Potency Of Bacteriocin For Animal Health And Food Safety

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Siti Chotiah

2013-06-01

Full Text Available The emergence of antibiotic resistance in many bacteria related to animal and public health stresses the importance of decreasing the use of antibiotics in animal production. The reduction of antibiotic application in livestock can only be achieved if alternative antimicrobial strategies are available. A number of strategies have been explored to control microbial pathogens and to improve growth and feed efficiency in livestock without the use of antibiotics. Bacteriocins have been more extensively studied and proposed as potential alternatives to conventional antibiotics in animal husbandry. Bacteriocins are antimicrobial peptides ribosomally synthesized by many species of Bacteria and some strains of Archaea. In general, bacteriocins just exhibited bactericidal or bacteriostatic activity against other bacteria that are closely related to the producing strain. The main mechanisms of bacteriocin activity vary from pore formation in cytoplasmic membranes to the inhibition of cell wall biosynthesis and enzyme activities (RNAse or DNAse in target cells. The use of bacteriocins in probiotic applications, as preservatives, and most excitingly as alternatives to conventional antibiotics is being broadly explored and studied. This review will describe the bacteriocins potency for animal health and food safety, as well as the results of bacteriocin study that had been conducted in Indonesia.

Spontaneous bacteriocin resistance in Listeria monocytogenes as a susceptibility screen for identifying different mechanisms of resistance and modes of action by bacteriocins of lactic acid bacteria.

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Macwana, Sunita; Muriana, Peter M

2012-01-01

A practical system was devised for grouping bacteriocins of lactic acid bacteria (LAB) based on mode of action as determined by changes in inhibitory activity to spontaneously-acquired bacteriocin resistance (Bac(R)). Wild type Listeria monocytogenes 39-2 was sensitive to five bacteriocins produced by 3 genera of LAB: pediocin PA-1 and pediocin Bac3 (Pediococcus), lacticin FS97 and lacticin FS56 (Lactococcus), and curvaticin FS47 (Lactobacillus). A spontaneous Bac(R) derivative of L. monocytogenes 39-2 obtained by selective recovery against lacticin FS56 provided complete resistance to the bacteriocin made by Lactococcus lactis FS56. The lacticin FS56-resistant strain of L. monocyotgenes 39-2 was also cross-resistant to curvaticin FS47 and pediocin PA-1, but not to lacticin FS97 or pediocin Bac3. The same pattern of cross-resistance was also observed with Bac(R) isolates obtained with L. monocytogenes Scott A-2. A spontaneous mutation that renders a strain cross-resistant to different bacteriocins indicates that they share a common mechanism of resistance due to similar modes of action of the bacteriocins. Spontaneous resistance was acquired to other bacteriocins (in aggregate) by following the same procedure against which the Bac(R) strain was still sensitive. In subsequent challenge assays, mixtures of bacteriocins of different modes of action provided greater inhibition than mixtures of bacteriocins of the same mode of action (as determined by our screening method). This study identifies a methodical approach to classify bacteriocins into functional groups based on mechanism of resistance (i.e., mode of action) that could be used for identifying the best mixture of bacteriocins for use as biopreservatives. Copyright © 2011 Elsevier B.V. All rights reserved.

Biochemical and genetic characterization of enterocin A from Enterococcus faecium, a new antilisterial bacteriocin in the pediocin family of bacteriocins.

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Aymerich, T; Holo, H; Håvarstein, L S; Hugas, M; Garriga, M; Nes, I F

1996-01-01

A new bacteriocin has been isolated from an Enterococcus faecium strain. The bacteriocin, termed enterocin A, was purified to homogeneity as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, N-terminal amino acid sequencing, and mass spectrometry analysis. By combining the data obtained from amino acid and DNA sequencing, the primary structure of enterocin A was determined. It consists of 47 amino acid residues, and the molecular weight was calculated to be 4,829, assuming that the four cysteine residues form intramolecular disulfide bridges. This molecular weight was confirmed by mass spectrometry analysis. The amino acid sequence of enterocin A shared significant homology with a group of bacteriocins (now termed pediocin-like bacteriocins) isolated from a variety of lactic acid-producing bacteria, which include members of the genera Lactobacillus, Pediococcus, Leuconostoc, and Carnobacterium. Sequencing of the structural gene of enterocin A, which is located on the bacterial chromosome, revealed an N-terminal leader sequence of 18 amino acid residues, which was removed during the maturation process. The enterocin A leader belongs to the double-glycine leaders which are found among most other small nonlantibiotic bacteriocins, some lantibiotics, and colicin V. Downstream of the enterocin A gene was located a second open reading frame, encoding a putative protein of 103 amino acid residues. This gene may encode the immunity factor of enterocin A, and it shares 40% identity with a similar open reading frame in the operon of leucocin AUL 187, another pediocin-like bacteriocin. PMID:8633865

Antibacterial Activities of Bacteriocins: Application in Foods and Pharmaceuticals

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Shih-Chun eYang

2014-05-01

Full Text Available Bacteriocins are a kind of ribosomal synthesized antimicrobial peptides produced by bacteria, which can kill or inhibit bacterial strains closely-related or non-related to produced bacteria, but will not harm the bacteria themselves by specific immunity proteins. Bacteriocins become one of the weapons against microorganisms due to the specific characteristics of large diversity of structure and function, natural resource, and being stable to heat. Many recent studies have purified and identified bacteriocins for application in food technology, which aims to extend food preservation time, treat pathogen disease and cancer therapy, and maintain human health. Therefore, bacteriocins may become a potential drug candidate for replacing antibiotics in order to treat multiple drugs resistance pathogens in the future. This review article summarizes different types of bacteriocins from bacteria. The latter half of this review focuses on the potential applications applications in food science and pharmaceutical industry.

Genetic identification of the bacteriocins produced by Enterococcus faecium IT62 and evidence that bacteriocin 32 is identical to enterocin IT.

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Izquierdo, Esther; Cai, Yimin; Marchioni, Eric; Ennahar, Saïd

2009-05-01

Enterococcus faecium IT62, a strain isolated from ryegrass in Japan, produces three bacteriocins (enterocins L50A, L50B, and IT) that have been previously purified and the primary structures of which have been determined by amino acid sequencing (E. Izquierdo, A. Bednarczyk, C. Schaeffer, Y. Cai, E. Marchioni, A. Van Dorsselaer, and S. Ennahar, Antimicrob. Agents Chemother., 52:1917-1923, 2008). Genetic analysis showed that the bacteriocins of E. faecium IT62 are plasmid encoded, but with the structural genes specifying enterocin L50A and enterocin L50B being carried by a plasmid (pTAB1) that is separate from the one (pTIT1) carrying the structural gene of enterocin IT. Sequencing analysis of a 1,475-bp region from pTAB1 identified two consecutive open reading frames corresponding, with the exception of 2 bp, to the genes entL50A and entL50B, encoding EntL50A and EntL50B, respectively. Both bacteriocins are synthesized without N-terminal leader sequences. Genetic analysis of a sequenced 1,380-bp pTIT1 fragment showed that the genes entIT and entIM, encoding enterocin IT and its immunity protein, respectively, were both found in E. faecium VRE200 for bacteriocin 32. Enterocin IT, a 6,390-Da peptide made up of 54 amino acids, has been previously shown to be identical to the C-terminal part of bacteriocin 32, a 7,998-Da bacteriocin produced by E. faecium VRE200 whose structure was deduced from its structural gene (T. Inoue, H. Tomita, and Y. Ike, Antimicrob. Agents Chemother., 50:1202-1212, 2006). By combining the biochemical and genetic data on enterocin IT, it was concluded that bacteriocin 32 is in fact identical to enterocin IT, both being encoded by the same plasmid-borne gene, and that the N-terminal leader peptide for this bacteriocin is 35 amino acids long and not 19 amino acids long as previously reported.

Bacteriocins of Non-aureus Staphylococci Isolated from Bovine Milk

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Carson, Domonique A.; Barkema, Herman W.; Naushad, Sohail

2017-01-01

ABSTRACT Non-aureus staphylococci (NAS), the bacteria most commonly isolated from the bovine udder, potentially protect the udder against infection by major mastitis pathogens due to bacteriocin production. In this study, we determined the inhibitory capability of 441 bovine NAS isolates (comprising 26 species) against bovine Staphylococcus aureus. Furthermore, inhibiting isolates were tested against a human methicillin-resistant S. aureus (MRSA) isolate using a cross-streaking method. We determined the presence of bacteriocin clusters in NAS whole genomes using genome mining tools, BLAST, and comparison of genomes of closely related inhibiting and noninhibiting isolates and determined the genetic organization of any identified bacteriocin biosynthetic gene clusters. Forty isolates from 9 species (S. capitis, S. chromogenes, S. epidermidis, S. pasteuri, S. saprophyticus, S. sciuri, S. simulans, S. warneri, and S. xylosus) inhibited growth of S. aureus in vitro, 23 isolates of which, from S. capitis, S. chromogenes, S. epidermidis, S. pasteuri, S. simulans, and S. xylosus, also inhibited MRSA. One hundred five putative bacteriocin gene clusters encompassing 6 different classes (lanthipeptides, sactipeptides, lasso peptides, class IIa, class IIc, and class IId) in 95 whole genomes from 16 species were identified. A total of 25 novel bacteriocin precursors were described. In conclusion, NAS from bovine mammary glands are a source of potential bacteriocins, with >21% being possible producers, representing potential for future characterization and prospective clinical applications. IMPORTANCE Mastitis (particularly infections caused by Staphylococcus aureus) costs Canadian dairy producers $400 million/year and is the leading cause of antibiotic use on dairy farms. With increasing antibiotic resistance and regulations regarding use, there is impetus to explore bacteriocins (bacterially produced antimicrobial peptides) for treatment and prevention of bacterial infections

Bacteriocin-mediated competition in cystic fibrosis lung infections

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Ghoul, Melanie; West, Stuart A.; Johansen, Helle Krogh

2015-01-01

Bacteriocins are toxins produced by bacteria to kill competitors of the same species. Theory and laboratory experiments suggest that bacteriocin production and immunity play a key role in the competitive dynamics of bacterial strains. The extent to which this is the case in natural populations......, especially human pathogens, remains to be tested. We examined the role of bacteriocins in competition using Pseudomonas aeruginosa strains infecting lungs of humans with cystic fibrosis (CF). We assessed the ability of different strains to kill each other using phenotypic assays, and sequenced their genomes...

Bacteriocin-producing Enterococci from Rabbit Meat

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Szabóová, R.

2012-01-01

Full Text Available Aims: Enterococci are lactic acid bacteria belonging to the division Firmicutes. They occur in different ecosystems, rabbits including. Enterococci can possess probiotic properties and produce antimicrobial substances-bacteriocins. Rabbit meat as nutritionally healthy food offers novel source to study bacteriocin-producing and/or probiotic enterococci. Methodology and results: Enterococci were detected from rabbit meat samples (42. Most of the isolates were allotted to the species Enterococcus faecium by PCR method. The isolates have possessed the structural genes for enterocins A, P, B production. The inhibitory substances produced by the isolated enterococci inhibited the growth of 12 indicators. Of 34 isolates, 15 strains have shown the antimicrobial activity against L. monocytogenes CCM 4699, 12 strains against S. aureus 3A3, 10 strains against S. aureus 5A2 as well as Salmonella enterica serovar Enteritidis PT4. Moreover, enterococci have tolerated 5 % bile, low pH; they have produced lactid acid in the amount from 0.740 ± 0.091 to 1.720 ± 0.095 mmol/l. The isolates were mostly sensitive to antibiotics. Conclusion, significance and impact of study: Bacteriocin-producing strain E. faecium M3a has been selected for more detail characterization of its bacteriocin and probiotic properties with the aim for its further application as an additive.

Partial Purification Characterization and Application of Bacteriocin from Bacteria Isolated Parkia biglobosa Seeds

OpenAIRE

Olorunjuwon, O. Bello; Olubukola, O. Babalola; Mobolaji, Adegboye; Muibat, O. Fashola; Temitope, K. Bello

2018-01-01

Bacteriocins are proteinaceous toxins produced by bacteria to inhibit the growth of similar or closely related bacterial strains. Fermented Parkia biglobosa seeds (African locust bean) were screened for bacteriocin-producing lactic acid bacteria (LAB) with the characterization of putative bacteriocins. Bacteriocin-producing lactic acid bacteria (LAB) were identified by 16s rDNA sequencing. Molecular sizes of the bacteriocins were determined using the tricine-sodium dodecyl sulphate-polyacryla...

Current state of purification, isolation and analysis of bacteriocins produced by lactic acid bacteria.

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Kaškonienė, Vilma; Stankevičius, Mantas; Bimbiraitė-Survilienė, Kristina; Naujokaitytė, Gintarė; Šernienė, Loreta; Mulkytė, Kristina; Malakauskas, Mindaugas; Maruška, Audrius

2017-02-01

The scientific interest for the search of natural means of microbial inhibitors has not faded for several years. A search of natural antibiotics, so-called bacteriocins which are produced by lactic acid bacteria (LAB), gains a huge attention of the scientists in the last century, in order to reduce the usage of synthetic food additives. Pure bacteriocins with wide spectra of antibacterial activity are promising among the natural biopreservatives. The usage of bacteriocin(s) producing LAB as starter culture for the fermentation of some food products, in order to increase their shelf-life, when synthetic preservatives are not allowable, is also possible. There are a lot of studies focusing on the isolation of new bacteriocins from traditional fermented food, dairy products and other foods or sometimes even from unusual non-food matrices. Bacteriocins producing bacteria have been isolated from different sources with the different antibacterial activity against food-borne microorganisms. This review covers the classification of bacteriocins, diversity of sources of bacteriocin(s) producing LAB, antibacterial spectra of isolated bacteriocins and analytical methods for the bacteriocin purification and analysis within the last 15 years.

Bacteriocins of Non-aureus Staphylococci Isolated from Bovine Milk.

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Carson, Domonique A; Barkema, Herman W; Naushad, Sohail; De Buck, Jeroen

2017-09-01

Non- aureus staphylococci (NAS), the bacteria most commonly isolated from the bovine udder, potentially protect the udder against infection by major mastitis pathogens due to bacteriocin production. In this study, we determined the inhibitory capability of 441 bovine NAS isolates (comprising 26 species) against bovine Staphylococcus aureus Furthermore, inhibiting isolates were tested against a human methicillin-resistant S. aureus (MRSA) isolate using a cross-streaking method. We determined the presence of bacteriocin clusters in NAS whole genomes using genome mining tools, BLAST, and comparison of genomes of closely related inhibiting and noninhibiting isolates and determined the genetic organization of any identified bacteriocin biosynthetic gene clusters. Forty isolates from 9 species ( S. capitis , S. chromogenes , S. epidermidis , S. pasteuri , S. saprophyticus , S. sciuri , S. simulans , S. warneri , and S. xylosus ) inhibited growth of S. aureus in vitro , 23 isolates of which, from S. capitis , S. chromogenes , S. epidermidis , S. pasteuri , S. simulans , and S. xylosus , also inhibited MRSA. One hundred five putative bacteriocin gene clusters encompassing 6 different classes (lanthipeptides, sactipeptides, lasso peptides, class IIa, class IIc, and class IId) in 95 whole genomes from 16 species were identified. A total of 25 novel bacteriocin precursors were described. In conclusion, NAS from bovine mammary glands are a source of potential bacteriocins, with >21% being possible producers, representing potential for future characterization and prospective clinical applications. IMPORTANCE Mastitis (particularly infections caused by Staphylococcus aureus ) costs Canadian dairy producers $400 million/year and is the leading cause of antibiotic use on dairy farms. With increasing antibiotic resistance and regulations regarding use, there is impetus to explore bacteriocins (bacterially produced antimicrobial peptides) for treatment and prevention of bacterial

Enterocin TW21, a novel bacteriocin from dochi-isolated Enterococcus faecium D081821.

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Chang, S-Y; Chen, Y-S; Pan, S-F; Lee, Y-S; Chang, C-H; Chang, C-H; Yu, B; Wu, H-C

2013-09-01

Purification and characterization of a novel bacteriocin produced by strain Enterococcus faecium D081821. Enterococcus faecium D081821, isolated from the traditional Taiwanese fermented food dochi (fermented black beans), was previously found to produce a bacteriocin against Listeria monocytogenes and some Gram-positive bacteria. This bacteriocin, termed enterocin TW21, was purified from culture supernatant by ammonium sulfate precipitation, Sep-Pak C18 cartridge, ion-exchange and gel filtration chromatography. Mass spectrometry analysis showed the mass of the peptide to be approximately 5300·6 Da. The N-terminal amino acid sequencing yielded a partial sequence NH2 -ATYYGNGVYxNTQK by Edman degradation, and it contains the consensus class IIa bacteriocin motif YGNGV in the N-terminal region. The open reading frame (ORF) encoding the bacteriocin was identified from the draft genome sequence of Enterococcus faecium D081821, and sequence analysis of this peptide indicated that enterocin TW21 is a novel bacteriocin. Enterococcus faecium D081821 produced a bacteriocin named enterocin TW21, the molecular weight and amino acid sequence both revealed it to be a novel bacteriocin. A new member of class IIa bacteriocin was identified. This bacteriocin shows great inhibitory ability against L. monocytogenes and could be applied as a natural food preservative. © 2013 The Society for Applied Microbiology.

Purification of bacteriocins produced by lactic acid bacteria.

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Saavedra, Lucila; Castellano, Patricia; Sesma, Fernando

2004-01-01

Bacteriocins are antibacterial substances of a proteinaceous nature that are produced by different bacterial species. Lactic acid bacteria (LAB) produce biologically active peptides or protein complexes that display a bactericidal mode of action almost exclusively toward Gram-positive bacteria and particularly toward closely related species. Generally they are active against food spoilage and foodborne pathogenic microorganisms including Bacillus cereus, Clostridium perfringens, Staphylococcus aureus, and Listeria monocytogenes. There is an increased tendency to use natural occurring metabolites to prevent the growth of undesirable flora in foodstuffs. These metabolites could replace the use of chemical additives such as sorbic acid, sulfur dioxide, nitrite, nitrate, and others. For instance, bacteriocins produced by LAB may be promising for use as bio-preservaties. Bacteriocins of lactic acid bacteria are typically cationic, hydrophobic peptides and differ widely in many characteristics including molecular weight, presence of particular groups of amino acids, pI, net positive charge, and post-translational modifications of certain amino acids. This heterogeneity within the LAB bacteriocins may explain the different procedures for isolation and purification developed so far. The methods most frequently used for isolation, concentration, and purification involve salt precipitation of bacteriocins from culture supernatants, followed by various combinations of gel filtration, ion-exchange chromatography, and reverse-phase high-performance liquid chromatography (RP-HPLC). In this chapter, a protocol is described that combines several methods used in our laboratory for the purification of two cationic bacteriocins, Lactocin 705AL and Enterocin CRL10, produced by Lactobacillus casei CRL705 and Enterococcus mundtii CRL10, respectively.

Antagonistic activity expressed by Shigella sonnei: identification of a putative new bacteriocin

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Mireille Angela Bernardes Sousa

2013-09-01

Full Text Available Bacteriocins are antibacterial, proteinaceous substances that mediate microbial dynamics. Bacteriocin production is a highly disseminated property among all major lineages of bacteria, including Shigella. In this paper, we addressed the purification and characterisation of a bacteriocin produced by a Shigella sonnei strain (SS9 isolated from a child with acute diarrhoea. The substance was purified through ammonium-sulphate precipitation and sequential steps of chromatography. The intracellular fraction obtained at 75% ammonium sulphate maintained activity following exposure to pH values from 1-11 and storage at -80ºC for more than two years and was inactivated by high temperatures and proteases. The molecular mass of the purified bacteriocin was determined by mass spectrometry to be 18.56 kDa. The N-terminal sequence of the bacteriocin did not match any other antibacterial proteins described. A putative new bacteriocin produced by S. sonnei has been detected. This bacteriocin may represent a newly described protein or a previously described protein with a newly detected function. Considering that SS9 expresses antagonism against other diarrhoeagenic bacteria, the bacteriocin may contribute to S. sonnei virulence and is potentially applicable to either preventing or controlling diarrhoeal disease.

Production and Characterization of Bacteriocin Produced by Lactobacillus Viridescence(NICM 2167

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Sure KP

Full Text Available The present study focused on the production optimization of bacteriocin by Lactobacillus viridescence NICM 2167 followed by its purification and characterization. The bacteriocins are antimicrobial peptides produced by many Gram positive and Gram negative bacteria.The bacteriocin produced by LAB (lactic acid bacteria received attention in recent years due to their potential application as natural preservatives in food. Bacteriocinproduced by Lactobacillus viridescence showed broad range of antimicrobial activity against food borne pathogens. Production parameters were optimized showing highest production of bacteriocinin MRS broth with pH= 7.0 incubated at 37°C for 48 h. Bacteriocin was purified in two steps involving ammonium sulphate precipitation followed by gel filtration using Sephadex G-100. Purified bacteriocin with single band on SDS-PAGE showed molecular weight of 8.3 kDa. This purified bacteriocin was stable over wide range of pH (4-10 as well as temperatures (4°C-121°C suggesting it as a potent candidate for preservation of various foods.

Novel bacteriocins from lactic acid bacteria (LAB): various structures and applications.

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Perez, Rodney H; Zendo, Takeshi; Sonomoto, Kenji

2014-08-29

Bacteriocins are heat-stable ribosomally synthesized antimicrobial peptides produced by various bacteria, including food-grade lactic acid bacteria (LAB). These antimicrobial peptides have huge potential as both food preservatives, and as next-generation antibiotics targeting the multiple-drug resistant pathogens. The increasing number of reports of new bacteriocins with unique properties indicates that there is still a lot to learn about this family of peptide antibiotics. In this review, we highlight our system of fast tracking the discovery of novel bacteriocins, belonging to different classes, and isolated from various sources. This system employs molecular mass analysis of supernatant from the candidate strain, coupled with a statistical analysis of their antimicrobial spectra that can even discriminate novel variants of known bacteriocins. This review also discusses current updates regarding the structural characterization, mode of antimicrobial action, and biosynthetic mechanisms of various novel bacteriocins. Future perspectives and potential applications of these novel bacteriocins are also discussed.

Partial Purification, Characterization and Application of Bacteriocin from Bacteria Isolated Parkia biglobosa Seeds

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Olorunjuwon O. Bello

2018-05-01

Full Text Available Bacteriocins are proteinaceous toxins produced by bacteria to inhibit the growth of similar or closely related bacterial strains. Fermented Parkia biglobosa seeds (African locust bean were screened for bacteriocin-producing lactic acid bacteria (LAB with the characterization of putative bacteriocins. Bacteriocin-producing lactic acid bacteria (LAB were identified by 16s rDNA sequencing. Molecular sizes of the bacteriocins were determined using the tricine-sodium dodecyl sulphate-polyacrylamide gel electrophoresis (tricine-SDS–PAGE and effects of enzymes, pH, detergents and temperature on bacteriocin activity investigated, using standard procedures. Bacteriocins production and activities were measured by spectrophotometric analysis. Statistical analysis was carried out using student t-test and Analyses of Variance. Bacteriocigenic LAB isolated were Lactobacillus plantarum Z1116, Enterococcus faecium AU02 and Leuconostoc lactis PKT0003. They inhibited the growth of both Gram-positive and Gram-negative bacteria. The sizes of bacteriocins Z1116, AU02 and PKT0003 were 3.2 kDa, 10 kDa and 10 kDa, respectively. The synergistic effects of characterized bacteriocins and rifampicin tested on organisms showed significant differences (P < 0.05, as compared with the effects of only one of the two. The antimicrobial activity of the three bacteriocins was deactivated after treatment of the cell-free supernatants with proteinase K, papain, pepsin and trypsin. Parkia biglobosa seeds are, therefore, rich in LAB bacteriocins which could be explored. The biosynthetic mechanisms of LAB bacteriocins could be employed in food safety and security, preservation, peptide design, infection control and pharmacotherapy. This should help in the control of undesirable bacteria and in designing more potent and selective antimicrobial peptides.

Enterocin T, a novel class IIa bacteriocin produced by Enterococcus sp. 812.

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Chen, Yi-Sheng; Yu, Chi-Rong; Ji, Si-Hua; Liou, Min-Shiuan; Leong, Kun-Hon; Pan, Shwu-Fen; Wu, Hui-Chung; Lin, Yu-Hsuan; Yu, Bi; Yanagida, Fujitoshi

2013-09-01

Enterococcus sp. 812, isolated from fresh broccoli, was previously found to produce a bacteriocin active against a number of Gram-positive bacteria, including Listeria monocytogenes. Bacteriocin activity decreased slightly after autoclaving (121 °C for 15 min), but was inactivated by protease K. Mass spectrometry analysis revealed the bacteriocin mass to be approximately 4,521.34 Da. N-terminal amino acid sequencing yielded a partial sequence, NH2-ATYYGNGVYXDKKKXWVEWGQA, by Edman degradation, which contained the consensus class IIa bacteriocin motif YGNGV in the N-terminal region. The obtained partial sequence showed high homology with some enterococcal bacteriocins; however, no identical peptide or protein was found. This peptide was therefore considered to be a novel bacteriocin produced by Enterococcus sp. 812 and was termed enterocin T.

Subspecies diversity in bacteriocin production by intestinal Lactobacillus salivarius strains.

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O' Shea, Eileen F; O' Connor, Paula M; Raftis, Emma J; O' Toole, Paul W; Stanton, Catherine; Cotter, Paul D; Ross, R Paul; Hill, Colin

2012-01-01

A recent comparative genomic hybridization study in our laboratory revealed considerable plasticity within the bacteriocin locus of gastrointestinal strains of Lactobacillus salivarius. Most notably, these analyses led to the identification of two novel unmodified bacteriocins, salivaricin L and salivaricin T, produced by the neonatal isolate L. salivarius DPC6488 with immunity, regulatory and export systems analogous to those of abp118, a two-component bacteriocin produced by the well characterized reference strain L. salivarius UCC118. In this addendum we discuss the intraspecific diversity of our seven bacteriocin-producing L. salivarius isolates on a genome-wide level, and more specifically, with respect to their salivaricin loci.

LAB bacteriocin applications in the last decade

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Ma. del Rocío López-Cuellar

2016-11-01

Full Text Available In the early 2000s, the expectations about bacteriocins produced by lactic acid bacteria (LABs were aimed at food applications. However, the effectiveness of bacteriocins against undesirable micro-organisms opened endless possibilities for innovative research. In the present review, we collected a database including 429 published papers and 245 granted patents (from 2004 to 2015. Based on bibliometric analysis, the progress of bacteriocin research in the last 11 years was discussed in detail. It was found that 164 patents were granted in 2010–2015, which is equivalent to 60% in comparison with previous years (i.e. only 81 patents were granted in 2004–2009. Currently, the research on bacteriocins is still gaining importance. In the realm of therapeutic strategies, about a 37% of the published research was focused on biomedical applications in the last decade. This vein of research is currently seeking for alternative solutions to problems such as cancer, systemic infections, oral-care, vaginal infections, contraception and skincare. On the other hand, food preservation, bio-nanomaterial and veterinary applications represent 29%, 25% and 9%, respectively. All this technology is being applied and will surely grow in the future, since about 31% of the patents granted since 2004 are focused on the biomedical area, 29% on food preservation, 5% on veterinary use; whereas 13% and 16% correspond to patents granted on production–purification systems and recombinant proteins or molecular modifications in the producer strains. This review contributes to the analysis of recent LAB bacteriocin applications and their role in safety, quality and improvement of human health.

Purification and partial characterization of bacteriocin produced by Lactococcus lactis ssp. lactis LL171.

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Kumari, Archana; Akkoç, Nefise; Akçelik, Mustafa

2012-04-01

Lactic acid bacteria (LAB) are possessing ability to synthesize antimicrobial compounds (like bacteriocin) during their growth. In this regard, novel bacteriocin compound secreting capability of LAB isolated from Tulum Cheese in Turkey was demonstrated. The synthesized bacteriocin was purified by ammonium sulphate precipitation, dialysis and gel filtration. The molecular weight (≈3.4 kDa) of obtained bacteriocin was confirmed by SDS-PAGE, which revealed single peptide band. Molecular identification of LAB strain isolated from Tulum Cheese was conducted using 16S rDNA gene sequencing as Lactococcus lactis ssp. lactis LL171. The amino acid sequences (KKIDTRTGKTMEKTEKKIELSLKNMKTAT) of the bacteriocin from Lactococcus lactis ssp. lactis LL171 was found unique and novel than reported bacteriocins. Further, the bacteriocin was possessed the thermostable property and active at wide range of pH values from 1 to 11. Thus, bacteriocin reported in this study has the potential applications property as food preservative agent.

Bacteriocins produced by lactic acid bacteria: A review

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Vesković-Moračanin Slavica M.

2014-01-01

Full Text Available Lactic acid bacteria (LAB have an essential role in the production of fermented products. With their metabolic activity, they influence the ripening processes - leading to desired sensory qualities while at the same time inhibiting the growth of undesired microorganisms. Because of their dominant role during fermentation and because of a long tradition of utilization, Lhave been designated as “safe microbiota”. Biological protection of LAB, as a naturally present and/or selected and intentionally added microflora, is realized through the production of non-specific (lactic acid, acetic acid and other volatile organic acids, hydrogen peroxide, diacetyl, etc and specific metabolites, bacteriocins. Bacteriocins are extracellularly released proteins or peptides which possess certain antibacterial activity towards certain types of microorganisms, usually related to the producing bacteria. Today, bacteriocins represent a very interesting potential for their application in the food industry. Their application can reduce the use of synthetic preservatives and/or the intensity of thermal treatment during food production consumer’s need for safe, fresh and minimally-processed food. With the intention of realizing this potential to the fullest, it is necessary to understand the nature of bacteriocins, their production mechanisms, regulations and actions, as well as the influence of external factors on the their antimicrobial activity. The composition of food, i.e. its characteristics (pH, temperature, ingredients and additives, types and quantities of epiphytic microbiota and the actual technological process used in production, can all influence the stability and activity of the added bacteriocins. The future research in this field should also aim to clarify this unknown aspect of the application of bacteriocins, to provide the necessary knowledge about the optimization of the external conditions and open up the possibility of discovering their new

Identification and characterization of novel multiple bacteriocins produced by Leuconostoc pseudomesenteroides QU 15.

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Sawa, N; Okamura, K; Zendo, T; Himeno, K; Nakayama, J; Sonomoto, K

2010-07-01

To characterize novel multiple bacteriocins produced by Leuconostoc pseudomesenteroides QU 15. Leuconostoc pseudomesenteroides QU 15 isolated from Nukadoko (rice bran bed) produced novel bacteriocins. By using three purification steps, four antimicrobial peptides termed leucocin A (ΔC7), leucocin A-QU 15, leucocin Q and leucocin N were purified from the culture supernatant. The amino acid sequences of leucocin A (ΔC7) and leucocin A-QU 15 were identical to that of leucocin A-UAL 187 belonging to class IIa bacteriocins, but leucocin A (ΔC7) was deficient in seven C-terminal residues. Leucocin Q and leucocin N are novel class IId bacteriocins. Moreover, the DNA sequences encoding three bacteriocins, leucocin A-QU 15, leucocin Q and leucocin N were obtained. These bacteriocins including two novel bacteriocins were identified from Leuc. pseudomesenteroides QU 15. They showed similar antimicrobial spectra, but their intensities differed. The C-terminal region of leucocin A-QU 15 was important for its antimicrobial activity. Leucocins Q and N were encoded by adjacent open reading frames (ORFs) in the same operon, but leucocin A-QU 15 was not. These leucocins were produced concomitantly by the same strain. Although the two novel bacteriocins were encoded by adjacent ORFs, a characteristic of class IIb bacteriocins, they did not show synergistic activity. © 2010 The Authors. Journal compilation © 2010 The Society for Applied Microbiology.

Using the overlay assay to qualitatively measure bacterial production of and sensitivity to pneumococcal bacteriocins.

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Maricic, Natalie; Dawid, Suzanne

2014-09-30

Streptococcus pneumoniae colonizes the highly diverse polymicrobial community of the nasopharynx where it must compete with resident organisms. We have shown that bacterially produced antimicrobial peptides (bacteriocins) dictate the outcome of these competitive interactions. All fully-sequenced pneumococcal strains harbor a bacteriocin-like peptide (blp) locus. The blp locus encodes for a range of diverse bacteriocins and all of the highly conserved components needed for their regulation, processing, and secretion. The diversity of the bacteriocins found in the bacteriocin immunity region (BIR) of the locus is a major contributor of pneumococcal competition. Along with the bacteriocins, immunity genes are found in the BIR and are needed to protect the producer cell from the effects of its own bacteriocin. The overlay assay is a quick method for examining a large number of strains for competitive interactions mediated by bacteriocins. The overlay assay also allows for the characterization of bacteriocin-specific immunity, and detection of secreted quorum sensing peptides. The assay is performed by pre-inoculating an agar plate with a strain to be tested for bacteriocin production followed by application of a soft agar overlay containing a strain to be tested for bacteriocin sensitivity. A zone of clearance surrounding the stab indicates that the overlay strain is sensitive to the bacteriocins produced by the pre-inoculated strain. If no zone of clearance is observed, either the overlay strain is immune to the bacteriocins being produced or the pre-inoculated strain does not produce bacteriocins. To determine if the blp locus is functional in a given strain, the overlay assay can be adapted to evaluate for peptide pheromone secretion by the pre-inoculated strain. In this case, a series of four lacZ-reporter strains with different pheromone specificity are used in the overlay.

Thermostable Bacteriocin BL8 from Bacillus licheniformis isolated from marine sediment.

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Smitha, S; Bhat, S G

2013-03-01

To isolate and characterize bacteriocin, BL8, from the bacteria identified as Bacillus licheniformis from marine environment. One-hundred and twelve bacterial isolates from sediment and water samples collected off the coast of Cochin, India, were screened for antibacterial activity. Strain BTHT8, identified as Bacillus licheniformis, inhibited the growth of Gram-positive test organisms. The active component labelled as bacteriocin BL8 was partially purified by ammonium sulphate fractionation and was subjected to glycine SDS-PAGE. The band exhibiting antimicrobial activity was electroeluted and analysed using MALDI-TOF mass spectrometry, and the molecular mass was determined as 1.4 kDa. N-terminal amino acid sequencing of BL8 gave a 13 amino acid sequence stretch. Bacteriocin BL8 was stable even after boiling at 100 °C for 30 min and over a wide pH range of 1-12. A novel, pH-tolerant and thermostable bacteriocin BL8, active against the tested Gram-positive bacteria, was isolated from Bacillus licheniformis. This study reports a stable, low molecular weight bacteriocin from Bacillus licheniformis. This bacteriocin can be used to address two important applications: as a therapeutic agent and as a biopreservative in food processing industry. © 2012 The Society for Applied Microbiology.

Impact of Environmental Factors on Bacteriocin Promoter Activity in Gut-Derived Lactobacillus salivarius.

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Guinane, Caitriona M; Piper, Clare; Draper, Lorraine A; O'Connor, Paula M; Hill, Colin; Ross, R Paul; Cotter, Paul D

2015-11-01

Bacteriocin production is regarded as a desirable probiotic trait that aids in colonization and persistence in the gastrointestinal tract (GIT). Strains of Lactobacillus salivarius, a species associated with the GIT, are regarded as promising probiotic candidates and have a number of associated bacteriocins documented to date. These include multiple class IIb bacteriocins (salivaricin T, salivaricin P, and ABP-118) and the class IId bacteriocin bactofencin A, which show activity against medically important pathogens. However, the production of a bacteriocin in laboratory media does not ensure production under stressful environmental conditions, such as those encountered within the GIT. To allow this issue to be addressed, the promoter regions located upstream of the structural genes encoding the L. salivarius bacteriocins mentioned above were fused to a number of reporter proteins (green fluorescent protein [GFP], red fluorescent protein [RFP], and luciferase [Lux]). Of these, only transcriptional fusions to GFP generated signals of sufficient strength to enable the study of promoter activity in L. salivarius. While analysis of the class IIb bacteriocin promoter regions indicated relatively weak GFP expression, assessment of the promoter of the antistaphylococcal bacteriocin bactofencin A revealed a strong promoter that is most active in the absence of the antimicrobial peptide and is positively induced in the presence of mild environmental stresses, including simulated gastric fluid. Taken together, these data provide information on factors that influence bacteriocin production, which will assist in the development of strategies to optimize in vivo and in vitro production of these antimicrobials. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Characterization, production, and purification of carnocin H, a bacteriocin produced by Carnobacterium 377.

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Blom, H; Katla, T; Nissen, H; Holo, H

2001-10-01

Carnocin H, a bacteriocin produced by a Carnobacterium sp., inhibited lactic acid bacteria, clostridia, enterococci, and some Staphylococcus aureus strains. Some strains of Listeria and Pediococcus were also sensitive to carnocin H. The bacteriocin was produced during the late stationary growth phase. Carnocin H was purified by cation exchange chromatography and reverse phase chromatography. Amino acid sequence and composition indicate that carnocin H is a novel bacteriocin belonging to the class II bacteriocins. The bacteriocin consists of approximately 75 amino acid residues with a highly cationic N-terminal containing six succeeding lysines. Activity, as measured by agar diffusion zones, was reduced at increased pH values, levels of indicator bacteria, NaCl, agar, and soy oil.

Sec-mediated secretion of bacteriocin enterocin P by Lactococcus lactis

NARCIS (Netherlands)

Herranz, C; Driessen, AJM

Most lactic acid bacterium bacteriocins utilize specific leader peptides and dedicated machineries for secretion. In contrast, the enterococcal bacteriocin enterocin P (EntP) contains a typical signal peptide that directs its secretion when heterologously expressed in Lactococcus lactis. Signal

Fungicidal effect of bacteriocins harvested from Bacillus spp.

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Adetunji, V. O.

2013-01-01

Full Text Available Aims: This study investigated the ability of bacteriocins isolated from Bacillus spp. (Bacillus species to inhibit fourdifferent yeast isolates obtained from common food products (nono, yoghurt, ogi and cheese commonly consumed byNigerians with minimal heat treatment.Methodology and results: Forty-five Bacillus spp. was isolated and identified from common food products usingcultural, morphological, physiological and biochemical characteristics. These isolates were tested for antimicrobialactivity against Salmonella enteritidis (3, Micrococcus luteus (1 and Staphylococcus aureus (2. Eight bacteriocinproducing strains were identified from an over- night broth culture centrifugated at 3500 revolutions for five minutes.Fungicidal effects of these bacteriocins were tested against four yeast strains using the Agar Well Diffusion method. Thebacteriocins produced wide zones of inhibition ranging from 5.9±0.000 to 24.00±0.000 mm against the 4 yeast strainstested. There was a significant difference (at p<0.05 between the yeast organisms and the bacteriocins from theBacillus spp.Conclusion, significance and impact of study: The study reveals the antifungal property of bacteriocins from Bacillusspp. and serves therefore as a base for further studies in its use in the control of diseases and extension of shelf-life ofproducts prone to fungi contamination.

Interactions between oral bacteria: inhibition of Streptococcus mutans bacteriocin production by Streptococcus gordonii.

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Wang, Bing-Yan; Kuramitsu, Howard K

2005-01-01

Streptococcus mutans has been recognized as an important etiological agent in human dental caries. Some strains of S. mutans also produce bacteriocins. In this study, we sought to demonstrate that bacteriocin production by S. mutans strains GS5 and BM71 was mediated by quorum sensing, which is dependent on a competence-stimulating peptide (CSP) signaling system encoded by the com genes. We also demonstrated that interactions with some other oral streptococci interfered with S. mutans bacteriocin production both in broth and in biofilms. The inhibition of S. mutans bacteriocin production by oral bacteria was stronger in biofilms than in broth. Using transposon Tn916 mutagenesis, we identified a gene (sgc; named for Streptococcus gordonii challisin) responsible for the inhibition of S. mutans bacteriocin production by S. gordonii Challis. Interruption of the sgc gene in S. gordonii Challis resulted in attenuated inhibition of S. mutans bacteriocin production. The supernatant fluids from the sgc mutant did not inactivate the exogenous S. mutans CSP as did those from the parent strain Challis. S. gordonii Challis did not inactivate bacteriocin produced by S. mutans GS5. Because S. mutans uses quorum sensing to regulate virulence, strategies designed to interfere with these signaling systems may have broad applicability for biological control of this caries-causing organism.

Bacteriocin Isolated From Halomon sp.: A Bacterial Ding Protein?

International Nuclear Information System (INIS)

Atirah Azemin; Klappa, P.; Mohd Shahir Shamsir Omar

2015-01-01

A marine halophile, Halomonas sp. strain M3 was isolated from Straits of Johor, Malaysia and produce bacteriocin CC that acts as bacteriostatic agent. Characterisation of the bacterium showed that optimal growth and bacteriocin production is at ambient temperature, pH of 8-8.5 in nutrient broth medium supplemented with 2.9 % w/v NaCI to mimic saltwater conditions. The stability studies indicated that bacteriocin CC is heat-labile (35-50 degree Celsius) and was stable over 2 years when stored in 0.02 M Tris- HCI with 30-60 % glycerol at 4 degree Celsius. A loss of activity was detected after proteolytic enzymes treatment, indicating the proteinaceous nature of the antimicrobial compound. The amino acid sequence of bacteriocin CC was obtained by Edman degradation and MALDI-TOF analysis, showed the characteristic sequence of a DING protein (D-I-N-G-G-G-A-T-L-P-Q-A-LY- Q) in size 38.9-kDa at pI of 6.8. These proteins constitute a conserved and widely distributed set of proteins found in all kingdoms with ligand-binding activities and hydrolytic enzyme, suggesting a possible role in cell signalling and bio mineralization in DING isolates. Intriguingly, DING proteins also have been involved as an anti-tumour agent in humans. Thus, bacteriocin CC as DING protein family members should be further studied to investigate its potential as a novel antimicrobial agent. (author)

Influence of cultural conditions on the production of bacteriocin by ...

African Journals Online (AJOL)

Bacteriocin produced by Lactobacillus brevis OG1 has large spectrum of inhibition against pathogenic, food spoilage microorganisms and various Lactic acid bacteria employed as test strains. The bacteriocin inhibited E coli NCTC 10418 and Enterococcus faecalis, but did not inhibit Candida albicans ATCC 10231 and ...

PHYSICHOCHEMICAL CHACTERIZATION OF BACTERIOCIN PRODUCING ENTEROCOCCUS DURANS ISOLATED FROM COLON’S BALI CATTLE

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I Wayan Suardana

2017-08-01

Full Text Available Lactic acid bacteria can excrete antimicrobial compounds like bacteriocins. The study  aimed  to  find  out  the  characteristic  of  physic-chemical  of  bacteriocin  producing Enterococcus durans          isolate 18A isolated from  colon’s bali cattle. The study initiated by Gram staining and catalase test, followed by isolation and purification  of bacteriocin.       The result of the research showed  that bacteriocins of    Enterococcus durans    isolate 18A as a protein with it’s concentrationis  0,272?g/ml  and  it  does  not  contain  carbohydrate.  On  the  other  hand,  the bacteriocins was not showed a band while tested on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE. The result showed, the bacteriocin producing            Enterococcus durans isolate 18A has antimicrobial activity to Bacillus cereus  as 23,88%.

