocin 226NWC is a protein with an apparent molecular weight of about 5800; it is relatively heat-stable and has a bactericidal mode of action. Listeria monocytogenes, growing in the presence of the enterocin 226NWC producer strain in broth and in reconstituted skim milk, was inhibited.     

Effectiveness of cell-adsorbed bacteriocin produced by Lactobacillus curvatus CWBI-B28 and selected essential oils to control Listeria monocytogenes in pork meat during cold storage

Aims:  To study the effectiveness of a combination of cell-adsorbed bacteriocin (CAB; a suspension of producer cells on which maximum bacteriocin has been immobilized by pH adjustments) of a Lactobacillus curvatus strain with oregano or savory essential oil to control Listeria monocytogenes in pork meat at 4°C.
Methods and Results:  The antimicrobial activity of the CAB and six different essential oils was tested by the well diffusion assay against L. monocytogenes M, Escherichia coli 10536 and Salmonella serotype Typhi CWBI-H1. The anti- Listeria activity of the CAB and oregano or savory essential oils was also investigated in pork meat. The results of the well diffusion assay showed that CAB was only inhibitory to L. monocytogenes while savory and oregano essential oils were the most active against the three indicator bacteria. In pork meat, Listeria counts have declined from c. 10² CFU g⁻¹ to below the detectable limit during the first week of storage in samples treated with CAB or oregano essential oil and in those treated with CAB combined with oregano or savory essential oil. However, the counts of L. monocytogenes have increased after the third week of storage in all samples with the exception of those treated with the combination of CAB and oregano essential oil. The combination of CAB with savory essential oil resulted in a 2-week delay of the growth rebound compared with samples treated with CAB alone.
Conclusions:  Addition of oregano or savory essential oil exhibited a synergistic effect with CAB to control L. monocytogenes in pork meat during storage at 4°C.
Significance and Impact of the Study:  The combination of CAB with oregano or savory essential oil may be effectively used in meat industry to enhance the safety and stability of meat products.     

Factors influencing immunity and resistance of Pediococcus acidilactici to the bacteriocin, pediocin AcH

In pediocin AcH producing Pediococcus acidilactici strains the genes for both the production of pediocin and immunity against it are encoded in an 8.9 kb plasmid pSMB74. Following loss of this plasmid, the variants lost the ability to produce pediocin AcH, but some retained the resistance against it. This resistance was a transient trait, acquired while nonproducing cells grew in the presence of pediocin AcH but lost when the cells were grown in the absence of it.     

Isolation and characterization of pediocin L50, a new bacteriocin from Pediococcus acidilactici with a broad inhibitory spectrum.

Lactic acid bacteria were isolated from Spanish dry-fermented sausages and screened for bacteriocin production. About 10% of the isolates produced antimicrobial substances when grown on solid media, but only 2% produced detectable activity in liquid media. Strain L50, identified as Pediococcus acidilactici, showed the strongest inhibitory activity and was active against members of all of the gram-positive genera tested. The strain produced a heat-stable bacteriocin when grown at 8 to 32 degrees C but not at 45 degrees C. The bacteriocin was purified to homogeneity. Its mass was determined to be 5,250.11 +/- 0.30 by electrospray mass spectrometry. The N terminus of the bacteriocin was blocked for sequencing by Edman degradation, but a partial sequence of 42 amino acids was obtained after cleavage of the peptide by cyanogen bromide. The sequence showed no similarity to those of other bacteriocins. Pediocin L50 appears to contain modified amino acids but not lanthionine or methyl-lanthionine.