Importance in dairy technology of bacteriocins produced by dairy starter cultures

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Bedia Şimşek

2002-03-01

Full Text Available Bacteriocins produced by Lactic acid bacteria (LAB and propionic acid bacteria (PAB are heterogeneous group of peptide inhibitors which include lantibiotics (class I, e. g. nisin, small heat-stable peptides (class II, e. g. pediocin PA-1 and large heat-labile proteins (class III, e. g. helveticin J. Many bacteriocins belonging to the first two groups can be successfully used to inhibit undesirable microorganisms in foods, but only nisin is produced industrially and is used as a food preservative. LAB and PAB develops easily in milk and milk products. LAB and PAB growth in dairy products can cause microbial interference to spoilage and pathogenic bacteria through several metabolits, specially bacteriocins. The review deals with the description of milk-borne bacteriocins and their application in milk and milk products either to extend the shelf life or to inhibit milk pathogens.

Bacteriocins from the rhizosphere microbiome – from an agriculture perspective

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Sowmyalakshmi eSubramanian

2015-10-01

Full Text Available Bacteria produce and excrete a versatile and dynamic suit of compounds to defend against microbial competitors and mediate local population dynamics. These include a wide range of broad-spectrum non-ribosomally synthesized antibiotics, lytic enzymes, metabolic by-products, proteinaceous exotoxins and ribosomally produced antimicrobial peptides (bacteriocins. Most bacteria produce at least one bacteriocin. Bacteriocins are of interest in the food industry as natural preservatives and in the probiotics industry, leading to extensive studies on lactic acid bacteria (colicin produced by Escherichia coli is a model bacteriocin. Recent studies have projected use of bacteriocins in veterinary medicine and in agriculture, as a biostimulants of plant growth and development and as biocontrol agents. For example, bacteriocins such as Cerein 8A, Bac-GM17, putidacin, Bac 14B, amylocyclicin have been studied for their mechanisms of anti-microbial activity. Bac IH7 promotes tomato and musk melon plant growth. Thuricin 17 (Th17 is the only bacteriocin studied extensively for plant growth promotion and at the molecular level. Th17 functions as a bacterial signal compound, promoting plant growth in legumes and non-legumes. In Arabidopsis thaliana and Glycine max Th17 increased phytohormones IAA and SA at 24 h post treatment. At the proteome level Th17 treatment of 3-week-old A. thaliana rosettes led to > 2-fold changes in activation of the carbon and energy metabolism pathway proteins, 24 h post treatment. At 250 mM NaCl stress, the control plants under osmotic-shock shut down most of carbon-metabolism and activated energy-metabolism and antioxidant pathways. Th17 treated plants, at 250 mM NaCl, retained meaningful levels of the light harvesting complex, photosystem I and II proteins and energy and antioxidant pathways were activated, so that rosettes could better withstand the salt stress. In Glycine max, Th17 helped seeds germinate in the presence of Na

Lactococcus lactis LMG2081 Produces Two Bacteriocins, a Nonlantibiotic and a Novel Lantibiotic.

Science.gov (United States)

Mirkovic, Nemanja; Polovic, Natalija; Vukotic, Goran; Jovcic, Branko; Miljkovic, Marija; Radulovic, Zorica; Diep, Dzung B; Kojic, Milan

2016-04-01

Bacteriocin producers normally possess dedicated immunity systems to protect themselves from their own bacteriocins.Lactococcus lactis strains LMG2081 and BGBM50 are known as lactococcin G producers. However, BGBM50 was sensitive to LMG2081, which indicated that LMG2081 might produce additional bacteriocins that are not present in BGBM50. Therefore, whole-genome sequencing of the two strains was performed, and a lantibiotic operon (called lctLMG) was identified in LMG2081 but not in BGBM50. The lctLMG operon contains six open reading frames; the first three genes,lmgA ,lmgM, and lmgT, are involved in the biosynthesis and export of bacteriocin, while the other three genes,lmgF,lmgE, and lmgG, are involved in lantibiotic immunity. Mutational analysis confirmed that the lctLMG operon is responsible for the additional antimicrobial activity. Specifically, site-directed mutation within this operon rendered LMG2081 inactive toward BGBM50. Subsequent purification and electrospray ionization-time of flight mass spectrometric analysis confirmed that the lantibiotic bacteriocin called lacticin LMG is exported as a 25-amino-acid peptide. Lacticin LMG is highly similar to the lacticin 481 group. It is interesting that a bacteriocin producer produces two different classes of bacteriocins, whose operons are located in the chromosome and a plasmid. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Antibacterial efficacy of nisin, pediocin 34 and enterocin FH99 against L. monocytogenes, E. faecium and E. faecalis and bacteriocin cross resistance and antibiotic susceptibility of their bacteriocin resistant variants.

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Kaur, Gurpreet; Singh, Tejinder Pal; Malik, Ravinder Kumar; Bhardwaj, Arun; De, Sachinandan

2014-02-01

The bacteriocin susceptibility of Listeria monocytogenes MTCC 657, Enterococcus faecium DSMZ 20477, E. faecium VRE, and E. faecalis ATCC 29212 and their corresponding bacteriocin resistant variants was assessed. The single and combined effect of nisin and pediocin 34 and enterocin FH99 bacteriocins produced by Pediococcus pentosaceus 34, and E. faecium FH99, respectively, was determined. Pediocin34 proved to be more effective in inhibiting L. monocytogenes MTCC 657. A greater antibacterial effect was observed against E. faecium DSMZ 20477 and E. faecium (VRE) when the a combination of nisin, pediocin 34 and enterocin FH99 were used whereas in case of L. monocytogenes MTCC 657 a combination of pediocin 34 and enterocin FH99 was more effective in reducing the survival of pathogen. Bacteriocin cross-resistance and the antibiotic susceptibility of wild type and their corresponding resistant variants were assessed and results showed that resistance to a bacteriocin may extend to other bacteriocins within the same class and also the acquired resistance to bacteriocins can modify the antibiotic susceptibility/resistance profile of the bacterial species used in the study. According to the hydrophobicity nisin resistant variant of L. monocytogenes was more hydrophobic (p enterocin FH99 resistant variants were less hydrophobic than the wild type strain. Nisin, pediocin 34 and enterocin FH99 resistant variants of E. faecium DSMZ 20477 and E. faecium VRE were less hydrophobic than their wild type counterparts. Nisin resistant E. faecalis ATCC 29212 was less hydrophobic than its wild type counterpart.

Antilisterial Activity of Bacteriocin Isolated from Leuconostoc mesenteroides ssp. mesenteroides IMAU:10231 in the Production of Sremska Sausages: Lactic Acid Bacteria Isolation, Bacteriocin Identification and Meat Application Experiments

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Marija Škrinjar

2013-01-01

Full Text Available Lactic acid bacteria (LAB have an essential role in the production of fermented meat products. The metabolic activity of LAB affects the ripening process, leading to the formation of the desired sensory characteristics of the products, while inhibiting the growth of undesirable microorganisms. Bacteriocins are extracellular peptides or protein molecules, produced by some LAB, which possess bactericidal properties against specific species or genera of microorganisms, usually related bacteria. Bacteriocin production by LAB can act in a selective and competitive way against the surrounding microbiota, which may contain spoilage bacteria or pathogenic microorganisms including Listeria monocytogenes. This pathogen is widely distributed in raw products, it survives in different production areas, and human infections have a high mortality rate, all of which makes the control of this microorganism important in food production. The aim of this work is to determine the possibilities of utilizing a novel bacteriocin isolated from Leuconostoc mesenteroides ssp. mesenteroides IMAU:10231 in order to prevent the survival of Listeria monocytogenes in the production of traditional Serbian Sremska sausages. The bacteriocin-producing strain of Leuconostoc originated from the same sausage, which had been produced in the traditional manner. Bacteriocin was isolated using precipitation procedures with ammonium sulphate, and then its properties (strength and range of activities, relationship to high temperatures and proteolytic enzymes were determined under laboratory conditions. Also, based on the obtained laboratory results, the antilisterial effect of bacteriocin, included as an additive, was examined in the production of traditional Sremska sausages. Expressed antilisterial activity of bacteriocin has an interesting food safety potential which can be used in the meat industry in the production of fermented sausages. Further research will contribute to a better

Recent Progress in the Chemical Synthesis of Class II and S-Glycosylated Bacteriocins

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François Bédard

2018-05-01

Full Text Available A wide variety of antimicrobial peptides produced by lactic acid bacteria (LAB have been identified and studied in the last decades. Known as bacteriocins, these ribosomally synthesized peptides inhibit the growth of a wide range of bacterial species through numerous mechanisms and show a great variety of spectrum of activity. With their great potential as antimicrobial additives and alternatives to traditional antibiotics in food preservation and handling, animal production and in veterinary and medical medicine, the demand for bacteriocins is rapidly increasing. Bacteriocins are most often produced by fermentation but, in several cases, the low isolated yields and difficulties associated with their purification seriously limit their use on a large scale. Chemical synthesis has been proposed for their production and recent advances in peptide synthesis methodologies have allowed the preparation of several bacteriocins. Moreover, the significant cost reduction for peptide synthesis reagents and building blocks has made chemical synthesis of bacteriocins more attractive and competitive. From a protein engineering point of view, the chemical approach offers many advantages such as the possibility to rapidly perform amino acid substitution, use unnatural or modified residues, and make backbone and side chain modifications to improve potency, modify the activity spectrum or increase the stability of the targeted bacteriocin. This review summarized synthetic approaches that have been developed and used in recent years to allow the preparation of class IIa bacteriocins and S-linked glycopeptides from LAB. Synthetic strategies such as the use of pseudoprolines, backbone protecting groups, microwave irradiations, selective disulfide bridge formation and chemical ligations to prepare class II and S-glycosylsated bacteriocins are discussed.

Bacteriocin Production with Lactobacillus amylovorus DCE 471 Is Improved and Stabilized by Fed-Batch Fermentation

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Callewaert, Raf; De Vuyst, Luc

2000-01-01

Amylovorin L471 is a small, heat-stable, and hydrophobic bacteriocin produced by Lactobacillus amylovorus DCE 471. The nutritional requirements for amylovorin L471 production were studied with fed-batch fermentations. A twofold increase in bacteriocin titer was obtained when substrate addition was controlled by the acidification rate of the culture, compared with the titers reached with constant substrate addition or pH-controlled batch cultures carried out under the same conditions. An interesting feature of fed-batch cultures observed under certain culture conditions (constant feed rate) is the apparent stabilization of bacteriocin activity after obtaining maximum production. Finally, a mathematical model was set up to simulate cell growth, glucose and complex nitrogen source consumption, and lactic acid and bacteriocin production kinetics. The model showed that bacterial growth was dependent on both the energy and the complex nitrogen source. Bacteriocin production was growth associated, with a simultaneous bacteriocin adsorption on the producer cells dependent on the lactic acid accumulated and hence the viability of the cells. Both bacteriocin production and adsorption were inhibited by high concentrations of the complex nitrogen source. PMID:10653724

Production of multiple bacteriocins from a single locus by gastrointestinal strains of Lactobacillus salivarius.

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O'Shea, Eileen F; O'Connor, Paula M; Raftis, Emma J; O'Toole, Paul W; Stanton, Catherine; Cotter, Paul D; Ross, R Paul; Hill, Colin

2011-12-01

Bacteriocins produced by Lactobacillus salivarius isolates derived from a gastrointestinal origin have previously demonstrated efficacy for in vivo protection against Listeria monocytogenes infection. In this study, comparative genomic analysis was employed to investigate the intraspecies diversity of seven L. salivarius isolates of human and porcine intestinal origin, based on the genome of the well-characterized bacteriocin-producing strain L. salivarius UCC118. This revealed a highly conserved megaplasmid-borne gene cluster in these strains involved in the regulation and secretion of two-component class IIb bacteriocins. However, considerable intraspecific variation was observed in the structural genes encoding the bacteriocin peptides. They ranged from close relatives of abp118, such as salivaricin P, which differs by 2 amino acids, to completely novel bacteriocins, such as salivaricin T, which is characterized in this study. Salivaricin T inhibits closely related lactobacilli and bears little homology to previously characterized salivaricins. Interestingly, the two peptides responsible for salivaricin T activity, SalTα and SalTβ, share considerable identity with the component peptides of thermophilin 13, a bacteriocin produced by Streptococcus thermophilus. Furthermore, the salivaricin locus of strain DPC6488 also encodes an additional novel one-component class IId anti-listerial bacteriocin, salivaricin L. These findings suggest a high level of redundancy in the bacteriocins that can be produced by intestinal L. salivarius isolates using the same enzymatic production and export machinery. Such diversity may contribute to their ability to dominate and compete within the complex microbiota of the mammalian gut.

Bacteriocins from lactic acid bacteria: production, purification, and food applications.

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De Vuyst, Luc; Leroy, Frédéric

2007-01-01

In fermented foods, lactic acid bacteria (LAB) display numerous antimicrobial activities. This is mainly due to the production of organic acids, but also of other compounds, such as bacteriocins and antifungal peptides. Several bacteriocins with industrial potential have been purified and characterized. The kinetics of bacteriocin production by LAB in relation to process factors have been studied in detail through mathematical modeling and positive predictive microbiology. Application of bacteriocin-producing starter cultures in sourdough (to increase competitiveness), in fermented sausage (anti-listerial effect), and in cheese (anti-listerial and anti-clostridial effects), have been studied during in vitro laboratory fermentations as well as on pilot-scale level. The highly promising results of these studies underline the important role that functional, bacteriocinogenic LAB strains may play in the food industry as starter cultures, co-cultures, or bioprotective cultures, to improve food quality and safety. In addition, antimicrobial production by probiotic LAB might play a role during in vivo interactions occurring in the human gastrointestinal tract, hence contributing to gut health.

Bacteriocins from lactic acid bacteria: purification, properties and use as biopreservatives

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José Luis Parada

2007-05-01

Full Text Available Biopreservation systems in foods are of increasing interest for industry and consumers. Bacteriocinogenic lactic acid bacteria and/or their isolated bacteriocins are considered safe additives (GRAS, useful to control the frequent development of pathogens and spoiling microorganisms in foods and feed. The spreading of bacterial antibiotic resistance and the demand for products with fewer chemicals create the necessity of exploring new alternatives, in order to reduce the abusive use of therapeutic antibiotics. In this context, bacteriocins are indicated to prevent the growth of undesirable bacteria in a food-grade and more natural way, which is convenient for health and accepted by the community. According to their properties, structure, molecular weight (MW, and antimicrobial spectrum, bacteriocins are classified in three different groups: lantibiotics and non-lantibiotics of low MW, and those of higher MW. Several strategies for isolation and purification of bacteriocins from complex cultivation broths to final products were described. Biotechnological procedures including salting-out, solvent extraction, ultrafiltration, adsorption-desortion, ion-exchange, and size exclusion chromatography are among the most usual methods. Peptide structure-function studies of bacteriocins and bacterial genetic advances will help to understand the molecular basis of their specificity and mode of action. Nisin is a good example of commercial success, and a good perspective is open to continue the study and development of new bacteriocins and their biotechnological applications. These substances in appropriate concentrations may be used in veterinary medicine and as animal growth promoter instead usual antibiotics, as well as an additional hurdle factor for increasing the shelf life of minimal processed foods.

Purification Techniques of Bacteriocins from Lactic Acid Bacteria and Other Gram-Positive Bacteria

Science.gov (United States)

Saavedra, Lucila; Sesma, Fernando

The search for new antimicrobial peptides produced by lactic acid ­bacteria and other Gram-positive microorganisms has become an interesting field of research in the past decades. The fact that bacteriocins are active against numerous foodborne and human pathogens, are produced by generally regarded as safe (GRAS) microorganisms, and are readily degraded by proteolytic host systems makes them attractive candidates for biotechnological applications. However, before suggesting or choosing a new bacteriocin for future technology developments, it is necessary to elucidate its biochemical structure and its mode of action, which may be carried out once the bacteriocin is purified to homogeneity. This chapter focuses on describing the main strategies used for the purification of numerous bacteriocins.

Purification and characterization of plantaricin Y, a novel bacteriocin produced by Lactobacillus plantarum 510.

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Chen, Yi-sheng; Wang, Yan-chong; Chow, Yiou-shing; Yanagida, Fujitoshi; Liao, Chen-chung; Chiu, Chi-ming

2014-03-01

Lactobacillus plantarum 510, previously isolated from a koshu vineyard in Japan, was found to produce a bacteriocin-like inhibitory substance which was purified and characterized. Mass spectrometry analysis showed that the mass of this bacteriocin is 4,296.65 Da. A partial sequence, NH2- SSSLLNTAWRKFG, was obtained by N-terminal amino acid sequence analysis. A BLAST search revealed that this is a unique sequence; this peptide is thus a novel bacteriocin produced by Lactobacillus plantarum 510 and was termed plantaricin Y. Plantaricin Y shows strong inhibitory activity against Listeria monocytogenes BCRC 14845, but no activity against other pathogens tested. Bacteriocin activity decreased slightly after autoclaving (121 °C for 15 min), but was completely inactivated by protease K. Furthermore, trypsin-digested bacteriocin product fragments retained activity against L. monocytogenes BCRC 14845 and exhibited a different inhibitory spectrum.

Identification, purification and characterization of laterosporulin, a novel bacteriocin produced by Brevibacillus sp. strain GI-9.

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Pradip Kumar Singh

Full Text Available BACKGROUND: Bacteriocins are antimicrobial peptides that are produced by bacteria as a defense mechanism in complex environments. Identification and characterization of novel bacteriocins in novel strains of bacteria is one of the important fields in bacteriology. METHODOLOGY/FINDINGS: The strain GI-9 was identified as Brevibacillus sp. by 16 S rRNA gene sequence analysis. The bacteriocin produced by strain GI-9, namely, laterosporulin was purified from supernatant of the culture grown under optimal conditions using hydrophobic interaction chromatography and reverse-phase HPLC. The bacteriocin was active against a wide range of Gram-positive and Gram-negative bacteria. MALDI-TOF experiments determined the precise molecular mass of the peptide to be of 5.6 kDa and N-terminal sequencing of the thermo-stable peptide revealed low similarity with existing antimicrobial peptides. The putative open reading frame (ORF encoding laterosporulin and its surrounding genomic region was fished out from the draft genome sequence of GI-9. Sequence analysis of the putative bacteriocin gene did not show significant similarity to any reported bacteriocin producing genes in database. CONCLUSIONS: We have identified a bacteriocin producing strain GI-9, belonging to the genus Brevibacillus sp. Biochemical and genomic characterization of laterosporulin suggests it as a novel bacteriocin with broad spectrum antibacterial activity.

Identification, purification and characterization of laterosporulin, a novel bacteriocin produced by Brevibacillus sp. strain GI-9.

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Singh, Pradip Kumar; Chittpurna; Ashish; Sharma, Vikas; Patil, Prabhu B; Korpole, Suresh

2012-01-01

Bacteriocins are antimicrobial peptides that are produced by bacteria as a defense mechanism in complex environments. Identification and characterization of novel bacteriocins in novel strains of bacteria is one of the important fields in bacteriology. The strain GI-9 was identified as Brevibacillus sp. by 16 S rRNA gene sequence analysis. The bacteriocin produced by strain GI-9, namely, laterosporulin was purified from supernatant of the culture grown under optimal conditions using hydrophobic interaction chromatography and reverse-phase HPLC. The bacteriocin was active against a wide range of Gram-positive and Gram-negative bacteria. MALDI-TOF experiments determined the precise molecular mass of the peptide to be of 5.6 kDa and N-terminal sequencing of the thermo-stable peptide revealed low similarity with existing antimicrobial peptides. The putative open reading frame (ORF) encoding laterosporulin and its surrounding genomic region was fished out from the draft genome sequence of GI-9. Sequence analysis of the putative bacteriocin gene did not show significant similarity to any reported bacteriocin producing genes in database. We have identified a bacteriocin producing strain GI-9, belonging to the genus Brevibacillus sp. Biochemical and genomic characterization of laterosporulin suggests it as a novel bacteriocin with broad spectrum antibacterial activity.

Optimization of Fermentation Conditions for the Production of Bacteriocin Fermentate

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2015-03-30

FERMENTATION CONDITIONS FOR THE PRODUCTION OF BACTERIOCIN “ FERMENTATE ” by Anthony Sikes Wayne Muller and Claire Lee March 2015...From - To) October 2010 – November 2013 4. TITLE AND SUBTITLE OPTIMIZATION OF FERMENTATION CONDITIONS FOR THE PRODUCTION OF BACTERIOCIN “ FERMENTATE ...nisin and pediocin. Whey + yeast extract was the best performing whey fermentation media. The nisin producer strain Lactococcus. lactis ssp. lactis was

Use of synthetic genes for cloning, production and functional expression of the bacteriocins enterocin A and bacteriocin E 50-52 by Pichia pastoris and Kluyveromyces lactis.

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Jiménez, Juan J; Borrero, Juan; Gútiez, Loreto; Arbulu, Sara; Herranz, Carmen; Cintas, Luis M; Hernández, Pablo E

2014-06-01

The use of synthetic genes may constitute a successful approach for the heterologous production and functional expression of bacterial antimicrobial peptides (bacteriocins) by recombinant yeasts. In this work, synthetic genes with adapted codon usage designed from the mature amino acid sequence of the bacteriocin enterocin A (EntA), produced by Enterococcus faecium T136, and the mature bacteriocin E 50-52 (BacE50-52), produced by E. faecium NRRL B-32746, were synthesized. The synthetic entA and bacE50-52 were cloned into the protein expression vectors pPICZαA and pKLAC2 for transformation of derived vectors into Pichia pastoris X-33 and Kluyveromyces lactis GG799, respectively. The recombinant vectors were linearized and transformed into competent cells selecting for P. pastoris X-33EAS (entA), P. pastoris X-33BE50-52S (bacE50-52), K. lactis GG799EAS (entA), and K. lactis GG799BE50-52S (bacE50-52). P. pastoris X-33EAS and K. lactis GG799EAS, but not P. pastoris X-33BE50-52S and K. lactis GG799BE50-52S, showed antimicrobial activity in their supernatants. However, purification of the supernatants of the producer yeasts permitted recovery of the bacteriocins EntA and BacE50-52. Both purified bacteriocins were active against Gram-positive bacteria such as Listeria monocytogenes but not against Gram-negative bacteria, including Campylobacter jejuni.

Semi-preparative scale purification of enterococcal bacteriocin enterocin EJ97, and evaluation of substrates for its production.

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López, Rosario Lucas; García, Ma Teresa; Abriouel, Hikmate; Ben Omar, Nabil; Grande, Ma José; Martínez-Cañamero, Magdalena; Gálvez, Antonio

2007-12-01

The influence of substrate composition on the production of enterocin EJ97 and the conditions for semi-preparative bacteriocin recovery have been studied. Final bacteriocin concentrations of 12.5 or 15.6 mg/l were obtained in the commercial media brain heart infusion broth (BHI) and tryptic soya broth, respectively. The bacteriocin was also produced in the complex medium CM (8.75 mg/l), in which the vitamin supplement was essential for production. Some combinations of meat peptone and yeast extract plus either soy peptone or BHI also supported bacteriocin production, at concentrations of 6.25-7.5 mg/l. In cow milk (whole, half-skimmed, and skimmed), the final bacteriocin concentrations obtained ranged from 7.5 to 11.25 mg/l. Highest bacteriocin activity was obtained by using pasteurised milk whey as growth substrate (up to 25 mg/l), suggesting that this bacteriocin can be obtained on a large scale by using this cheap food-grade industrial by-product. Highest bacteriocin titres were always obtained after 8 h of incubation at 37 degrees C. Semi-preparative concentration and purification of enterocin EJ97 produced in a complex medium was achieved by bulk cation exchange chromatography without previous cell separation, followed by reversed-phase chromatography. This two-step procedure allowed preparation of milligram quantities of purified bacteriocin, which is an improvement compared to purification procedures established for most other bacteriocins (35). The availability of purified enterocin EJ97 will facilitate other studies such as the elucidation of its molecular structure and its interaction with target bacteria.

Effects of Streptococcus sanguinis Bacteriocin on Cell Surface Hydrophobicity, Membrane Permeability, and Ultrastructure of Candida Thallus

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Shengli Ma

2015-01-01

Full Text Available Candida albicans (C.a and Candida tropicalis (C.t were treated with Streptococcus sanguinis bacteriocin (S.s bacteriocin, respectively; the bacteriostatic dynamics of S.s bacteriocin, their effects on cell surface hydrophobicity, leakage of inorganic phosphorus and macromolecular substance, cytosolic calcium concentration, and ultrastructure changes of Candida thallus were detected and analyzed. The results showed that inhibitory effect of S.s bacteriocin on C.a and C.t reached peak level at 24 h, the cell-surface hydrophobicity decreased significantly (P < 0.05 after S.s bacteriocin treatment, and there was leakage of cytoplasmic inorganic phosphorus and macromolecular substance from C.a and C.t; cytosolic calcium concentration decreased greatly. After 24 h treatment by S.s bacteriocin, depressive deformity and defect could be found in the cell surface of C.a and C.t; the thallus displayed irregular forms: C.a was shrunken, there was unclear margins abutting upon cell wall and cell membrane, nucleus disappeared, and cytoplasm was inhomogeneous; likewise, C.t was first plasmolysis, and then the cytoplasm was shrunk, the ultrastructure of cell wall and cell membrane was continuously damaged, and the nucleus was karyolysis. It was illustrated that S.s bacteriocin had similar antifungal effect on C.a and C.t; their cell surface hydrophobicity, membrane permeability, and ultrastructure were changed significantly on exposure to S.s bacteriocin.

Enterocin 96, a Novel Class II Bacteriocin Produced by Enterococcus faecalis WHE 96, Isolated from Munster Cheese▿

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Izquierdo, Esther; Wagner, Camille; Marchioni, Eric; Aoude-Werner, Dalal; Ennahar, Saïd

2009-01-01

Enterococcus faecalis WHE 96, a strain isolated from soft cheese based on its anti-Listeria activity, produced a 5,494-Da bacteriocin that was purified to homogeneity by ultrafiltration and cation-exchange and reversed-phase chromatographies. The amino acid sequence of this bacteriocin, named enterocin 96, was determined by Edman degradation, and its structural gene was sequenced, revealing a double-glycine leader peptide. After a comparison with other bacteriocins, it was shown that enterocin 96 was a new class II bacteriocin that showed very little similarity with known structures. Enterocin 96 was indeed a new bacteriocin belonging to class II bacteriocins. The activity spectrum of enterocin 96 covered a wide range of bacteria, with strong activity against most gram-positive strains but very little or no activity against gram-negative strains. PMID:19411428

Enterocin 96, a novel class II bacteriocin produced by Enterococcus faecalis WHE 96, isolated from Munster cheese.

Science.gov (United States)

Izquierdo, Esther; Wagner, Camille; Marchioni, Eric; Aoude-Werner, Dalal; Ennahar, Saïd

2009-07-01

Enterococcus faecalis WHE 96, a strain isolated from soft cheese based on its anti-Listeria activity, produced a 5,494-Da bacteriocin that was purified to homogeneity by ultrafiltration and cation-exchange and reversed-phase chromatographies. The amino acid sequence of this bacteriocin, named enterocin 96, was determined by Edman degradation, and its structural gene was sequenced, revealing a double-glycine leader peptide. After a comparison with other bacteriocins, it was shown that enterocin 96 was a new class II bacteriocin that showed very little similarity with known structures. Enterocin 96 was indeed a new bacteriocin belonging to class II bacteriocins. The activity spectrum of enterocin 96 covered a wide range of bacteria, with strong activity against most gram-positive strains but very little or no activity against gram-negative strains.

Purification and Molecular Characterization of the Novel Highly Potent Bacteriocin TSU4 Produced by Lactobacillus animalis TSU4.

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Sahoo, Tapasa Kumar; Jena, Prasant Kumar; Patel, Amiya Kumar; Seshadri, Sriram

2015-09-01

Bacterial infections causing fish diseases and spoilage during fish food processing and storage are major concerns in aquaculture. Use of bacteriocins has recently been considered as an effective strategy for prevention of bacterial infections. A novel bacteriocin produced by Catla catla gut isolates, Lactobacillus animalis TSU4, designated as bacteriocin TSU4 was purified to homogeneity by a three-step protocol. The molecular mass of bacteriocin TSU4 was 4117 Da determined by Q-TOF LC/MS analysis. Its isoelectric point was ~9. Secondary conformation obtained by circular dichroism spectroscopy showed molecular conformation with significant proportions of the structure in α-helix (23.7 %) and β-sheets (17.1 %). N-terminal sequencing was carried out by the Edman degradation method; partial sequence identified was NH2-SMSGFSKPHD. Bacteriocin TSU4 exhibited a wide range of antimicrobial activity, pH and thermal stability. It showed a bacteriocidal mode of action against the indicator strain Aeromonas hydrophila MTCC 646. Bacteriocin TSU4 is the first reported bacteriocin produced by fish isolate Lactobacillus animalis. The characterization of bacteriocin TSU4 suggested that it is a novel bacteriocin with potential value against infections of bacteria such as A. hydrophila MTCC 646 and Pseudomonas aeruginosa MTCC 1688 and application to prevent spoilage during food preservation.

A Natural Chimeric Pseudomonas Bacteriocin with Novel Pore-Forming Activity Parasitizes the Ferrichrome Transporter.

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Ghequire, Maarten G K; Kemland, Lieselore; Anoz-Carbonell, Ernesto; Buchanan, Susan K; De Mot, René

2017-02-21

Modular bacteriocins represent a major group of secreted protein toxins with a narrow spectrum of activity, involved in interference competition between Gram-negative bacteria. These antibacterial proteins include a domain for binding to the target cell and a toxin module at the carboxy terminus. Self-inhibition of producers is provided by coexpression of linked immunity genes that transiently inhibit the toxin's activity through formation of bacteriocin-immunity complexes or by insertion in the inner membrane, depending on the type of toxin module. We demonstrate strain-specific inhibitory activity for PmnH, a Pseudomonas bacteriocin with an unprecedented dual-toxin architecture, hosting both a colicin M domain, potentially interfering with peptidoglycan synthesis, and a novel colicin N-type domain, a pore-forming module distinct from the colicin Ia-type domain in Pseudomonas aeruginosa pyocin S5. A downstream-linked gene product confers PmnH immunity upon susceptible strains. This protein, ImnH, has a transmembrane topology similar to that of Pseudomonas colicin M-like and pore-forming immunity proteins, although homology with either of these is essentially absent. The enhanced killing activity of PmnH under iron-limited growth conditions reflects parasitism of the ferrichrome-type transporter for entry into target cells, a strategy shown here to be used as well by monodomain colicin M-like bacteriocins from pseudomonads. The integration of a second type of toxin module in a bacteriocin gene could offer a competitive advantage against bacteria displaying immunity against only one of both toxic activities. IMPORTANCE In their continuous struggle for ecological space, bacteria face a huge load of contenders, including phylogenetically related strains that compete for the same niche. One important group of secreted antibacterial proteins assisting in eliminating these rivals are modular bacteriocins of Gram-negative bacteria, comprising a domain for docking onto the

Isolation and purification of two bacteriocins 3D produced by Enterococcus faecium with inhibitory activity against Listeria monocytogenes.

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Bayoub, Kaoutar; Mardad, Ilham; Ammar, Emna; Serrano, Aurelio; Soukri, Abdelaziz

2011-02-01

Strain 3D, isolated from fermented traditional Moroccan dairy product, and identified as Enterococcus faecium, was studied for its capability to produce two bacteriocins acting against Listeria monocytogenes. Bacteriocins 3 Da and 3Db were heat stable inactivated by proteinase K, pepsin, and trypsin but not when treated with catalase. The evidenced bacteriocins were stable in a wide pH range from 2 to 11 and bactericidal activity was kept during storage at 4°C. However, the combination of temperature and pH exhibited a stability of the bacteriocins. RP-HPLC purification of the anti-microbial compounds shows two active fractions eluted at 16 and 30.5 min, respectively. Mass spectrometry analysis showed that E. faecium 3D produce two bacteriocins Enterocin 3 Da (3893.080 Da) and Enterocin 3Db (4203.350 Da). This strain is food-grade organism and its bacteriocins were heat-stable peptides at basic, neutral, and acid pH: such bacteriocins may be of interest as food preservatives.

Bacteriocins produced by L. fermentum and L .acidophilus can inhibit cephalosporin resistant E .coli.

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Saba Riaz

2010-10-01

Full Text Available Reemerging infections occur due to resistant bacteria. Such infections create restrictions for clinicians and microbiologists in drug selection. Such problems demand new strategies for solution. Use of bacteriocins for this purpose may be fruitful. In the present research work, the inhibitory effects of bactericins on cephalosporin resistant Escherichia coli are used as model system for the control of antibiotic resistant pathogenic bacteria. Cephalosporin resistant Escherichia coli strain was isolated from pus by using conventional methodology. For bacteriocin production, Lactobacilli strains were selected by using selective media. Out of seventy two strains isolated from yogurt, fecal materials of human, chick, parrot and cat, only two strains (strain 45 and strain 52 were found to produce bacteriocins having antimicrobial potential against cephalosporin resistant Escherichia coli. Biochemical characterization showed that strain 45 belonged to group of Lactobacillus fermentum and strain 52 to Lactobacillus acidophilus. Both strains showed maximum growth at 25°C and 35°C respectively. Suitable pH was 5.5 and 6.0 for Lactobacillus fermentum and Lactobacillus acidophilus respectively. Bacteriocins produced by both strains were found stable at 50, 75 and 100°C for 60min. Function of bacteriocin was also not disturbed due to change in pH. These findings suggest that bacteriocin produced by Lactobacillus fermentum and Lactobacillus acidophilus can be used for the infection control of cephalosporin resistant Escherichia coli.

Production of bacteriocin by Leuconostoc mesenteroides 406 isolated from Mongolian fermented mare's milk, airag.

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Wulijideligen; Asahina, Takayuki; Hara, Kazushi; Arakawa, Kensuke; Nakano, Hiroyuki; Miyamoto, Taku

2012-10-01

The purification and characterization of a bacteriocin produced by Leuconostoc mesenteroides strain 406 that was isolated from traditional Mongolian fermented mare's milk, airag, were carried out. Leuconostoc mesenteroides strain 406 was identified on the basis of its morphological and biochemical characteristics and carbohydrate fermentation profile and by API 50 CH kit and 16S ribosomal DNA analyses. The neutral-pH cell-free supernatant of this bacterium inhibited the growth of several lactic acid bacteria and food spoilage and pathogenic organisms, including Listeria monocytogenes and Clostridium botulinum. The bacteriocin was heat-stable and not sensitive to acid and alkaline conditions, but was sensitive to several proteolytic enzymes such as pepsin, pronase E, proteinase K, trypsin, and α-chymotrypsin, but not catalase. Optimum bacteriocin production (4000 activity units/mL) was achieved when the strain was cultured at 25°C for 24-36 h in Man Rogosa Sharpe medium. The bacteriocin was partially purified by ammonium sulfate precipitation (80% saturation), dialysis (cut-off MW: 1000), and gel filtration chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that the bacteriocin had a molecular weight of approximately 3.3 kDa. To our knowledge, this is the first report of the isolation of a bacteriocin-producing Leuconostoc strain from airag. An application to fermented milks would be desired. © 2012 The Authors. Animal Science Journal © 2012 Japanese Society of Animal Science.

A counterselection method for Lactococcus lactis genome editing based on class IIa bacteriocin sensitivity.

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Wan, Xing; Usvalampi, Anne M; Saris, Per E J; Takala, Timo M

2016-11-01

In this paper, we present a new counterselection method for deleting fragments from Lactococcus lactis chromosome. The method uses a non-replicating plasmid vector, which integrates into the chromosome and makes the cell sensitive to bacteriocins. The integration vector carries pUC ori functional in Escherichia coli but not in L. lactis, an erythromycin resistance gene for selecting single crossover integrants, and two fragments from L. lactis chromosome for homologous recombinations. In addition, the integration vector is equipped with the Listeria monocytogenes gene mptC encoding the mannose-phosphotransferase system component IIC, the receptor for class IIa bacteriocins. Expression of mptC from the integration vector renders the naturally resistant L. lactis sensitive to class IIa bacteriocins. This sensitivity is then used to select the double crossover colonies on bacteriocin agar. Only the cells which have regained the endogenous bacteriocin resistance through the loss of the mptC plasmid will survive. The colonies carrying the desired deletion can then be distinguished from the wild-type revertants by PCR. By using the class IIa bacteriocins leucocin A, leucocin C or pediocin AcH as the counterselective agents, we deleted 22- and 33-kb chromosomal fragments from the wild-type nisin producing L. lactis strain N8. In conclusion, this counterselection method presented here is a convenient, efficient and inexpensive technique to generate successive deletions in L. lactis chromosome.

Characterization of Partially Purified Bacteriocin Like Substance (BLIS Produced by Probiotic Lactobacillus Strains

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Saeed Ismail Khanian

2014-05-01

Full Text Available Background: There is an increasing interest in search for antimicrobial peptides (bacteriocins and bacteriocin-like compounds produced by lactic acid bacteria (LAB because of their potential to be used as antimicrobial agents for improving the safety of food products. Objectives: The main objective of study was to evaluate the antibacterial potential of locally isolated Lactic Acid bacteria (LAB and determine their bacteriocin producing ability in in-vitro conditions. Materials and Methods: The antibacterial activity of 77 isolated LAB strains was tested against a number of pathogens by well-diffusion method. The isolates demonstrating antimicrobial potential were selected and tested for the production of bacteriocin or bacteriocin like substance. The bacteriocin produced by two of the isolates were partially purified and characterized. Results: The results indicated the neutralized supernatant fluid of two of the isolates identified as L. brevis LB32 and L. pentosus LP05, were active against the growth of Listeria monocytogenes, Salmonella enteritidis, Shigella dysenteriae, Staphylococcus aureus and Streptococcus pneumoniae. Additionally, L. brevis LB32 was able to inhibit the growth of Salmonella typhi and Klebsiella pneumoniae, while, S. pnuemoniae and L. monocytogenes appeared to be the most sensitive strain as apparent by highest zone of inhibition against these pathogens, respectively. The antimicrobial activity in the supernatant fluids of the mentioned strains remained unaffected after treating with enzymes catalase, lipase and lysozyme, while were strongly sensitive to the action of proteolytic enzymes, suggesting the presence of bacteriocin like inhibitory substance (BLIS in the two isolates. The inhibitory substance produced by the two isolates appeared heat resistant and tolerated 100˚C and 121˚C for 55 minutes and 20 minutes, respectively. Partial purification of the concentrated culture supernatant fluids of L. brevis LB32 and L

Elimination of Listeria monocytogenes in sausage meat by combination treatment: Radiation and radiation-resistant bacteriocins

International Nuclear Information System (INIS)

Turgis, Mélanie; Stotz, Viviane; Dupont, Claude; Salmieri, Stéphane; Khan, Ruhul A.; Lacroix, Monique

2012-01-01

Two new bacteria were isolated from human feces and were designated MT 104 and MT 162. They were able to produce bacteriocins that are active against five strains of Listeria monocytogenes. Bacteriocins produced by these isolated strains had 100% and 82.35% residual activity when they were treated by gamma radiation at doses of 4 and 40 kGy, respectively. A reduction of 1.0, 1.5 and 3 log CFU/g of L. monocytogenes was observed in sausage meat when treated with bacteriocins from MT 104, MT 162, and nisin, respectively. For synergic effect, the D 10 value in presence of the bacteriocins produced by MT 104 showed a 1.08 fold increased relative sensitivity of L. monocytogenes as compared to control after 5 days. The highest synergic effect was observed in presence of nisin which led to 1.61 fold increased relative sensitivity. Combined treatments with nisin and γ-irradiation showed a synergic antimicrobial effect in meat after 24 h and 5 days of storage. A synergic effect was observed only after 5 days at 4 °C for the bacteriocin from MT 104, as compared to the bacteriocin produced by MT 162 that had only an additive antimicrobial effect in all conditions.

Elimination of Listeria monocytogenes in sausage meat by combination treatment: Radiation and radiation-resistant bacteriocins

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Turgis, Mélanie; Stotz, Viviane; Dupont, Claude; Salmieri, Stéphane; Khan, Ruhul A.; Lacroix, Monique

2012-08-01

Two new bacteria were isolated from human feces and were designated MT 104 and MT 162. They were able to produce bacteriocins that are active against five strains of Listeria monocytogenes. Bacteriocins produced by these isolated strains had 100% and 82.35% residual activity when they were treated by gamma radiation at doses of 4 and 40 kGy, respectively. A reduction of 1.0, 1.5 and 3 log CFU/g of L. monocytogenes was observed in sausage meat when treated with bacteriocins from MT 104, MT 162, and nisin, respectively. For synergic effect, the D10 value in presence of the bacteriocins produced by MT 104 showed a 1.08 fold increased relative sensitivity of L. monocytogenes as compared to control after 5 days. The highest synergic effect was observed in presence of nisin which led to 1.61 fold increased relative sensitivity. Combined treatments with nisin and γ-irradiation showed a synergic antimicrobial effect in meat after 24 h and 5 days of storage. A synergic effect was observed only after 5 days at 4 °C for the bacteriocin from MT 104, as compared to the bacteriocin produced by MT 162 that had only an additive antimicrobial effect in all conditions.

Improving safety of salami by application of bacteriocins produced by an autochthonous Lactobacillus curvatus isolate.

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de Souza Barbosa, Matheus; Todorov, Svetoslav Dimitrov; Ivanova, Iskra; Chobert, Jean-Marc; Haertlé, Thomas; de Melo Franco, Bernadette Dora Gombossy

2015-04-01

The aims of this study were to isolate LAB with anti-Listeria activity from salami samples, characterize the bacteriocin/s produced by selected isolates, semi-purify them and evaluate their effectiveness for the control of Listeria monocytogenes during manufacturing of salami in a pilot scale. Two isolates (differentiated by RAPD-PCR) presented activity against 22 out of 23 L. monocytogenes strains for bacteriocin MBSa2, while the bacteriocin MBSa3 inhibited all 23 strains in addition to several other Gram-positive bacteria for both antimicrobials and were identified as Lactobacillus curvatus based on 16S rRNA sequencing. A three-step purification procedure indicated that both strains produced the same two active peptides (4457.9 Da and 4360.1 Da), homlogous to sakacins P and X, respectively. Addition of the semi-purified bacteriocins produced by Lb. curvatus MBSa2 to the batter for production of salami, experimentally contaminated with L. monocytogenes (10(4)-10(5) CFU/g), caused 2 log and 1.5 log reductions in the counts of the pathogen in the product after 10 and 20 days respectively, highlighting the interest for application of these bacteriocins to improve safety of salami during its manufacture. Copyright © 2014 Elsevier Ltd. All rights reserved.

The structure of pyogenecin immunity protein, a novel bacteriocin-like immunity protein from streptococcus pyogenes.

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Chang, C.; Coggill, P.; Bateman, A.; Finn, R.; Cymborowski, M.; Otwinowski, Z.; Minor, W.; Volkart, L.; Joachimiak, A.; Wellcome Trust Sanger Inst.; Univ. of Virginia; UT Southwestern Medical Center

2009-12-17

Many Gram-positive lactic acid bacteria (LAB) produce anti-bacterial peptides and small proteins called bacteriocins, which enable them to compete against other bacteria in the environment. These peptides fall structurally into three different classes, I, II, III, with class IIa being pediocin-like single entities and class IIb being two-peptide bacteriocins. Self-protective cognate immunity proteins are usually co-transcribed with these toxins. Several examples of cognates for IIa have already been solved structurally. Streptococcus pyogenes, closely related to LAB, is one of the most common human pathogens, so knowledge of how it competes against other LAB species is likely to prove invaluable. We have solved the crystal structure of the gene-product of locus Spy-2152 from S. pyogenes, (PDB: 2fu2), and found it to comprise an anti-parallel four-helix bundle that is structurally similar to other bacteriocin immunity proteins. Sequence analyses indicate this protein to be a possible immunity protein protective against class IIa or IIb bacteriocins. However, given that S. pyogenes appears to lack any IIa pediocin-like proteins but does possess class IIb bacteriocins, we suggest this protein confers immunity to IIb-like peptides. Combined structural, genomic and proteomic analyses have allowed the identification and in silico characterization of a new putative immunity protein from S. pyogenes, possibly the first structure of an immunity protein protective against potential class IIb two-peptide bacteriocins. We have named the two pairs of putative bacteriocins found in S. pyogenes pyogenecin 1, 2, 3 and 4.

Novel bacteriocins from lactic acid bacteria (LAB): various structures and applications

OpenAIRE

Perez, Rodney H; Zendo, Takeshi; Sonomoto, Kenji

2014-01-01

Bacteriocins are heat-stable ribosomally synthesized antimicrobial peptides produced by various bacteria, including food-grade lactic acid bacteria (LAB). These antimicrobial peptides have huge potential as both food preservatives, and as next-generation antibiotics targeting the multiple-drug resistant pathogens. The increasing number of reports of new bacteriocins with unique properties indicates that there is still a lot to learn about this family of peptide antibiotics. In this review, we...

Screening and characterization of bacteriocins produced by some Strains of Lactobacillus spp isolated from Iranian Dairy products

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S Mirdamadi

2011-11-01

Full Text Available In this study, the inhibitory effects of bacteriocins of lactobacilli which were isolated from Iranian traditional dairy products was determined against known gram positive, gram negative and yeast by well diffusion technique. Among 8 isolates with higher capability of bacteriocin production, 2 isolates were selected for further investigations. The bacteriocins were purified by iso-propanol and ammonium sulfate precipitation following by dialysis and chromatography technique. The molecular weight of bacteriocins was determined as 45 to 66/2 KDa. by SDS-page electrophoresis. According to the results, the produced bacteriocins had more inhibition effect on Micrococcus luteus PTCC1169, Staphylococcus epidermidis PTCC1435 as well as Bacillus cereus PTCC1247 and with lesser degree of extent on Listeria monocytogenes PTCC 1301. Results also revealed that, Micrococcus luteus  was the most sensitive bacterium among indicator bacteria, while Candid albicans PTCC 5027 identified as the most resistance organism. This research showed that, bacteriocins produced by lactobacilli isolated from traditional dairy products have high potency to be used against microbial pathogens and could be applied as bio-preservative in food products.

Method for Rapid Purification of Class IIa Bacteriocins and Comparison of Their Activities

OpenAIRE

Guyonnet, D.; Fremaux, C.; Cenatiempo, Y.; Berjeaud, J. M.

2000-01-01

A three-step method was developed for the purification of mesentericin Y105 (60% yield) from the culture supernatant of Leuconostoc mesenteroides Y105. The same procedure was successfully applied to the purification of five other anti-Listeria bacteriocins identified by mass spectrometry. Specific activities of the purified bacteriocins were compared.

Optimization and partial characterization of bacteriocin produced by Lactobacillus bulgaricus -TLBFT06 isolated from Dahi.

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Mahmood, Talat; Masud, Tariq; Ali, Sartaj; Abbasi, Kashif Sarfraz; Liaquat, Muhammad

2015-03-01

Lactobacillus bulgaricus is one of the predominant lactic acid bacteria of dahi, conferring technological and functional attributes. In the present study thirty dahi samples were investigated for bacteriocin producing L. bulgaricus. Fourteen different isolates were obtained and five were scrutinized for antibacterial activities against food born pathogens. Amongst, a strain TLB06FT was found to have a wide array of antibacterial activities against Gram positive and negative bacteria was selected for further characterization. Growth media optimization for this strain revealed maximum bacteriocin production on MRS media supplemented with glucose (2%), sodium chloride (1%), Tween-80 (0.5%) and yeast extract (1 %). In addition, optimization of growth conditions revealed maximum bacteriocin production at pH 5.5 and temperature of 30-37°C. Bacteriocin showed thermo stability at 90°C and remained highly active in the pH range of 3.5-7.5, inactive by protein catalyzing enzymes and showed no change in activity (800AumL(-1)) when treated with organic solvents and surfactants. The obtained bacteriocin was purified to 1600AU mL(-1) by ammonium sulfate precipitation (80%) by using dialyzing tubing. In the same way, a single peak was obtained by RP-HPLC having antibacterial activity of 6400AU mL(-1). Thus, wild strains of L. bulgaricus have great potential for the production new and novel type of bacteriocins.

Role of acetate in production of an autoinducible Class IIa Bacteriocin in Carnobacterium piscicola A9b

DEFF Research Database (Denmark)

Nilsson, Lilian; Nielsen, Michael Krogsgaard; Ng, Yin

2002-01-01

was to purify the compound and describe factors affecting its production, with particular emphasis on food-relevant factors. Amino acid sequencing showed that the compound is a class IIa bacteriocin with an N-terminal amino acid sequence identical to that of carnobacteriocin B2. The production....... The induction of bacteriocin production showed a dose-dependent relationship at acetate concentrations of up to 10 to 20 mM (depending on the growth medium) and at a concentration of 1.9 x 10(-8) M for the bacteriocin itself; a saturation level of bacteriocin specific activity was reached...

Gene Cluster Responsible for Secretion of and Immunity to Multiple Bacteriocins, the NKR-5-3 Enterocins

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Ishibashi, Naoki; Himeno, Kohei; Masuda, Yoshimitsu; Perez, Rodney Honrada; Iwatani, Shun; Wilaipun, Pongtep; Leelawatcharamas, Vichien; Nakayama, Jiro; Sonomoto, Kenji

2014-01-01

Enterococcus faecium NKR-5-3, isolated from Thai fermented fish, is characterized by the unique ability to produce five bacteriocins, namely, enterocins NKR-5-3A, -B, -C, -D, and -Z (Ent53A, Ent53B, Ent53C, Ent53D, and Ent53Z). Genetic analysis with a genome library revealed that the bacteriocin structural genes (enkA [ent53A], enkC [ent53C], enkD [ent53D], and enkZ [ent53Z]) that encode these peptides (except for Ent53B) are located in close proximity to each other. This NKR-5-3ACDZ (Ent53ACDZ) enterocin gene cluster (approximately 13 kb long) includes certain bacteriocin biosynthetic genes such as an ABC transporter gene (enkT), two immunity genes (enkIaz and enkIc), a response regulator (enkR), and a histidine protein kinase (enkK). Heterologous-expression studies of enkT and ΔenkT mutant strains showed that enkT is responsible for the secretion of Ent53A, Ent53C, Ent53D, and Ent53Z, suggesting that EnkT is a wide-range ABC transporter that contributes to the effective production of these bacteriocins. In addition, EnkIaz and EnkIc were found to confer self-immunity to the respective bacteriocins. Furthermore, bacteriocin induction assays performed with the ΔenkRK mutant strain showed that EnkR and EnkK are regulatory proteins responsible for bacteriocin production and that, together with Ent53D, they constitute a three-component regulatory system. Thus, the Ent53ACDZ gene cluster is essential for the biosynthesis and regulation of NKR-5-3 enterocins, and this is, to our knowledge, the first report that demonstrates the secretion of multiple bacteriocins by an ABC transporter. PMID:25149515

ANTIMICROBIAL ACTIVITY OF BACTERIOCIN FROM INDIGENOUS Lactobacillus plantarum 2C12 AND ITS APPLICATION ON BEEF MEATBALL AS BIOPRESERVATIVE

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I.I. Arief

2014-10-01

Full Text Available One purpose of food preservation is to extend the shelf life of foods. Biological preservations canbe conducted by adding antimicrobial substances, such as bacteriocin produced by lactic acid bacteriaand has been characterized as biopreservatives. The aims of this research were to evaluate antimicrobialactivity of bacteriocin produced by indigenous lactic acid bacteria Lactobacillus plantarum 2C12isolated from local beef and to study the quality of beef meatball with 0.3% bacteriocin asbiopreservative at different storage times (0, 3, and 6 days in cold temperature (4oC, compared to 0.3%nitrite and control (without preservative. The results showed that bacteriocin from L. plantarum 2C12could inhibit pathogenic bacteria such as Escherichia coli, Staphylococcus aureus and SalmonellaTyphimurium. Bacteriocin was effective as well as nitrite as biopreservatives of meatballs by inhibitingthe growth of total microbes and E. coli. The addition of bacteriocin did not lead the physical andnutritional changes in the meatballs. The quality of meatball with bacteriocin treatment conformed withIndonesia National Standard of meatball.

ANTIMICROBIAL ACTIVITY OF BACTERIOCIN FROM INDIGENOUS Lactobacillus plantarum 2C12 AND ITS APPLICATION ON BEEF MEATBALL AS BIOPRESERVATIVE

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T. Suryati

2012-06-01

Full Text Available One purpose of food preservation is to extend the shelf life of foods. Biological preservations can be conducted by adding antimicrobial substances, such as bacteriocin produced by lactic acid bacteria and has been characterized as biopreservatives. The aims of this research were to evaluate antimicrobial activity of bacteriocin produced by indigenous lactic acid bacteria Lactobacillus plantarum 2C12 isolated from local beef and to study the quality of beef meatball with 0.3% bacteriocin as biopreservative at different storage times (0, 3, and 6 days in cold temperature (4oC, compared to 0.3% nitrite and control (without preservative. The results showed that bacteriocin from L. plantarum 2C12 could inhibit pathogenic bacteria such as Escherichia coli, Staphylococcus aureus and Salmonella Typhimurium. Bacteriocin was effective as well as nitrite as biopreservatives of meatballs by inhibiting the growth of total microbes and E. coli. The addition of bacteriocin did not lead the physical and nutritional changes in the meatballs. The quality of meatball with bacteriocin treatment conformed with Indonesia National Standard of meatball.

Method for Rapid Purification of Class IIa Bacteriocins and Comparison of Their Activities

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Guyonnet, D.; Fremaux, C.; Cenatiempo, Y.; Berjeaud, J. M.

2000-01-01

A three-step method was developed for the purification of mesentericin Y105 (60% yield) from the culture supernatant of Leuconostoc mesenteroides Y105. The same procedure was successfully applied to the purification of five other anti-Listeria bacteriocins identified by mass spectrometry. Specific activities of the purified bacteriocins were compared. PMID:10742275

Coculture-inducible bacteriocin biosynthesis of different probiotic strains by dairy starter culture Lactococcus lactis

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Blaženka Kos

2011-12-01

Full Text Available Bacteriocins produced by probiotic strains effectively contribute to colonization ability of probiotic strains and facilitate their establishment in the competitive gut environment and also protect the gut from gastrointestinal pathogens. Moreover, bacteriocins have received considerable attention due to their potential application as biopreservatives, especially in dairy industry. Hence, the objective of this research was to investigate antimicrobial activity of probiotic strains Lactobacillus helveticus M92, Lactobacillus plantarum L4 and Enterococcus faecium L3, with special focus on their bacteriocinogenic activity directed towards representatives of the same or related bacterial species, and towards distant microorganisms including potential food contaminants or causative agents of gut infections. In order to induce bacteriocin production, probiotic cells were cocultivated with Lactococcus lactis subsp. lactis LMG 9450, one of the most important starter cultures in cheese production. The presence of bacteriocin coding genes was investigated by PCR amplification with sequence-specific primers for helveticin and was confirmed for probiotic strain L. helveticus M92. All examined probiotic strains have shown bacteriocinogenic activity against Staphylococcus aureus 3048, Staphylococcus aureus K-144, Escherichia coli 3014, Salmonella enterica serovar Typhimurium FP1, Bacillus subtilis ATCC 6633, Bacillus cereus TM2, which is an important functional treat of probiotic strains significant in competitive exclusion mechanism which provides selective advantage of probiotic strains against undesirable microorganisms in gastrointestinal tract of the host. According to obtained results, living cells of starter culture Lc. lactis subsp. lactis LMG 9450 induced bacteriocin production by examined probiotic strains but starter culture itself was not sensitive to bacteriocin activity.

Purification and Characterization of Bacteriocin Produced by Weissella confusa A3 of Dairy Origin.

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Goh, Hweh Fen; Philip, Koshy

2015-01-01

A dramatic increase in bacterial resistance towards currently available antibiotics has raised worldwide concerns for public health. Therefore, antimicrobial peptides (AMPs) have emerged as a promisingly new group of therapeutic agents for managing infectious diseases. The present investigation focusses on the isolation and purification of a novel bacteriocin from an indigenous sample of cow milk and it's mode of action. The bacteriocin was isolated from Weissella confusa A3 that was isolated from the sample and was shown to have inhibitory activity towards pathogenic bacteria namely Bacillus cereus, Escherichia coli, Pseudomonas aeruginosa and Micrococcus luteus. The bacteriocin was shown to be heat stable and functioned well at low pH (2 to 6). Reduction of activity was shown after treatment with proteinase K, trypsin and peptidase that confirmed the proteinaceous nature of the compound. MALDI-TOF analysis of the sample gave a mass approximating 2.7 kDa. The membrane of the bacteria was disrupted by the bacteriocin causing SYTOX® green dye to enter the cell and bind to the bacterial DNA giving fluorescence signal. Bacterial cell treated with the bacteriocin also showed significant morphological changes under transmission electron microscope. No virulence and disease related genes can be detected from the genome of the strain.

Purification and Characterization of Bacteriocin Produced by Weissella confusa A3 of Dairy Origin

Science.gov (United States)

Goh, Hweh Fen; Philip, Koshy

2015-01-01

A dramatic increase in bacterial resistance towards currently available antibiotics has raised worldwide concerns for public health. Therefore, antimicrobial peptides (AMPs) have emerged as a promisingly new group of therapeutic agents for managing infectious diseases. The present investigation focusses on the isolation and purification of a novel bacteriocin from an indigenous sample of cow milk and it’s mode of action. The bacteriocin was isolated from Weissella confusa A3 that was isolated from the sample and was shown to have inhibitory activity towards pathogenic bacteria namely Bacillus cereus, Escherichia coli, Pseudomonas aeruginosa and Micrococcus luteus. The bacteriocin was shown to be heat stable and functioned well at low pH (2 to 6). Reduction of activity was shown after treatment with proteinase K, trypsin and peptidase that confirmed the proteinaceous nature of the compound. MALDI-TOF analysis of the sample gave a mass approximating 2.7 kDa. The membrane of the bacteria was disrupted by the bacteriocin causing SYTOX® green dye to enter the cell and bind to the bacterial DNA giving fluorescence signal. Bacterial cell treated with the bacteriocin also showed significant morphological changes under transmission electron microscope. No virulence and disease related genes can be detected from the genome of the strain. PMID:26474074

Coculture-inducible bacteriocin activity of Lactobacillus plantarum strain J23 isolated from grape must.

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Rojo-Bezares, Beatriz; Sáenz, Yolanda; Navarro, Laura; Zarazaga, Myriam; Ruiz-Larrea, Fernanda; Torres, Carmen

2007-08-01

Detection and characterization of bacteriocin production by Lactobacillus plantarum strain J23, recovered from a grape must sample in Spain, have been carried out. Bacteriocin activity was degraded by proteolytic enzymes (trypsin, alfa-chymotrypsin, papaine, protease, proteinase K and acid proteases), and it was stable at high temperatures (121 degrees C, 20min), in a wide range of pH (1-12), and after treatment with organic solvents. L. plantarum J23 showed antimicrobial activity against Oenococcus oeni, and a range of Lactobacillus and Pediococcus species. Bacteriocin production was detected in liquid media only when J23 was cocultivated with some inducing bacteria, and induction took place when intact cells or 55 degrees C heated cells of the inducer were cocultivated with J23, but not with their autoclaved cells. Bacteriocin activity of J23 was not induced by high initial J23 inocula, and it was detected in cocultures during the exponential phase. The presence of ethanol or acidic pH in the media reduced bacteriocin production in the cocultures of J23 with the inducing bacteria. The presence of plantaricin-related plnEF and plnJ genes was detected by PCR and sequencing. Nevertheless, negative results were obtained for plnA, plnK, plNC8, plS and plW genes.

Antimicrobial Effect of Bacteriocin produced Pediococcus pentosaceus on some clinical isolates

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Nehad A. Taher

2017-07-01

Full Text Available About 10 isolates of Pediococcus sp were isolated from different cheese made in Iraq, These isolates were identified morphologically and biochemically and Api20 kit, thus there was only 6 isolate were identified as Pediococcus pentosaceus (60%.In this study, we investigate, the effect of crude Bacteriocin from Pediococcus pentosaceus on 30 clinical isolates (5 E.coli, 5 Klepsiella pneumoniae, 5 Staphylococcus aureus, 5 Pseudomonas aeroginosa, 5 Bacillus subtilis, 5 Candida albicans. The protein concentration of this Bacteriocin was measured 67mg\\ml by Bradford method and used as (1:2 by vol during the measuring the antimicrobial activity against the above clinical isolates by two methods wells and  agar plug assay. The results showed that  the inhibitory activity of this Bacteriocin was higher by wells method than agar pluq assay against Gram–positive bacteria or Gram-negative bacteria and yeast under this study.

EFFECT OF CULTURE MEDIUM ON BACTERIOCIN PRODUCTION BY LACTOBACILLUS RHAMNOSUS HN001 AND LACTOBACILLUS REUTERI ATCC 53608

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Aguilar-Uscanga B. R.

2013-06-01

Full Text Available The aim of this study was to evaluate the effect of media on bacteriocin production by Lactobacillus rhamnosus HN001 and Lactobacillus reuteri ATCC 53608 using three different media: YPM, YPF and MRS supplemented with glucose and K2HPO4. The optimum temperature was 37°C and initial pH 6.5. Bacteriocin-like substances produced by tested bacteria in MRS medium supplemented with glucose and K2HPO4 exhibited a broad antimicrobial spectrum determined by well diffusion assay against indicator bacteria Listeria monocytogenes, Lactobacillus sakei, Enterococcus faecium, Lactobacillus delbrueckii, Lactobacillus acidophilus, but no antimicrobial spectrum against E. coli O157:H7, Salmonella typhimurium, Staphylococcus aureus, Bacillus cereus was detected. Bacteriocin was sensitive to protease IV, trypsin, pepsin and -amylases, but resistant to lipase. It was also resistant to detergents such as Tween 80, Triton-X and SDS. This bacteriocin was thermo-stable (resistant at 60°C, 90°C and 100°C for 30 min. Tested bacteria showed the best antimicrobial (bacteriocin-like activity after growth in MRS medium. Bacteriocin substances produced by tested bacteria showed promising thermo-stable technological properties.

Interactions of meat-associated bacteriocin-producing Lactobacilli with Listeria innocua under stringent sausage fermentation conditions.

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Leroy, Frédéric; Lievens, Kristoff; De Vuyst, Luc

2005-10-01

The kinetics of the antilisterial effect of meat-associated lactobacilli on Listeria innocua LMG 13568 were investigated during laboratory batch fermentations. During these fermentations, which were performed in a liquid meat simulation medium, a combination of process factors typical for European-style sausage fermentations was applied, such as a temperature of 20 degrees C and a representative pH and salting profile. Two bacteriocin-producing sausage isolates (Lactobacillus sakei CTC 494 and Lactobacillus curvatus LTH 1174), which have already proven efficacy in sausage trials, and one nonbacteriocinogenic, industrial strain (Lactobacillus sakei I), were evaluated. Staphylococcus carnosus 833 was included in the experiment because of its role in flavor and color development. When grown as a monoculture or upon cocultivation with L. sakei I and S. carnosus 833, L. innocua LMG 13568 developed slightly, despite the stress of low temperature, pH, lactic acid, salt, and nitrite. In contrast, when either of the bacteriocin producers was used, the L. innocua LMG 13568 population was rapidly inactivated with more than 3 log CFU ml(-1) after 2 days of fermentation. A bacteriocin-tolerant L. innocua LMG 13568 subpopulation (4 X 10(-4)) remained after bacteriocin inactivation. Thus, when the initial level of L. innocua LMG 13568 equaled 3 log CFU ml(-1), all cells were inactivated and no bacteriocin-tolerant cells were detected, even after 7 days of incubation. S. carnosus was not inactivated by the Lactobacillus bacteriocins and displayed slight growth.

Influence of Growth Medium on Hydrogen Peroxide and Bacteriocin Production of Lactobacillus Strains

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Edina Németh

2005-01-01

Full Text Available This study was conducted to investigate the inhibitory effect of bacteriocin and the production of hydrogen peroxide by four non-starter lactic acid bacteria, Lactobacillus plantarum 2142, Lactobacillus curvatus 2770, Lactobacillus curvatus 2775, Lactobacillus casei subsp. pseudoplantarum 2750 and the probiotic strain Lactobacillus casei Shirota, propagated in de Man Rogosa Sharpe (MRS and tomato juice (TJ broth. The methods were a commonly used agar diffusion technique and a microtiter assay method. The best peroxide-producing Lactobacillus strain was selected for screening the inhibitory activity against Listeria monocytogenes, Bacillus cereus, Escherichia coli and the activity of bacteriocins against Lactobacillus sakei and Candida glabrata. All of the investigated lactic acid bacteria (LAB strains grown in MRS broth produced the highest concentration of hydrogen peroxide ranging from 2–6 g/mL after 72 h of storage. L. plantarum 2142 produced enough hydrogen peroxide already after 24 h at 5 °C in phosphate buffer to inhibit the growth of L. monocytogenes and B. cereus. Crude bacteriocin suspension from the investigated LAB inhibited only slightly the growth of L. sakei, however, the same suspension from MRS completely inhibited the 6-fold diluted yeast suspension. The concentrated bacteriocin suspensions from the both broths inhibited the growth of L. sakei completely. Among the strains, L. plantarum 2142 seemed to be the best peroxide and bacteriocin producer, and the antimicrobial metabolite production was better in MRS than in TJ broth.

Cloning, overexpression, purification of bacteriocin enterocin-B and structural analysis, interaction determination of enterocin-A, B against pathogenic bacteria and human cancer cells.

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Ankaiah, Dasari; Palanichamy, Esakkiraj; Antonyraj, Christian Bharathi; Ayyanna, Repally; Perumal, Venkatesh; Ahamed, Syed Ibrahim Basheer; Arul, Venkatesan

2018-05-02

In this present study, a gene (ent-B) encoding the bacteriocin enterocin-B was cloned, overexpressed and purified from Enterococcus faecium por1. The molecular weight of the bacteriocin enterocin-B was observed around 7.2 kDa and exhibited antimicrobial activity against several human pathogenic bacteria. The antimicrobial activity of cloned enterocin-B was increased effectively by combining with another bacteriocin enterocin-A from the same microorganism. Protein-protein docking and molecular dynamics simulation studies revealed that the bacteriocin enterocin-B is interacting with enterocin-A and formation of a heterodimer (enterocin A + B). The heterodimer of bacteriocin enterocin-A + B exhibited potential anti-bacterial, anti-biofilm activity against Staphylococcus aureus, Acinetobacter baumannii, Listeria monocytogenes and Escherichia coli. The bacteriocin enterocin-B, A and heterodimer of bacteriocin enterocin A + B showed no haemolysis on human RBC cells. This is the first report that the cell growth inhibitory activity of the bacteriocin enterocin B against HeLa, HT-29 and AGS human cancer cells and this cell growth inhibitory activity was significantly increased when cancer cells treated with the heterodimer of bacteriocins enterocin-A + B. The cell growth inhibitory activity of the bacteriocin enterocin-B and the heterodimer of bacteriocin enterocin-A + B were not observed in non-cancerous INT-407 cells (intestinal epithelial cells). Copyright © 2018. Published by Elsevier B.V.

HPLC purification and re-evaluation of chemical identity of two circular bacteriocins, gassericin A and reutericin 6.

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Arakawa, K; Kawai, Y; Ito, Y; Nakamura, K; Chujo, T; Nishimura, J; Kitazawa, H; Saito, T

2010-04-01

The study aimed for the complete purification and recharacterization of the highly hydrophobic circular bacteriocins, gassericin A and reutericin 6. Gassericin A and reutericin 6 were purified to homogeneity using previously described method and reverse-phase HPLC with an octyl column and eluents of aqueous acetonitrile and 2-propanol. Mass analysis, N-terminal sequencing and bacteriocin assay of the HPLC-purified bacteriocins showed the two bacteriocins had identical seamless circular structures with the same m/z value (5651) of [M + H](+) and both had the same specific activity. D/L-amino acid composition analysis using two distinct methods with the chiral fluorescent derivatization reagents (+)-1-(9-fluorenyl)ethyl chloroformate and O-phthalaldehyde/N-acetyl-L-cystein revealed neither gassericin A nor reutericin 6 contained D-alanine residues contrary to our previous results. Purified gassericin A and reutericin 6 are chemically identical circular molecules containing no D-alanine residues. The HPLC conditions developed in this study will facilitate advanced purification and correct characterization of other highly hydrophobic bacteriocins.

Modeling Bacteriocin Resistance and Inactivation of Listeria innocua LMG 13568 by Lactobacillus sakei CTC 494 under Sausage Fermentation Conditions

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Leroy, Frédéric; Lievens, Kristoff; De Vuyst, Luc

2005-01-01

In mixed cultures, bacteriocin production by the sausage isolate Lactobacillus sakei CTC 494 rapidly inactivated sensitive Listeria innocua LMG 13568 cells, even at low bacteriocin activity levels. A small fraction of the listerial population was bacteriocin resistant. However, sausage fermentation conditions inhibited regrowth of resistant cells. PMID:16269805

Screening and characterization of bacteriocins produced by some Strains of Lactobacillus spp isolated from Iranian Dairy products

OpenAIRE

S Mirdamadi; M Tangestani

2011-01-01

In this study, the inhibitory effects of bacteriocins of lactobacilli which were isolated from Iranian traditional dairy products was determined against known gram positive, gram negative and yeast by well diffusion technique. Among 8 isolates with higher capability of bacteriocin production, 2 isolates were selected for further investigations. The bacteriocins were purified by iso-propanol and ammonium sulfate precipitation following by dialysis and chromatography technique. The molecular we...

Growing insights into the safety of bacteriocins: the case of enterocin S37.

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Belguesmia, Yanath; Madi, Amar; Sperandio, Daniel; Merieau, Annabelle; Feuilloley, Marc; Prévost, Hervé; Drider, Djamel; Connil, Nathalie

2011-01-01

Very few studies have been reported on the cytotoxicity and impact of bacteriocins, and especially enterocins, upon eukaryotic cells. In order to gain more information on the safety of bacteriocins, we focused this study on enterocin S37, a bacteriocin produced by Enterococcus faecalis S37. We observed dose-dependent cytotoxicity toward undifferentiated Caco-2/TC7 cells. Moreover, no significant effect on differentiated monolayer Caco-2/TC7 and no apoptotic features were observed when cells were treated with 10 μg/ml of enterocin S37. The results obtained indicate possible safe use of enterocin S37 in the gastrointestinal tract of animals to prevent pathogen invasion and/or infection. Copyright © 2010 Institut Pasteur. Published by Elsevier SAS. All rights reserved.

Bacteriocinogenic Lactococcus lactis subsp. lactis DF04Mi isolated from goat milk: characterization of the bacteriocin

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Danielle N. Furtado

2014-12-01

Full Text Available Lactic acid bacteria capable of producing bacteriocins and presenting probiotic potential open innovative technological applications in the dairy industry. In this study, a bacteriocinogenic strain (Lactococcus lactis subsp. lactis DF4Mi was isolated from goat milk, and studied for its antimicrobial activity. The bacteriocin presented a broad spectrum of activity, was sensitive to proteolytic enzymes, resistant to heat and pH extremes, and not affected by the presence of SDS, Tween 20, Tween 80, EDTA or NaCl. Bacteriocin production was dependent on the components of the culture media, especially nitrogen source and salts. When tested by PCR, the bacteriocin gene presented 100% homology to nisin Z gene. These properties indicate that this L. lactis subsp. lactis DF4Mi can be used for enhancement of dairy foods safety and quality.

Bacteriocinogenic Lactococcus lactis subsp. lactis DF04Mi isolated from goat milk: Characterization of the bacteriocin

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Furtado, Danielle N.; Todorov, Svetoslav D.; Landgraf, Mariza; Destro, Maria T.; Franco, Bernadette D.G.M.

2014-01-01

Lactic acid bacteria capable of producing bacteriocins and presenting probiotic potential open innovative technological applications in the dairy industry. In this study, a bacteriocinogenic strain (Lactococcus lactis subsp. lactis DF4Mi) was isolated from goat milk, and studied for its antimicrobial activity. The bacteriocin presented a broad spectrum of activity, was sensitive to proteolytic enzymes, resistant to heat and pH extremes, and not affected by the presence of SDS, Tween 20, Tween 80, EDTA or NaCl. Bacteriocin production was dependent on the components of the culture media, especially nitrogen source and salts. When tested by PCR, the bacteriocin gene presented 100% homology to nisin Z gene. These properties indicate that this L. lactis subsp. lactis DF4Mi can be used for enhancement of dairy foods safety and quality. PMID:25763065

Sensitivity to the two-peptide bacteriocin lactococcin G is dependent on UppP, an enzyme involved in cell-wall synthesis

NARCIS (Netherlands)

Kjos, Morten; Oppegård, Camilla; Diep, Dzung B; Nes, Ingolf F; Veening, Jan-Willem; Nissen-Meyer, Jon; Kristensen, Tom

Most bacterially produced antimicrobial peptides (bacteriocins) are thought to kill target cells by a receptor-mediated mechanism. However, for most bacteriocins the receptor is unknown. For instance, no target receptor has been identified for the two-peptide bacteriocins (class IIb), whose activity

Distinct colicin M-like bacteriocin-immunity pairs in Burkholderia.

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Ghequire, Maarten G K; De Mot, René

2015-11-27

The Escherichia coli bacteriocin colicin M (ColM) acts via degradation of the cell wall precursor lipid II in target cells. ColM producers avoid self-inhibition by a periplasmic immunity protein anchored in the inner membrane. In this study, we identified colM-like bacteriocin genes in genomes of several β-proteobacterial strains belonging to the Burkholderia cepacia complex (Bcc) and the Burkholderia pseudomallei group. Two selected Burkholderia ambifaria proteins, designated burkhocins M1 and M2, were produced recombinantly and showed antagonistic activity against Bcc strains. In their considerably sequence-diverged catalytic domain, a conserved aspartate residue equally proved pivotal for cytotoxicity. Immunity to M-type burkhocins is conferred upon susceptible strains by heterologous expression of a cognate gene located either upstream or downstream of the toxin gene. These genes lack homology with currently known ColM immunity genes and encode inner membrane-associated proteins of two distinct types, differing in predicted transmembrane topology and moiety exposed to the periplasm. The addition of burkhocins to the bacteriocin complement of Burkholderia reveals a wider phylogenetic distribution of ColM-like bacteriotoxins, beyond the γ-proteobacterial genera Escherichia, Pectobacterium and Pseudomonas, and illuminates the diversified nature of immunity-providing proteins.

A Microplate Growth Inhibition Assay for Screening Bacteriocins against Listeria monocytogenes to Differentiate Their Mode-of-Action

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Paul Priyesh Vijayakumar

2015-06-01

Full Text Available Lactic acid bacteria (LAB have historically been used in food fermentations to preserve foods and are generally-recognized-as-safe (GRAS by the FDA for use as food ingredients. In addition to lactic acid; some strains also produce bacteriocins that have been proposed for use as food preservatives. In this study we examined the inhibition of Listeria monocytogenes 39-2 by neutralized and non-neutralized bacteriocin preparations (Bac+ preps produced by Lactobacillus curvatus FS47; Lb. curvatus Beef3; Pediococcus acidilactici Bac3; Lactococcus lactis FLS1; Enterococcus faecium FS56-1; and Enterococcus thailandicus FS92. Activity differences between non-neutralized and neutralized Bac+ preps in agar spot assays could not readily be attributed to acid because a bacteriocin-negative control strain was not inhibitory to Listeria in these assays. When neutralized and non-neutralized Bac+ preps were used in microplate growth inhibition assays against L. monocytogenes 39-2 we observed some differences attributed to acid inhibition. A microplate growth inhibition assay was used to compare inhibitory reactions of wild-type and bacteriocin-resistant variants of L. monocytogenes to differentiate bacteriocins with different modes-of-action (MOA whereby curvaticins FS47 and Beef3, and pediocin Bac3 were categorized to be in MOA1; enterocins FS92 and FS56-1 in MOA2; and lacticin FLS1 in MOA3. The microplate bacteriocin MOA assay establishes a platform to evaluate the best combination of bacteriocin preparations for use in food applications as biopreservatives against L. monocytogenes.

A Microplate Growth Inhibition Assay for Screening Bacteriocins against Listeria monocytogenes to Differentiate Their Mode-of-Action.

Science.gov (United States)

Vijayakumar, Paul Priyesh; Muriana, Peter M

2015-06-11

Lactic acid bacteria (LAB) have historically been used in food fermentations to preserve foods and are generally-recognized-as-safe (GRAS) by the FDA for use as food ingredients. In addition to lactic acid; some strains also produce bacteriocins that have been proposed for use as food preservatives. In this study we examined the inhibition of Listeria monocytogenes 39-2 by neutralized and non-neutralized bacteriocin preparations (Bac+ preps) produced by Lactobacillus curvatus FS47; Lb. curvatus Beef3; Pediococcus acidilactici Bac3; Lactococcus lactis FLS1; Enterococcus faecium FS56-1; and Enterococcus thailandicus FS92. Activity differences between non-neutralized and neutralized Bac+ preps in agar spot assays could not readily be attributed to acid because a bacteriocin-negative control strain was not inhibitory to Listeria in these assays. When neutralized and non-neutralized Bac+ preps were used in microplate growth inhibition assays against L. monocytogenes 39-2 we observed some differences attributed to acid inhibition. A microplate growth inhibition assay was used to compare inhibitory reactions of wild-type and bacteriocin-resistant variants of L. monocytogenes to differentiate bacteriocins with different modes-of-action (MOA) whereby curvaticins FS47 and Beef3, and pediocin Bac3 were categorized to be in MOA1; enterocins FS92 and FS56-1 in MOA2; and lacticin FLS1 in MOA3. The microplate bacteriocin MOA assay establishes a platform to evaluate the best combination of bacteriocin preparations for use in food applications as biopreservatives against L. monocytogenes.

Examination of Lactic Acid Bacteria to Secretion of Bacteriocins

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Maira Urazova

2014-01-01

Full Text Available Introduction: Bacteriocins produced by lactic acid bacteria (LAB have the potential to cover a very broad field of applications, including the food industry and the medical sector. In the food industry, bacteriocinogenic LAB strains can be used as starter cultures, co-cultures, and bioprotective cultures, which would be used to improve food quality and safety. In the medical sector, bacteriocins of probiotic LAB might play a role in interactions, which take place in human gastrointestinal tract, and contribute to gut health. The aim of this study was the examine the effect of LAB antimicrobial activity. Methods: LAB were isolated from different commercial and home made products, such as kazy and sour cream. To screen for bacteriocin producing LAB, we used an agar diffusion bioassay, described in a previous study by Dr. Yang, with three modifications in cell-free supernatant (CFS. First we had a clear supernatant, second we adjusted the CFS to pH 6.0 to eliminate acids antimicrobial effects, and third the CFS pH 6.0 was treated with catalase to exclude the action of H2O2 and confirm action of bacteriocin-like substances. Pathogenic S.marcescens, E. coli, S.aureus cultures were used as indicators. Results: Screening of 95 strains of LAB through deferred antagonism to six indicator cultures showed that all of the selected strains had a high value of antibacterial activity. However, CFS of only 50 strains retained their antimicrobial activity, and 10 of them lost this activity in the second modification of CFS with pH 6.0 to test culture S.marcescens, which confirmed the acidic nature of antimicrobial activity of CFS. Lb.rhamnosus (P-1, Lb.fermentum (N-6, and Lc.lactis (7M lost antibacterial activity in the presence of the catalase. All modifications of CFS of three strains: Lb.pentosus (16al, Lb.pentosus (P-2, and Pediococcusacidilactici (8 retained inhibitory activity to E.coli and S. aureus. Supernatants of only Lactococcusgarvieae (10a and

Effect of Lactobacillus salivarius bacteriocin Abp118 on the mouse and pig intestinal microbiota.

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Eliette Riboulet-Bisson

Full Text Available Lactobacilli are gram-positive bacteria that are a subdominant element in the human gastrointestinal microbiota, and which are commonly used in the food industry. Some lactobacilli are considered probiotic, and have been associated with health benefits. However, there is very little culture-independent information on how consumed probiotic microorganisms might affect the entire intestinal microbiota. We therefore studied the impact of the administration of Lactobacillus salivarius UCC118, a microorganism well characterized for its probiotic properties, on the composition of the intestinal microbiota in two model animals. UCC118 has anti-infective activity due to production of the bacteriocin Abp118, a broad-spectrum class IIb bacteriocin, which we hypothesized could impact the microbiota. Mice and pigs were administered wild-type (WT L. salivarius UCC118 cells, or a mutant lacking bacteriocin production. The microbiota composition was determined by pyrosequencing of 16S rRNA gene amplicons from faeces. The data show that L. salivarius UCC118 administration had no significant effect on proportions of major phyla comprising the mouse microbiota, whether the strain was producing bacteriocin or not. However, L. salivarius UCC118 WT administration led to a significant decrease in Spirochaetes levels, the third major phylum in the untreated pig microbiota. In both pigs and mice, L. salivarius UCC118 administration had an effect on Firmicutes genus members. This effect was not observed when the mutant strain was administered, and was thus associated with bacteriocin production. Surprisingly, in both models, L. salivarius UCC118 administration and production of Abp118 had an effect on gram-negative microorganisms, even though Abp118 is normally not active in vitro against this group of microorganisms. Thus L. salivarius UCC118 administration has a significant but subtle impact on mouse and pig microbiota, by a mechanism that seems at least partially

Inhibitory effect of bacteriocin-producing lactic acid bacteria against histamine-forming bacteria isolated from Myeolchi-jeot

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Eun-Seo Lim

2016-12-01

Full Text Available Abstract The objectives of this study were to identify the histamine-forming bacteria and bacteriocin- producing lactic acid bacteria (LAB isolated from Myeolchi-jeot according to sequence analysis of the 16S rRNA gene, to evaluate the inhibitory effects of the bacteriocin on the growth and histamine accumulation of histamine-forming bacteria, and to assess the physico-chemical properties of the bacteriocin. Based on 16S rRNA gene sequences, histamine-forming bacteria were identified as Bacillus licheniformis MCH01, Serratia marcescens MCH02, Staphylococcus xylosus MCH03, Aeromonas hydrophila MCH04, and Morganella morganii MCH05. The five LAB strains identified as Pediococcus acidilactici MCL11, Leuconostoc mesenteroides MCL12, Enterococcus faecium MCL13, Lactobacillus sakei MCL14, and Lactobacillus acidophilus MCL15 were found to produce an antibacterial compound with inhibitory activity against the tested histamine-producing bacteria. The inhibitory activity of these bacteriocins obtained from the five LAB remained stable after incubation at pH 4.0–8.0 and heating for 10 min at 80 °C; however, the bacteriocin activity was destroyed after treatment with papain, pepsin, proteinase K, α-chymotrypsin, or trypsin. Meanwhile, these bacteriocins produced by the tested LAB strains also exhibited histamine-degradation ability. Therefore, these antimicrobial substances may play a role in inhibiting histamine formation in the fermented fish products and preventing seafood-related food-borne disease caused by bacterially generated histamine.

A Novel High-Molecular-Mass Bacteriocin Produced by Enterococcus faecium: Biochemical Features and Mode of Action.

Science.gov (United States)

Vasilchenko, A S; Vasilchenko, A V; Valyshev, A V; Rogozhin, E A

2018-02-08

Discovery of a novel bacteriocin is always an event in sciences, since cultivation of most bacterial species is a general problem in microbiology. This statement is reflected by the fact that number of bacteriocins is smaller for tenfold comparing to known antimicrobial peptides. We cultivated Enterococcus faecium on simplified medium to reduce amount of purification steps. This approach allows to purify the novel heavy weight bacteriocin produced by E. faecium ICIS 7. The novelty of this bacteriocin, named enterocin-7, was confirmed by N-terminal sequencing and by comparing the structural-functional properties with available data. Purified enterocin-7 is characterized by a sequence of amino acid residues having no homology in UniProt/SwissProt/TrEMBL databases: NH2 - Asp - Ala - His - Leu - Ser - Glu - Val - Ala - Glu - Arg - Phe - Glu - Asp - Leu - Gly. Isolated thermostable protein has a molecular mass of 65 kDa, which allows it to be classified into class III in bacteriocin classification schemes. Enterocin-7 displayed a broad spectrum of activity against some Gram-positive and Gram-negative microorganisms. Fluorescent microscopy and spectroscopy showed the permeabilizing mechanism of the action of enterocin-7, which is realized within a few minutes.

Improved adsorption-desorption extraction applied to the partial characterization of the antilisterial bacteriocin produced by Carnobacterium maltaromaticum C2

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F. L Tulini

2010-06-01

Full Text Available Bacteriocins are ribosomally produced peptides useful for food biopreservation. An improved adsorption-desorption process is proposed for the partial purification of the bacteriocin produced by the fish isolate Carnobacterium maltaromaticum C2. Analyzis of extract by SDS-PAGE indicated this method may offer an alternative to improve the yield of purification of bacteriocins.

Purification and antibacterial mechanism of fish-borne bacteriocin and its application in shrimp (Penaeus vannamei) for inhibiting Vibrio parahaemolyticus.

Science.gov (United States)

Lv, Xinran; Du, Jingfang; Jie, Yu; Zhang, Bolin; Bai, Fengling; Zhao, Hongfei; Li, Jianrong

2017-08-01

Vibrio parahaemolyticus: is recognized as the main cause of gastroenteritis associated with consumption of seafood. Bacteriocin-producing Lactobacillus plantarum FGC-12 isolated from golden carp intestine had strong antibacterial activity toward V. parahaemolyticus. The fish-borne bacteriocin was purified by a three-step procedure consisting of ethyl acetate extraction, gel filtration chromatography and high performance liquid chromatography. Its molecular weight was estimated at 4.1 kDa using SDS-PAGE. The fish-borne bacteriocin reached the maximum production at stationary phase after 20 h. It was heat-stable (30 min at 121 °C) and remained active at pH range from 3.0 to 5.5, but was sensitive to nutrasin, papain and pepsin. Its minimum inhibitory concentration for V. parahaemolyticus was 6.0 mg/ml. Scanning electron microscopy analysis showed that the fish-borne bacteriocin disrupted cell wall of V. parahaemolyticus. The antibacterial mechanism of the fish-borne bacteriocin against V. parahaemolyticus might be described as action on membrane integrity in terms of the leakage of electrolytes, the losses of Na + K + -ATPase, AKP and proteins. The addition of the fish-borne bacteriocin to shrimps leaded V. parahaemolyticus to reduce 1.3 log units at 4 °C storage for 6 day. Moreover, a marked decline in total volatile base nitrogen and total viable counts was observed in bacteriocin treated samples than the control. It is clear that this fish-borne bacteriocin has promising potential as biopreservation for the control of V. parahaemolyticus in aquatic products.

Rational design of syn-safencin, a novel linear antimicrobial peptide derived from the circular bacteriocin safencin AS-48.

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Fields, Francisco R; Carothers, Katelyn E; Balsara, Rashna D; Ploplis, Victoria A; Castellino, Francis J; Lee, Shaun W

2018-06-01

Bacteriocins hold unprecedented promise as a largely untapped source of antibiotic alternatives in the age of multidrug resistance. Here, we describe the first approach to systematically design variants of a novel AS-48 bacteriocin homologue, which we have termed safencin AS-48, from Bacillus safensis, to gain insights into engineering improved activity of bacteriocins. A library of synthetic peptides in which systematic amino acid substitutions to vary the periodicity and abundance of polar, acidic, aliphatic, and hydrophobic residues were generated for a total of 96 novel peptide variants of a single bacteriocin candidate. Using this method, we identified nine synthetic safencin (syn-safencin) variants with broad and potent antimicrobial activities with minimal inhibitory concentrations (MIC) as low as 250 nM against E. coli, P. aeruginosa, X. axonopodis, and S. pyogenes with minimal cytotoxicity to mammalian cells. It is anticipated that the strategies we have developed will serve as general guides for tuning the specificity of a given natural bacteriocin compound for therapeutic specificity.

Inhibitory activity of bacteriocin produced from Lactobacillus SCG 1223 toward L. monocytogenes, S. thypimurium and E. coli

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Marwati, T.; Cahyaningrum, N.; Widodo, S.; Januarsyah, T.; Purwoko

2018-01-01

Bacteriocin is a protein compound which has bactericidal ability against pathogen bacteria. This research aims to study the inhibitory activity of bacteriocin produced from Lactobacillus SCG 1223 against Listeria monocytogenes, Salmonella thypimuruim and Escherchia coli. The bacteriocin produce from Lactobacillus SCG 1223 in the MRS broth media The experimental design used was Completely Randomized Design. The variations used in this design were percentage of inoculum (5%, 10%), medium pH (4, 6), incubation temperature (27°C, 40°C), and incubation time (4, 10, 14 hours). Result showed that bacteriocin from Lactobacillus SCG 1223 had wide spectrum toward L. monocytogenes, S. thypimuruim and E. coli. The highest bacteriocin activity toward L. monocytogenes produced by Lactobacillus SCG 1223 with 10% inoculum in media with initial pH 6, incubation temperature 27°C for 14 hour, toward S. thypimurium produced by Lactobacillus SCG 1223 with in media with initial pH 6, incubation temperature 40°C for 14 hour, and toward E. coli was 1085.81 AU/ml, produced by Lactobacillus SCG 1223 in MRS broth with initial pH 4, incubation temperature 40°C for 14 hour. This study is expected to find a new food preservative that can inhibit the growth of pathogenic bacteria and extend the shelf life of food. From the economic prospective of view, bacteriocin is very promising natural alternative biopreservatives.

Antimicrobial activities of the bacteriocin-like substances produced ...

African Journals Online (AJOL)

Administrator

2011-09-07

Sep 7, 2011 ... bacteriocins (subclass IIc); the large (>30 kDa) heat-labile non-lantibiotics (class III) and ... LAB from cow and goat milk and dairy products were ... (Oxoid, Hapshire, UK) was used to determine hydrolysis of esculin and growth ...

Simultaneous Production of Biosurfactants and Bacteriocins by Probiotic Lactobacillus casei MRTL3

Science.gov (United States)

Sharma, Deepansh; Singh Saharan, Baljeet

2014-01-01

Lactic acid bacteria (LAB) are ubiquitous and well-known commensal bacteria in the human and animal microflora. LAB are extensively studied and used in a variety of industrial and food fermentations. They are widely used for humans and animals as adjuvants, probiotic formulation, and dietary supplements and in other food fermentation applications. In the present investigation, LAB were isolated from raw milk samples collected from local dairy farms of Haryana, India. Further, the isolates were screened for simultaneous production of biosurfactants and bacteriocins. Biosurfactant produced was found to be a mixture of lipid and sugar similar to glycolipids. The bacteriocin obtained was found to be heat stable (5 min at 100°C). Further, DNA of the strain was extracted and amplified by the 16S rRNA sequencing using universal primers. The isolate Lactobacillus casei MRTL3 was found to be a potent biosurfactant and bacteriocin producer. It seems to have huge potential for food industry as a biopreservative and/or food ingredient. PMID:24669225

Selection of bacteriocin producer strains of lactic acid bacteria from a dairy environment.

Science.gov (United States)

Lasagno, M; Beoleito, V; Sesma, F; Raya, R; Font de Valdez, G; Eraso, A

2002-01-01

Two strains showing bacteriocin production were selected from a total of 206 lactic acid bacteria isolated from samples of milk, milk serum, whey and homemade cheeses in Southern Cordoba, Argentina. This property was detected by means of well diffusion assays. The strains were identified as Enterococcus hirae and Enterococcus durans. The protein nature of those substances was proved by showing their sensitivity to type IV and XXV proteases, papaine, trypsin, pepsin and K proteinase. The bacteriocins inhibited the growth of Listeria monocytogenes, Bacillus cereus, Clostridium perfringes and two strains of Staphylococcus aureus, an A-enterotoxin and a B-enterotoxin producers. All of these bacteria are common pathogens usually associated with food borne diseases (ETA). These lactic acid bacteria or their bacteriocins could be suitable candidates for food preservation and specially useful in the our regional dairy industry.

BAGEL2 : mining for bacteriocins in genomic data

NARCIS (Netherlands)

de Jong, Anne; van Heel, Auke J.; Kok, Jan; Kuipers, Oscar P.

Mining bacterial genomes for bacteriocins is a challenging task due to the substantial structure and sequence diversity, and generally small sizes, of these antimicrobial peptides. Major progress in the research of antimicrobial peptides and the ever-increasing quantities of genomic data, varying

Determination of bacteriocin activity with bioassays carried out on solid and liquid substrates: assessing the factor "indicator microorganism"

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Ambrosiadis Ioannis

2006-10-01

Full Text Available Abstract Background Successful application of growth inhibition techniques for quantitative determination of bacteriocins relies on the sensitivity of the applied indicator microorganism to the bacteriocin to which is exposed. However, information on indicator microorganisms' performance and comparisons in bacteriocin determination with bioassays is almost non-existing in the literature. The aim of the present work was to evaluate the parameter "indicator microorganism" in bioassays carried out on solid -agar diffusion assay- and liquid -turbidometric assay- substrates, applied in the quantification of the most studied bacteriocin nisin. Results The performance of characterized microorganisms of known sources, belonging to the genera of Lactobacillus, Pediococcus, Micrococcus and Leuconostoc, has been assessed in this work in the assays of plate agar diffusion and turbidometry. Dose responses and sensitivities were examined and compared over a range of assay variables in standard bacteriocin solutions, fermentation broth filtrates and processed food samples. Measurements on inhibition zones produced on agar plates were made by means of digital image analysis. The data produced were analyzed statistically using the ANOVA technique and pairwise comparisons tests. Sensitivity limits and linearity of responses to bacteriocin varied significantly among different test-microorganisms in both applied methods, the lower sensitivity limits depending on both the test-microorganism and the applied method. In both methods, however, only two of the nine tested microorganisms (Lactobacillus curvatus ATCC 51436 and Pediococcus acidilactici ATCC 25740 were sensitive to very low concentrations of the bacteriocin and produced a linear-type of response in all kinds of samples used in this work. In all cases, very low bacteriocin concentrations, e.g. 1 IU/ml nisin, were more accurately determined in the turbidometric assay. Conclusion The present work shows that in

Purification of bacteriocins using size-exclusion chromatography

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Vivek K. Bajpai

2016-06-01

Full Text Available The bacteriocin purification involves following main steps. a. Extraction of cell-free-supernatant of bacteria. b. Ammonium sulfate precipitation. c. Dialysis. d. Diafiltration using PVP and e. Size-exclusion chromatography. However, depending on the nature of work, the compound could be further analyzed by reverse-phase HPLC, NMR, mass spectrometry and sequencing.

Solution Structure of Enterocin HF, an Antilisterial Bacteriocin Produced by Enterococcus faecium M3K31.

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Arbulu, Sara; Lohans, Christopher T; van Belkum, Marco J; Cintas, Luis M; Herranz, Carmen; Vederas, John C; Hernández, Pablo E

2015-12-16

The solution structure of enterocin HF (EntHF), a class IIa bacteriocin of 43 amino acids produced by Enterococcus faecium M3K31, was evaluated by CD and NMR spectroscopy. Purified EntHF was unstructured in water, but CD analysis supports that EntHF adopts an α-helical conformation when exposed to increasing concentrations of trifluoroethanol. Furthermore, NMR spectroscopy indicates that this bacteriocin adopts an antiparallel β-sheet structure in the N-terminal region (residues 1-17), followed by a well-defined central α-helix (residues 19-30) and a more disordered C-terminal end (residues 31-43). EntHF could be structurally organized into three flexible regions that might act in a coordinated manner. This is in agreement with the absence of long-range nuclear Overhauser effect signals between the β-sheet domain and the C-terminal end of the bacteriocin. The 3D structure recorded for EntHF fits emerging facts regarding target recognition and mode of action of class IIa bacteriocins.

Detection of bacteriocins produced by plant pathogenic bacteria from the general Erwinia, Pseudomonas and Xanthomonas

International Nuclear Information System (INIS)

Biagi, C.M.R. de

1992-01-01

Detection of bacteriocin production was studied under distinct conditions using strains of plant pathogenic bacteria from the genera Erwinia, Pseudomonas and Xanthomonas. 58.06%, 79.31% and 40.00% of producing strains were found respectively in the three groups of bacteria using the 523 medium which was the best for the detection of bacteriocin production. Increasing agar concentrations added to the medium up to 1,5% improved the detection. The amount of medium added to the Petri dishes did not affect bacteriocin production. The longest incubation time (72 h.) improved the detection of haloes production. Ultra-violet irradiation in low dosages seems to improve the visualization of haloes production but this is dependent on the tested strains. (author)

Partial purification and characterization of bacteriocin produced by Enterococcus faecalis DU10 and its probiotic attributes.

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Perumal, Venkatesh; Repally, Ayyanna; Dasari, Ankaiah; Venkatesan, Arul

2016-10-02

A novel bacteriocin produced by avian duck isolated lactic acid bacterium Enterococcus faecalis DU10 was isolated. This bacteriocin showed a broad spectrum of antibacterial activity against important food-borne pathogens and was purified by size exclusion chromatography followed by reverse-phase high-performance liquid chromatography in a C-18 column. Tricine-SDS PAGE revealed the presence of a band with an estimated molecular mass of 6.3 kDa. The zymogram clearly linked the antimicrobial activity with this band. This result was further confirmed by mass-assisted laser desorption ionization time-of-flight mass spectrometry, since a sharp peak corresponding to 6.313 kDa was detected and the functional groups were revealed by Fourier transform infrared spectroscopy. Bacteriocin DU10 activity was found sensitive to proteinase-K and pepsin and partially affected by trypsin and α-chymotrypsin. The activity of bacteriocin DU10 was partially resistant to heat treatments ranging from 30 to 90°C for 30 min. It also withstood a treatment at 121°C for 10 min. Cytotoxicity of bacteriocin DU10 by methyl-thiazolyl-diphenyl-tetrazolium bromide assay showed that the viability of HT-29 and HeLa cells decreased 60 ± 0.7% and 43 ± 4.8%, respectively, in the presence of 3,200 AU/mL of bacteriocin. The strain withstood 0.3% w/v of bile oxgall and pH 2 affected the bacterial growth between 2 and 4 hr of incubation. Adhesion properties examined with HT-29 cell line showed 69.85% initial population of strain E. faecalis DU10, which was found to be strongly adhered to this cell line. These results conclude bacteriocin DU10 may be used as a potential biopreservative and E. faecalis DU10 may be used as a potential probiont to control Salmonella infections.

Purification and characteristics of a novel bacteriocin produced by Enterococcus faecalis L11 isolated from Chinese traditional fermented cucumber.

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Gao, Yurong; Li, Benling; Li, Dapeng; Zhang, Liyuan

2016-05-01

To purify and characterize a novel bacteriocin with broad inhibitory spectrum produced by an isolate of Enterococcus faecalis from Chinese fermented cucumber. E. faecalis L11 produced a bacteriocin with antimicrobial activity against both Escherichia coli and Staphylococcus aureus. The amino acid sequence of the purified bacteriocin, enterocin L11, was assayed by Edman degradation method. It differs from other class II bacteriocins and exhibited a broad antimicrobial activity against not only Gram-positive bacteria, including Bacillus subtilis, S. aureus, Listeria monocytogenes, Sarcina flava, Lactobacillus acidophilus, L. plantarum, L. delbrueckii subsp. delbrueckii, L. delbrueckii subsp. bulgaricus and Streptococcus thermophilus, but also some Gram-negative bacteria including Salmonella typhimurium, E. coli and Shigella flexneri. Enterocin L11 retained 91 % of its activity after holding at 121 °C for 30 min. It was also resistant to acids and alkalis. Enterocin L11 is a novel broad-spectrum Class II bacteriocin produced by E. faecalis L11, and may have potential as a food biopreservative.

Partial characterization of bacteriocin induced by irradiated and non-irradiated strain of yersinia enterocolitical

International Nuclear Information System (INIS)

Awny, N.M.

1991-01-01

Twenty isolates of yersinia enterocolitica were tested for the inhibition of the growth of different strains of yersinia. The screening tests revealed three possible bacteriocinogenic strains. One of them was selected for additional studies after it was shown that its inhibitory substances differed in their activity spectra. The gamma irradiated strain lost the ability to produce bacteriocin at 0.6 kGy level. Crude preparation of bacteriocin obtained from the wild strain were not affected by chloroform or other organic solvents but inactivated by trypsin and heating at 80 C for 45 min. Bacteriocin induced by irradiated strain was easily inactivated by thermal treatment. Exposure of agar fragments containing the inhibitory active component to a pH value ranging between 2 to 11 did not affect bactericidal activity.4 tab

Interactions between Oral Bacteria: Inhibition of Streptococcus mutans Bacteriocin Production by Streptococcus gordonii

OpenAIRE

Wang, Bing-Yan; Kuramitsu, Howard K.

2005-01-01

Streptococcus mutans has been recognized as an important etiological agent in human dental caries. Some strains of S. mutans also produce bacteriocins. In this study, we sought to demonstrate that bacteriocin production by S. mutans strains GS5 and BM71 was mediated by quorum sensing, which is dependent on a competence-stimulating peptide (CSP) signaling system encoded by the com genes. We also demonstrated that interactions with some other oral streptococci interfered with S. mutans bacterio...

Variable characteristics of bacteriocin-producing Streptococcus salivarius strains isolated from Malaysian subjects.

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Abdelahhad Barbour

Full Text Available BACKGROUND: Salivaricins are bacteriocins produced by Streptococcus salivarius, some strains of which can have significant probiotic effects. S. salivarius strains were isolated from Malaysian subjects showing variable antimicrobial activity, metabolic profile, antibiotic susceptibility and lantibiotic production. METHODOLOGY/PRINCIPAL FINDINGS: In this study we report new S. salivarius strains isolated from Malaysian subjects with potential as probiotics. Safety assessment of these strains included their antibiotic susceptibility and metabolic profiles. Genome sequencing using Illumina's MiSeq system was performed for both strains NU10 and YU10 and demonstrating the absence of any known streptococcal virulence determinants indicating that these strains are safe for subsequent use as probiotics. Strain NU10 was found to harbour genes encoding salivaricins A and 9 while strain YU10 was shown to harbour genes encoding salivaricins A3, G32, streptin and slnA1 lantibiotic-like protein. Strain GT2 was shown to harbour genes encoding a large non-lantibiotic bacteriocin (salivaricin-MPS. A new medium for maximum biomass production buffered with 2-(N-morpholinoethanesulfonic acid (MES was developed and showed better biomass accumulation compared with other commercial media. Furthermore, we extracted and purified salivaricin 9 (by strain NU10 and salivaricin G32 (by strain YU10 from S. salivarius cells grown aerobically in this medium. In addition to bacteriocin production, S. salivarius strains produced levan-sucrase which was detected by a specific ESI-LC-MS/MS method which indicates additional health benefits from the developed strains. CONCLUSION: The current study established the bacteriocin, levan-sucrase production and basic safety features of S. salivarius strains isolated from healthy Malaysian subjects demonstrating their potential for use as probiotics. A new bacteriocin-production medium was developed with potential scale up application for

Plantaricyclin A, a Novel Circular Bacteriocin Produced by Lactobacillus plantarum NI326: Purification, Characterization, and Heterologous Production.

Science.gov (United States)

Borrero, Juan; Kelly, Eoin; O'Connor, Paula M; Kelleher, Philip; Scully, Colm; Cotter, Paul D; Mahony, Jennifer; van Sinderen, Douwe

2018-01-01

Bacteriocins from lactic acid bacteria (LAB) are of increasing interest in recent years due to their potential as natural preservatives against food and beverage spoilage microorganisms. In a screening study for LAB, we isolated from olives a strain, Lactobacillus plantarum NI326, with activity against the beverage-spoilage bacterium Alicyclobacillus acidoterrestris Genome sequencing of NI326 enabled the identification of a gene cluster (designated plc ) encoding a putative circular bacteriocin and proteins involved in its modification, transport, and immunity. This novel bacteriocin, named plantaricyclin A (PlcA), was grouped into the circular bacteriocin subgroup II due to its high degree of similarity with other gassericin A-like bacteriocins. Purification of PlcA from the supernatant of Lb. plantarum NI326 resulted in an active peptide with a molecular mass of 5,570 Da, corresponding to that predicted from the (processed) PlcA amino acid sequence. The plc gene cluster was cloned and expressed in Lactococcus lactis NZ9000, resulting in the production of an active 5,570-Da bacteriocin in the supernatant. PlcA is believed to be produced as a 91-amino-acid precursor with a 33-amino-acid leader peptide, which is predicted to be removed, followed by joining of the N and C termini via a covalent linkage to form the mature 58-amino-acid circular bacteriocin PlcA. We report the characterization of a circular bacteriocin produced by Lb. plantarum The inhibition displayed against A. acidoterrestris highlights its potential use as a preservative in food and beverages. IMPORTANCE In this work, we describe the purification and characterization of an antimicrobial peptide, termed plantaricyclin A (PlcA), produced by a Lactobacillus plantarum strain isolated from olives. This peptide has a circular structure, and all genes involved in its production, circularization, and secretion were identified. PlcA shows antimicrobial activity against different strains, including

Cultivation, isolation and characterization of bacteriocin from fresh ...

African Journals Online (AJOL)

This study focus on cultivation, isolation and characterization of Bacteriocin from fresh cow milk (FCM) and fresh cow meat (FMS) samples obtained from Lapai Market in Niger State, Nigeria. Potential bacteriocinogenic bacteria were screened with agar diffusion method on culture plates seeded with Staphylococcus and ...

Lactobacillus pentosus B231 Isolated from a Portuguese PDO Cheese: Production and Partial Characterization of Its Bacteriocin.

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Guerreiro, Joana; Monteiro, Vitor; Ramos, Carla; Franco, Bernadette Dora Gombossy de Melo; Martinez, Rafael Chacon Ruiz; Todorov, Svetoslav Dimitrov; Fernandes, Paulo

2014-06-01

Bacteriocin B231 produced by Lactobacillus pentosus, isolated from an artisanal raw cow's milk protected designation of origin Portuguese cheese, is a small protein with an apparent relative mass of about 5 kDa and active against a large number of Listeria monocytogenes wild-type strains, Listeria ivanovii and Listeria innocua. Bacteriocin B231 production is highly dependent on the type of the culture media used for growth of Lact. pentosus B231. Replacement of glucose with maltose yielded the highest bacteriocin production from eight different carbon sources. Similar results were recorded in the presence of combination of glucose and maltose or galactose. Production of bacteriocin B231 reached maximal levels of 800 AU/ml during the stationary phase of growth of Lact. pentosus B231 in MRS broth at 30 °C. Bacteriocin B231 (in cell-free supernatant) was sensitive to treatment with trypsin and proteinase K, but not affected by the thermal treatment in range of 55-121 °C, or freezing (-20 °C). Bacteriocin production and inhibitory spectrum were evaluated. Gene encoding plantaricin S has been detected in the genomic DNA. Virulence potential and safety of Lact. pentosus B231 were assessed by PCR targeted the genes gelE, hyl, asa1, esp, cylA, efaA, ace, vanA, vanB, hdc1, hdc2, tdc and odc. The Lact. pentosus B231 strains harbored plantaricin S gene, while the occurrence of virulence, antibiotic resistance and biogenic amine genes was limited to cytolysin, hyaluronidase, aggregation substance, adhesion of collagen protein, gelatinase, tyrosine decarboxylase and vancomycin B genes.

Maltaricin CPN, a new class IIa bacteriocin produced by Carnobacterium maltaromaticum CPN isolated from mould-ripened cheese.

Science.gov (United States)

Hammi, I; Delalande, F; Belkhou, R; Marchioni, E; Cianferani, S; Ennahar, S

2016-11-01

The purpose of this study was to isolate, characterize and determine the structure and the antibacterial activities of a bacteriocin produced by Carnobacterium maltaromaticum CPN, a strain isolated from unpasteurized milk Camembert cheese. This bacteriocin, termed maltaricin CPN, was produced at higher amounts in MRS broth at temperatures between 15°C and 25°C. It was purified to homogeneity from culture supernatant by using a simple method consisting of cation-exchange and reversed-phase chromatographies. Mass spectrometry showed that maltaricin was a 4427·29 Da bacteriocin. Its amino acid sequence was determined by Edman degradation which showed that it had close similarity with bacteriocins of the class IIa. Maltaricin CPN consisted in fact of 44 unmodified amino acids including two cysteine residues at positions 9 and 14 linked by a disulphide bond. The antimicrobial activity of maltaricin CPN covered a range of bacteria, with strong activity against many species of Gram-positive bacteria, especially the food-borne pathogen Listeria monocytogenes, but no activity against Gram-negative ones. In the studied conditions, C. maltaromaticum CPN produced a new class IIa bacteriocin with strong anti-Listeria activity. The study covers the purification and the structural characterization of a new bacteriocin produced by strain C. maltaromaticum CPN isolated from Camembert cheese. Its activity against strains of L. monocytogenes and higher production rates at relatively low temperatures show potential technological applications to improve the safety of refrigerated food. © 2016 The Society for Applied Microbiology.

Antimicrobial potential of bacteriocins in poultry and swine production.

Science.gov (United States)

Ben Lagha, Amel; Haas, Bruno; Gottschalk, Marcelo; Grenier, Daniel

2017-04-11

The routine use of antibiotics in agriculture has contributed to an increase in drug-resistant bacterial pathogens in animals that can potentially be transmitted to humans. In 2000, the World Health Organization identified resistance to antibiotics as one of the most significant global threats to public health and recommended that the use of antibiotics as additives in animal feed be phased out or terminated, particularly those used to treat human infections. Research is currently being carried out to identify alternative antimicrobial compounds for use in animal production. A number of studies, mostly in vitro, have provided evidence indicating that bacteriocins, which are antimicrobial peptides of bacterial origin, may be promising alternatives to conventional antibiotics in poultry and swine production. This review provides an update on bacteriocins and their potential for use in the poultry and swine industries.

Partial Diversity Generates Effector Immunity Specificity of the Bac41-Like Bacteriocins of Enterococcus faecalis Clinical Strains.

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Kurushima, Jun; Ike, Yasuyoshi; Tomita, Haruyoshi

2016-09-01

Bacteriocin 41 (Bac41) is the plasmid-encoded bacteriocin produced by the opportunistic pathogen Enterococcus faecalis Its genetic determinant consists of bacL1 (effector), bacL2 (regulator), bacA (effector), and bacI (immunity). The secreted effectors BacL1 and BacA coordinate to induce the lytic cell death of E. faecalis Meanwhile, the immunity factor BacI provides self-resistance to the Bac41 producer, E. faecalis, against the action of BacL1 and BacA. In this study, we demonstrated that more than half of the 327 clinical strains of E. faecalis screened had functional Bac41 genes. Analysis of the genetic structure of the Bac41 genes in the DNA sequences of the E. faecalis strains revealed that the Bac41-like genes consist of a relatively conserved region and a variable region located downstream from bacA Based on similarities in the variable region, the Bac41-like genes could be classified into type I, type IIa, and type IIb. Interestingly, the distinct Bac41 types had specific immunity factors for self-resistance, BacI1 or BacI2, and did not show cross-immunity to the other type of effector. We also demonstrated experimentally that the specificity of the immunity was determined by the combination of the C-terminal region of BacA and the presence of the unique BacI1 or BacI2 factor. These observations suggested that Bac41-like bacteriocin genes are extensively disseminated among E. faecalis strains in the clinical environment and can be grouped into at least three types. It was also indicated that the partial diversity results in specificity of self-resistance which may offer these strains a competitive advantage. Bacteriocins are antibacterial effectors produced by bacteria. In general, a bacteriocin-coding gene is accompanied by a cognate immunity gene that confers self-resistance on the bacteriocin-producing bacterium itself. We demonstrated that one of the bacteriocins, Bac41, is disseminated among E. faecalis clinical strains and the Bac41 subtypes with

Rapid Two-Step Procedure for Large-Scale Purification of Pediocin-Like Bacteriocins and Other Cationic Antimicrobial Peptides from Complex Culture Medium

Science.gov (United States)

Uteng, Marianne; Hauge, Håvard Hildeng; Brondz, Ilia; Nissen-Meyer, Jon; Fimland, Gunnar

2002-01-01

A rapid and simple two-step procedure suitable for both small- and large-scale purification of pediocin-like bacteriocins and other cationic peptides has been developed. In the first step, the bacterial culture was applied directly on a cation-exchange column (1-ml cation exchanger per 100-ml cell culture). Bacteria and anionic compounds passed through the column, and cationic bacteriocins were subsequently eluted with 1 M NaCl. In the second step, the bacteriocin fraction was applied on a low-pressure, reverse-phase column and the bacteriocins were detected as major optical density peaks upon elution with propanol. More than 80% of the activity that was initially in the culture supernatant was recovered in both purification steps, and the final bacteriocin preparation was more than 90% pure as judged by analytical reverse-phase chromatography and capillary electrophoresis. PMID:11823243

Isolation of a bacteriocin-producing Lactococcus lactis subsp. lactis and application to control Listeria monocytogenes in Moroccan jben.

Science.gov (United States)

Benkerroum, N; Oubel, H; Zahar, M; Dlia, S; Filali-Maltouf, A

2000-12-01

Use of a bacteriocin-producing lactococcal strain to control Listeria monocytogenes in jben. A Lactococcus lactis strain isolated from lben was shown, by the spot technique, to produce a bacteriocin different from nisin. Inhibitory activity of the bacteriocin-producing strain against Listeria monocytogenes was investigated in jben, made from cow's milk fermented with the producer organism and contaminated with 104 or 107 cfu ml-1. Listeria counts were monitored during manufacture, and during conservation at room and at refrigeration temperatures. Results showed that the pathogen was reduced by 2.7 logarithmic units after 30 h of jben processing when the initial inoculum of 107 cfu ml(-1) was used. For the initial inoculum of 104 cfu ml(-1), the bacterium was completely eliminated at 24 h. Furthermore, the use of the bacteriocin-producing starter culture extended the shelf-life of jben by 5 days. In situ production of the lactococcal bacteriocin is an efficient biological means of controlling L. monocytogenes in jben and of allowing shelf-life extension. The proposed technology will essentially benefit minimally processed dairy products and those made with raw milk.

Evaluation of leader peptides that affect the secretory ability of a multiple bacteriocin transporter, EnkT.

Science.gov (United States)

Sushida, Hirotoshi; Ishibashi, Naoki; Zendo, Takeshi; Wilaipun, Pongtep; Leelawatcharamas, Vichien; Nakayama, Jiro; Sonomoto, Kenji

2018-02-13

EnkT is a novel ATP-binding cassette (ABC) transporter responsible for secretion of four bacteriocins, enterocins NKR-5-3A, C, D, and Z (Ent53A, C, D, and Z), produced by Enterococcus faecium NKR-5-3. It is generally recognized that the secretion of a bacteriocin requires a dedicated ABC transporter, although molecular mechanisms of this secretion are yet to be revealed. In order to characterize the unique ability of EnkT to secrete multiple bacteriocins, the role of N-terminal leader peptides of bacteriocin precursors was evaluated using Ent53C precursor as a model. The 18-amino acid leader peptide of Ent53C (Lc) was modified by site-directed mutagenesis to generate various point mutations, truncations, or extensions, and substitutions with other leader peptides. The impact of these Lc mutations on Ent53C secretion was evaluated using a quantitative antimicrobial activity assay. We observed that Ent53C production increased with Ala substitution of the highly conserved C-terminal double glycine residues that are recognized as the cleavage site. In contrast, Ent53C antimicrobial activity decreased, with decrease in the length of the putative α-helix-forming region of Lc. Furthermore, EnkT recognized and transported Ent53C of the transformants possessing heterologous leader peptides of enterocin A, pediocin PA-1, brochocins A and B, and lactococcins Qα and Qβ. These results indicated that EnkT shows significant tolerance towards the sequence and length of leader peptides, to secrete multiple bacteriocins. This further demonstrates the functional diversity of bacteriocin ABC transporters and the importance of leader peptides as their recognition motif. Copyright © 2018 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

The effect of bacteriocin-producing Lactobacillus plantarum strains on the intracellular pH of sessile and planktonic Listeria monocytongenes single cells

DEFF Research Database (Denmark)

Nielsen, Dennis Sandris; Cho, Gyu-Sung; Hanak, Alexander

2010-01-01

and/or bacteriocin-producing LAB as “natural” food preservatives in foods such as cheese, meat and ready-to-eat products. Some strains of Lactobacillus plantarum produce bacteriocins termed plantaricins. Using a single-cell based approach, the effect on the intracellular pH as a measure......A wide range of lactic acid bacteria (LAB) produce bacteriocins mainly active against other closely related LAB, but some bacteriocins are also active against the food-borne pathogen Listeria monocytogenes. With the aim of increasing food safety it has thus been considered to utilise bacteriocins...

Effect of Bacteriocin-like Inhibitory Substances Produced by Vaginal ...

African Journals Online (AJOL)

Reduction of vaginal Lactobacillus population leads to overgrowth of opportunistic organisms such as Streptococcus agalactiae (Group B Streptococcus, GBS), which causes life threatening neonatal infections. The activities of bacteriocin-like inhibitory substances (BLIS) produced by Lactobacillus species isolated from the ...

Role of plnB gene in the regulation of bacteriocin production in Lactobacillus paraplantarum L-XM1.

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Zhang, Xiangmei; Shang, Nan; Zhang, Xu; Gui, Meng; Li, Pinglan

2013-06-12

Homologues of plnB gene have been shown to participate in regulation of bacteriocin production through quorum sensing system in other organisms, to investigate the possible role of plnB gene in Lactobacillus paraplantarum L-XM1, we cloned and insertionally inactivated the plnB gene. The plnB knockout mutant ΔplnB21 showed loss of bacteriocin production, its Bac⁺ phenotype could not be restored even after the addition of PlnA. Furthermore, reverse transcription-PCR analysis from total RNA preparations showed that the bacteriocin structural genes of the plnEF and plnJK were not transcribed in the plnB knockout mutant compared with the wild-type strain. It was therefore concluded that plnB is invovled in a quorum sensing based bacteriocin production. This is the first demonstration of a role for plnB by gene knockout in L. paraplantarum. Copyright © 2012 Elsevier GmbH. All rights reserved.

[A comparison of the properties of bacteriocins formed by Lactococcus lactis subsp. lactis strains of diverse origin].

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Stoianova, L G; Egorov, N S; Fedorova, G B; Katrukha, G S; Netrusov, A I

2007-01-01

Bacteriocins formed by four strains of Lactococcus lactis subsp. lactis have been studied and compared: 729 (a natural strain isolated from milk), 1605 (a mutant of strain 729), F-116 (a recombinant obtained by fusing of protoplasts of the two related strain 729 and 1605), and a nisin-forming strain obtained by adaptive selection at Moscow State University. Antimicrobial activity studies revealed differences between the strains in the effects on individual groups of microorganisms; the activities of the strains were also distinct from that of Nisaplin (a commercial preparation of the bacteriocin nisin). Methods for isolation and purification of bacteriocins have been developed, making it possible to obtain individual components of antibiotic complexes as chromatographically pure preparations. Bacteriocins formed by the strains of Lactococcus lactis subsp. lactis have been identified and differences in their biological and physicochemical properties, established. A novel potent broad-spectrum antibiotic substance distinct from nisin has been isolated from the recombinant strain F-116.

Improved antimicrobial activities of synthetic-hybrid bacteriocins designed from enterocin E50-52 and pediocin PA-1.

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Tiwari, Santosh Kumar; Sutyak Noll, Katia; Cavera, Veronica L; Chikindas, Michael L

2015-03-01

Two hybrid bacteriocins, enterocin E50-52/pediocin PA-1 (EP) and pediocin PA-1/enterocin E50-52 (PE), were designed by combining the N terminus of enterocin E50-52 and the C terminus of pediocin PA-1 and by combining the C terminus of pediocin PA-1 and the N terminus of enterocin E50-52, respectively. Both hybrid bacteriocins showed reduced MICs compared to those of their natural counterparts. The MICs of hybrid PE and EP were 64- and 32-fold lower, respectively, than the MIC of pediocin PA-1 and 8- and 4-fold lower, respectively, than the MIC of enterocin E50-52. In this study, the effect of hybrid as well as wild-type (WT) bacteriocins on the transmembrane electrical potential (ΔΨ) and their ability to induce the efflux of intracellular ATP were investigated. Enterocin E50-52, pediocin PA-1, and hybrid bacteriocin PE were able to dissipate ΔΨ, but EP was unable to deplete this component. Both hybrid bacteriocins caused a loss of the intracellular concentration of ATP. EP, however, caused a faster efflux than PE and enterocin E50-52. Enterocin E50-52 and hybrids PE and EP were active against the Gram-positive and Gram-negative bacteria tested, such as Micrococcus luteus, Salmonella enterica serovar Enteritidis 20E1090, and Escherichia coli O157:H7. The hybrid bacteriocins designed and described herein are antimicrobial peptides with MICs lower those of their natural counterparts. Both hybrid peptides induce the loss of intracellular ATP and are capable of inhibiting Gram-negative bacteria, and PE dissipates the electrical potential. In this study, the MIC of hybrid bacteriocin PE decreased 64-fold compared to the MIC of its natural peptide counterpart, pediocin PA-1. Inhibition of Gram-negative pathogens confers an additional advantage for the application of these peptides in therapeutics. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Isolation and Characterization of a Broad Spectrum Bacteriocin from Bacillus amyloliquefaciens RX7

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Kong Boon Lim

2016-01-01

Full Text Available We isolated a Bacillus strain, RX7, with inhibitory activity against Listeria monocytogenes from soil and identified it as Bacillus amyloliquefaciens based on 16S rRNA gene sequencing. The inhibitory activity was stable over a wide range of pH and was fully retained after 30 min at 80°C, after which it decreased gradually at higher temperatures. The activity was sensitive to the proteolytic action of α-chymotrypsin, proteinase-K, and trypsin, indicating its proteinaceous nature. This bacteriocin was active against a broad spectrum of bacteria and the fungus Candida albicans. Direct detection of antimicrobial activity on a sodium dodecyl sulfate-polyacrylamide gel suggested an apparent molecular mass of approximately 5 kDa. Ammonium sulfate precipitation and anion-exchange and gel permeation chromatography integrated with reverse phase-high-performance liquid chromatography were used for bacteriocin purification. Automated N-terminal Edman degradation of the purified RX7 bacteriocin recognized the first 15 amino acids as NH2-X-Ala-Trp-Tyr-Asp-Ile-Arg-Lys-Leu-Gly-Asn-Lys-Gly-Ala, where the letter X in the sequence indicates an unknown or nonstandard amino acid. Based on BLAST similarity search and multiple alignment analysis, the obtained partial sequence showed high homology with the two-peptide lantibiotic haloduracin (HalA1 from Bacillus halodurans, although at least two amino acids differed between the sequences. A time-kill study demonstrated a bactericidal mode of action of RX7 bacteriocin.

Bacteriocin and cellulose production by lactic acid bacteria isolated ...

African Journals Online (AJOL)

SERVER

2007-11-19

after 20 and 25 min) and E. coli 0157:H7 (after 15, 20 and 25 ... in a coculture with various Lactobacillus mali strains showed that cell-cell .... were found to be capable of producing viable bacteriocin can be purified and charac-.

Identification of Structural and Immunity Genes of a Class IIb Bacteriocin Encoded in the Enterocin A Operon of Enterococcus faecium Strain MXVK29.

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Escamilla-Martínez, E E; Cisneros, Y M Álvarez; Fernández, F J; Quirasco-Baruch, M; Ponce-Alquicira, E

2017-10-09

The Enterococcus faecium strain MXVK29, isolated from fermented sausages, produces a bacteriocin with a molecular mass of 3.5 kDa that belongs to the class of enterocins II.1, according to the terminal amino acid sequence, and has been identified as enterocin A. This bacteriocin is active against selected strains of Listeria, Staphylococcus, Pediococcus, and Enterococcus. In this study, we identified the genes adjacent to the structural gene for this bacteriocin, such as the immunity gene (entI) and the inducer gene (entF). Accessory genes for this bacteriocin, such as entK, entR, and entT, were identified as well, in addition to the orf2 and orf3, showing a high identity with class IIb peptides bacteriocins. The orf2 shows the consensus motif GxxxG, similar to those shown by bacteriocins such as PlnNC8α, EntCα, and Ent1071A, whereas orf3 shows a consensus motif SxxxS similar to that present in PlnNC8β (AxxxA). PlnNC8 is expressed only in bacterial cocultures, so there is the possibility that the expression of this two-peptide bacteriocin can be induced by a similar mechanism. So far, only the expression of enterocin A has been found in this strain; however, the presence of the genes ent29α and ent29β opens the possibility for further research on its induction, functionality, and origin. Although there are reports on this type of bacteriocin (EntX, EntC, and Ent1071) in other strains of E. faecium, no report exists yet on an Enterococcus strain producing two different classes of bacteriocin.

Induction of bacteriocin production by coculture is widespread among plantaricin-producing Lactobacillus plantarum strains with different regulatory operons.

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Maldonado-Barragán, Antonio; Caballero-Guerrero, Belén; Lucena-Padrós, Helena; Ruiz-Barba, José Luis

2013-02-01

We describe the bacteriocin-production phenotype in a group of eight singular bacteriocinogenic Lactobacillus plantarum strains with three distinct genotypes regarding the plantaricin locus. Genotyping of these strains revealed the existence of two different plantaricin-production regulatory operons, plNC8-plNC8HK-plnD or plnABCD, involving three-component systems controlled each of them by a specific autoinducer peptide (AIP), i.e. PLNC8IF or PlnA. While all of the strains produced antimicrobial activity when growing on solid medium, most of them halted this production when cultured in broth, thus reflecting the functionality of regulatory mechanisms. Antimicrobial activity in broth cultures was re-established or enhanced when the specific AIP was added to the culture or by coculturing with specific bacterial strains. The latter trait appeared to be widespread in bacteriocinogenic L. plantarum strains independently of the regulatory system used to regulate bacteriocin production or the specific bacteriocins produced. The induction spectrum through coculture, i.e. the pattern of bacterial strains able to induce bacteriocin production, was characteristic of each individual L. plantarum strain. Also, the ability of some bacteria to induce bacteriocin production in L. plantarum by coculture appeared to be strain specific. The fact that induction of bacteriocin production by coculturing appeared to be a common feature in L. plantarum can be exploited accordingly to enhance the viability of this species in food and feed fermentations, as well as to contribute to probiotic functionality when colonising the gastrointestinal tract. Copyright © 2012 Elsevier Ltd. All rights reserved.

Purification and amino acid sequence of a bacteriocins produced by Lactobacillus salivarius K7 isolated from chicken intestine

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Kenji Sonomoto

2006-03-01

Full Text Available A bacteriocin-producing strain, Lactobacillus K7, was isolated from a chicken intestine. The inhibitory activity was determined by spot-on-lawn technique. Identification of the strain was performed by morphological, biochemical (API 50 CH kit and molecular genetic (16S rDNA basis. Bacteriocin purification processes were carried out by amberlite adsorption, cation exchange and reverse-phase high perform- ance liquid chromatography. N-terminal amino acid sequences were performed by Edman degradation. Molecular mass was determined by electrospray-ionization (ESI mass spectrometry (MS. Lactobacillus K7 showed inhibitory activity against Lactobacillus sakei subsp. sakei JCM 1157T, Leuconostoc mesenteroides subsp. mesenteroides JCM 6124T and Bacillus coagulans JCM 2257T. This strain was identified as Lb. salivarius. The antimicrobial substance was destroyed by proteolytic enzymes, indicating its proteinaceous structure designated as a bacteriocin type. The purification of bacteriocin by amberlite adsorption, cation exchange, and reverse-phase chromatography resulted in only one single active peak, which was designated FK22. Molecular weight of this fraction was 4331.70 Da. By amino acid sequence, this peptide was homology to Abp 118 beta produced by Lb. salivarius UCC118. In addition, Lb. salivarius UCC118 produced 2-peptide bacteriocin, which was Abp 118 alpha and beta. Based on the partial amino acid sequences of Abp 118 beta, specific primers were designed from nucleotide sequences according to data from GenBank. The result showed that the deduced peptide was high homology to 2-peptide bacteriocin, Abp 118 alpha and beta.

Purification and partial characterization of bacillocin 490, a novel bacteriocin produced by a thermophilic strain of Bacillus licheniformis

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De Felice Maurilio

2002-04-01

Full Text Available Abstract Background Applications of bacteriocins as food preservatives have been so far limited, principally because of their low antimicrobial activity in foods. Nisin is the only bacteriocin of significant use, but applications are restricted principally because of its very low activity at neutral or alkaline pH. Thus the isolation of new bacteriocins active in foods is desirable. Results We isolated a Bacillus licheniformis thermophilic strain producing a bacteriocin with some novel features, named here bacillocin 490. This bacteriocin was inactivated by pronase E and proteinase K and was active against closely related Bacillus spp. both in aerobic and in anaerobic conditions. Bactericidal activity was kept during storage at 4°C and was remarkably stable in a wide pH range. The bacteriocin was partially purified by elution after adhesion to cells of the food-isolated strain Bacillus smithii and had a rather low mass (2 KDa. Antimicrobial activity against B. smithii was observed also when this organism was grown in water buffalo milk. Conclusions Bacillocin 490 is a novel candidate as a food anti-microbial agent since it displays its activity in milk, is stable to heat treatment and during storage, is active in a wide pH range and has bactericidal activity also at high temperature. These features may allow the use of bacillocin 490 during processes performed at high temperature and as a complementary antimicrobial agent of nisin against some Bacillus spp. in non-acidic foods. The small size suggests its use on solid foods.

Comparison of two methods for purification of enterocin B, a bacteriocin produced by Enterococcus faecium W3.

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Dündar, Halil; Atakay, Mehmet; Çelikbıçak, Ömür; Salih, Bekir; Bozoğlu, Faruk

2015-01-01

This study aimed to compare two different approaches for the purification of enterocin B from Enterococcus faecium strain W3 based on the observation that the bacteriocin was found both in cell associated form and in culture supernatant. The first approach employed ammonium sulfate precipitation, cation-exchange chromatography, and sequential reverse-phase high-performance liquid chromatography. The latter approach exploited a pH-mediated cell adsorption-desorption method to extract cell-bound bacteriocin, and one run of reverse-phase chromatography. The first method resulted in purification of enterocin B with a recovery of 4% of the initial bacteriocin activity found in culture supernatant. MALDI-TOF MS analysis and de novo peptide sequencing of the purified bacteriocin confirmed that the active peptide was enterocin B. The second method achieved the purification of enterocin B with a higher recovery (16%) and enabled us to achieve pure bacteriocin within a shorter period of time by avoiding time consuming purification protocols. The purity and identity of the active peptide were confirmed again by matrix-assisted laser desorption/ionization time-of flight (MALDI-TOF) mass spectrometry (MS) analysis. Although both approaches were satisfactory to obtain a sufficient amount of enterocin B for use in MS and amino acid sequence analysis, the latter was proved to be applicable in large-scale and rapid purification of enterocin B.

A strain-variable bacteriocin in Bacillus anthracis and Bacillus cereus with repeated Cys-Xaa-Xaa motifs

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Haft Daniel H

2009-04-01

Full Text Available Abstract Bacteriocins are peptide antibiotics from ribosomally translated precursors, produced by bacteria often through extensive post-translational modification. Minimal sequence conservation, short gene lengths, and low complexity sequence can hinder bacteriocin identification, even during gene calling, so they are often discovered by proximity to accessory genes encoding maturation, immunity, and export functions. This work reports a new subfamily of putative thiazole-containing heterocyclic bacteriocins. It appears universal in all strains of Bacillus anthracis and B. cereus, but has gone unrecognized because it is always encoded far from its maturation protein operon. Patterns of insertions and deletions among twenty-four variants suggest a repeating functional unit of Cys-Xaa-Xaa. Reviewers This article was reviewed by Andrei Osterman and Lakshminarayan Iyer.

Solution structures of the linear leaderless bacteriocins enterocin 7A and 7B resemble carnocyclin A, a circular antimicrobial peptide.

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Lohans, Christopher T; Towle, Kaitlyn M; Miskolzie, Mark; McKay, Ryan T; van Belkum, Marco J; McMullen, Lynn M; Vederas, John C

2013-06-11

Leaderless bacteriocins are a class of ribosomally synthesized antimicrobial peptides that are produced by certain Gram-positive bacteria without an N-terminal leader section. These bacteriocins are of great interest due to their potent inhibition of many Gram-positive organisms, including food-borne pathogens such as Listeria and Clostridium spp. We now report the NMR solution structures of enterocins 7A and 7B, leaderless bacteriocins recently isolated from Enterococcus faecalis 710C. These are the first three-dimensional structures to be reported for bacteriocins of this class. Unlike most other linear Gram-positive bacteriocins, enterocins 7A and 7B are highly structured in aqueous conditions. Both peptides are primarily α-helical, adopting a similar overall fold. The structures can be divided into three separate α-helical regions: the N- and C-termini are both α-helical, separated by a central kinked α-helix. The overall structures bear an unexpected resemblance to carnocyclin A, a 60-residue peptide that is cyclized via an amide bond between the C- and N-termini and has a saposin fold. Because of synergism observed for other two-peptide leaderless bacteriocins, it was of interest to probe possible binding interactions between enterocins 7A and 7B. However, despite synergistic activity observed between these peptides, no significant binding interaction was observed based on NMR and isothermal calorimetry.

Antimicrobial and Anti-Swarming Effects of Bacteriocins and Biosurfactants from Probiotic Bacterial Strains against Proteus spp.

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Laila Goudarzi

2017-02-01

Full Text Available Background:   Proteus spp. belongs to the family of Enterobacteriaceae. These bacteria are Gram-negative and motile microorganisms and known as the third most common causes of urinary tract infections. The aim of the current study was to investigate the effects of some secondary metabolites from probiotic strains of Lactobacillus spp. on swarming and growth of Proteus mirabilis and P. vulgaris. Methods:   After determination of optimal conditions for the growth and production of antimicrobials, bacteriocins and biosurfactants were partially purified from Lactobacillus culture supernatants. Then, effects of the purified compounds on growth and swarming migration of Proteus spp. were examined in the presence of various concentrations of semi-purified compounds. Results:  Results showed that the partially purified bacteriocins inhibited Proteus spp. swarming distance and had a significant reduction on the bacterial growth curves. Biosurfactants in a solvent form did not have any considerable effects on factors produced by Proteus spp. Conclusion:  According to the results, the secondary metabolites, especially bacteriocins or bacteriocin-like substances derived from Lactobacillus strains, can inhibit or reduce growth and swarming migration of Proteus spp. which are considered as the bacteria major virulence factors.

Bacteriocin and cellulose production by lactic acid bacteria isolated ...

African Journals Online (AJOL)

Sixteen colonies of lactic acid bacteria (LAB) were selected and screened for their ability to produce bacteriocin by agar well diffusion method using the supernatant of centrifuged test cultures. Four isolates inhibited the growth of Listeria monocytogenes and Escherichia coli. Lactobacillus plantarum (6) and Lactobacillus ...

Influence of culture media, pH and temperature on growth and bacteriocin production of bacteriocinogenic lactic acid bacteria.

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Yang, En; Fan, Lihua; Yan, Jinping; Jiang, Yueming; Doucette, Craig; Fillmore, Sherry; Walker, Bradley

2018-01-24

There has been continued interest in bacteriocins research from an applied perspective as bacteriocins have potential to be used as natural preservative. Four bacteriocinogenic lactic acid bacteria (LAB) strains of Lactobacillus curvatus (Arla-10), Enterococcus faecium (JFR-1), Lactobacillus paracasei subsp. paracasei (JFR-5) and Streptococcus thermophilus (TSB-8) were previously isolated and identified in our lab. The objective of this study was to determine the optimal growth conditions for both LAB growth and bacteriocins production. In this study, various growth conditions including culture media (MRS and BHI), initial pH of culture media (4.5, 5.5, 6.2, 7.4 and 8.5), and incubation temperatures (20, 37 and 44 °C) were investigated for LAB growth measured as optical density (OD), bacteriocin activity determined as arbitrary unit and viability of LAB expressed as log CFU ml -1 . Growth curves of the bacteriocinogenic LAB were generated using a Bioscreen C. Our results indicated that Arla-10, JFR-1, and JFR-5 strains grew well on both MRS and BHI media at growth temperature tested whereas TSB-8 strain, unable to grow at 20 °C. LAB growth was significantly affected by the initial pH of culture media (p < 0.001) and the optimal pH was found ranging from 6.2 to 8.5. Bacteriocin activity was significantly different in MRS versus BHI (p < 0.001), and the optimal condition for LAB to produce bacteriocins was determined in MRS broth, pH 6.2 at 37 °C. This study provides useful information on potential application of bacteriocinogenic LAB in food fermentation processes.

Isolation of Lactic Acid Bacteria That Produce Protease and Bacteriocin-Like Substance From Mud Crab (Scylla sp. Digestive Tract (Isolasi Bakteri Asam Laktat yang Menghasilkan Protease dan Senyawa Bacteriocin-Like dari Saluran Pencernaan Kepiting

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Heru Pramono

2015-03-01

Kata kunci: Bakteri Asam Laktat, Bakteriosin-like substance, Protease, Scylla  sp. Digestive tract is complex environment consist of large amount of bacteria’s species. Fish intestine bacteria consist of aerobic or facultative anaerob bacteria which can produce antibacterial and enzym. The objectives of this research were to isolated lactic acid bacteria that produce bacteriocin-like and protease from mud crab digestive tract. Isolation and characterization of isolates were conducted employing media MRS.  Neutralized cell free supernatant of isolates were tested using disc diffusion agar of against pathogenic and spoilage bacteria to indicate bacteriocin-like-producing lactic acid bacteria. Protease-producing isolate was tested using disc diffusion method in casein agar. Among a hundred isolates, 96 isolates were showed clear zone in MRS+CaCO3,, catalase negative, and Gram positive bacteria. Thirty four isolates produced protease and only four isolates (i.e. IKP29, IKP30, IKP52, and IKP94 showed strong inhibition against pathogenic and spoilage bacteria. There were three patterns of inhibition among three isolates against Bacillus subtilis, Staphylococcus aureus, Eschericia coli, and Salmonella sp. All three isolates showed potential uses for produce starter culture for fishery product fermentation purpose. This is the first report of isolation lactic acid bacteria that produced protease and bacteriocin-like from digestive tract of mud crab. Keywords: Lactic acid bacteria, Bacteriocin-like substance, Protease, Scylla  sp.

Growth of Enterococcus durans E204 producing bacteriocin-like ...

African Journals Online (AJOL)

Bacteriocin-like substance E204 is an antimicrobial compound produced by Enterococcus durans E204 isolated from camel milk of Morocco that shows a broad spectrum of inhibitory activity against taxonomically related microorganisms. It is sensitive to digestive proteases. In the first study, de Man, Regosa and Sharpe ...

Purification and characterization of two bacteriocins produced by lactic acid bacteria isolated from Mongolian airag.

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Batdorj, B; Dalgalarrondo, M; Choiset, Y; Pedroche, J; Métro, F; Prévost, H; Chobert, J-M; Haertlé, T

2006-10-01

The aim of this study was to isolate and identify bacteriocin-producing lactic acid bacteria (LAB) issued from Mongolian airag (traditional fermented mare's milk), and to purify and characterize bacteriocins produced by these LAB. Identification of the bacteria (Enterococcus durans) was carried out on the basis of its morphological, biochemical characteristics and carbohydrate fermentation profile and by API50CH kit and 16S rDNA analyses. The pH-neutral cell-free supernatant of this bacterium inhibited the growth of several Lactobacillus spp. and food-borne pathogens including Escherichia coli, Staphylococcus aureus and Listeria innocua. The antimicrobial agent (enterocin A5-11) was heat stable and was not sensitive to acid and alkaline conditions (pH 2-10), but was sensitive to several proteolytic enzymes. Its inhibitory activity was completely eliminated after treatment with proteinase K and alpha-chymotrypsin. The activity was however not completely inactivated by other proteases including trypsin and pepsin. Three-step purification procedure with high recovery yields was developed to separate two bacteriocins. The applied procedure allowed the recovery of 16% and 64% of enterocins A5-11A and A5-11B, respectively, present in the culture supernatant with purity higher than 99%. SDS-PAGE analyses revealed that enterocin A5-11 has a molecular mass of 5000 Da and mass spectrometry analyses demonstrates molecular masses of 5206 and 5218 Da for fractions A and B, respectively. Amino acid analyses of both enterocins indicated significant quantitative difference in their contents in threonine, alanine, isoleucine and leucine. Their N-termini were blocked hampering straightforward Edman degradation. Bacteriocins A5-11A and B from Ent. durans belong to the class II of bacteriocins. Judging from molecular masses, amino acid composition and spectrum of activities, bacteriocins A5-11A and B from Ent. durans show high degree of similarity with enterocins L50A and L50B

Production, purification, and characterization of micrococcin GO5, a bacteriocin produced by Micrococcus sp. GO5 isolated from kimchi.

Science.gov (United States)

Kim, Mi-Hee; Kong, Yoon-Jung; Baek, Hong; Hyun, Hyung-Hwan

2005-01-01

Strain GO5, a bacteriocin-producing bacterium, was isolated from green onion kimchi and identified as Micrococcus sp. The bacteriocin, micrococcin GO5, displayed a broad spectrum of inhibitory activity against a variety of pathogenic and nonpathogenic microorganisms, as tested by the spot-on-lawn method; its activity spectrum was almost identical to that of nisin. Micrococcin GO5 was inactivated by trypsin (whereas nisin was not) and was completely stable at 100 degrees C for 30 min and in the pH range of 2.0 to 7.0. Micrococcin GO5 exhibited a typical mode of bactericidal activity against Micrococcus flavus ATCC 10240. It was purified to homogeneity through ammonium sulfate precipitation, ultrafiltration, and CM-Sepharose column chromatography. The molecular mass of micrococcin GO5 was estimated to be about 5.0 kDa by tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis and in situ activity assay with the indicator organism. The amino acid sequence of micrococcin GO5 lacks lanthionine and beta-methyllanthionine and is rich in hydrophobic amino acids and glycine, providing the basis for the high heat stability of this bacteriocin. The N-terminal amino acid sequence of micrococcin GO5 is Lys-Lys-Ser-Phe-Cys-Gln-Lys, and no homology to bacteriocins reported previously was observed in the amino acid composition or N-terminal amino acid sequence. Based on the physicochemical properties, small molecular size, and inhibition of Listeria monocytogenes, micrococcin GO5 has been placed with the class II bacteriocins, but its broad spectrum of activity differs from that of other bacteriocins in this class.

Active polymers containing Lactobacillus curvatus CRL705 bacteriocins: effectiveness assessment in Wieners.

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Blanco Massani, M; Molina, V; Sanchez, M; Renaud, V; Eisenberg, P; Vignolo, G

2014-05-16

Bacteriocins from lactic acid bacteria have potential as natural food preservatives. In this study two active (synthetic and gluten) films were obtained by the incorporation of lactocin 705 and lactocin AL705, bacteriocins produced by Lactobacillus curvatus CRL705 with antimicrobial activity against spoilage lactic acid bacteria and Listeria. Antimicrobial film effectiveness was determined in Wieners inoculated with Lactobacillus plantarum CRL691 and Listeria innocua 7 (10(4)CFU/g) stored at 5°C during 45days. Active and control (absence of bacteriocins) packages were prepared and bacterial counts in selective media were carried out. Visual inspection and pH measurement of Wieners were also performed. Typical growth of both inoculated microorganisms was observed in control packages which reached 10(6)-10(7)CFU/g at the end of storage period. In the active packages, L. innocua 7 was effectively inhibited (2.5 log cycles reduction at day 45), while L. plantarum CRL691 was only slightly inhibited (0.5 log cycles) up to the second week of storage, then counts around 10(6)-10(7)CFU/g were reached. Changes in pH values from 6.3 to 5.8 were produced and gas formation was observed in active and control packages. The low inhibitory effectiveness against lactic acid bacteria is in correlation with the low activity observed for lactocin 705 in the presence of fat; Wieners fat content (20-30%) may adversely affect antimicrobial activity. This study supports the feasibility of using polymers activated with L. curvatus CRL705 bacteriocins to control Listeria on the surface of Wieners and highlights the importance of evaluating antimicrobial packaging systems for each particular food application. Copyright © 2014 Elsevier B.V. All rights reserved.

Purification and genetic characterisation of the novel bacteriocin LS2 produced by the human oral strain Lactobacillus salivarius BGHO1.

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Busarcevic, Milos; Dalgalarrondo, Michèle

2012-08-01

The aim of this study was to investigate the antimicrobial potential of Lactobacillus salivarius BGHO1, a human oral strain with probiotic characteristics and a broad inhibitory spectrum both against Gram-positive and Gram-negative pathogens. Here we present the bacteriocin LS2, an extremely pH- and heat-stable peptide with antilisterial activity. LS2 is a novel member of the class IId bacteriocins, unique among all currently characterised bacteriocins. It is somewhat similar to putative bacteriocins from several oral streptococci, including the cariogenic Streptococcus mutans. LS2 is a 41-amino-acid, highly hydrophobic cationic peptide of 4115.1Da that is sensitive to proteolytic enzymes. LS2 was purified from cells of strain BGHO1 by solvent extraction and reverse-phase chromatography. Mass spectrometry was used to determine the molecular mass of the purified peptide. N-terminal amino acid sequencing enabled identification of the LS2 structural gene bacls2 by a reverse genetics approach. Downstream of the bacls2 gene, two bacteriocin-like genes were found, named blp1a and blp1b, and one putative bacteriocin immunity gene named bimlp. We also present the identification of the 242-kb megaplasmid pMPHO1 by pulsed-field gel electrophoresis, which harbours the genes bacls2, blp1a, blp1b and bimlp. Two peptides with antimicrobial activity, whose approximate sizes corresponded to those of blp1a and blp1b, were identified only after culturing strain BGHO1 in a chemically defined medium. This study demonstrated the capacity of Lactobacillus salivarius BGHO1 to produce multiple bacteriocins and further established this strain as a promising probiotic candidate. Copyright © 2012 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

Description of durancin TW-49M, a novel enterocin B-homologous bacteriocin in carrot-isolated Enterococcus durans QU 49.

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Hu, C-B; Zendo, T; Nakayama, J; Sonomoto, K

2008-09-01

To characterize the novel bacteriocin produced by Enterococcus durans. Enterococcus durans QU 49 was isolated from carrot and expressed bactericidal activity over 20-43 degrees C. Bacteriocins were purified to homogeneity using the three-step purification method, one of which, termed durancin TW-49M, was an enterocin B-homologous peptide with most identical residues occurring in the N-terminus. Durancin TW-49M was more tolerant in acidic than in alkali. DNA sequencing analysis revealed durancin TW-49M was translated as a prepeptide of the double-glycine type. Durancin TW-49M and enterocin B expressed similar antimicrobial spectra, in which no significant variation due to the diversity in their C-termini was observed. Durancin TW-49M, a novel nonpediocin-like class II bacteriocin, was characterized to the amino acid and genetic levels. The diverse C-terminal parts of durancin TW-49M and enterocin B were hardly to be suggested as the place determining the target cell specificity. This is the first and comprehensive study of a novel bacteriocin produced by Ent. durans. The high homology at the N-terminal halves between durancin TW-49M and enterocin B makes them suitable to study the structure-function relationship of bacteriocins and their immunity proteins.

The study of Bacteriocin of Pseudomonas fluorescens and Citrus limon effects against Propionibacterium acnes and Staphylococcus epidermidis in acne patients

Science.gov (United States)

Ahmed, Mais E.

2018-05-01

Research was carried out on the antibacterial effect of (Citrus limon) juice on Acnevulgaris. Samples were obtained from individuals with pimples by swabbing their faces. Natural substances that derive from plants are promising to treat disease cause Acnevulgaris, the study in vitro biological activity of the juice, as well as bacterocin cultivated and fruits was investigated on two strains of bacteria (Propionibacterium acnes, Staphylococcus epidermidis). The new antimicrobial (bacteriocin and Citrus juice) is an ongoing search. This study used juice at different concentrations at (20%, 30%, 40%, 60%, 80% and 100%). The bacteriocin produced from local P. fluorescens isolates from wound infection and majority of isolates were found to produce crude bacteriocin were (P1 and P2) in Pseudomonas agar at 37°C for 24 hrs. Crude bacteriocin and Citrus limon juice against some pathogenic skin bacteria was find to be effective juice Citrus limon aganist S. epidermidis at 100% Concentrations with a range of inhibition zone (18) mm. The isolates of P. fluorescens (P2) was positive as producer of bacteriocin with a wide inhibition growth against gram positive pathogenic bacteria with a range between (10-12) mm.

Purification and Characterization of Bacteriocin Produced by Bacillus subtilis R75 Isolated from Fermented Chunks of Mung Bean (Phaseolus radiatus

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Riti Kapoor

2011-01-01

Full Text Available Food-grade bacteria capable of producing bacteriocin with desirable preservation attributes have been isolated from traditional Indian fermented food dal vari, which has not been investigated so far. Among different isolates, Bacillus subtilis R75, isolated on MRS agar, exhibited antagonism against a wide range of foodborne pathogens that cause serious spoilage. Extracellularly produced bacteriocin was purified by single step gel exclusion column chromatography. The purity rate and molecular mass of 12 kDa of this compound were determined using SDS-PAGE. Activity units (AU of bacteriocin were increased in each step of purification, reaching up to 5·10^6 AU/mL. The increase in the activity units directly affected the antimicrobial activity of purified bacteriocin, resulting in an increase up to 200, 333 and 175 % of the inhibition zones against indicator bacteria. Continuous decrease in the number of viable cells of microorganisms within 10 h after adding purified bacteriocin proved its bactericidal action. It withstood very high temperature, up to 121 °C, for 10 min, wider pH range, from 4.0 to 11.0, complete inactivation in the presence of proteolytic enzymes and storage stability up to 2.5 months.

Spatial attributes of the four-helix bundle group of bacteriocins – The high-resolution structure of BacSp222 in solution

KAUST Repository

Nowakowski, Michał

2017-11-01

BacSp222 is a multifunctional bacteriocin produced by Staphylococcus pseudintermedius strain 222, an opportunistic pathogen of domestic animals. At micromolar concentrations, BacSp222 kills Gram-positive bacteria and is cytotoxic toward mammalian cells, while at nanomolar doses, it acts as an immunomodulatory factor, enhancing nitric oxide release in macrophage-like cell lines. The bacteriocin is a cationic, N-terminally formylated, 50-amino-acid-long linear peptide that is rich in tryptophan residues.In this study, the solution structure of BacSp222 was determined and compared to the currently known structures of similar bacteriocins. BacSp222 was isolated from a liquid culture medium in a uniformly 13C- and 15N-labeled form, and NMR data were collected. The structure was calculated based on NMR-derived constraints and consists of a rigid and tightly packed globular bundle of four alpha-helices separated by three short turns.Although the amino acid sequence of BacSp222 has no significant similarity to any known peptide or protein, a 3D structure similarity search indicates a close relation to other four-helix bundle-motif bacteriocins, such as aureocin A53, lacticin Q and enterocins 7A/7B. Assuming similar functions, biology, structure and physicochemical properties, we propose to distinguish the four-helix bundle bacteriocins as a new Type A in subclass IId of bacteriocins, containing linear, non-pediocin-like peptides.

Purification and genetic characterization of gassericin E, a novel co-culture inducible bacteriocin from Lactobacillus gasseri EV1461 isolated from the vagina of a healthy woman.

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Maldonado-Barragán, Antonio; Caballero-Guerrero, Belén; Martín, Virginia; Ruiz-Barba, José Luis; Rodríguez, Juan Miguel

2016-03-12

Lactobacillus gasseri is one of the dominant Lactobacillus species in the vaginal ecosystem. Some strains of this species have a high potential for being used as probiotics in order to maintain vaginal homeostasis, since they may confer colonization resistance against pathogens in the vagina by direct inhibition through production of antimicrobial compounds, as bacteriocins. In this work we have studied bacteriocin production of gassericin E (GasE), a novel bacteriocin produced by L. gasseri EV1461, a strain isolated from the vagina of a healthy woman, and whose production was shown to be promoted by the presence of certain specific bacteria in co-culture. Biochemical and genetic characterization of this novel bacteriocin are addressed. We found that the inhibitory spectrum of L. gasseri EV1461 was broad, being directed to species both related and non-related to the producing strain. Interestingly, L. gasseri EV1461 inhibited the grown of pathogens usually associated with bacterial vaginosis (BV). The antimicrobial activity was due to the production of a novel bacteriocin, gassericin E (GasE). Production of this bacteriocin in broth medium only was achieved at high cell densities. At low cell densities, bacteriocin production ceased and only was restored after the addition of a supernatant from a previous bacteriocin-producing EV1461 culture (autoinduction), or through co-cultivation with several other Gram-positive strains (inducing bacteria). DNA sequence of the GasE locus revealed the presence of two putative operons which could be involved in biosynthesis and immunity of this bacteriocin (gaeAXI), and in regulation, transport and processing (gaePKRTC). The gaePKR encodes a putative three-component regulatory system, involving an autoinducer peptide (GaeP), a histidine protein kinase (GaeK) and a response regulator (GaeR), while the gaeTC encodes for an ABC transporter (GaeT) and their accessory protein (GaeC), involved in transport and processing of the

Quorum Sensing Regulation of Competence and Bacteriocins in Streptococcus pneumoniae and mutans

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Shanker, Erin; Federle, Michael J.

2017-01-01

The human pathogens Streptococcus pneumoniae and Streptococcus mutans have both evolved complex quorum sensing (QS) systems that regulate the production of bacteriocins and the entry into the competent state, a requirement for natural transformation. Natural transformation provides bacteria with a mechanism to repair damaged genes or as a source of new advantageous traits. In S. pneumoniae, the competence pathway is controlled by the two-component signal transduction pathway ComCDE, which directly regulates SigX, the alternative sigma factor required for the initiation into competence. Over the past two decades, effectors of cellular killing (i.e., fratricides) have been recognized as important targets of the pneumococcal competence QS pathway. Recently, direct interactions between the ComCDE and the paralogous BlpRH pathway, regulating bacteriocin production, were identified, further strengthening the interconnections between these two QS systems. Interestingly, a similar theme is being revealed in S. mutans, the primary etiological agent of dental caries. This review compares the relationship between the bacteriocin and the competence QS pathways in both S. pneumoniae and S. mutans, and hopes to provide clues to regulatory pathways across the genus Streptococcus as a potential tool to efficiently investigate putative competence pathways in nontransformable streptococci. PMID:28067778

Cloning, purification, and functional characterization of Carocin S2, a ribonuclease bacteriocin produced by Pectobacterium carotovorum

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Tzeng Kuo-Ching

2011-05-01

Full Text Available Abstract Background Most isolates of Pectobacterium carotovorum subsp. carotovorum (Pcc produce bacteriocins. In this study, we have determined that Pcc strain F-rif-18 has a chromosomal gene encoding the low-molecular-weight bacteriocin, Carocin S2, and that this bacteriocin inhibits the growth of a closely related strain. Carocin S2 is inducible by ultraviolet radiation but not by mutagenic agents such as mitomycin C. Results A carocin S2-defective mutant, TF1-2, was obtained by Tn5 insertional mutagenesis using F-rif-18. A 5706-bp DNA fragment was detected by Southern blotting, selected from a genomic DNA library, and cloned to the vector, pMS2KI. Two adjacent complete open reading frames within pMS2KI were sequenced, characterized, and identified as caroS2K and caroS2I, which respectively encode the killing protein and immunity protein. Notably, carocin S2 could be expressed not only in the mutant TF1-2 but also in Escherichia coli DH5α after entry of the plasmid pMS2KI. Furthermore, the C-terminal domain of CaroS2K was homologous to the nuclease domains of colicin D and klebicin D. Moreover, SDS-PAGE analysis showed that the relative mass of CaroS2K was 85 kDa and that of CaroS2I was 10 kDa. Conclusion This study shown that another nuclease type of bacteriocin was found in Pectobacterium carotovorum. This new type of bacteriocin, Carocin S2, has the ribonuclease activity of CaroS2K and the immunity protein activity of CaroS2I.

Rapid Two-Step Procedure for Large-Scale Purification of Pediocin-Like Bacteriocins and Other Cationic Antimicrobial Peptides from Complex Culture Medium

OpenAIRE

Uteng, Marianne; Hauge, Håvard Hildeng; Brondz, Ilia; Nissen-Meyer, Jon; Fimland, Gunnar

2002-01-01

A rapid and simple two-step procedure suitable for both small- and large-scale purification of pediocin-like bacteriocins and other cationic peptides has been developed. In the first step, the bacterial culture was applied directly on a cation-exchange column (1-ml cation exchanger per 100-ml cell culture). Bacteria and anionic compounds passed through the column, and cationic bacteriocins were subsequently eluted with 1 M NaCl. In the second step, the bacteriocin fraction was applied on a lo...

PRODUCTION OF BACTERIOCIN EC2 AND ITS INTERFERENCE IN THE GROWTH OF SALMONELLA TYPHI IN A MILK MATRIX

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Yuri de Jesus Lopes de Abreu

2013-08-01

Full Text Available Bacterial interference can occur through various mechanisms, including the production of peroxides, acids, ammonia, bacteriolytic enzymes or bacteriocins. The strain Escherichia coli EC2 produces the antimicrobial substance (AMS EC2, able to inhibit different strains of Gram-negative bacteria isolated from food, as E. coli and Salmonella sp. The activity of AMS EC2 was lost after treatment with proteolytic enzymes, indicating the presence of an active proteinaceous compound, suggesting that it is a bacteriocin. The substance, renamed bacteriocin EC2, has its better production when the producer strain is grown on Casoy medium, at 37ºC and pH 6.0, without NaCl addition, but it is also able to be produced in milk. When co-cultivated in UHT milk with the producer strain E. coli EC2, the growth of the indicator strain Salmonella Typhi is totally inhibited within the first 4 hours of incubation, suggesting a potential application of bacteriocin EC2 in the control of Salmonella sp. e.g. in foods.

Class IIa bacteriocin resistance in Enterococcus faecalis V583: The mannose PTS operon mediates global transcriptional responses

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Opsata Mona

2010-08-01

Full Text Available Abstract Background The class IIa bacteriocin, pediocin PA-1, has clear potential as food preservative and in the medical field to be used against Gram negative pathogen species as Enterococcus faecalis and Listeria monocytogenes. Resistance towards class IIa bacteriocins appear in laboratory and characterization of these phenotypes is important for their application. To gain insight into bacteriocin resistance we studied mutants of E. faecalis V583 resistant to pediocin PA-1 by use of transcriptomic analyses. Results Mutants of E. faecalis V583 resistant to pediocin PA-1 were isolated, and their gene expression profiles were analyzed and compared to the wild type using whole-genome microarray. Significantly altered transcription was detected from about 200 genes; most of them encoding proteins involved in energy metabolism and transport. Glycolytic genes were down-regulated in the mutants, but most of the genes showing differential expression were up-regulated. The data indicate that the mutants were relieved from glucose repression and putative catabolic responsive elements (cre could be identified in the upstream regions of 70% of the differentially expressed genes. Bacteriocin resistance was caused by reduced expression of the mpt operon encoding the mannose-specific phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS, and the same transcriptional changes were seen in a mptD-inactivated mutant. This mutant also had decreased transcription of the whole mpt operon, showing that the PTS is involved in its own transcriptional regulation. Conclusion Our data confirm the important role of mannose PTS in class IIa bacteriocin sensitivity and we demonstrate its importance involving global carbon catabolite control.

Rapid and Efficient Purification Method for Small, Hydrophobic, Cationic Bacteriocins: Purification of Lactococcin B and Pediocin PA-1

OpenAIRE

Venema, K.; Chikindas, M. L.; Seegers, J.; Haandrikman, A. J.; Leenhouts, K. J.; Venema, G.; Kok, J.

1997-01-01

The bacteriocins lactococcin B and pediocin PA-1 were purified by ethanol precipitation, preparative isoelectric focusing, and ultrafiltration. The procedure reproducibly leads to high final yields in comparison to the generally low yields obtained by column chromatography. Specifically, during isoelectric focusing no loss of activity occurs. The method, in general, should be applicable to small, hydrophobic, cationic bacteriocins.

Genetic features of circular bacteriocins produced by Gram-positive bacteria

NARCIS (Netherlands)

Maqueda, Mercedes; Sánchez-Hidalgo, Marina; Fernández, Matilde; Montalbán-López, Manuel; Valdivia, Eva; Martínez-Bueno, Manuel

This review highlights the main genetic features of circular bacteriocins, which require the co-ordinated expression of several genetic determinants. In general terms, it has been demonstrated that the expression of such structural genes must be combined with the activity of proteins involved in

Identification of the Propionicin F Bacteriocin Immunity Gene (pcfI) and Development of a Food-Grade Cloning System for Propionibacterium freudenreichii▿ †

OpenAIRE

Brede, Dag Anders; Lothe, Sheba; Salehian, Zhian; Faye, Therese; Nes, Ingolf F.

2007-01-01

This report describes the first functional analysis of a bacteriocin immunity gene from Propionibacterium freudenreichii and its use as a selection marker for food-grade cloning. Cloning of the pcfI gene (previously orf5 [located as part of the pcfABC propionicin F operon]) rendered the sensitive host 1,000-fold more tolerant to the propionicin F bacteriocin. The physiochemical properties of the 127-residue large PcfI protein resemble those of membrane-bound immunity proteins from bacteriocin...

LAB Bacteriocins Controlling the Food Isolated (Drug-Resistant Staphylococci

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Jesús Perales-Adán

2018-06-01

Full Text Available Staphylococci are a group of microorganisms that can be often found in processed food and they might pose a risk for human health. In this study we have determined the content of staphylococci in 7 different fresh goat-milk cheeses. These bacteria were present in all of them, ranging from 103 to 106 CFU/g based on growth on selective media. Thus, a set of 97 colonies was randomly picked for phenotypic and genotypic identification. They could be clustered by RAPD-PCR in 10 genotypes, which were assigned by 16S rDNA sequencing to four Staphylococcus species: Staphylococcus aureus, Staphylococcus chromogenes, S. simulans, and S. xylosus. Representative strains of these species (n = 25 were tested for antibiotic sensitivity, and 11 of them were resistant to at least one of the antibiotics tested, including erythromycin, amoxicillin-clavulanic acid and oxacillin. We also tested two bacteriocins produced by lactic acid bacteria (LAB, namely the circular bacteriocin AS-48 and the lantibiotic nisin. These peptides have different mechanism of action at the membrane level. Nevertheless, both were able to inhibit staphylococci growth at low concentrations ranging between 0.16–0.73 μM for AS-48 and 0.02–0.23 μM for nisin, including the strains that displayed antibiotic resistance. The combined effect of these bacteriocins were tested and the fractional inhibitory concentration index (FICI was calculated. Remarkably, upon combination, they were active at the low micromolar range with a significant reduction of the minimal inhibitory concentration. Our data confirms synergistic effect, either total or partial, between AS-48 and nisin for the control of staphylococci and including antibiotic resistant strains. Collectively, these results indicate that the combined use of AS-48 and nisin could help controlling (pathogenic staphylococci in food processing and preventing antibiotic-resistant strains reaching the consumer in the final products.

Alternatives for biosurfactants and bacteriocins extraction from Lactococcus lactis cultures produced under different pH conditions.

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Rodríguez, N; Salgado, J M; Cortés, S; Domínguez, J M

2010-08-01

Study of the potential of Lactococcus lactis CECT-4434 as a biosurfactants and nisin (the only bacteriocin allowed to be used in the food industry) producer for industrial applications, exploiting the possibility of recovering separately both metabolites, taking into account that L. lactis is an interesting micro-organism with several applications in the food industry because it is recognized as GRAS. The results showed the ability of this strain to produce cell-bound biosurfactants, under controlled pH, and cell-bound biosurfactants and bacteriocins, when pH was not controlled. Three extraction procedures were designed to separately recover these substances. The strain L. lactis CECT-4434 showed to be a cell-bound biosurfactants and bacterocins producer when fermentations were carried out under uncontrolled pH. Both products can be recovered separately. Development of a convenient tool for the extraction of cell-bound biosurfactants and bacteriocins from the fermentation broth.

Purification and Characterization of a Novel Cold Shock Protein-Like Bacteriocin Synthesized by Bacillus thuringiensis.

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Huang, Tianpei; Zhang, Xiaojuan; Pan, Jieru; Su, Xiaoyu; Jin, Xin; Guan, Xiong

2016-10-20

Bacillus thuringiensis (Bt), one of the most successful biopesticides, may expand its potential by producing bacteriocins (thuricins). The aim of this study was to investigate the antimicrobial potential of a novel Bt bacteriocin, thuricin BtCspB, produced by Bt BRC-ZYR2. The results showed that this bacteriocin has a high similarity with cold-shock protein B (CspB). BtCspB lost its activity after proteinase K treatment; however it was active at 60 °C for 30 min and was stable in the pH range 5-7. The partial loss of activity after the treatments of lipase II and catalase were likely due to the change in BtCspB structure and the partial degradation of BtCspB, respectively. The loss of activity at high temperatures and the activity variation at different pHs were not due to degradation or large conformational change. BtCspB did not inhibit four probiotics. It was only active against B. cereus strains 0938 and ATCC 10987 with MIC values of 3.125 μg/mL and 0.781 μg/mL, and MBC values of 12.5 μg/mL and 6.25 μg/mL, respectively. Taken together, these results provide new insights into a novel cold shock protein-like bacteriocin, BtCspB, which displayed promise for its use in food preservation and treatment of B. cereus-associated diseases.

Purification and characterization of plantaricin 163, a novel bacteriocin produced by Lactobacillus plantarum 163 isolated from traditional Chinese fermented vegetables.

Science.gov (United States)

Hu, Meizhong; Zhao, Haizhen; Zhang, Chong; Yu, Jiansheng; Lu, Zhaoxin

2013-11-27

Presumptive lactic acid bacteria (LAB) strains isolated from traditional Chinese fermented vegetables were screened for bacteriocin production. A novel bacteriocin-producing strain, Lactobacillus plantarum 163, was identified on the basis of its physiobiochemical characteristics and characterized by 16S rDNA sequencing. The novel bacteriocin, plantaricin 163, produced by Lb. plantarum 163 was purified by salt precipitation, gel filtration, and reverse-phase high-performance liquid chromatography (RP-HPLC). Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis of plantaricin 163 revealed the molecular weight to be 3553.2 Da. The complete amino acid sequence showed VFHAYSARGNYYGNCPANWPSCRNNYKSAGGK, and no similarity to known bacteriocins was found. Plantaricin 163 was highly thermostable (20 min, 121 °C), active in the presence of acidic pH (3-5), sensitive to protease, and exhibited broad-spectrum antimicrobial activity against LAB and other tested Gram-positive and Gram-negative bacteria. The results suggest that plantaricin 163 may be employed as a biopreservative in the food industry.

Antibacterial efficacy of Nisin, Pediocin 34 and Enterocin FH99 against Listeria monocytogenes and cross resistance of its bacteriocin resistant variants to common food preservatives.

Science.gov (United States)

Kaur, G; Singh, T P; Malik, R K

2013-01-01

Antilisterial efficiency of three bacteriocins, viz, Nisin, Pediocin 34 and Enterocin FH99 was tested individually and in combination against Listeria mononcytogenes ATCC 53135. A greater antibacterial effect was observed when the bacteriocins were combined in pairs, indicating that the use of more than one LAB bacteriocin in combination have a higher antibacterial action than when used individually. Variants of Listeria monocytogenes ATCC 53135 resistant to Nisin, Pediocin 34 and Enterocin FH99 were developed. Bacteriocin cross-resistance of wild type and their corresponding resistant variants were assessed and results showed that resistance to a bacteriocin may extend to other bacteriocins within the same class. Resistance to Pediocin 34 conferred cross resistance to Enterocin FH 99 but not to Nisin. Similarly resistance to Enterocin FH99 conferred cross resistance to Pediocin 34 but not to Nisin. Also, the sensitivity of Nisin, Pediocin 34 and Enterocin FH99 resistant variants of Listeria monocytogenes to low pH, salt, sodium nitrite, and potassium sorbate was assayed in broth and compared to the parental wild-type strain. The Nisin, Pediocin 34 and Enterocin FH99 resistant variants did not have intrinsic resistance to low pH, sodium chloride, potassium sorbate, or sodium nitrite. In no case were the bacteriocin resistant Listeria monocytogenes variants examined were more resistant to inhibitors than the parental strains.

Antibacterial efficacy of Nisin, Pediocin 34 and Enterocin FH99 against Listeria monocytogenes and cross resistance of its bacteriocin resistant variants to common food preservatives

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G. Kaur

2013-01-01

Full Text Available Antilisterial efficiency of three bacteriocins, viz, Nisin, Pediocin 34 and Enterocin FH99 was tested individually and in combination against Listeria mononcytogenes ATCC 53135. A greater antibacterial effect was observed when the bacteriocins were combined in pairs, indicating that the use of more than one LAB bacteriocin in combination have a higher antibacterial action than when used individually. Variants of Listeria monocytogenes ATCC 53135 resistant to Nisin, Pediocin 34 and Enterocin FH99 were developed. Bacteriocin cross-resistance of wild type and their corresponding resistant variants were assessed and results showed that resistance to a bacteriocin may extend to other bacteriocins within the same class. Resistance to Pediocin 34 conferred cross resistance to Enterocin FH 99 but not to Nisin. Similarly resistance to Enterocin FH99 conferred cross resistance to Pediocin 34 but not to Nisin. Also, the sensitivity of Nisin, Pediocin 34 and Enterocin FH99 resistant variants of Listeria monocytogenes to low pH, salt, sodium nitrite, and potassium sorbate was assayed in broth and compared to the parental wild-type strain. The Nisin, Pediocin 34 and Enterocin FH99 resistant variants did not have intrinsic resistance to low pH, sodium chloride, potassium sorbate, or sodium nitrite. In no case were the bacteriocin resistant Listeria monocytogenes variants examined were more resistant to inhibitors than the parental strains.

Technological properties of bacteriocin-producing lactic acid bacteria isolated from Pico cheese an artisanal cow's milk cheese.

Science.gov (United States)

Ribeiro, S C; Coelho, M C; Todorov, S D; Franco, B D G M; Dapkevicius, M L E; Silva, C C G

2014-03-01

Evaluate technologically relevant properties from bacteriocin-producing strains to use as starter/adjunct cultures in cheese making. Eight isolates obtained from Pico cheese produced in Azores (Portugal) were found to produce bacteriocins against Listeria monocytogenes and three isolates against Clostridium perfringens. They were identified as Lactococcus lactis and Enterococcus faecalis and submitted to technological tests: growth at different conditions of temperature and salt, acid production, proteolysis, lipolysis, coexistence, enzymatic profile and autolytic capacity. Safety evaluation was performed by evaluating haemolytic, gelatinase and DNase activity, resistance to antibiotics and the presence of virulence genes. Some isolates presented good technological features such as high autolytic activity, acid and diacetyl production. Lactococcus lactis was negative for all virulence genes tested and inhibit the growth of all Lactic acid bacteria (LAB) isolates. Enterococci were positive for the presence of some virulence genes, but none of the isolates were classified as resistant to important antibiotics. The bacteriocin-producing Lc. lactis present good potential for application in food as adjunct culture in cheese production. The study also reveals good technological features for some Enterococcus isolates. Bacteriocin-producing strains presented important technological properties to be exploited as new adjunct culture for the dairy industry, influencing flavour development and improve safety. © 2013 The Society for Applied Microbiology.

Characterization of Bacteriocin like inhibitory substance produced by a new Strain Brevibacillus borstelensis AG1 Isolated from 'Marcha'.

Science.gov (United States)

Sharma, Nivedita; Gupta, Anupama; Gautam, Neha

2014-01-01

In the present study, a bacterium isolated from Marcha- a herbal cake used as traditional starter culture to ferment local wine in North East India, was evaluated for bacteriocin like inhibitory substance production and was tested against six food borne/spoilage causing pathogens viz. Listeria monocytogenes MTCC 839, Bacillus subtilis MTCC 121, Clostridium perfringens MTCC 450, Staphylococcus aureus, Lactobacillus plantarum and Leuconostoc mesenteroides MTCC 107 by using bit/disc method followed by well diffusion method. The bacterial isolate was identified as Brevibacillus borstelensis on the basis of phenotypic, biochemical and molecular characteristics using 16Sr RNA gene technique. Bacteriocin like inhibitory substance produced by Brevibacillus borstelensis AG1 was purified by gel exclusion chromatography. The molecular mass of the Brevibacillus borstelensis AG1 was found to be 12 kDa. Purified bacteriocin like inhibitory substance of Brevibacillus borstelensis was further characterized by studying the effect of temperature, pH, proteolytic enzyme and stability. Bacteriocin like inhibitory substance was found to be thermostable upto 100 °C, active at neutral pH, sensitive to trypsin, and partially stable till third week of storage thus showing a bright prospective to be used as a potential food biopreservative.

Partial purification and characterization of a bacteriocin produced by Enterococcus faecium 130 isolated from mozzarella cheese

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Fabrício Luiz Tulini

2011-03-01

Full Text Available Lactic acid bacteria are important in foods as potential probiotics and also due to the ability to produce antimicrobial compounds that can contribute for biopreservation. In this work, the bacteriocin produced by the food isolate Enterococcus faecium 130 was partially purified and characterized. The compound was active against Gram-positive bacteria, including Listeria monocytogenes. It was produced after 4 days of storage at a broad temperature range (4 to 37 °C; it was stable at pH ranging from 2 to 10 with no loss of activity after heating at 100 °C for 15 minutes. Bacteriocin was partially purified by the adsorption-desorption technique, and the analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE showed a molecular mass of 3.5 to 6.5 kDa. These data encourage studies on application of this bacteriocin in food systems as an additional hurdle to microbial growth.

Purification and characterization of a bacteriocin from an oenological strain of Leuconostoc mesenteroides subsp. cremoris.

Science.gov (United States)

Dündar, Halil; Salih, Bekir; Bozoğlu, Faruk

2016-05-18

Malolactic fermentation (MLF), which improves organoleptic properties and biologic stability of some wines, may cause wine spoilage if uncontrolled. Bacteriocins were reported as efficient preservatives to control MLF through their bactericidal effect on malolactic bacteria. Leuconostoc mesenteroides subsp. cremoris W3 isolated from wine produces an inhibitory substance that is bactericidal against malolactic bacteria in model wine medium. Treatment of the culture supernatant of strain W3 with proteases eliminated the inhibitory activity, which proved that it is a true bacteriocin and we tentatively termed it mesentericin W3. The bacteriocin inhibited the growth of food-borne pathogenic bacteria such as Enterococcus faecalis, Listeria monocytogenes, and malolactic bacteria. It was active over a wide pH range and stable to organic solvents and heat. Mesentericin W3 was purified to homogeneity by a pH-mediated cell adsorption-desorption method, cation exchange, hydrophobic interaction, and reverse-phase chromatography. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectroscopy (MS) and partial amino acid sequence analysis revealed that mesentericin W3 was identical to mesentericin Y105.

In vitro evaluation of the antimicrobial effect of a raw bacteriocin extract in combination with chemical preservatives employed in meat industry

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Luis A. Aguado Bautista

2010-12-01

Full Text Available Biopreservation can be defined as the foods shelf life extension employing antibacterial products like bacteriocins. The objective of this work was to determinate the efficacy of E. faecium MXVK29 bacteriocin in combination with chemical preservatives against spoilage and pathogens microorganisms. Bacteriocin raw extrac antimicrobial activity was 46.34 UA/g of protein. Growth of Pseudomonas putida was not affected by the preservatives employed at the conditions employed. Antimicrobial response was different for other microorganisms since a synergetic effect of the preservatives combination inhibited Brochothrix thermosphacta and Escherichia coli growth. Sodium lactate had additive effect only against Listeria innocua.

Relationships between MDR proteins, bacteriocin production and proteolysis in Lactococcus lactis

NARCIS (Netherlands)

Gajic, Olivera

2003-01-01

The Gram-positive lactic acid bacterium Lactococcus lactis can harbour a wide variety of circular extrachromosomal DNA molecules, so-called plasmids. Many of the traits that make them useful for manufacturing of fermented food products (e.g. bacteriophage resistance, bacteriocin and proteinase

Coordinated Bacteriocin Expression and Competence in Streptococcus pneumoniae Contributes to Genetic Adaptation through Neighbor Predation.

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Wei-Yun Wholey

2016-02-01

Full Text Available Streptococcus pneumoniae (pneumococcus has remained a persistent cause of invasive and mucosal disease in humans despite the widespread use of antibiotics and vaccines. The resilience of this organism is due to its capacity for adaptation through the uptake and incorporation of new genetic material from the surrounding microbial community. DNA uptake and recombination is controlled by a tightly regulated quorum sensing system that is triggered by the extracellular accumulation of competence stimulating peptide (CSP. In this study, we demonstrate that CSP can stimulate the production of a diverse array of blp bacteriocins. This cross stimulation occurs through increased production and secretion of the bacteriocin pheromone, BlpC, and requires a functional competence regulatory system. We show that a highly conserved motif in the promoter of the operon encoding BlpC and its transporter mediates the upregulation by CSP. The accumulation of BlpC following CSP stimulation results in augmented activation of the entire blp locus. Using biofilm-grown organisms as a model for competition and genetic exchange on the mucosal surface, we demonstrate that DNA exchange is enhanced by bacteriocin secretion suggesting that co-stimulation of bacteriocins with competence provides an adaptive advantage. The blp and com regulatory pathways are believed to have diverged and specialized in a remote ancestor of pneumococcus. Despite this, the two systems have maintained a regulatory connection that promotes competition and adaptation by targeting for lysis a wide array of potential competitors while simultaneously providing the means for incorporation of their DNA.

Cysteine S-glycosylation, a new post-translational modification found in glycopeptide bacteriocins

Czech Academy of Sciences Publication Activity Database

Stepper, J.; Shastri, S.; Loo, T. S.; Preston, J. C.; Novák, Petr; Man, Petr; Moore, Ch. H.; Havlíček, Vladimír; Patchett, M. L.; Norris, G. E.

2011-01-01

Roč. 585, č. 4 (2011), s. 645-650 ISSN 0014-5793 Institutional research plan: CEZ:AV0Z50200510 Keywords : Post-translational modification * Glycosylation * Bacteriocin Subject RIV: CE - Biochemistry Impact factor: 3.538, year: 2011

Exploration and conservation of bacterial genetic resources as bacteriocin producing inhibitory microorganisms to pathogen bacteria in livestock

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Chotiah S

2013-06-01

Full Text Available Exploration and conservation of microorganisms producing bacteriocin was done as the primary study towards the collection of potential bacteria and its application in improving livestock health condition and inhibit food borne pathogens. Diferent kinds of samples such as beef cattle rectal swab, rumen fluids, cow’s milk, chicken gut content, goat’s milk were collected at Bogor cattle slaughter houses, poultry slaughter houses, dairy cattle and goat farms. A total of 452 bacterial isolates consisted of 73 Gram negative bacteria and 379 Gram positive bacteria were isolated from samples collected and screened for bacteriocin activity. Determination of bacteriocin activity with bioassay using agar spot tests were carried out on liquid and semisolid medium assessing 8 kins of indicators of pathogenic bacteria and food borne pathogens. A total of 51 bacteriocin producing strains were collected and some of the strains had high inhibitory zone such as Lactobacillus casei SS14C (26 mm, Enterobacter cloacae SRUT (24mm, Enterococcus faecalis SK39 (21mm and Bifidobacterium dentium SS14T (20mm respectively, to Salmonella typhimurium BCC B0046/ATCC 13311, E. coli O157 hemolytic BCC B2717, Listeria monocytogenes BCC B2767/ATCC 7764 and Escherichia coli VTEC O157 BCC B2687. Evaluation after conservation ex situ to all bacterocin producing strain at 5oC for 1 year in freeze drying ampoules in vacuum and dry condition revealed the decreasing viability starting from log 0.8 CFU/ml for Lactococcus and Leuconostoc to log 2.2. CFU/ml for Streptococcus. Result of the study showed that the bacteriocin producing strains obtained were offered a potential resource for preventing disease of livestock and food borne diseases.

Bacteriocins with a broader antimicrobial spectrum prevail in enterococcal symbionts isolated from the hoopoe's uropygial gland.

Science.gov (United States)

Ruiz-Rodríguez, Magdalena; Martínez-Bueno, Manuel; Martín-Vivaldi, Manuel; Valdivia, Eva; Soler, Juan J

2013-09-01

The use of compounds produced by symbiotic bacteria against pathogens in animals is one of the most exciting discoveries in ecological immunology. The study of those antibiotic metabolites will enable an understanding of the defensive strategies against pathogenic infections. Here, we explore the role of bacteriocins explaining the antimicrobial properties of symbiotic bacteria isolated from the uropygial gland of the hoopoe (Upupa epops). The antagonistic activity of 187 strains was assayed against eight indicator bacteria, and the presence of six bacteriocin genes was detected in the genomic DNA. The presence of bacteriocin genes correlated with the antimicrobial activity of isolates. The most frequently detected bacteriocin genes were those encoding for the MR10 and AS-48 enterocins, which confer the highest inhibition capacity. All the isolates belonged to the genus Enterococcus, with E. faecalis as the most abundant species, with the broadest antimicrobial spectrum and the highest antagonistic activity. The vast majority of E. faecalis strains carried the genes of MR10 and AS-48 in their genome. Therefore, we suggest that fitness-related benefits for hoopoes associated with harbouring the most bactericidal symbionts cause the highest frequency of strains carrying MR10 and AS-48 genes. The study of mechanisms associated with the acquisition and selection of bacterial symbionts by hoopoes is necessary, however, to reach further conclusions. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Factors affecting production of an antilisterial bacteriocin by Carnobacterium piscicola strain A9b in laboratory media and model fish systems

DEFF Research Database (Denmark)

Himelbloom, B.; Nilsson, Lilian; Gram, Lone

2001-01-01

in APT broth when a low inoculum level (0.001%) was used. In contrast, inoculum level did not influence bacteriocin production in BHI and MRS7 without glucose. Bacteriocin production in APT was induced by the presence of an extracellular compound present in the sterile, filtered, cell- free supernatant...

Characterization of Bacteriocin like inhibitory substance produced by a new Strain Brevibacillus borstelensis AG1 Isolated from ‘Marcha’

Science.gov (United States)

Sharma, Nivedita; Gupta, Anupama; Gautam, Neha

2014-01-01

In the present study, a bacterium isolated from Marcha- a herbal cake used as traditional starter culture to ferment local wine in North East India, was evaluated for bacteriocin like inhibitory substance production and was tested against six food borne/spoilage causing pathogens viz. Listeria monocytogenes MTCC 839, Bacillus subtilis MTCC 121, Clostridium perfringens MTCC 450, Staphylococcus aureus, Lactobacillus plantarum and Leuconostoc mesenteroides MTCC 107 by using bit/disc method followed by well diffusion method. The bacterial isolate was identified as Brevibacillus borstelensis on the basis of phenotypic, biochemical and molecular characteristics using 16Sr RNA gene technique. Bacteriocin like inhibitory substance produced by Brevibacillus borstelensis AG1 was purified by gel exclusion chromatography. The molecular mass of the Brevibacillus borstelensis AG1 was found to be 12 kDa. Purified bacteriocin like inhibitory substance of Brevibacillus borstelensis was further characterized by studying the effect of temperature, pH, proteolytic enzyme and stability. Bacteriocin like inhibitory substance was found to be thermostable upto 100 °C, active at neutral pH, sensitive to trypsin, and partially stable till third week of storage thus showing a bright prospective to be used as a potential food biopreservative. PMID:25477937

Antagonistic Activity of Lactobacillus plantarum C11: Two New Two-Peptide Bacteriocins, Plantaricins EF and JK, and the Induction Factor Plantaricin A

Science.gov (United States)

Anderssen, Erlend L.; Diep, Dzung Bao; Nes, Ingolf F.; Eijsink, Vincent G. H.; Nissen-Meyer, Jon

1998-01-01

Six bacteriocinlike peptides (plantaricin A [PlnA], PlnE, PlnF, PlnJ, PlnK, and PlnN) produced by Lactobacillus plantarum C11 were detected by amino acid sequencing and mass spectrometry. Since purification to homogeneity was problematic, all six peptides were obtained by solid-phase peptide synthesis and were tested for bacteriocin activity. It was found that L. plantarum C11 produces two two-peptide bacteriocins (PlnEF and PlnJK); a strain-specific antagonistic activity was detected at nanomolar concentrations when PlnE and PlnF were combined and when PlnJ and PlnK were combined. Complementary peptides were at least 103 times more active when they were combined than when they were present individually, and optimal activity was obtained when the complementary peptides were present in approximately equal amounts. The interaction between complementary peptides was specific, since neither PlnE nor PlnF could complement PlnJ or PlnK, and none of these peptides could complement the peptides constituting the two-peptide bacteriocin lactococcin G. Interestingly, PlnA, which acts as an extracellular signal (pheromone) that triggers bacteriocin production, also possessed a strain-specific antagonistic activity. No bacteriocin activity could be detected for PlnN. PMID:9603847

Leuconostoc carnosum 4010 has the potential for use as a protective culture for vacuum-packed meats: culture isolation, bacteriocin identification, and meat application experiments

DEFF Research Database (Denmark)

Budde, B.B.; Hornbæk, T.; Jacobsen, T.

2003-01-01

A new culture, Leuconostoc carnosum 4010, for biopreservation of vacuum-packed meats is described. The culture originated from bacteriocin-producing lactic acid bacteria (LAB) naturally present in vacuum-packed meat products. Approximately, 72,000 colonies were isolated from 48 different vacuum......-packed meat products and examined for antibacterial activity. Bacteriocin-producing colonies were isolated from 46% of the packages examined. Leuc. carnosum was the predominant bacteriocin-producing strain and Leuc. carnosum 4010 was selected for further experiments because it showed strong antilisterial...... activity corresponding to molecular sizes of 4.6 and 5.3 kDa. N-terminal amino acid sequencing showed that Leuc. carnosum 4010 produced two bacteriocins highly similar or identical to leucocin A and leucocin C. Application experiments showed that the addition of 10(7) cfu/g Leuc. carnosum 4010 to a vacuum...

Salivaricin P, One of a Family of Two-Component Antilisterial Bacteriocins Produced by Intestinal Isolates of Lactobacillus salivarius▿

Science.gov (United States)

Barrett, Eoin; Hayes, Maria; O'Connor, Paula; Gardiner, Gillian; Fitzgerald, Gerald F.; Stanton, Catherine; Ross, R. Paul; Hill, Colin

2007-01-01

Lactobacillus salivarius DPC6005, a porcine intestinal isolate, produces a two-component bacteriocin, salivaricin P, with homology to ABP-118 produced by a human probiotic L. salivarius strain. Indeed, molecular characterization revealed that while the peptides Sln1 and ABP-118α are identical, their companion peptides (Sln2 and ABP-118β, respectively) differ by two amino acids. This observation suggests that two-component bacteriocins may be a common feature of intestinal L. salivarius strains. PMID:17416691

The Curing Agent Sodium Nitrite, Used in the Production of Fermented Sausages, Is Less Inhibiting to the Bacteriocin-Producing Meat Starter Culture Lactobacillus curvatus LTH 1174 under Anaerobic Conditions

Science.gov (United States)

Verluyten, Jurgen; Messens, Winy; De Vuyst, Luc

2003-01-01

Curvacin A is a listericidal bacteriocin produced by Lactobacillus curvatus LTH 1174, a strain isolated from fermented sausage. The response of this strain to an added curing agent (sodium nitrite) in terms of cell growth and bacteriocin production was investigated in vitro by laboratory fermentations with modified MRS broth. The strain was highly sensitive to nitrite; even a concentration of 10 ppm of curing agent inhibited its growth and both volumetric and specific bacteriocin production. A meat simulation medium containing 5 ppm of sodium nitrite was tested to investigate the influence of the gas phase on the growth and bacteriocin production of L. curvatus LTH 1174. Aerating the culture during growth had no effect on biomass formation, but the oxidative stress caused a higher level of specific bacteriocin production and led to a metabolic shift toward acetic acid production. Anaerobic conditions, on the other hand, led to an increased biomass concentration and less growth inhibition. Also, higher maximum volumetric bacteriocin activities and a higher level of specific bacteriocin production were obtained in the presence of sodium nitrite than in fermentations under aerobic conditions or standard conditions of air supply. These results indicate that the inhibitory effect of the curing agent is at least partially masked under anaerobic conditions. PMID:12839751

The curing agent sodium nitrite, used in the production of fermented sausages, is less inhibiting to the bacteriocin-producing meat starter culture Lactobacillus curvatus LTH 1174 under anaerobic conditions.

Science.gov (United States)

Verluyten, Jurgen; Messens, Winy; De Vuyst, Luc

2003-07-01

Curvacin A is a listericidal bacteriocin produced by Lactobacillus curvatus LTH 1174, a strain isolated from fermented sausage. The response of this strain to an added curing agent (sodium nitrite) in terms of cell growth and bacteriocin production was investigated in vitro by laboratory fermentations with modified MRS broth. The strain was highly sensitive to nitrite; even a concentration of 10 ppm of curing agent inhibited its growth and both volumetric and specific bacteriocin production. A meat simulation medium containing 5 ppm of sodium nitrite was tested to investigate the influence of the gas phase on the growth and bacteriocin production of L. curvatus LTH 1174. Aerating the culture during growth had no effect on biomass formation, but the oxidative stress caused a higher level of specific bacteriocin production and led to a metabolic shift toward acetic acid production. Anaerobic conditions, on the other hand, led to an increased biomass concentration and less growth inhibition. Also, higher maximum volumetric bacteriocin activities and a higher level of specific bacteriocin production were obtained in the presence of sodium nitrite than in fermentations under aerobic conditions or standard conditions of air supply. These results indicate that the inhibitory effect of the curing agent is at least partially masked under anaerobic conditions.

Anti-bacterial Efficacy of Bacteriocin Produced by Marine Bacillus subtilis Against Clinically Important Extended Spectrum Beta-Lactamase Strains and Methicillin-Resistant Staphylococcus aureus

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Suresh Mickymaray

2018-02-01

Full Text Available Objective: To investigate the anti-bacterial efficacy of bacteriocin produced by Bacillus subtilis SM01 (GenBank accession no: KY612347, a Gram-positive marine bacterium, against Extended Spectrum Beta-Lactamase (ESBL producing Gram-negative pathogens Acinetobacter baumannii, Pseudomonas aeruginosa, and Escherichia coli, and Gram-positive pathogen Methicillin-Resistant Staphylococcus aureus (MRSA. Methods: A marine bacterium was isolated from mangrove sediment from the Red Sea coast of Jeddah, Kingdom of Saudi Arabia, and identified based on its morphological, biochemical, and molecular characteristics. The bacteriocin production using this isolate was carried out in brain heart infusion broth (BHIB medium. The Anti-bacterial activity of bacteriocin was evaluated against selected ESBL strains and MRSA by the well agar method. The effects of incubation time, pH, and temperature on the Anti-bacterial activity were studied. Results: The bacteriocin Bac-SM01 produced by B. subtilis SM01 demonstrated broad-spectrum Anti-bacterial activity against both Gram-negative and -positive bacteria. The present study is the first report that the bacteriocin Bac-SM01 inhibits the growth of ESBL producing Gram-negative strains A. baumannii, P. aeruginosa, and E. coli, and a Gram-positive MRSA strain. The optimum incubation time, pH, and temperature for the Anti-bacterial activity of Bac-SM01 was 24 h, 7, and 37°C respectively. Conclusion: The overall investigation can conclude that the bacteriocin Bac-SM01 from the marine isolate Bacillus subtilis SM01 could be used as an alternative Anti-bacterial agent in pharmaceutical products.

An application in cheddar cheese manufacture for a strain of Lactococcus lactis producing a novel broad-spectrum bacteriocin, lacticin 3147.

Science.gov (United States)

Ryan, M P; Rea, M C; Hill, C; Ross, R P

1996-01-01

Lactococcus lactis DPC3147, a strain isolated from an Irish kefir grain, produces a bacteriocin with a broad spectrum of inhibition. The bacteriocin produced is heat stable, particularly at a low pH, and inhibits nisin-producing (Nip+) lactococci. On the basis of the observation that the nisin structural gene (nisA) does not hybridize to DPC3147 genomic DNA, the bacteriocin produced was considered novel and designated lacticin 3147. The genetic determinants which encode lacticin 3147 are contained on a 63-kb plasmid, which was conjugally mobilized to a commercial cheese starter, L. lactis subsp. cremoris DPC4268. The resultant transconjugant, DPC4275, both produces and is immune to lacticin 3147. The ability of lacticin 3147-producing lactococci to perform as cheddar cheese starters was subsequently investigated in cheesemaking trials. Bacteriocin-producing starters (which included the transconjugant strain DPC4275) produced acid at rates similar to those of commercial strains. The level of lacticin 3147 produced in cheese remained constant over 6 months of ripening and correlated with a significant reduction in the levels of nonstarter lactic acid bacteria. Such results suggest that these starters provide a means of controlling developing microflora in ripened fermented products. PMID:8593062

Purification and Partial Characterization of a Novel Bacteriocin Synthesized by Lactobacillus paracasei HD1-7 Isolated from Chinese Sauerkraut Juice.

Science.gov (United States)

Ge, Jingping; Sun, Yanyang; Xin, Xing; Wang, Ying; Ping, Wenxiang

2016-01-14

Bacteriocins have antimicrobial activities against food-spoiling bacteria and food-borne pathogens. Paracin 1.7, a bacteriocin synthesized by Lactobacillus paracasei HD1-7 isolated from Chinese sauerkraut juice, was studied. Following partial purification with ammonium sulfate precipitation, CM Sepharose Fast Flow, and Sephadex G-10 chromatography, the molecular weight of Paracin 1.7 was about 10 kDa based on Tricine-SDS-PAGE results. A 2.87 fold purified bacteriocin was produced, reaching a final yield of 39.93% and the specific activity of 1.56 × 10(3) AU/mg. The N-terminal amino acid sequence of Paracin 1.7 was VSNTFFA, and the LC/LTQ results revealed that the N-terminal amino acid sequence was similar to that of ABC-type oligopeptide transport system protein and N-acetylmuramoyl-L-alanine amidase. Paracin 1.7 was sensitive to protease K, had antimicrobial activities at a broad pH range (3.0-8.0), and was heat resistant (121 °C for 20 min). Paracin 1.7 from Lactobacillus paracasei HD1-7 is a novel bacteriocin that has potential applications in food preservation.

Purification and Genetic Characterization of Enterocin I from Enterococcus faecium 6T1a, a Novel Antilisterial Plasmid-Encoded Bacteriocin Which Does Not Belong to the Pediocin Family of Bacteriocins

Science.gov (United States)

Floriano, Belén; Ruiz-Barba, José L.; Jiménez-Díaz, Rufino

1998-01-01

Enterocin I (ENTI) is a novel bacteriocin produced by Enterococcus faecium 6T1a, a strain originally isolated from a Spanish-style green olive fermentation. The bacteriocin is active against many olive spoilage and food-borne gram-positive pathogenic bacteria, including clostridia, propionibacteria, and Listeria monocytogenes. ENTI was purified to homogeneity by ammonium sulfate precipitation, binding to an SP-Sepharose fast-flow column, and phenyl-Sepharose CL-4B and C2/C18 reverse-phase chromatography. The purification procedure resulted in a final yield of 954% and a 170,000-fold increase in specific activity. The primary structure of ENTI was determined by amino acid and nucleotide sequencing. ENTI consists of 44 amino acids and does not show significant sequence similarity with any other previously described bacteriocin. Sequencing of the entI structural gene, which is located on the 23-kb plasmid pEF1 of E. faecium 6T1a, revealed the absence of a leader peptide at the N-terminal region of the gene product. A second open reading frame, ORF2, located downstream of entI, encodes a putative protein that is 72.7% identical to ENTI. entI and ORF2 appear to be cotranscribed, yielding an mRNA of ca. 0.35 kb. A gene encoding immunity to ENTI was not identified. However, curing experiments demonstrated that both enterocin production and immunity are conferred by pEF1. PMID:9835578

Evaluation of Potential Probiotic Properties of Enterococcus mundtii, Its Survival in Boza and in situ Bacteriocin Production

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Svetoslav D. Todorov

2009-01-01

Full Text Available Boza is a low-pH and low-alcohol cereal-based beverage produced in the Balkan Peninsula. Barley was cooked and prepared according to a traditional recipe and inoculated with Enterococcus mundtii ST4V (a potential probiotic and bacteriocin-producing strain, commercially produced boza, Saccharomyces cerevisiae, and a combination of strain E. mundtii ST4V and Saccharomyces cerevisiae. Fermentation was carried out at 37 °C for 3 h. The organoleptic properties of fermented products were evaluated by a qualified taste panel. No significant differences in rheological properties were observed, suggesting that E. mundtii ST4V had no effect on the quality of the final product. Microbial cell numbers remained relatively unchanged during one week of storage. The preservative properties of bacteriocin ST4V were evaluated by contaminating boza with Lactobacillus sakei DSM 20017. Changes in microbial populations were monitored by using classical microbiological methods, PCR with species-specific primers and denaturing gradient gel electrophoresis (DGGE. Adsorption of bacteriocin ST4V to target cells is pH-dependent, with the highest adsorption (88 % recorded at pH=8.0 and pH=10.0. Maximum adsorption of bacteriocin ST4V (75 % to Enterococcus faecalis and Listeria innocua was recorded at 25 to 37 °C. Growth of E. mundtii ST4V was inhibited only by a few antibiotics and anti-inflammatory medicaments, suggesting that the strain may be used as a probiotic by individuals receiving medical treatment.

Antibacterial activity of bacteriocin-like substance P34 on Listeria monocytogenes in chicken sausage

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Voltaire Sant'Anna

2013-12-01

Full Text Available The antimicrobial activity of the bacteriocin-like substance (BLS P34 against Listeria monocytogenes was investigated in chicken sausage. The BLS was applied to chicken sausages (256 AU g-1 previously inoculated with a suspension of 10² cfu g-1 of L. monocytogenes. BLS P34 inhibited the indicator microorganism in situ in all incubation times for up to 10 days at 5 °C. The effectiveness of BLS P34 was increased when it was added in combination with nisin. The bacteriocin was also tested in natural eatable natural bovine wrapping (salty semi-dried tripe against the same indicator microorganism, also showing inhibitory capability in vitro. BLS P34 showed potential to control L. monocytogenes in refrigerated meat products.

Pediocins: The bacteriocins of Pediococci. Sources, production, properties and applications

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Anastasiadou Sofia

2009-01-01

Full Text Available Abstract Class IIa bacteriocins from lactic acid bacteria are small, cationic proteins with antilisterial activity. Within this class, the pediocins are those bacteriocins that share a highly conserved hydrophilic and charged N-terminal part harboring the consensus sequence -YGNGV- and a more variable hydrophobic and/or amphiphilic C-terminal part. Several pediocins have been isolated and characterized. Despite the structural similarities, their molecular weight varies, as well as their spectrum of antimicrobial activity. They exhibit important technological properties, e.g. thermostability and retaining of activity at a wide pH range, which along with the bactericidal action against Gram-positive food spoilage and pathogenic bacteria, make them an important class of biopreservatives. Much new information regarding the pediocins has emerged during the last years. In this review, we summarize and discuss all the available information regarding the sources of pediocins, the characteristics of their biosynthesis and production in fermentation systems, the characteristics of the known pediocin molecules, and their antibacterial action. The advances made by genetic engineering in improving the features of pediocins are also discussed, as well as their perspectives for future applications.

Partial Characterisation of Bacteriocins Produced by Bacillus cereus Isolates from Milk and Milk Products

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Bojana Bogović Matijašić

2003-01-01

Full Text Available Thirty one (19.2 % out of 161 Bacillus cereus isolates from raw milk and milk products were found to produce proteinaceous substances which inhibit the growth of other B. cereus isolates. The detection of antibacterial activity depended on medium and method used. Bactericidal activity was detected in 23 (14 % or 19 (12 % of the tested strains on the triptic soya agar and brain-heart infusion with glucose, respectively, while 11 (7 % of the strains produced bactericidal substances on both media. Nineteen percent of isolates from raw milk and 20 % of isolates from milk products were found to produce bacteriocins. Four B. cereus isolates inhibited the growth of individual test strains belonging to B. licheniformis, B. subtilis, Enterococcus faecalis, Escherichia coli, Staphylococcus aureus, Lactobacillus helveticus and L. casei species. The bacteriocins of four B. cereus isolates were studied in more detail. The production and activity of these substances were detected in stationary- phase of bacterial culture. Two of them were stable after heating at 60 °C, while only one was stable after heating at 75 °C for 15 minutes. All of them were active over a range of pH=3–10. The apparent molecular weights of four bacteriocins detected by SDS-PAGE electrophoresis were in the range of 1 to 8 kDa.

Identification of the propionicin F bacteriocin immunity gene (pcfI) and development of a food-grade cloning system for Propionibacterium freudenreichii.

Science.gov (United States)

Brede, Dag Anders; Lothe, Sheba; Salehian, Zhian; Faye, Therese; Nes, Ingolf F

2007-12-01

This report describes the first functional analysis of a bacteriocin immunity gene from Propionibacterium freudenreichii and its use as a selection marker for food-grade cloning. Cloning of the pcfI gene (previously orf5 [located as part of the pcfABC propionicin F operon]) rendered the sensitive host 1,000-fold more tolerant to the propionicin F bacteriocin. The physiochemical properties of the 127-residue large PcfI protein resemble those of membrane-bound immunity proteins from bacteriocin systems found in lactic acid bacteria. The high level of immunity conferred by pcfI allowed its use as a selection marker for plasmid transformation in P. freudenreichii. Electroporation of P. freudenreichii IFO12426 by use of the pcfI expression plasmid pSL102 and propionicin F selection (200 bacteriocin units/ml) yielded 10(7) transformants/microg DNA. The 2.7-kb P. freudenreichii food-grade cloning vector pSL104 consists of the pLME108 replicon, a multiple cloning site, and pcfI expressed from the constitutive P(pampS) promoter for selection. The pSL104 vector efficiently facilitated cloning of the propionicin T1 bacteriocin in P. freudenreichii. High-level propionicin T1 production (640 BU/ml) was obtained with the IFO12426 strain, and the food-grade propionicin T1 expression plasmid pSL106 was maintained by approximately 91% of the cells over 25 generations in the absence of selection. To the best of our knowledge this is the first report of an efficient cloning system that facilitates the generation of food-grade recombinant P. freudenreichii strains.

Purification, Characterization, and Optimum Conditions of Fermencin SD11, a Bacteriocin Produced by Human Orally Lactobacillus fermentum SD11.

Science.gov (United States)

Wannun, Phirawat; Piwat, Supatcharin; Teanpaisan, Rawee

2016-06-01

Fermencin SD11, a bacteriocin produced by human orally Lactobacillus fermentum SD11, was purified, characterized, and optimized in conditions for bacterial growth and bacteriocin production. Fermencin SD11 was purified using three steps of ammonium sulfate precipitation, gel filtration chromatography, and reverse-phase high-performance liquid chromatography. The molecular weight was found to be 33,000 Da using SDS-PAGE and confirmed as 33,593.4 Da by liquid chromatography-mass spectrometry. Fermencin SD11 exhibited activity against a wide range of oral pathogens including cariogenic and periodontogenic pathogens and Candida. The active activity was stable between 60 - 80 °C in a pH range of 3.0 to 7.0. It was sensitive to proteolytic enzymes (proteinase K and trypsin), but it was not affected by α-amylase, catalase, lysozyme, and saliva. The optimum conditions for growth and bacteriocin production of L. fermentum SD11 were cultured at acidic with pH of 5.0-6.0 at 37 or 40 °C under aerobic or anaerobic conditions for 12 h. It is promising that L. fermentum SD11 and its bacteriocin may be an alternative approach for promoting oral health or prevention of oral diseases, e.g., dental caries and periodontitis, which would require further clinical trials.

Topology of a type I secretion system for bacteriocins of Lactococcus lactis

NARCIS (Netherlands)

Franke, Christian Marc

1998-01-01

This thesis describes the analysis of a number of aspects of the secretion and muturation machinery of the bacteriocin lactococcin A (LcnA) from Lactococcus lactis, whick is initially synthesized as a precursor protein (preLcnA), containing an N-terminal extension of 20 amino acids (the leader)....

Bacteriocin formation by dominant aerobic sporeformers isolated from traditional maari

DEFF Research Database (Denmark)

Kaboré, Donatien; Thorsen, Line; Nielsen, Dennis Sandris

2012-01-01

that the antimicrobial substances produced were heat stable, in-sensitive to catalase, sensitive to protease and trypsin but resistant to the proteolytic action of papain and proteinase K and equally active at pH values ranging from 3 to 11. Bacteriocin secretion started in late exponential growth phase and maximum...

Time-dependent fermentation control strategies for enhancing synthesis of marine bacteriocin 1701 using artificial neural network and genetic algorithm.

Science.gov (United States)

Peng, Jiansheng; Meng, Fanmei; Ai, Yuncan

2013-06-01

The artificial neural network (ANN) and genetic algorithm (GA) were combined to optimize the fermentation process for enhancing production of marine bacteriocin 1701 in a 5-L-stirred-tank. Fermentation time, pH value, dissolved oxygen level, temperature and turbidity were used to construct a "5-10-1" ANN topology to identify the nonlinear relationship between fermentation parameters and the antibiotic effects (shown as in inhibition diameters) of bacteriocin 1701. The predicted values by the trained ANN model were coincided with the observed ones (the coefficient of R(2) was greater than 0.95). As the fermentation time was brought in as one of the ANN input nodes, fermentation parameters could be optimized by stages through GA, and an optimal fermentation process control trajectory was created. The production of marine bacteriocin 1701 was significantly improved by 26% under the guidance of fermentation control trajectory that was optimized by using of combined ANN-GA method. Copyright © 2013 Elsevier Ltd. All rights reserved.

Cloning strategies for heterologous expression of the bacteriocin enterocin A by Lactobacillus sakei Lb790, Lb. plantarum NC8 and Lb. casei CECT475.

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Jiménez, Juan J; Diep, Dzung B; Borrero, Juan; Gútiez, Loreto; Arbulu, Sara; Nes, Ingolf F; Herranz, Carmen; Cintas, Luis M; Hernández, Pablo E

2015-10-15

Bacteriocins produced by lactic acid bacteria (LAB) attract considerable interest as natural and nontoxic food preservatives and as therapeutics whereas the bacteriocin-producing LAB are considered potential probiotics for food, human and veterinary applications, and in the animal production field. Within LAB the lactobacilli are increasingly used as starter cultures for food preservation and as probiotics. The lactobacilli are also natural inhabitants of the gastrointestinal (GI) tract and attractive vectors for delivery of therapeutic peptides and proteins, and for production of bioactive peptides. Research efforts for production of bacteriocins in heterologous hosts should be performed if the use of bacteriocins and the LAB bacteriocin-producers is ever to meet the high expectations deposited in these antimicrobial peptides. The recombinant production and functional expression of bacteriocins by lactobacilli would have an additive effect on their probiotic functionality. The heterologous production of the bacteriocin enterocin A (EntA) was evaluated in different Lactobacillus spp. after fusion of the versatile Sec-dependent signal peptide (SP usp45 ) to mature EntA plus the EntA immunity gene (entA + entiA) (fragment UAI), and their cloning into plasmid vectors that permitted their inducible (pSIP409 and pSIP411) or constitutive (pMG36c) production. The amount, antimicrobial activity (AA) and specific antimicrobial activity (SAA) of the EntA produced by Lactobacillus sakei Lb790, Lb. plantarum NC8 and Lb. casei CECT475 transformed with the recombinant plasmids pSIP409UAI, pSIP411UAI and pMGUAI varied depending of the expression vector and the host strain. The Lb. casei CECT475 recombinant strains produced the largest amounts of EntA, with the highest AA and SAA. Supernatants from Lb. casei CECT (pSIP411UAI) showed a 4.9-fold higher production of EntA with a 22.8-fold higher AA and 4.7-fold higher SAA than those from Enterococcus faecium T136, the natural

Antibacterial activity and optimisation of bacteriocin producing lactic acid bacteria isolated from beef (red meat) samples

International Nuclear Information System (INIS)

Ali, N.M.; Mazhar, B.; Khadija, I.; Kalim, B.

2016-01-01

Bacteriocin producing bacteria are commonly found in meat products to enhance their shelf-life. In the present study, bacterial species were isolated from meat samples (beef) from different localities of Lahore, Pakistan. MRS agar medium was used to isolate lactic acid bacteria (LAB) through spread and streak methods (incubated for 72 h at 37 degree C). Identification of bacteriocinogenic LAB strains was done by using staining techniques, morphology based characteristics and biochemical tests. These strains were BSH 1b, BSH 3a, BIP 4a, BIP 3a, BIP 1b and BRR 3a. Antibacterial activity of LAB was performed against food borne pathogens viz., Escherichia coli and Staphylococcus aureus through paper disc diffusion method. Three bacterial strains showed maximum inhibition and characterised by ribotyping viz., BIP 4a was identified as Lactobacillus curvatures, BIP 3a was Staphylococcus warneri and BIP 1b was Lactobacillus graminis . Optimum pH 5-6.5 and 30-37 degree C temperature for isolated bacterial strains was recorded. Protein concentration measured was 0.07 mg/mL for BSH 1b, 0.065 mg/mL for BSH 3a, 0.057 mg/mL for BIP 4a, 0.062 mg/mL for BIP 1b, 0.065 mg/mL for BIP 3a and for BRR 3a 0.078 mg/mL, respectively. Bacteriocin of all isolates except BIP 3a was found to be sensitive towards pepsin and resistant towards Rnase. Bacteriocin production was stable at between pH 5.0 and 6.0 and resistant temperature was 40 degree C. It was concluded that lactic acid bacteria (LAB) from meat can be helpful as antibacterial agents against food-borne bacterial pathogens because of thermostable producing bacteriocin. (author)

The Leaderless Bacteriocin Enterocin K1 Is Highly Potent against Enterococcus faecium: A Study on Structure, Target Spectrum and Receptor.

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Ovchinnikov, Kirill V; Kristiansen, Per Eugen; Straume, Daniel; Jensen, Marianne S; Aleksandrzak-Piekarczyk, Tamara; Nes, Ingolf F; Diep, Dzung B

2017-01-01

Enterocin K1 (EntK1), enterocin EJ97 (EntEJ97), and LsbB are three sequence related leaderless bacteriocins. Yet LsbB kills only lactococci while EntK1 and EntEJ97 target wider spectra with EntK1 being particularly active against Enterococcus faecium , including nosocomial multidrug resistant isolates. NMR study of EntK1 showed that it had a structure very similar to LsbB - both having an amphiphilic N-terminal α-helix and an unstructured C-terminus. The α-helix in EntK1 is, however, about 3-4 residues longer than that of LsbB. Enterococcal mutants highly resistant to EntEJ97 and EntK1 were found to have mutations within rseP , a gene encoding a stress response membrane-bound Zn-dependent protease. Heterologous expression of the enterococcal rseP rendered resistant cells of Streptococcus pneumoniae sensitive to EntK1 and EntEJ97, suggesting that RseP likely serves as the receptor for EntK1 and EntEJ97. It was also shown that the conserved proteolytic active site in E. faecalis RseP is partly required for EntK1 and EntEJ97 activity, since alanine substitutions of its conserved residues (HExxH) reduced the sensitivity of the clones to the bacteriocins. RseP is known to be involved in bacterial stress response. As expected, the growth of resistant mutants with mutations within rseP was severely affected when they were exposed to higher (stressing) growth temperatures, e.g., at 45°C, at which wild type cells still grew well. These findings allow us to design a hurdle strategy with a combination of the bacteriocin(s) and higher temperature that effectively kills bacteriocin sensitive bacteria and prevents the development of resistant cells.

The Leaderless Bacteriocin Enterocin K1 Is Highly Potent against Enterococcus faecium: A Study on Structure, Target Spectrum and Receptor

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Kirill V. Ovchinnikov

2017-05-01

Full Text Available Enterocin K1 (EntK1, enterocin EJ97 (EntEJ97, and LsbB are three sequence related leaderless bacteriocins. Yet LsbB kills only lactococci while EntK1 and EntEJ97 target wider spectra with EntK1 being particularly active against Enterococcus faecium, including nosocomial multidrug resistant isolates. NMR study of EntK1 showed that it had a structure very similar to LsbB – both having an amphiphilic N-terminal α-helix and an unstructured C-terminus. The α-helix in EntK1 is, however, about 3–4 residues longer than that of LsbB. Enterococcal mutants highly resistant to EntEJ97 and EntK1 were found to have mutations within rseP, a gene encoding a stress response membrane-bound Zn-dependent protease. Heterologous expression of the enterococcal rseP rendered resistant cells of Streptococcus pneumoniae sensitive to EntK1 and EntEJ97, suggesting that RseP likely serves as the receptor for EntK1 and EntEJ97. It was also shown that the conserved proteolytic active site in E. faecalis RseP is partly required for EntK1 and EntEJ97 activity, since alanine substitutions of its conserved residues (HExxH reduced the sensitivity of the clones to the bacteriocins. RseP is known to be involved in bacterial stress response. As expected, the growth of resistant mutants with mutations within rseP was severely affected when they were exposed to higher (stressing growth temperatures, e.g., at 45°C, at which wild type cells still grew well. These findings allow us to design a hurdle strategy with a combination of the bacteriocin(s and higher temperature that effectively kills bacteriocin sensitive bacteria and prevents the development of resistant cells.

Application of anti-listerial bacteriocins: monitoring enterocin expression by multiplex relative reverse transcription-PCR.

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Williams, D Ross; Chanos, Panagiotis

2012-12-01

Listeriosis is a deadly food-borne disease, and its incidence may be limited through the biotechnological exploitation of a number of anti-listerial biocontrol agents. The most widely used of these agents are bacteriocins and the Class II enterocins are characterized by their activity against Listeria. Enterocins are primarily produced by enterococci, particularly Enterococcus faecium and many strains have been described, often encoding multiple bacteriocins. The use of these strains in food will require that they are free of virulence functions and that they exhibit a high level expression of anti-listerial enterocins in fermentation conditions. Multiplex relative RT (reverse transcription)-PCR is a technique that is useful in the discovery of advantageous expression characteristics among enterocin-producing strains. It allows the levels of individual enterocin gene expression to be monitored and determination of how expression is altered under different growth conditions.

Assessment of hemolytic activity, enzyme production and bacteriocin characterization of Bacillus subtilis LR1 isolated from the gastrointestinal tract of fish.

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Banerjee, Goutam; Nandi, Ankita; Ray, Arun Kumar

2017-01-01

In the present investigation, probiotic potential (antagonistic activity, enzyme production, hemolytic activity, biosafety, antibiotic sensitivity and bile tolerance level) of Bacillus subtilis LR1 was evaluated. Bacteriocin produced by the bacterial strain B. subtilis LR1 isolated from the gastrointestinal tract of Labeo rohita was purified and characterized. The molecular weight of the purified bacteriocin was ~50 kDa in 12 % Native PAGE and showed inhibitory activity against four fish pathogens such as Bacillus mycoides, Aeromonas salmonicida, Pseudomonas fluorescens and Aeromonas hydrophila. The purified bacteriocin was maximally active at temperature 40 °C and pH 7.0, while none of the tested surfactants affect the bacteriocin activity. Extracellular enzyme activity of the selected bacterial strain was also evaluated. Amylase activity was estimated to be highest (38.23 ± 1.15 µg of maltose liberated mg -1  protein ml -1 of culture filtrate) followed by cellulase and protease activity. The selected bacterium was sensitive to most of the antibiotics used in this experiment, can tolerate 0.25 % bile salt and non-hemolytic in nature. Finally, the efficiency of the proposed probiotic candidate was evaluated in in vivo condition. It was detected that the bacterial strain can effectively reduce bacterial pathogenicity in Indian major carps.

Monitoring of multiple bacteriocins through a developed dual extraction protocol and comparison of HPLC-DAD with turbidometry as their quantification system.

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Katharopoulos, Efstathios; Touloupi, Katerina; Touraki, Maria

2016-08-01

The present study describes the development of a simple and efficient screening system that allows identification and quantification of nine bacteriocins produced by Lactococcus lactis. Cell-free L. lactis extracts presented a broad spectrum of antibacterial activity, including Gram-negative bacteria, Gram-positive bacteria, and fungi. The characterization of their sensitivity to pH, and heat, showed that the extracts retained their antibacterial activity at extreme pH values and in a wide temperature range. The loss of antibacterial activity following treatment of the extracts with lipase or protease suggests a lipoproteinaceous nature of the produced antimicrobials. The extracts were subjected to a purification protocol that employs a two phase extraction using ammonium sulfate precipitation and organic solvent precipitation, followed by ion exchange chromatography, solid phase extraction and HPLC. In the nine fractions that presented antimicrobial activity, bacteriocins were quantified by the turbidometric method using a standard curve of nisin and by the HPLC method with nisin as the external standard, with both methods producing comparable results. Turbidometry appears to be unique in the qualitative determination of bacteriocins but the only method suitable to both separate and quantify the bacteriocins providing increased sensitivity, accuracy, and precision is HPLC. Copyright © 2016 Elsevier B.V. All rights reserved.

Purification and characterization of Plantaricin ZJ5, a new bacteriocin produced by Lactobacillus plantarum ZJ5.

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Song, Da-Feng; Zhu, Mu-Yuan; Gu, Qing

2014-01-01

The aim of this study is to investigate the antimicrobial potential of Lactobacillus plantarum ZJ5, a strain isolated from fermented mustard with a broad range of inhibitory activity against both Gram-positive and Gram-negative bacteria. Here we present the peptide plantaricin ZJ5 (PZJ5), which is an extreme pH and heat-stable. However, it can be digested by pepsin and proteinase K. This peptide has strong activity against Staphylococcus aureus. PZJ5 has been purified using a multi-step process, including ammonium sulfate precipitation, cation-exchange chromatography, hydrophobic interactions and reverse-phase chromatography. The molecular mass of the peptide was found to be 2572.9 Da using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The primary structure of this peptide was determined using amino acid sequencing and DNA sequencing, and these analyses revealed that the DNA sequence translated as a 44-residue precursor containing a 22-amino-acid N-terminal extension that was of the double-glycine type. The bacteriocin sequence exhibited no homology with known bacteriocins when compared with those available in the database, indicating that it was a new class IId bacteriocin. PZJ5 from a food-borne strain may be useful as a promising probiotic candidate.

Purification and characterization of Plantaricin ZJ5, a new bacteriocin produced by Lactobacillus plantarum ZJ5.

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Da-Feng Song

Full Text Available The aim of this study is to investigate the antimicrobial potential of Lactobacillus plantarum ZJ5, a strain isolated from fermented mustard with a broad range of inhibitory activity against both Gram-positive and Gram-negative bacteria. Here we present the peptide plantaricin ZJ5 (PZJ5, which is an extreme pH and heat-stable. However, it can be digested by pepsin and proteinase K. This peptide has strong activity against Staphylococcus aureus. PZJ5 has been purified using a multi-step process, including ammonium sulfate precipitation, cation-exchange chromatography, hydrophobic interactions and reverse-phase chromatography. The molecular mass of the peptide was found to be 2572.9 Da using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS. The primary structure of this peptide was determined using amino acid sequencing and DNA sequencing, and these analyses revealed that the DNA sequence translated as a 44-residue precursor containing a 22-amino-acid N-terminal extension that was of the double-glycine type. The bacteriocin sequence exhibited no homology with known bacteriocins when compared with those available in the database, indicating that it was a new class IId bacteriocin. PZJ5 from a food-borne strain may be useful as a promising probiotic candidate.

In vitro evaluation of bacteriocin-like inhibitory substances produced by lactic acid bacteria isolated during traditional Sicilian cheese making

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Giusi Macaluso

2016-02-01

Full Text Available Bacteriocins are antimicrobial proteins produced by bacteria that inhibit the growth of other bacteria with a bactericidal or bacteriostatic mode of action. Many lactic acid bacteria (LAB produce a high diversity of different bacteriocins. Bacteriocinogenic LAB are generally recognised as safe (GRAS and useful to control the frequent development of pathogens and spoilage microorganisms. For this reason they are commonly used as starter cultures in food fermentations. In this study, the authors describe the results of a screening on 699 LAB isolated from wooden vat surfaces, raw milk and traditional Sicilian cheeses, for the production of bacteriocin-like inhibitory substances, by comparing two alternative methods. The antagonistic activity of LAB and its proteinaceous nature were evaluated using the spot-on-the-lawn and the well-diffusion assay (WDA and the sensitivity to proteolytic (proteinase K, protease B and trypsin, amylolytic (α-amylase and lipolytic (lipase enzymes. The indicator strains used were: Listeria monocytogenes, Staphylococcus aureus, Escherichia coli, Salmonella enteritidis. A total of 223 strains (belonging to the species Enterococcus spp., Lactobacillus spp., Pediococcus spp., Streptococcus spp., Leuconostoc spp. and Lactococcus lactis were found to inhibit the growth of Listeria monocytogenes by using the spot-on-the-lawn method; only 37 of these were confirmed by using the WDA. The direct addition of bacteriocin-producing cultures into dairy products can be a more practical and economic option for the improvement of the safety and quality of the final product.

Bacteriocin-like inhibitory activities of seven Lactobacillus delbrueckii subsp. bulgaricus strains against antibiotic susceptible and resistant Helicobacter pylori strains.

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Boyanova, L; Gergova, G; Markovska, R; Yordanov, D; Mitov, I

2017-12-01

The aim of the study was to detect anti-Helicobacter pylori activity of seven Lactobacillus delbrueckii subsp. bulgaricus (GLB) strains by four cell-free supernatant (CFS) types. Activity of non-neutralized and non-heat-treated (CFSs1), non-neutralized and heat-treated (CFSs2), pH neutralized, catalase-treated and non-heat-treated (CFSs3), or neutralized, catalase- and heat-treated (CFSs4) CFSs against 18 H. pylori strains (11 of which with antibiotic resistance) was evaluated. All GLB strains produced bacteriocin-like inhibitory substances (BLISs), the neutralized CFSs of two GLB strains inhibited >81% of test strains and those of four GLB strains were active against >71% of antibiotic resistant strains. Two H. pylori strains were BLIS resistant. The heating did not reduce the CFS activity. Briefly, all GLB strains evaluated produced heat-stable BLISs, although GLB and H. pylori strain susceptibility patterns exhibited differences. Bacteriocin-like inhibitory substance activity can be an advantage for the probiotic choice for H. pylori infection control. In this study, anti-Helicobacter pylori activity of seven Lactobacillus delbrueckii subsp. bulgaricus (GLB) strains was evaluated by four cell-free supernatant (CFS) types. The GLB strains produced heat-stable bacteriocin-like inhibitory substances (BLISs) with a strong anti-H. pylori activity and some neutralized, catalase- and heat-treated CFSs inhibited >83% of the test strains. Bacteriocin-like inhibitory substance production of GLB strains can render them valuable probiotics in the control of H. pylori infection. © 2017 The Society for Applied Microbiology.

Release of Bacteriocins from Nanofibers Prepared with Combinations of Poly(D,L-lactide (PDLLA and Poly(Ethylene Oxide (PEO

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Leon Dicks

2011-03-01

Full Text Available Plantaricin 423, produced by Lactobacillus plantarum, and bacteriocin ST4SA produced by Enterococcus mundtii, were electrospun into nanofibers prepared from different combinations of poly(D,L-lactide (PDLLA and poly(ethylene oxide (PEO dissolved in N,N-dimethylformamide (DMF. Both peptides were released from the nanofibers with a high initial burst and retained 88% of their original antimicrobial activity at 37 °C. Nanofibers have the potential to serve as carrier matrix for bacteriocins and open a new field in developing controlled antimicrobial delivery systems for various applications.

Bacteriocins - exploring alternatives to antibiotics in mastitis treatment.

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Pieterse, Reneé; Todorov, Svetoslav D

2010-07-01

Mastitis is considered to be the most costly disease affecting the dairy industry. Management strategies involve the extensive use of antibiotics to treat and prevent this disease. Prophylactic dosages of antibiotics used in mastitis control programmes could select for strains with resistance to antibiotics. In addition, a strong drive towards reducing antibiotic residues in animal food products has lead to research in finding alternative antimicrobial agents. In this review we have focus on the pathogenesis of the mastitis in dairy cows, existing antibiotic treatments and possible alternative for application of bacteriocins from lactic acid bacteria in the treatment and prevention of this disease.

PRESERVATIVE POTENTIAL OF PURIFIED BACTERIOCIN PRODUCED FROM BREVIBACILLUS BORSTELENSIS AG1 ISOLATED FROM MARCHA – A TRADITIONAL WINE STARTER CULTURE CAKE IN TOMATO PASTE

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Anupama Gupta

2015-04-01

Full Text Available Purified bacteriocin produced from Brevibacillus borstelensis AG1 isolated from Marcha a local wine starter herbal cake, was used to enhance the shelf life of tomato paste. Preservative effect of purified bacteriocin was studied for nine days in tomato paste inoculated with food borne pathogens and was compared to commercial biopreservative – nisin and chemical preservative – sodium benzoate. The indicator strains i.e. Listeria monocytogenes MTCC839, Bacillus subtilis CRI and Clostridium perfringens MTCC1739 were used at the amount 8.16, 8.13 and 8.18 log CFU/ml. Viable cells were counted periodically and a consistent reduction in number of viable cells of each tested pathogen was observed. It was found antagonistic against L. monocytogenes MTCC839, B. subtilis CRI and C. perfringes MTCC1739 which are the most challengeable and food borne pathogens found in processed vegetables products. Purified bacteriocin was found active over a wide pH range i.e. 3.0 to 11.0 and was able to withstand temperature up to 100oC. It showed a better preservative potential by reducing pathogenic load of the tested strains (by 2.02, 2.05 and 2.02 log cycles (CFU/ml of L. monocytogenes MTCC839, B. subtilis CRI and C. perfringes MTCC1739, respectively in tomato paste as compared to control (without bacteriocin. This proves efficiency of bacteriocin produced by B. borstelensis AG1 as biopreservative to enhance the safety and shelf life of acidic foods.

Identification of the Propionicin F Bacteriocin Immunity Gene (pcfI) and Development of a Food-Grade Cloning System for Propionibacterium freudenreichii▿ †

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Brede, Dag Anders; Lothe, Sheba; Salehian, Zhian; Faye, Therese; Nes, Ingolf F.

2007-01-01

This report describes the first functional analysis of a bacteriocin immunity gene from Propionibacterium freudenreichii and its use as a selection marker for food-grade cloning. Cloning of the pcfI gene (previously orf5 [located as part of the pcfABC propionicin F operon]) rendered the sensitive host 1,000-fold more tolerant to the propionicin F bacteriocin. The physiochemical properties of the 127-residue large PcfI protein resemble those of membrane-bound immunity proteins from bacteriocin systems found in lactic acid bacteria. The high level of immunity conferred by pcfI allowed its use as a selection marker for plasmid transformation in P. freudenreichii. Electroporation of P. freudenreichii IFO12426 by use of the pcfI expression plasmid pSL102 and propionicin F selection (200 bacteriocin units/ml) yielded 107 transformants/μg DNA. The 2.7-kb P. freudenreichii food-grade cloning vector pSL104 consists of the pLME108 replicon, a multiple cloning site, and pcfI expressed from the constitutive PpampS promoter for selection. The pSL104 vector efficiently facilitated cloning of the propionicin T1 bacteriocin in P. freudenreichii. High-level propionicin T1 production (640 BU/ml) was obtained with the IFO12426 strain, and the food-grade propionicin T1 expression plasmid pSL106 was maintained by ∼91% of the cells over 25 generations in the absence of selection. To the best of our knowledge this is the first report of an efficient cloning system that facilitates the generation of food-grade recombinant P. freudenreichii strains. PMID:17933941

Mechanism of protection of transepithelial barrier function by Lactobacillus salivarius: strain dependence and attenuation by bacteriocin production.

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Miyauchi, Eiji; O'Callaghan, John; Buttó, Ludovica F; Hurley, Gráinne; Melgar, Silvia; Tanabe, Soichi; Shanahan, Fergus; Nally, Kenneth; O'Toole, Paul W

2012-11-01

Enhanced barrier function is one mechanism whereby commensals and probiotic bacteria limit translocation of foreign antigens or pathogens in the gut. However, barrier protection is not exhibited by all probiotic or commensals and the strain-specific molecules involved remain to be clarified. We evaluated the effects of 33 individual Lactobacillus salivarius strains on the hydrogen peroxide (H(2)O(2))-induced barrier impairment in human epithelial Caco-2 cells. These strains showed markedly different effects on H(2)O(2)-induced reduction in transepithelial resistance (TER). The effective strains such as UCC118 and CCUG38008 attenuated H(2)O(2)-induced disassembly and relocalization of tight junction proteins, but the ineffective strain AH43324 did not. Strains UCC118 and CCUG38008 induced phosphorylation of extracellular signal-regulated kinase (ERK) in Caco-2 cells, and the ERK inhibitor U0126 attenuated the barrier-protecting effect of these strains. In contrast, the AH43324 strain induced phosphorylation of Akt and p38, which was associated with an absence of a protective effect. Global transcriptome analysis of UCC118 and AH43324 revealed that some genes in a bacteriocin gene cluster were upregulated in AH43324 under TER assay conditions. A bacteriocin-negative UCC118 mutant displayed significantly greater suppressive effect on H(2)O(2)-induced reduction in TER compared with wild-type UCC118. The wild-type strain augmented H(2)O(2)-induced phosphorylation of Akt and p38, whereas a bacteriocin-negative UCC118 mutant did not. These observations indicate that L. salivarius strains are widely divergent in their capacity for barrier protection, and this is underpinned by differences in the activation of intracellular signaling pathways. Furthermore, bacteriocin production appears to have an attenuating influence on lactobacillus-mediated barrier protection.

Toxicity of bovicin HC5 against mammalian cell lines and the role of cholesterol in bacteriocin activity.

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Paiva, Aline Dias; de Oliveira, Michelle Dias; de Paula, Sérgio Oliveira; Baracat-Pereira, Maria Cristina; Breukink, Eefjan; Mantovani, Hilário Cuquetto

2012-11-01

Bacteriocins are ribosomally synthesized antimicrobial peptides produced by Bacteria and some Archaea. The assessment of the toxic potential of antimicrobial peptides is important in order to apply these peptides on an industrial scale. The aim of the present study was to investigate the in vitro cytotoxic and haemolytic potential of bovicin HC5, as well as to determine whether cholesterol influences bacteriocin activity on model membranes. Nisin, for which the mechanism of action is well described, was used as a reference peptide in our assays. The viability of three distinct eukaryotic cell lines treated with bovicin HC5 or nisin was analysed by using the MTT assay and cellular morphological changes were determined by light microscopy. The haemolytic potential was evaluated by using the haemoglobin liberation assay and the role of cholesterol on bacteriocin activity was examined by using model membranes composed of DOPC (1,2-dioleoyl-sn-glycero-3-phosphocholine) and DPoPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine). The IC(50) of bovicin HC5 and nisin against Vero cells was 65.42 and 13.48 µM, respectively. When the MTT assay was performed with MCF-7 and HepG2 cells, the IC(50) obtained for bovicin HC5 was 279.39 and 289.30 µM, respectively, while for nisin these values were 105.46 and 112.25 µM. The haemolytic activity of bovicin HC5 against eukaryotic cells was always lower than that determined for nisin. The presence of cholesterol did not influence the activity of either bacteriocin on DOPC model membranes, but nisin showed reduced carboxyfluorescein leakage in DPoPC membranes containing cholesterol. In conclusion, bovicin HC5 only exerted cytotoxic effects at concentrations that were greater than the concentration needed for its biological activity, and the presence of cholesterol did not affect its interaction with model membranes.

Potential of bacteriocin-producing lactic acid bacteria for safety improvements of traditional Thai fermented meat and human health.

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Swetwiwathana, Adisorn; Visessanguan, Wonnop

2015-11-01

Lactic acid bacteria (LAB) are very important in converting of agricultural products into safe, delicious and shelf stable foods for human consumption. The preservative activity of LAB in foods is mainly attributed to the production of anti-microbial metabolites such as organic acids and bacteriocins which enables them to grow and control the growth of pathogens and spoilage microorganisms. Besides ensuring safety, bacteriocin-producing LAB with their probiotic potentials could also be emerging as a means to develop functional meat products with desirable health benefits. Nevertheless, to be qualified as a candidate probiotic culture, other prerequisite probiotic properties of bacteriocin-producing LAB have to be assessed according to regulatory guidelines for probiotics. Nham is an indigenous fermented sausage of Thailand that has gained popularity and acceptance among Thais. Since Nham is made from raw meat and is usually consumed without cooking, risks due to undesirable microorganisms such as Salmonella spp., Staphylococcus aureus, and Listeria monocytogenes, are frequently observed. With an ultimate goal to produce safer and healthier product, our research attempts on the development of a variety of new Nham products are discussed. Copyright © 2015 Elsevier Ltd. All rights reserved.

Characterization of a noncytotoxic bacteriocin from probiotic Lactobacillus plantarum DM5 with potential as a food preservative.

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Das, Deeplina; Goyal, Arun

2014-10-01

The aim of this work was to purify and characterize the bacteriocin produced by probiotic Lactobacillus plantarum DM5 in order to evaluate its potential as nutraceuticals. Lb. plantarum DM5 exhibited in vitro probiotic properties such as high resistance to gastric juice and bile salt, adherence to human adenocarcinoma (HT-29) cells, bile salt hydrolase and cholesterol assimilation activity. Moreover, Lb. plantarum DM5 showed bacteriocin activity against several major food borne pathogens. Zymogram analysis of purified bacteriocin (plantaricin DM5) showed a molecular size of ∼15.2 kDa. Plantaricin DM5 was sensitive to proteolytic enzymes but stable in the pH range of 2.0-10.0, and it was heat resistant (121 °C for 15 min) and remained active upon treatment with surfactants and detergents. Cytotoxicity analysis of plantaricin DM5 on human embryonic kidney 293 (HEK 293) and human cervical cancer (HeLa) cell lines revealed its nontoxic and biocompatible nature. To the best of our knowledge, this is the first study on the isolated strain expressing probiotic properties and broad antimicrobial activity without any cytotoxic effect on mammalian cells from indigenous fermented beverage Marcha from India, and thus contributes to the food industry as a novel bio-preservant.

Bacteriocins from lactic acid bacteria: purification, properties and use as biopreservatives

OpenAIRE

Parada,José Luis; Caron,Carolina Ricoy; Medeiros,Adriane Bianchi P.; Soccol,Carlos Ricardo

2007-01-01

Biopreservation systems in foods are of increasing interest for industry and consumers. Bacteriocinogenic lactic acid bacteria and/or their isolated bacteriocins are considered safe additives (GRAS), useful to control the frequent development of pathogens and spoiling microorganisms in foods and feed. The spreading of bacterial antibiotic resistance and the demand for products with fewer chemicals create the necessity of exploring new alternatives, in order to reduce the abusive use of therap...

Complete genome sequence of Lactobacillus plantarum LZ95, a potential probiotic strain producing bacteriocins and B-group vitamin riboflavin.

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Li, Ping; Gu, Qing

2016-07-10

Lactobacillus plantarum LZ95 is a potential probiotic isolated from newborn infant fecal and it is identified to produce riboflavin with great antimicrobial activity. The complete genome sequence of this strain was reported in the present study. The genome contains a 3,261,418-bp chromosome and two plasmids. Genes, related to the biosynthesis of bacteriocins and riboflavin, were identified. This work will facilitate to reveal the biosynthetic mechanism of bacteriocins and B-group vitamins in lactic acid bacteria and provide evidence for its potential application in food industry. Copyright © 2016 Elsevier B.V. All rights reserved.

Bacteriocins: exploring alternatives to antibiotics in mastitis treatment

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Reneé Pieterse

2010-10-01

Full Text Available Mastitis is considered to be the most costly disease affecting the dairy industry. Management strategies involve the extensive use of antibiotics to treat and prevent this disease. Prophylactic dosages of antibiotics used in mastitis control programmes could select for strains with resistance to antibiotics. In addition, a strong drive towards reducing antibiotic residues in animal food products has lead to research in finding alternative antimicrobial agents. In this review we have focus on the pathogenesis of the mastitis in dairy cows, existing antibiotic treatments and possible alternative for application of bacteriocins from lactic acid bacteria in the treatment and prevention of this disease.

Expression of Genes Involved in Bacteriocin Production and Self-Resistance in Lactobacillus brevis 174A Is Mediated by Two Regulatory Proteins.

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Noda, Masafumi; Miyauchi, Rumi; Danshiitsoodol, Narandalai; Matoba, Yasuyuki; Kumagai, Takanori; Sugiyama, Masanori

2018-04-01

We have previously shown that the lactic acid bacterium Lactobacillus brevis 174A, isolated from Citrus iyo fruit, produces a bacteriocin designated brevicin 174A, which is comprised of two antibacterial polypeptides (designated brevicins 174A-β and 174A-γ). We have also found a gene cluster, composed of eight open reading frames (ORFs), that contains genes for the biosynthesis of brevicin 174A, self-resistance to its own bacteriocin, and two transcriptional regulatory proteins. Some lactic acid bacterial strains have a system to start the production of bacteriocin at an adequate stage of growth. Generally, the system consists of a membrane-bound histidine protein kinase (HPK) that senses a specific environmental stimulus and a corresponding response regulator (RR) that mediates the cellular response. We have previously shown that although the HPK- and RR-encoding genes are not found on the brevicin 174A biosynthetic gene cluster in the 174A strain, two putative regulatory genes, designated breD and breG , are in the gene cluster. In the present study, we demonstrate that the expression of brevicin 174A production and self-resistance is positively controlled by two transcriptional regulatory proteins, designated BreD and BreG. BreD is expressed together with BreE as the self-resistance determinant of L. brevis 174A. DNase I footprinting analysis and a promoter assay demonstrated that BreD binds to the breED promoter as a positive autoregulator. The present study also demonstrates that BreG, carrying a transmembrane domain, binds to the common promoter of breB and breC , encoding brevicins 174A-β and 174A-γ, respectively, for positive regulation. IMPORTANCE The problem of the appearance of bacteria that are resistant to practical antibiotics and the increasing demand for safe foods have increased interest in replacing conventional antibiotics with bacteriocin produced by the lactic acid bacteria. This antibacterial substance can inhibit the growth of pathogenic

In silico Prediction, in vitro Antibacterial Spectrum, and Physicochemical Properties of a Putative Bacteriocin Produced by Lactobacillus rhamnosus Strain L156.4

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Letícia de C. Oliveira

2017-05-01

Full Text Available A bacteriocinogenic Lactobacillus rhamnosus L156.4 strain isolated from the feces of NIH mice was identified by 16S rRNA gene sequencing and MALDI-TOF mass spectrometry. The entire genome was sequenced using Illumina, annotated in the PGAAP, and RAST servers, and deposited. Conserved genes associated with bacteriocin synthesis were predicted using BAGEL3, leading to the identification of an open reading frame (ORF that shows homology with the L. rhamnosus GG (ATCC 53103 prebacteriocin gene. The encoded protein contains a conserved protein motif associated a structural gene of the Enterocin A superfamily. We found ORFs related to the prebacteriocin, immunity protein, ABC transporter proteins, and regulatory genes with 100% identity to those of L. rhamnosus HN001. In this study, we provide evidence of a putative bacteriocin produced by L. rhamnosus L156.4 that was further confirmed by in vitro assays. The antibacterial activity of the substances produced by this strain was evaluated using the deferred agar-spot and spot-on-the lawn assays, and a wide antimicrobial activity spectrum against human and foodborne pathogens was observed. The physicochemical characterization of the putative bacteriocin indicated that it was sensitive to proteolytic enzymes, heat stable and maintained its antibacterial activity in a pH ranging from 3 to 9. The activity against Lactobacillus fermentum, which was used as an indicator strain, was detected during bacterial logarithmic growth phase, and a positive correlation was confirmed between bacterial growth and production of the putative bacteriocin. After a partial purification from cell-free supernatant by salt precipitation, the putative bacteriocin migrated as a diffuse band of approximately 1.0–3.0 kDa by SDS-PAGE. Additional studies are being conducted to explore its use in the food industry for controlling bacterial growth and for probiotic applications.

In silico Prediction, in vitro Antibacterial Spectrum, and Physicochemical Properties of a Putative Bacteriocin Produced by Lactobacillus rhamnosus Strain L156.4

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Oliveira, Letícia de C.; Silveira, Aline M. M.; Monteiro, Andréa de S.; dos Santos, Vera L.; Nicoli, Jacques R.; Azevedo, Vasco A. de C.; Soares, Siomar de C.; Dias-Souza, Marcus V.; Nardi, Regina M. D.

2017-01-01

A bacteriocinogenic Lactobacillus rhamnosus L156.4 strain isolated from the feces of NIH mice was identified by 16S rRNA gene sequencing and MALDI-TOF mass spectrometry. The entire genome was sequenced using Illumina, annotated in the PGAAP, and RAST servers, and deposited. Conserved genes associated with bacteriocin synthesis were predicted using BAGEL3, leading to the identification of an open reading frame (ORF) that shows homology with the L. rhamnosus GG (ATCC 53103) prebacteriocin gene. The encoded protein contains a conserved protein motif associated a structural gene of the Enterocin A superfamily. We found ORFs related to the prebacteriocin, immunity protein, ABC transporter proteins, and regulatory genes with 100% identity to those of L. rhamnosus HN001. In this study, we provide evidence of a putative bacteriocin produced by L. rhamnosus L156.4 that was further confirmed by in vitro assays. The antibacterial activity of the substances produced by this strain was evaluated using the deferred agar-spot and spot-on-the lawn assays, and a wide antimicrobial activity spectrum against human and foodborne pathogens was observed. The physicochemical characterization of the putative bacteriocin indicated that it was sensitive to proteolytic enzymes, heat stable and maintained its antibacterial activity in a pH ranging from 3 to 9. The activity against Lactobacillus fermentum, which was used as an indicator strain, was detected during bacterial logarithmic growth phase, and a positive correlation was confirmed between bacterial growth and production of the putative bacteriocin. After a partial purification from cell-free supernatant by salt precipitation, the putative bacteriocin migrated as a diffuse band of approximately 1.0–3.0 kDa by SDS-PAGE. Additional studies are being conducted to explore its use in the food industry for controlling bacterial growth and for probiotic applications. PMID:28579977

Effects of curing sodium nitrite additive and natural meat fat on growth control of Listeria monocytogenes by the bacteriocin-producing Lactobacillus curvatus strain CWBI-B28.

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Kouakou, P; Ghalfi, H; Destain, J; Dubois-Dauphin, R; Evrard, P; Thonart, P

2009-09-01

In realistic model meat systems, the separate and combined effects of fat content and sodium nitrite on the antilisterial activity of the bacteriocin of Lactobacillus curvatus CWBI-B28 were studied. In laboratory fermentations where Listeria monocytogenes was co-cultured at 4 degrees C with bacteriocin-producing CWBI-B28 in lean pork meat (fat content: 13%) without added nitrite, a strong antilisterial effect was observed after one week. The effect was maintained for an additional week, after which a slight and very gradual rebound was observed. Both added nitrite (20 ppm) and a high-fat content (43%) were found to antagonise this antilisterial effect, the Listeria cfu count reached after six weeks being 200 times as high in high-fat meat with added nitrite than in lean meat without nitrite. This antagonism could not be attributed to slower growth of the bacteriocin-producing strain, since CWBI-B28 grew optimally in fat-rich meat with 20 ppm sodium nitrite. Bacteriocin activity was also measured in the samples. The observed activity levels are discussed in relation to the degree of antilisterial protection conferred.

Sonorensin: an antimicrobial peptide, belonging to the heterocycloanthracin subfamily of bacteriocins, from a new marine isolate, Bacillus sonorensis MT93.

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Chopra, Lipsy; Singh, Gurdeep; Choudhary, Vikas; Sahoo, Debendra K

2014-05-01

Marine environments are the greatest fronts of biodiversity, representing a resource of unexploited or unknown microorganisms and new substances having potential applications. Among microbial products, antimicrobial peptides (AMPs) have received great attention recently due to their applications as food preservatives and therapeutic agents. A new marine soil isolate producing an AMP was identified as Bacillus sonorensis based on 16S rRNA gene sequence analysis. It produced an AMP that showed a broad spectrum of activity against both Gram-positive and Gram-negative bacteria. The peptide, named sonorensin, was purified to homogeneity using a combination of chromatographic techniques. The intact molecular mass of the purified peptide, 6,274 Da, as revealed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF), was in agreement with Tricine-SDS-PAGE analysis. A PCR array of primers was used to identify AMP structural genes, which allowed the successful amplification of the related genes from strain MT93. The putative open reading frame of sonorensin was amplified, cloned into the pET-32a(+) vector, expressed as a thioredoxin (Trx) fusion protein in Escherichia coli, and then purified. Sequence alignment analysis revealed that the bacteriocin being reported could belong to new subfamily of bacteriocins, heterocycloanthracin. The peptide indicated its potential as a biocontrol agent or food antimicrobial agent, due to its antimicrobial activity against bacteria such as Listeria monocytogenes and Staphylococcus aureus. This is the first report of the production, purification, and characterization of wild-type and recombinant bacteriocin by B. sonorensis and the first bacteriocin of the heterocycloanthracin subfamily to be characterized.

Enterocin F4-9, a Novel O-Linked Glycosylated Bacteriocin

OpenAIRE

Maky, Mohamed Abdelfattah; Ishibashi, Naoki; Zendo, Takeshi; Perez, Rodney Honrada; Doud, Jehan Ragab; Karmi, Mohamed; Sonomoto, Kenji

2015-01-01

Enterococcus faecalis F4-9 isolated from Egyptian salted-fermented fish produces a novel bacteriocin, termed enterocin F4-9. Enterocin F4-9 was purified from the culture supernatant by three steps, and its molecular mass was determined to be 5,516.6 Da by mass spectrometry. Amino acid and DNA sequencing showed that the propeptide consists of 67 amino acid residues, with a leader peptide containing a double glycine cleavage site to produce a 47-amino-acid mature peptide. Enterocin F4-9 is modi...

Characterization, production, and purification of leucocin H, a two-peptide bacteriocin from Leuconostoc MF215B.

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Blom, H; Katla, T; Holck, A; Sletten, K; Axelsson, L; Holo, H

1999-07-01

Leuconostoc MF215B was found to produce a two-peptide bacteriocin referred to as leucocin H. The two peptides were termed leucocin Halpha and leucocin Hbeta. When acting together, they inhibit, among others, Listeria monocytogenes, Bacillus cereus, and Clostridium perfringens. Production of leucocin H in growth medium takes place at temperatures down to 6 degrees C and at pH below 7. The highest activity of leucocin H in growth medium was demonstrated in the late exponential growth phase. The bacteriocin was purified by precipitation with ammonium sulfate, ion-exchange (SP Sepharose) and reverse phase chromatography. Upon purification, specific activity increased 10(5)-fold, and the final specific activity was 2 x 10(7) BU/OD280. Amino acid composition analyses of leucocin Halpha and leucocin Hbeta indicated that both peptides consisted of around 40 amino acid residues. Their N-termini were blocked for Edman degradation, and the methionin residues of leucocin Hbeta did not respond to Cyanogen Bromide (CNBr) cleavage. Absorbance at 280 nm indicated the presence of tryptophan residues and tryptophan-fracturing opened for partial sequencing by Edman degradation. From leucocin Halpha, the sequence of 20 amino acids was obtained; from leucocin Hbeta the sequence of 28 amino acid residues was obtained. No sequence homology to other known bacteriocins could be demonstrated. It also appeared that the two peptides themselves shared little or no sequence homology. The presence of soy oil did not affect the activity of leucocin H in agar.

Inhibition of Listeria monocytogenes on Ready-to-Eat Meats Using Bacteriocin Mixtures Based on Mode-of-Action

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Paul Priyesh Vijayakumar

2017-03-01

Full Text Available Bacteriocin-producing (Bac+ lactic acid bacteria (LAB comprising selected strains of Lactobacillus curvatus, Lactococcus lactis, Pediococcus acidilactici, and Enterococcus faecium and thailandicus were examined for inhibition of Listeria monocytogenes during hotdog challenge studies. The Bac+ strains, or their cell-free supernatants (CFS, were grouped according to mode-of-action (MOA as determined from prior studies. Making a mixture of as many MOAs as possible is a practical way to obtain a potent natural antimicrobial mixture to address L. monocytogenes contamination of RTE meat products (i.e., hotdogs. The heat resistance of the bacteriocins allowed the use of pasteurization to eliminate residual producer cells for use as post-process surface application or their inclusion into hotdog meat emulsion during cooking. The use of Bac+ LAB comprising 3× MOAs directly as co-inoculants on hotdogs was not effective at inhibiting L. monocytogenes. However, the use of multiple MOA Bac+ CFS mixtures in a variety of trials demonstrated the effectiveness of this approach by showing a >2-log decrease of L. monocytogenes in treatment samples and 6–7 log difference vs. controls. These data suggest that surface application of multiple mode-of-action bacteriocin mixtures can provide for an Alternative 2, and possibly Alternative 1, process category as specified by USDA-FSIS for control of L. monocytogenes on RTE meat products.

Enterocin X, a novel two-peptide bacteriocin from Enterococcus faecium KU-B5, has an antibacterial spectrum entirely different from those of its component peptides.

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Hu, Chih-Bo; Malaphan, Wanna; Zendo, Takeshi; Nakayama, Jiro; Sonomoto, Kenji

2010-07-01

Enterocin X, composed of two antibacterial peptides (Xalpha and Xbeta), is a novel class IIb bacteriocin from Enterococcus faecium KU-B5. When combined, Xalpha and Xbeta display variably enhanced or reduced antibacterial activity toward a panel of indicators compared to each peptide individually. In E. faecium strains that produce enterocins A and B, such as KU-B5, only one additional bacteriocin had previously been known.

Purification and characterization of enterocin 62-6, a two-peptide bacteriocin produced by a vaginal strain of Enterococcus faecium: Potential significance in bacterial vaginosis

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Dezwaan, Diane C.; Mequio, Michael J.; Littell, Julia S.; Allen, Jonathan P.; Rossbach, Silvia; Pybus, Vivien

2009-01-01

A bacteriocin produced by a vaginal isolate of Enterococcus faecium strain 62-6, designated enterocin 62-6, was characterized following purification and DNA sequence analysis and compared to previously described bacteriocins. Enterocin 62-6 was isolated from brain heart infusion (BHI) culture supernatants using ammonium sulfate precipitation followed by elution from a Sepharose cation exchange column using a continuous salt gradient (0.1–0.7 M NaCl). SDS-PAGE of an active column fraction resulted in an electrophoretically pure protein, which corresponded to the growth inhibition of the sensitive Lactobacillus indicator strain in the gel overlay assay. Purified enterocin 62-6 was shown to be heat- and pH-stable, and sensitive to the proteolytic enzymes α-chymotrypsin and pepsin. Results from mass spectrometry suggested that it comprised two peptides of 5206 and 5219±1 Da, which was confirmed by DNA sequence analysis. The characteristics of enterocin 62-6 as a small, heat- and pH-stable, cationic, hydrophobic, two-peptide, plasmid-borne bacteriocin, with an inhibitory spectrum against a broad range of Gram-positive but not Gram-negative bacteria, were consistent with its classification as a class IIc bacteriocin. Furthermore, its wide spectrum of growth inhibitory activity against Gram-positive bacteria of vaginal origin including lactobacilli, and stability under the acidic conditions of the vagina, are consistent with our hypothesis that it could have potential significance in disrupting the ecology of the vaginal tract and pave the way for the establishment of the abnormal microbiota associated with the vaginal syndrome bacterial vaginosis. This is the first class IIc bacteriocin produced by a strain of E. faecium of vaginal origin to be characterized. PMID:19578555

Expression of Streptococcus pneumoniae Bacteriocins Is Induced by Antibiotics via Regulatory Interplay with the Competence System

NARCIS (Netherlands)

Kjos, Morten; Miller, Eric; Slager, Jelle; Lake, Frank B; Gericke, Oliver; Roberts, Ian S; Rozen, Daniel E; Veening, Jan-Willem

2016-01-01

Pneumococcal bacteriocins (pneumocins) are antibacterial toxins that mediate intra-species competition within the human host. However, the triggers of pneumocin expression are poorly understood. Using RNA-sequencing, we mapped the regulon of the pneumocin cluster (blp) of Streptococcus pneumoniae

PRESERVATIVE POTENTIAL OF PURIFIED BACTERIOCIN PRODUCED FROM BREVIBACILLUS BORSTELENSIS AG1 ISOLATED FROM MARCHA – A TRADITIONAL WINE STARTER CULTURE CAKE IN TOMATO PASTE

OpenAIRE

Anupama Gupta; Nivedita Sharma; Neha Gautam

2015-01-01

Purified bacteriocin produced from Brevibacillus borstelensis AG1 isolated from Marcha a local wine starter herbal cake, was used to enhance the shelf life of tomato paste. Preservative effect of purified bacteriocin was studied for nine days in tomato paste inoculated with food borne pathogens and was compared to commercial biopreservative – nisin and chemical preservative – sodium benzoate. The indicator strains i.e. Listeria monocytogenes MTCC839, Bacillus subtilis CRI and Clostridium perf...

Analysis of Lactobacillus Products for Phages and Bacteriocins That Inhibit Vaginal Lactobacilli

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Lin Tao

1997-01-01

Full Text Available Objective: Bacterial vaginosis is associated with an unexplained loss of vaginal lactobacilli. Previously, we have identified certain vaginal lactobacilli-released phages that can inhibit in vitro other vaginal lactobacilli. However, there is no apparent route for phages to be transmitted among women. The purpose of this study was to identify whether certain Lactobacillus products commonly used by women release phages or bacteriocins that can inhibit vaginal lactobacilli.

Structure-activity relationships of an antimicrobial peptide plantaricin s from two-peptide class IIb bacteriocins.

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Soliman, Wael; Wang, Liru; Bhattacharjee, Subir; Kaur, Kamaljit

2011-04-14

Class IIb bacteriocins are ribosomally synthesized antimicrobial peptides comprising two different peptides synergistically acting in equal amounts for optimal potency. In this study, we demonstrate for the first time potent (nanomolar) antimicrobial activity of a representative class IIb bacteriocin, plantaricin S (Pls), against four pathogenic gram-positive bacteria, including Listeria monocytogenes. The structure-activity relationships for Pls were studied using activity assays, circular dichroism (CD), and molecular dynamics (MD) simulations. The two Pls peptides and five Pls derived fragments were synthesized. The CD spectra of the Pls and selected fragments revealed helical conformations in aqueous 2,2,2-trifluoroethanol. The MD simulations showed that when the two Pls peptides are in antiparallel orientation, the helical regions interact and align, mediated by strong attraction between conserved GxxxG/AxxxA motifs. The results strongly correlate with the antimicrobial activity suggesting that helix-helix alignment of the two Pls peptides and interaction between the conserved motifs are crucial for interaction with the target cell membrane.

Enterocin X, a Novel Two-Peptide Bacteriocin from Enterococcus faecium KU-B5, Has an Antibacterial Spectrum Entirely Different from Those of Its Component Peptides▿

Science.gov (United States)

Hu, Chih-Bo; Malaphan, Wanna; Zendo, Takeshi; Nakayama, Jiro; Sonomoto, Kenji

2010-01-01

Enterocin X, composed of two antibacterial peptides (Xα and Xβ), is a novel class IIb bacteriocin from Enterococcus faecium KU-B5. When combined, Xα and Xβ display variably enhanced or reduced antibacterial activity toward a panel of indicators compared to each peptide individually. In E. faecium strains that produce enterocins A and B, such as KU-B5, only one additional bacteriocin had previously been known. PMID:20418437

THE IMPORTANCE OF BACTERIOCINS IN MEAT AND MEAT PRODUCTS

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Meltem SERDAROĞLU

2000-03-01

Full Text Available There is an increasing consumer demand for food products which are free of chemical additives, reduced in salt and processed as little as possible. These minimally processed foods require special application to assure their microbiological safety. The use of microorganisms and enzymes for food preservatives is called biopreservation. The most important group of microorganisms with antimicrobial effect used in the production of foods is the lactic acid bacteria. In meats although lactic acid bacteria constitue apart of the initial microflora, they become dominant during the processing of meats. In this research bacteriocins of lactic acid bacteria and their usage in meat and meat products for biopreservation are discussed.

Bacteriocins – Exploring Alternatives to Antibiotics in Mastitis Treatment

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Pieterse, Reneé; Todorov, Svetoslav D.

2010-01-01

Mastitis is considered to be the most costly disease affecting the dairy industry. Management strategies involve the extensive use of antibiotics to treat and prevent this disease. Prophylactic dosages of antibiotics used in mastitis control programmes could select for strains with resistance to antibiotics. In addition, a strong drive towards reducing antibiotic residues in animal food products has lead to research in finding alternative antimicrobial agents. In this review we have focus on the pathogenesis of the mastitis in dairy cows, existing antibiotic treatments and possible alternative for application of bacteriocins from lactic acid bacteria in the treatment and prevention of this disease. PMID:24031528

Isolation and characterization of large spectrum and multiple bacteriocin-producing Enterococcus faecium strain from raw bovine milk.

Science.gov (United States)

Gaaloul, N; ben Braiek, O; Hani, K; Volski, A; Chikindas, M L; Ghrairi, T

2015-02-01

To assess the antimicrobial properties of lactic acid bacteria from Tunisian raw bovine milk. A bacteriocin-producing Enterococcus faecium strain was isolated from raw cow milk with activity against Gram-positive and Gram-negative bacteria. Antimicrobial substances produced by this strain were sensitive to proteolytic enzymes and were thermostable and resistant to a broad range of pH (2-10). Mode of action of antimicrobial substances was determined as bactericidal. Maximum activity was reached at the end of the exponential growth phase when checked against Listeria ivanovii BUG 496 (2366.62 AU ml(-1)). However, maximum antimicrobial activity against Pseudomonas aeruginosa 28753 was recorded at the beginning of the exponential growth phase. Enterococcus faecium GGN7 was characterized as free from virulence factors and was susceptible to tested antibiotics. PCR analysis of the micro-organism's genome revealed the presence of genes coding for enterocins A and B. Mass spectrometry analysis of RP-HPLC active fractions showed molecular masses corresponding to enterocins A (4835.77 Da) and B (5471.56 Da), and a peptide with a molecular mass of 3215.5 Da active only against Gram-negative indicator strains. The latter was unique in the databases. Enterococcus faecium GGN7 produces three bacteriocins with different inhibitory spectra. Based on its antimicrobial properties and safety, Ent. faecium GGN7 is potentially useful for food biopreservation. The results suggest the bacteriocins from GGN7 strain could be useful for food biopreservation. © 2014 The Society for Applied Microbiology.

Bacteriocins: molecules of fundamental impact on the microbial ecology and potential food biopreservatives

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Evandro Leite de Souza

2005-07-01

Full Text Available Bacteriocins are proteic molecules synthesized for various lineages of Gram-positive and Gram-negative bacteria when exposed to stressful conditions. Bacteriocins have been characterized as molecules of high antimicrobial property even at low concentrations, provoking the microbial survival inhibition by antibiosis. These substances have their synthesis mediated for genetic mechanisms and develop their lethal action on the microbial cell by multiples mechanisms that can act of isolated or concomitant way culminating with microbial cell killing. This molecules class presents characteristic of stability to heat, low pH, refrigeration and freezing, and resistance to weak organics solvents, salts and enzymes. On the other hand, they are very sensitive to proteolytic enzymes action. Bacteriocins could appear as potential agents to be applied in food conservation systems in order to provide microbiologically stable foods.Bacteriocinas são moléculas protéicas sintetizadas por várias linhagens de bactérias Gram-positivas e Gram-negativas quando submetidas a condições de stress. São ainda caracterizadas como moléculas de alto poder antimicrobiano mesmo em baixas concentrações, provocando a inibição da sobrevivência microbiana através de uma ação de antibiose. As bacteriocinas têm seu processo de síntese mediado por mecanismos genéticos, e desenvolvem sua ação letal sobre a célula microbiana por intermédio de múltiplos mecanismos que podem agir de forma isolada ou concomitante culminando com a morte da célula microbiana. Estas moléculas apresentam características de estabilidade ao calor, baixo pH, refrigeração, congelamento, resistência a ácidos orgânicos fracos, sais e enzimas, porém são muito sensíveis à enzimas proteolíticas. Assim, as bacteriocinas podem aparecer como potenciais agentes para serem aplicados em sistemas de conservação de alimentos com objetivo de prover alimentos microbiologicamente estáveis.

Bacteriocin AS-48 binding to model membranes and pore formation as revealed by coarse-grained simulations

NARCIS (Netherlands)

Cruz, Victor L.; Ramos, Javier; Martinez-Salazar, Javier; Melo, Manuel N.

Bacteriocin AS-48 is a membrane-interacting peptide that acts as a broad-spectrum antimicrobial against Gram-positive and Gram-negative bacteria. Prior Nuclear Magnetic Resonance experiments and the high resolution crystal structure of AS-48 have suggested a mechanism for the molecular activity of

Functional Analysis of Genes Involved in the Biosynthesis of Enterocin NKR-5-3B, a Novel Circular Bacteriocin.

Science.gov (United States)

Perez, Rodney H; Ishibashi, Naoki; Inoue, Tomoko; Himeno, Kohei; Masuda, Yoshimitsu; Sawa, Narukiko; Zendo, Takeshi; Wilaipun, Pongtep; Leelawatcharamas, Vichien; Nakayama, Jiro; Sonomoto, Kenji

2016-01-15

A putative biosynthetic gene cluster of the enterocin NKR-5-3B (Ent53B), a novel circular bacteriocin, was analyzed by sequencing the flanking regions around enkB, the Ent53B structural gene, using a fosmid library. A region approximately 9 kb in length was obtained, and the enkB1, enkB2, enkB3, and enkB4 genes, encoding putative biosynthetic proteins involved in the production, maturation, and secretion of Ent53B, were identified. We also determined the identity of proteins mediating self-immunity against the effects of Ent53B. Heterologous expression systems in various heterologous hosts, such as Enterococcus faecalis and Lactococcus lactis strains, were successfully established. The production and secretion of the mature Ent53B required the cooperative functions of five genes. Ent53B was produced only by those heterologous hosts that expressed protein products of the enkB, enkB1, enkB2, enkB3, and enkB4 genes. Moreover, self-immunity against the antimicrobial action of Ent53B was conferred by at least two independent mechanisms. Heterologous hosts harboring the intact enkB4 gene and/or a combination of intact enkB1 and enkB3 genes were immune to the inhibitory action of Ent53B. In addition to their potential application as food preservatives, circular bacteriocins are now considered possible alternatives to therapeutic antibiotics due to the exceptional stability conferred by their circular structure. The successful practical application of circular bacteriocins will become possible only if the molecular details of their biosynthesis are fully understood. The results of the present study offer a new perspective on the possible mechanism of circular bacteriocin biosynthesis. In addition, since some enterococcal strains are associated with pathogenicity, virulence, and drug resistance, the establishment of the first multigenus host heterologous production of Ent53B has very high practical significance, as it widens the scope of possible Ent53B applications

Purification and Characterization of Plantaricin JLA-9: A Novel Bacteriocin against Bacillus spp. Produced by Lactobacillus plantarum JLA-9 from Suan-Tsai, a Traditional Chinese Fermented Cabbage.

Science.gov (United States)

Zhao, Shengming; Han, Jinzhi; Bie, Xiaomei; Lu, Zhaoxin; Zhang, Chong; Lv, Fengxia

2016-04-06

Bacteriocins are ribosomally synthesized peptides with antimicrobial activity produced by numerous bacteria. A novel bacteriocin-producing strain, Lactobacillus plantarum JLA-9, isolated from Suan-Tsai, a traditional Chinese fermented cabbage, was screened and identified by its physiobiochemical characteristics and 16S rDNA sequence analysis. A new bacteriocin, designated plantaricin JLA-9, was purified using butanol extraction, gel filtration, and reverse-phase high-performance liquid chromatography. The molecular mass of plantaricin JLA-9 was shown to be 1044 Da by MALDI-TOF-MS analyses. The amino acid sequence of plantaricin JLA-9 was predicted to be FWQKMSFA by MALDI-TOF-MS/MS, which was confirmed by Edman degradation. This bacteriocin exhibited broad-spectrum antibacterial activity against Gram-positive and Gram-negative bacteria, especially Bacillus spp., high thermal stability (20 min, 121 °C), and narrow pH stability (pH 2.0-7.0). It was sensitive to α-chymotrypsin, pepsin, alkaline protease, and papain. The mode of action of this bacteriocin responsible for outgrowth inhibition of Bacillus cereus spores was studied. Plantaricin JLA-9 had no detectable effects on germination initiation over 1 h on monitoring the hydration, heat resistance, and 2,6-pyridinedicarboxylic acid (DPA) release of spores. Rather, germination initiation is a prerequisite for the action of plantaricin JLA-9. Plantaricin JLA-9 inhibited growth by preventing the establishment of oxidative metabolism and disrupting membrane integrity in germinating spores within 2 h. The results suggest that plantaricin JLA-9 has potential applications in the control of Bacillus spp. in the food industry.

Production of a bacteriocin-like inhibitory substance by Leuconostoc mesenteroides subsp. dextranicum 213M0 isolated from Mongolian fermented mare milk, airag.

Science.gov (United States)

Arakawa, Kensuke; Yoshida, Saki; Aikawa, Hiroki; Hano, Chihiro; Bolormaa, Tsognemekh; Burenjargal, Sedkhuu; Miyamoto, Taku

2016-03-01

Strain 213M0 was selected with productivity of a bacteriocin-like inhibitory substance (BLIS) among 235 strains of lactic acid bacteria (LAB) isolated from Mongolian fermented milk 'airag'. Strain 213M0 was species-identified as Leuconostoc mesenteroides subsp. dextranicum by morphological observation, carbohydrate fermentation profiling and sequencing the 16S rRNA gene. Incubation temperature proper to produce the BLIS was 25°C rather than 30 and 37°C, and the production actively proceeded during the exponential growth phase of the producer cells. Antibacterial effect of BLIS 213M0 was limited to all nine strains of Listeria sp. bacteria and seven strains of LAB cocci among 53 tested strains, which corresponds to a typical feature of the class IIa pediocin-like bacteriocins. BLIS 213M0 was not inactivated in every broad pH range solution (pH 2.0-11.0), and was stable against storage at 25°C for 1 week and heating at 121°C for 15 min under pH 4.5. Peptide frame of BLIS 213M0 was confirmed by inactivation with some peptidases, and then its molecular weight was estimated to be 2.6-3.0 kDa using an in situ activity assay following sodium dodecyl sulfate polyacrylamide gel electrophoresis. The estimated size was different from the other Leuconostoc bacteriocins already reported. These results suggest that BLIS 213M0 would be a novel listericidal bacteriocin. © 2015 Japanese Society of Animal Science.

Characterization of the genetic locus responsible for the production of ABP-118, a novel bacteriocin produced by the probiotic bacterium Lactobacillus salivarius subsp. salivarius UCC118.

Science.gov (United States)

Flynn, Sarah; van Sinderen, Douwe; Thornton, Gerardine M; Holo, Helge; Nes, Ingolf F; Collins, J Kevin

2002-04-01

ABP-118, a small heat-stable bacteriocin produced by Lactobacillus salivarius subsp. salivarius UCC118, a strain isolated from the ileal-caecal region of the human gastrointestinal tract, was purified to homogeneity. Using reverse genetics, a DNA fragment specifying part of ABP-118 was identified on a 10769 bp chromosomal region. Analysis of this region revealed that ABP-118 was a Class IIb two-peptide bacteriocin composed of Abp118alpha, which exhibited the antimicrobial activity, and Abp118beta, which enhanced the antimicrobial activity. The gene conferring strain UCC118 immunity to the action of ABP-118, abpIM, was identified downstream of the abp118beta gene. Located further downstream of abp118beta, several ORFs were identified whose deduced proteins resembled those of proteins involved in bacteriocin regulation and secretion. Heterologous expression of ABP-118 was achieved in Lactobacillus plantarum, Lactococcus lactis and Bacillus cereus. In addition, the abp118 locus encoded an inducing peptide, AbpIP, which was shown to play a role in the regulation of ABP-118 production. This novel bacteriocin is, to the authors' knowledge, the first to be isolated from a known human probiotic bacterium and to be characterized at the genetic level.

Characterization and Application of Enterocin RM6, a Bacteriocin from Enterococcus faecalis

OpenAIRE

Huang, En; Zhang, Liwen; Chung, Yoon-Kyung; Zheng, Zuoxing; Yousef, Ahmed E.

2013-01-01

Use of bacteriocins in food preservation has received great attention in recent years. The goal of this study is to characterize enterocin RM6 from Enterococcus faecalis OSY-RM6 and investigate its efficacy against Listeria monocytogenes in cottage cheese. Enterocin RM6 was purified from E. faecalis culture supernatant using ion exchange column, multiple C18-silica cartridges, followed by reverse-phase high-performance liquid chromatography. The molecular weight of enterocin RM6 is 7145.0823 ...

A food-grade process for isolation and partial purification of bacteriocins of lactic acid bacteria that uses diatomite calcium silicate.

Science.gov (United States)

Coventry, M J; Gordon, J B; Alexander, M; Hickey, M W; Wan, J

1996-01-01

Bacteriocins, including nisin, pediocin PO2, brevicin 286, and piscicolin 126, were extracted from fermentation media by adsorption onto Micro-Cel (a food-grade diatomite calcium silicate anticaking agent) and subsequent desorption. The optimal conditions for desorption of piscicolin 126 were determined and applied to other bacteriocins, and the relative purities of the desorbed preparations were compared. Piscicolin was not successfully desorbed from Micro-Cel at pH 1.0 to 12.0, with organic solvents, or by increase of ionic strength up to 1 M NaCl. However, 25 and 75% of the bacteriocin activity was desorbed by using 1% sodium deoxycholate and 1% sodium dodecyl sulfate (SDS), respectively. Higher levels (up to 100%) of desorption were achieved by repeated elution or by an increase in surfactant concentration. Desorption of piscicolin with 1/10 volume of SDS solution resulted in a preparation with 10 times concentration in activity, equivalent to that of ammonium sulfate preparations (409,600 to 819,200 activity units/ml). Determination of organic nitrogen (N) content revealed that the desorbed piscicolin preparations were substantially free of proteinaceous substances (approximately 92 to 99%) compared with original culture supernatants and ammonium sulfate preparations. Nisin, pediocin, and brevicin were also desorbed with 1% SDS with a similar level of purification. PMID:8633875

Rapid and efficient purification method for small, hydrophobic, cationic bacteriocins : Purification of lactococcin B and pediocin PA-1

NARCIS (Netherlands)

Venema, Koen; Chikindas, Michael L.; Seegers, Jos F.M.L.; Haandrikman, Alfred J.; Leenhouts, Kees J.; Venema, Gerard; Kok, Jan

The bacteriocins lactococcin B and pediocin PA 1 were purified by ethanol precipitation, preparative isoelectric focusing, and ultrafiltration. The procedure reproducibly leads to high final yields in comparison to the generally low yields obtained by column chromatography. Specifically, during

Purification and characterization of enterocin 4, a bacteriocin produced by Enterococcus faecalis INIA 4.

Science.gov (United States)

Joosten, H M; Nunez, M; Devreese, B; Van Beeumen, J; Marugg, J D

1996-01-01

A simple two-step procedure was developed to obtain pure enterocin 4, a bacteriocin produced by Enterococcus faecalis INIA 4. Chemical and genetic characterization revealed that the primary structure of enterocin 4 is identical to that of peptide antibiotic AS-48 from Enterococcus faecalis S-48. In contrast to the reported inhibitory spectrum of AS-48, enterocin 4 displayed no activity against gram-negative bacteria. PMID:8900014

Purification and characterization of enterocin 4, a bacteriocin produced by Enterococcus faecalis INIA 4.

OpenAIRE

Joosten, H M; Nunez, M; Devreese, B; Van Beeumen, J; Marugg, J D

1996-01-01

A simple two-step procedure was developed to obtain pure enterocin 4, a bacteriocin produced by Enterococcus faecalis INIA 4. Chemical and genetic characterization revealed that the primary structure of enterocin 4 is identical to that of peptide antibiotic AS-48 from Enterococcus faecalis S-48. In contrast to the reported inhibitory spectrum of AS-48, enterocin 4 displayed no activity against gram-negative bacteria.

Optimization of growth and bacteriocin activity of the food bioprotective Carnobacterium divergens V41 in an animal origin protein free medium

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Anne BRILLET-VIEL

2016-08-01

Full Text Available Optimization of Carnobacterium divergens V41 growth and bacteriocin activity in a culture medium deprived of animal protein, needs for food bioprotection, was performed by using a statistical approach. In a screening experiment, twelve factors (pH, temperature, carbohydrates, NaCl, yeast extract, soy peptone, sodium acetate, ammonium citrate, magnesium sulphate, manganese sulphate, ascorbic acid and thiamine were tested for their influence on the maximal growth and bacteriocin activity using a two-level incomplete factorial design with 192 experiments performed in microtiter plate wells. Based on results, a basic medium was developed and three variables (pH, temperature and carbohydrates concentration were selected for a scale-up study in bioreactor. A 23 complete factorial design was performed, allowing the estimation of linear effects of factors and all the first order interactions. The best conditions for the cell production were obtained with a temperature of 15°C and a carbohydrates concentration of 20 g/l whatever the pH (in the range 6.5-8, and the best conditions for bacteriocin activity were obtained at 15°C and pH 6.5 whatever the carbohydrates concentration (in the range 2-20 g/l. The predicted final count of C. divergens V41 and the bacteriocin activity under the optimized conditions (15°C, pH 6.5, 20 g/l carbohydrates were 2.4 x 1010 CFU/ml and 819200 AU/ml respectively. C. divergens V41 cells cultivated in the optimized conditions were able to grow in cold-smoked salmon and totally inhibited the growth of Listeria monocytogenes (< 50 CFU g-1 during five weeks of vacuum storage at 4° and 8°C.

Characterization of a Bacteriocin-Like Substance Produced by a Vaginal Lactobacillus salivarius Strain

Science.gov (United States)

Ocaña, Virginia S.; Pesce de Ruiz Holgado, Aída A.; Nader-Macías, María Elena

1999-01-01

A novel bacteriocin-like substance produced by vaginal Lactobacillus salivarius subsp. salivarius CRL 1328 with activity against Enterococcus faecalis, Enterococcus faecium, and Neisseria gonorrhoeae was characterized. The highest level of production of this heat-resistant peptide or protein occurred during the late exponential phase. Its mode of action was shown to be bactericidal. L. salivarius subsp. salivarius CRL 1328 could be used for the design of a probiotic to prevent urogenital infections. PMID:10584033

Identification, Characterization, and Three-Dimensional Structure of the Novel Circular Bacteriocin, Enterocin NKR-5-3B, from Enterococcus faecium.

Science.gov (United States)

Himeno, Kohei; Rosengren, K Johan; Inoue, Tomoko; Perez, Rodney H; Colgrave, Michelle L; Lee, Han Siean; Chan, Lai Y; Henriques, Sónia Troeira; Fujita, Koji; Ishibashi, Naoki; Zendo, Takeshi; Wilaipun, Pongtep; Nakayama, Jiro; Leelawatcharamas, Vichien; Jikuya, Hiroyuki; Craik, David J; Sonomoto, Kenji

2015-08-11

Enterocin NKR-5-3B, one of the multiple bacteriocins produced by Enterococcus faecium NKR-5-3, is a 64-amino acid novel circular bacteriocin that displays broad-spectrum antimicrobial activity. Here we report the identification, characterization, and three-dimensional nuclear magnetic resonance solution structure determination of enterocin NKR-5-3B. Enterocin NKR-5-3B is characterized by four helical segments that enclose a compact hydrophobic core, which together with its circular backbone impart high stability and structural integrity. We also report the corresponding structural gene, enkB, that encodes an 87-amino acid precursor peptide that undergoes a yet to be described enzymatic processing that involves adjacent cleavage and ligation of Leu(24) and Trp(87) to yield the mature (circular) enterocin NKR-5-3B.

Purification, Characterization, and Mode of Action of Pentocin JL-1, a Novel Bacteriocin Isolated from Lactobacillus pentosus, against Drug-Resistant Staphylococcus aureus

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Han Jiang

2017-01-01

Full Text Available Staphylococcus aureus and its drug-resistant strains, which threaten public health and food safety, are in need of effective control by biopreservatives. A novel bacteriocin, pentocin JL-1, produced by Lactobacillus pentosus that was isolated from the intestinal tract of Chiloscyllium punctatum, was purified by a four-step chromatographic process. Mass spectrometry based on MALDI-TOF indicated that pentocin JL-1 has a molecular mass of 2987.23 Da. Only six of the twenty-five amino acids could be identified by Edman degradation. This bacteriocin is thermostable and tolerates a pH range of 5–7. Also, it is sensitive to proteinase K, trypsin, pepsin, and alkaline protease. This bacteriocin has a broad inhibitory spectrum against both Gram-positive and Gram-negative strains and in particular is effective against multidrug-resistant S. aureus. Additionally, we showed that the cell membrane is the target of pentocin JL-1 against methicillin-resistant S. aureus (MRSA, causing a loss of proton motive force. Furthermore, pentocin JL-1 has a drastic impact on the structure and integrity of MRSA cells. These results suggest that pentocin JL-1 has potential as a biopreservative in the food industry.

Characterization and purification of a bacteriocin from Lactobacillus paracasei subsp. paracasei BMK2005, an intestinal isolate active against multidrug-resistant pathogens.

Science.gov (United States)

Bendjeddou, Kamel; Fons, Michel; Strocker, Pierre; Sadoun, Djamila

2012-04-01

A strain of Lactobacillus paracasei subsp. paracasei BMK2005 isolated from healthy infant faeces has shown a remarkable antibacterial activity against 32 bacterial pathogenic strains of human clinical isolates. Among them, 13 strains belonging to species of Escherichia coli, Citrobacter freundii, Citrobacter diversus, Klebsiella oxytoca, Enterobacter cloacae and Pseudomonas aeruginosa were resistant to Cefotaxime (CTX) and Ceftazidime (CAZ), and 4 strains of Staphylococcus aureus were resistant to Methicillin (MRSA). This antibacterial activity was attributed to a bacteriocin designated as Paracaseicin A. It was heat-stable up to 120°C for 5 min and active within the pH range of 2-5. Its activity was lost when treated with proteases, which reveals its proteinaceous nature. This bacteriocin was successfully purified only by two steps of reversed phase chromatography. Its molecular mass, determined by mass spectrometry analysis, was 2,462.5 Da. To our knowledge, the present study is the first report on characterization and purification of a bacteriocin, produced by a L. paracasei subsp. paracasei strain exhibiting an antibacterial activity against various multidrug-resistant species of Gram-positive and Gram-negative bacteria, which reveals its potential for use in prevention or treatment of infections caused by multidrug-resistant species especially in cases of antibiotics-associated diarrhea (AAD).

Bacteriocin production by Lactobacillus pentosus ST712BZ isolated from boza Bacteriocina produzida por Lactobacillus pentosus ST712BZ isolad de boza

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Svetoslav D. Todorov

2007-03-01

Full Text Available Bacteriocin ST712BZ (14.0kDa in size inhibits the growth of Lactobacillus casei,Escherichia coli, Pseudomonas aeruginosa,Enterococcus faecalis, Klebsiella pneumoniae and Lactobacillus curvatus. Growth of strain ST712BZ in BHI, M17, soy milk and molasses was similar to growth in MRS, with optimal bacteriocin production (12800AU/mL recorded in MRS after 24h. The same level of bacteriocin production (12800AU/mL was recorded in MRS broth with an initial pH of 6.5, 6.0 and 5.5. However, MRS broth (pH 6.5 supplemented with 1mM EDTA, yielded only 6400AU/mL. Low levels of bacteriocin activity were recorded in MRS broth with an initial pH of 5.0 and 4.5. Of all media compositions tested, MRS supplemented with tryptone (20.0g/L, glucose (20.0 to 40.0g/L, mannose (20.0g/L, vitamin B12, or vitamin C yielded 12800AU/mL. Glycerol concentrations of 1.0g/L and higher repressed bacteriocin production. Maximal bacteriocin activity (25600AU/mL was recorded in MRS supplemented with Vit. B1 or DL-6,8-thioctic acid.A bacteriocina ST712BZ produzida par Lactobacillus pentosus (peso molecular de 14,0kDa inibe o crescimento de Lactobacillus casei,Escherichia coli, Pseudomonas aeruginosa,Enterococcus faecalis, Klebsiella pneumoniae e Lactobacillus curvatus. O crescimento de L. pentosus ST712BZ em BHI, M17, leite de soja e melaços foi semelhante ao observado em MRS, registando-se a produção máxima de bacteriocina (12800UA/mL em MRS após 24 h. Observou-se o mesmo nível de produção de bacteriocina (12800UA/mL em caldo MRS com pH inicial de 6,5, 6,0 e 5,5. No entanto, em caldo MRS (pH 6,5 suplementado com 1 mM de EDTA a produção apenas atingiu 6400UA/mL. Os níveis de atividade bacteriocinogênica detectados em caldo MRS com um pH inicial de 5,0 e 4,5 foram baixos. De todas as fórmulas de meios de cultura testadas a que apresentou a atividade máxima 12800UA/mL foi MRS suplemento de triptona (20,0g/L, glicose (20,0 e 40,0 g/L, manose (20.0 g/L, vitamina B12 e

Isolation and characterization of a new bacteriocin, termed enterocin M, produced by environmental isolate Enterococcus faecium AL41.

Science.gov (United States)

Mareková, Mária; Lauková, Andrea; Skaugen, Morten; Nes, Ingolf

2007-08-01

The new bacteriocin, termed enterocin M, produced by Enterococcus faecium AL 41 showed a wide spectrum of inhibitory activity against the indicator organisms from different sources. It was purified by (NH4)2SO4 precipitation, cation-exchange chromatography and reverse phase chromatography (FPLC). The purified peptide was sequenced by N-terminal amino acid Edman degradation and a mass spectrometry analysis was performed. By combining the data obtained from amino acid sequence (39 N-terminal amino acid residues was determined) and the molecular weight (determined to be 4628 Da) it was concluded that the purified enterocin M is a new bacteriocin, which is very similar to enterocin P. However, its molecular weight is different from enterocin P (4701.25). Of the first 39 N-terminal residues of enterocin M, valine was found in position 20 and a lysine in position 35, while enterocin P has tryptophane residues in these positions.

Purification and biochemical characterization of a highly thermostable bacteriocin isolated from Brevibacillus brevis strain GM100.

Science.gov (United States)

Ghadbane, Mouloud; Harzallah, Daoud; Laribi, Atef Ibn; Jaouadi, Bassem; Belhadj, Hani

2013-01-01

A bacteriocin-producing (11,000 AU mL(-1)) strain was isolated from the rhizosphere of healthy Algerian plants Ononis angustissima Lam., and identified as Brevibacillus brevis strain GM100. The bacteriocin, called Bac-GM100, was purified to homogeneity from the culture supernatant, and, based on MALDI-TOF/MS analysis, was a monomer protein with a molecular mass of 4375.66 Da. The 21 N-terminal residues of Bac-GM100 displayed 65% homology with thurincin H from Bacillus thuringiensis. Bac-GM100 was extremely heat-stable (20 min at 120 °C), and was stable within a pH range of 3-10. It proved sensitive to various proteases, which demonstrated its protein nature. It was also found to display a bactericidal mode of action against gram-negative (Salmonella enteric ATCC 43972, Pseudomonas aeruginosa ATCC 49189, and Agrobacterium tumefaciens C58) and gram-positive (Enterococcus faecalis ENSAIA 631 and Staphylococcus aureus ATCC 6538) bacteria, and a fungistatic mode of action against the pathogenic fungus Candida tropicalis R2 CIP 203.

Pentocin MQ1: A Novel, Broad-Spectrum, Pore-Forming Bacteriocin From Lactobacillus pentosus CS2 With Quorum Sensing Regulatory Mechanism and Biopreservative Potential

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Samson B. Wayah

2018-03-01

Full Text Available Micrococcus luteus, Listeria monocytogenes, and Bacillus cereus are major food-borne pathogenic and spoilage bacteria. Emergence of antibiotic resistance and consumer demand for foods containing less of chemical preservatives led to a search for natural antimicrobials. A study aimed at characterizing, investigating the mechanism of action and regulation of biosynthesis and evaluating the biopreservative potential of pentocin from Lactobacillus pentosus CS2 was conducted. Pentocin MQ1 is a novel bacteriocin isolated from L. pentosus CS2 of coconut shake origin. The purification strategy involved adsorption-desorption of bacteriocin followed by RP-HPLC. It has a molecular weight of 2110.672 Da as determined by MALDI-TOF mass spectrometry and a molar extinction value of 298.82 M−1 cm−1. Pentocin MQ1 is not plasmid-borne and its biosynthesis is regulated by a quorum sensing mechanism. It has a broad spectrum of antibacterial activity, exhibited high chemical, thermal and pH stability but proved sensitive to proteolytic enzymes. It is potent against M. luteus, B. cereus, and L. monocytogenes at micromolar concentrations. It is quick-acting and exhibited a bactericidal mode of action against its targets. Target killing was mediated by pore formation. We report for the first time membrane permeabilization as a mechanism of action of the pentocin from the study against Gram-positive bacteria. Pentocin MQ1 is a cell wall-associated bacteriocin. Application of pentocin MQ1 improved the microbiological quality and extended the shelf life of fresh banana. This is the first report on the biopreservation of banana using bacteriocin. These findings place pentocin MQ1 as a potential biopreservative for further evaluation in food and medical applications.

Purification and characterization of the bacteriocin Thuricin Bn1 produced by Bacillus thuringiensis subsp. kurstaki Bn1 isolated from a hazelnut pest.

Science.gov (United States)

Ugras, Serpil; Sezen, Kazim; Kati, Hatice; Demirbag, Zihni

2013-02-01

A novel bioactive molecule produced by Bacillus thuringiensis subsp. kurstaki Bn1 (Bt-Bn1), isolated from a common pest of hazelnut, Balaninus nucum L. (Coleoptera: Curculionidae), was determined, purified, and characterized in this study. The Bt-Bn1 strain was investigated for antibacterial activity with an agar spot assay and well diffusion assay against B. cereus, B. weinhenstephenensis, L. monocytogenes, P. savastanoi, P. syringae, P. lemoignei, and many other B. thuringiensis strains. The production of bioactive molecule was determined at the early logarithmic phase in the growth cycle of strain Bt-Bn1 and its production continued until the beginning of the stationary phase. The mode of action of this molecule displayed bacteriocidal or bacteriolytic effect depending on the concentration. The bioactive molecule was purified 78-fold from the bacteria supernatant with ammonium sulfate precipitation, dialysis, ultrafiltration, gel filtration chromatography, and HPLC, respectively. The molecular mass of this molecule was estimated via SDS-PAGE and confirmed by the ESI-TOFMS as 3,139 Da. The bioactive molecule was also determined to be a heat-stable, pH-stable (range 6-8), and proteinase K sensitive antibacterial peptide, similar to bacteriocins. Based on all characteristics determined in this study, the purified bacteriocin was named as thuricin Bn1 because of the similarities to the previously identified thuricin-like bacteriocin produced by the various B. thuringiensis strains. Plasmid elution studies showed that gene responsible for the production of thuricin Bn1 is located on the chromosome of Bt-Bn1. Therefore, it is a novel bacteriocin and the first recorded one produced by an insect originated bacterium. It has potential usage for the control of many different pathogenic and spoilage bacteria in the food industry, agriculture, and various other areas.

Description of two Enterococcus strains isolated from traditional Peruvian artisanal-produced cheeses with a bacteriocin-like inhibitory activity

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Aguilar Galvez A.

2009-01-01

Full Text Available The aim of this work was to isolate and to characterize strains of lactic acid bacteria (LAB with bacteriocin-like inhibitory activity from 27 traditional cheeses artisanal-produced obtained from different Peruvian regions. Twenty Gram+ and catalasenegative strains among 2,277 isolates exhibited bacteriocin-like inhibitory activity against Listeria monocytogenes CWBIB2232 as target strain. No change in inhibitory activity was observed after organic acid neutralization and treatment with catalase of the cell-free supernatant (CFS. The proteinic nature of the antimicrobial activity was confirmed for the twenty LAB strains by proteolytic digestion of the CFS. Two strains, CWBI-B1431 and CWBI-B1430, with the best antimicrobial activity were selected for further researches. These strains were taxonomically identified by phenotypic and genotypic analyses as Enterococcus mundtii (CWBI-B1431 and Enterococcus faecium (CWBI-B1430. The two strains were sensitive to vancomycin (MIC 2 μg.ml-1 and showed absence of haemolysis.

Lacticin LC14, a new bacteriocin produced by Lactococcus lactis BMG6.14: isolation, purification and partial characterization.

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Lasta, Samar; Ouzari, Hadda; Andreotti, Nicolas; Fajloun, Ziad; Mansuelle, Pascal; Boudabous, Abdellatif; Sampieri, Francois; Sabatier, Jean Marc

2012-08-01

A new bacteriocin, lacticin LC14, produced by Lactococcus lactis BMG6.14, was isolated and characterized. It was purified to homogeneity from overnight broth culture by ammonium sulfate precipitation, Sep-Pak chromatography, and two steps of reversed-phase HPLC. Lacticin LC14 showed bactericidal-type antimicrobial activity against several lactic acid bacteria and pathogenic strains including Listeria monocytogenes. It was inactivated by proteinase K and pronase E, but was resistant to papain, lysozyme, lipase and catalase. Lacticin LC14 was heat resistant, stable over a wide range of pH (2-10) and after treatment by solvents and detergents. Its N-terminal end was found unreactive towards Edman sequencing. Based on MALDI-TOF mass spectrometry, its molecular mass was 3333.7 Da. LC14 amino acid composition revealed a high proportion of hydrophobic residues, but no modified ones. LC14 may be able to challenge other well known other bacteriocins in probiotic and therapeutic applications.

Application of the broad-spectrum bacteriocin enterocin AS-48 to inhibit Bacillus coagulans in canned fruit and vegetable foods.

Science.gov (United States)

Lucas, R; Grande, M A J; Abriouel, H; Maqueda, M; Ben Omar, N; Valdivia, E; Martínez-Cañamero, M; Gálvez, A

2006-10-01

The enterococcal bacteriocin (enterocin) AS-48 is a broad-spectrum cyclic peptide. Enterocin AS-48 was tested against Bacillus coagulans in three vegetable canned foods: tomato paste (pH 4.64), syrup from canned peaches (pH 3.97), and juice from canned pineapple (pH 3.65). When vegetative cells of B. coagulans CECT (Spanish Type Culture Collection) 12 were inoculated in tomato paste supplemented with 6 microg/ml AS-48 and stored at different temperatures, viable cell counts were reduced by approximately 2.37 (4 degrees C), 4.3 (22 degrees C) and 3.0 (37 degrees C) log units within 24 h storage. After 15-days storage, no viable cells were detected in any sample. Strain B. coagulans CECT 561 showed a poor survival in tomato paste, but surviving cells were also killed by AS-48. The bacteriocin was also very active against B. coagulans CECT 12 vegetative cells in juice from canned pineapple stored at 22 degrees C, and slightly less active in syrup from canned peaches. In food samples supplemented with 1.5% lactic acid, enterocin AS-48 (6 microg/ml) rapidly reduced viable counts of vegetative cells below detection limits within 24 h storage. Addition of glucose and sucrose (10% and 20%) significantly increased bacteriocin activity against vegetative cells of B. coagulans CECT 12. Enterocin AS-48 had no significant effect on B. coagulans CECT 12 spores. However, the combined application of AS-48 and heat (80-95 degrees C for 5 min) significantly increased the effect of thermal treatments on spores.

Detection of Bacteriocins by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

OpenAIRE

Rose, Natisha L.; Sporns, Peter; McMullen, Lynn M.

1999-01-01

The use of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the detection of bacteriocins was investigated. A 30-s water wash of the sample on the MALDI-TOF MS probe was effective in removing contaminants of the analyte. This method was used for rapid detection of nisin, pediocin, brochocin A and B, and enterocin A and B from culture supernatants and for detection of enterocin B throughout its purification.

A peptide factor secreted by Staphylococcus pseudintermedius exhibits properties of both bacteriocins and virulence factors.

Science.gov (United States)

Wladyka, Benedykt; Piejko, Marcin; Bzowska, Monika; Pieta, Piotr; Krzysik, Monika; Mazurek, Łukasz; Guevara-Lora, Ibeth; Bukowski, Michał; Sabat, Artur J; Friedrich, Alexander W; Bonar, Emilia; Międzobrodzki, Jacek; Dubin, Adam; Mak, Paweł

2015-09-28

Staphylococcus pseudintermedius is a common commensal bacterium colonizing the skin and mucosal surfaces of household animals. However, it has recently emerged as a dangerous opportunistic pathogen, comparable to S. aureus for humans. The epidemiological situation is further complicated by the increasing number of methicillin-resistant S. pseudintermedius infections and evidence of gene transmission driving antibiotic resistance between staphylococci colonizing human and zoonotic hosts. In the present study, we describe a unique peptide, BacSp222, that possesses features characteristic of both bacteriocins and virulence factors. BacSp222 is secreted in high quantities by S. pseudintermedius strain 222 isolated from dog skin lesions. This linear, fifty-amino-acid highly cationic peptide is plasmid-encoded and does not exhibit significant sequence similarities to any other known peptides or proteins. BacSp222 kills gram-positive bacteria (at doses ranging from 0.1 to several micromol/l) but also demonstrates significant cytotoxic activities towards eukaryotic cells at slightly higher concentrations. Moreover, at nanomolar concentrations, the peptide also possesses modulatory properties, efficiently enhancing interferon gamma-induced nitric oxide release in murine macrophage-like cell lines. BacSp222 appears to be one of the first examples of multifunctional peptides that breaks the convention of splitting bacteriocins and virulence factors into two unrelated groups.

Genotypic and Phylogenic Analysis of Lactobacilli Producing Bacteriocin Isolated from Traditional Dairy Products and Food

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Frazaneh Tafvizi

2012-09-01

Full Text Available Background & Objective: Lactic acid bacteria (LAB are a group of Gram-positive, non-spore forming, cocci or rod shaped, catalase negative organisms, considered as Generally Recognized as Safe (GRAS organisms. These bacteria are used for thousands of years for production of fermented foods because of their ability to produce desirable changes in taste, flavor and texture. Different antimicrobial molecules such as bacteriocins produced by these bacteria that can inhibit food pathogens, so enhancing the shelf life and improving the safety of food products. Because of important role of LAB to improving the human health, molecular identification and phylogenic analysis of these bacteria based on 16S rRNA sequencing play the critical role in investigation of local sources of LAB in Iran. Materials & Methods: 5 isolates were selected from 20 isolates for molecular identification. These strains produced the high level of bacteriocin. Total genomic DNA was extracted by lysosyme extraction protocol. PCR-mediated amplification was carried out by degenerate primers. Sequencing was performed after purification of PCR product. Results: Isolates were deposited as novel strains of Lactobacillus casei and Entrococcus facium in GenBank. Conclusion: Because of high potential of local probiotic bacteria in Iran, these strains may be useful and could be used in the food industry.

Enterocin AS-48 as Evidence for the Use of Bacteriocins as New Leishmanicidal Agents.

Science.gov (United States)

Abengózar, María Ángeles; Cebrián, Rubén; Saugar, José María; Gárate, Teresa; Valdivia, Eva; Martínez-Bueno, Manuel; Maqueda, Mercedes; Rivas, Luis

2017-04-01

We report the feasibility of enterocin AS-48, a circular cationic peptide produced by Enterococcus faecalis , as a new leishmanicidal agent. AS-48 is lethal to Leishmania promastigotes as well as to axenic and intracellular amastigotes at low micromolar concentrations, with scarce cytotoxicity to macrophages. AS-48 induced a fast bioenergetic collapse of L. donovani promastigotes but only a partial permeation of their plasma membrane with limited entrance of vital dyes, even at concentrations beyond its full lethality. Fluoresceinated AS-48 was visualized inside parasites by confocal microscopy and seen to cause mitochondrial depolarization and reactive oxygen species production. Altogether, AS-48 appeared to have a mixed leishmanicidal mechanism that includes both plasma membrane permeabilization and additional intracellular targets, with mitochondrial dysfunctionality being of special relevance. This complex leishmanicidal mechanism of AS-48 persisted even for the killing of intracellular amastigotes, as evidenced by transmission electron microscopy. We demonstrated the potentiality of AS-48 as a new and safe leishmanicidal agent, expanding the growing repertoire of eukaryotic targets for bacteriocins, and our results provide a proof of mechanism for the search of new leishmanicidal bacteriocins, whose diversity constitutes an almost endless source for new structures at moderate production cost and whose safe use on food preservation is well established. Copyright © 2017 American Society for Microbiology.

Bacteriocins from Lactobacillus plantarum production, genetic organization and mode of action: produção, organização genética e modo de ação Bacteriocinas de Lactobacillus plantarum

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Svetoslav D. Todorov

2009-06-01

Full Text Available Bacteriocins are biologically active proteins or protein complexes that display a bactericidal mode of action towards usually closely related species. Numerous strains of bacteriocin producing Lactobacillus plantarum have been isolated in the last two decades from different ecological niches including meat, fish, fruits, vegetables, and milk and cereal products. Several of these plantaricins have been characterized and the aminoacid sequence determined. Different aspects of the mode of action, fermentation optimization and genetic organization of the bacteriocin operon have been studied. However, numerous of bacteriocins produced by different Lactobacillus plantarum strains have not been fully characterized. In this article, a brief overview of the classification, genetics, characterization, including mode of action and production optimization for bacteriocins from Lactic Acid Bacteria in general, and where appropriate, with focus on bacteriocins produced by Lactobacillus plantarum, is presented.Bacteriocinas são proteínas ou complexos protéicos biologicamente ativos que apresentam atividade bactericida contra espécies relacionadas. Nas ultimas duas décadas, várias cepas de Lactobacillus plantarum produtoras de bacteriocinas foram isoladas de diferentes nichos ecológicos como carnes, peixes, frutas, vegetais e produtos lácteos e de cereais. Várias plantaricinas foram caracterizadas e suas seqüências de aminoácidos determinadas. Diferentes aspectos do modo de ação, otimização da fermentação e organização genética já foram estudados. Entretanto, muitas bacteriocinas produzidas por diferentes cepas de Lactobacillus plantarum ainda não foram completamente caracterizadas.Nesse artigo, apresenta-se uma breve revisão sobre a classificação, genética, caracterização, modo de ação, e otimização da produção de bacteriocinas de bactérias láticas em geral, e, quando apropriado, de bacteriocinas de Lactobacillus plantarum.

Large-Scale Purification, Characterization, and Spore Outgrowth Inhibitory Effect of Thurincin H, a Bacteriocin Produced by Bacillus thuringiensis SF361.

Science.gov (United States)

Wang, Gaoyan; Manns, David C; Guron, Giselle K; Churey, John J; Worobo, Randy W

2014-06-01

Large-scale purification of the highly hydrophobic bacteriocin thurincin H was accomplished via a novel and simple two-step method: ammonia sulfate precipitation and C18 solid-phase extraction. The inhibition spectrum and stability of thurincin H as well as its antagonistic activity against Bacillus cereus F4552 spores were further characterized. In the purification method, secreted proteins contained in the supernatant of a 40 h incubated culture of B. thuringiensis SF361 were precipitated by 68 % ammonia sulfate and purified by reverse-phase chromatography, with a yield of 18.53 mg/l of pure thurincin H. Silver-stained SDS-PAGE, high-performance liquid chromatography, and liquid chromatography-mass spectrometry confirmed the high purity of the prepared sample. Thurincin H exhibited a broad antimicrobial activity against 22 tested bacterial strains among six different genera including Bacillus, Carnobacterium, Geobacillus, Enterococcus, Listeria, and Staphylococcus. There was no detectable activity against any of the selected yeast or fungi. The bacteriocin activity was stable for 30 min at 50 °C and decreased to undetectable levels within 10 min at temperatures above 80 °C. Thurincin H is also stable from pH 2-7 for at least 24 h at room temperature. Thurincin H is germicidal against B. cereus spores in brain heart infusion broth, but not in Tris-NaCl buffer. The efficient purification method enables the large-scale production of pure thurincin H. The broad inhibitory spectrum of this bacteriocin may be of interest as a potential natural biopreservative in the food industry, particularly in post-processed and ready-to-eat food.

Heterologous expression and purification of plantaricin NC8, a two-peptide bacteriocin against Salmonella spp. from Lactobacillus plantarum ZJ316.

Science.gov (United States)

Jiang, Han; Li, Ping; Gu, Qing

2016-11-01

Bacteriocin, which is produced by lactic acid bacteria (LAB), has the potential to act as natural preservatives in the food industry. To develop strategies to overproduce such peptides, plantaricin NC8, a class IIb LAB bacteriocin that consists of two peptides, PLNC8α and PLNC8β, was successfully heterologously expressed in Escherichia coli BL21 (DE3). PLNC8α and PLNC8β peptides were expressed as His6-tag fusion proteins and were separated by Ni(2+) chelating affinity chromatography. To get the PLNC8α and PLNC8β peptides without extra amino acids in the N-terminus, the fusion proteins were cleaved by enterokinase and further purified using the Ni-NTA Sefinose™ Resin Kit. The molecular masses of peptides were checked using Tricine-SDS-PAGE and MALDI-TOF-MS. The yield of purified PLNC8α was around 2-2.5 mg/L, and the yield of PLNC8β was around 1.5-2 mg/L. The antimicrobial spectrum of cleaved peptides was detected and the synergistic action of PLNC8α and PLNC8β was preliminarily confirmed. It was found that E. coli was a suitable host for heterologous expression of plantaricin NC8 with a significant yield. Importantly, the bacteriocin appeared to be very active for controlling and inhibiting the food-borne pathogenic Gram-negative bacteria Salmonella spp., and might be useful as a natural preservative candidate. Copyright © 2016 Elsevier Inc. All rights reserved.

Licheniocin 50.2 and Bacteriocins from Lactococcus lactis subsp. lactis biovar. diacetylactis BGBU1-4 Inhibit Biofilms of Coagulase Negative Staphylococci and Listeria monocytogenes Clinical Isolates.

Science.gov (United States)

Cirkovic, Ivana; Bozic, Dragana D; Draganic, Veselin; Lozo, Jelena; Beric, Tanja; Kojic, Milan; Arsic, Biljana; Garalejic, Eliana; Djukic, Slobodanka; Stankovic, Slavisa

2016-01-01

Coagulase negative staphylococci (CoNS) and Listeria monocytogenes have important roles in pathogenesis of various genital tract infections and fatal foetomaternal infections, respectively. The aim of our study was to investigate the inhibitory effects of two novel bacteriocins on biofilms of CoNS and L. monocytogenes genital isolates. The effects of licheniocin 50.2 from Bacillus licheniformis VPS50.2 and crude extract of bacteriocins produced by Lactococcus lactis subsp. lactis biovar. diacetylactis BGBU1-4 (BGBU1-4 crude extract) were evaluated on biofilm formation and formed biofilms of eight CoNS (four S. epidermidis, two S. hominis, one S. lugdunensis and one S. haemolyticus) and 12 L. monocytogenes genital isolates. Licheniocin 50.2 and BGBU1-4 crude extract inhibited the growth of both CoNS and L. monocytogenes isolates, with MIC values in the range between 200-400 AU/ml for licheniocin 50.2 and 400-3200 AU/ml for BGBU1-4 crude extract. Subinhibitory concentrations (1/2 × and 1/4 × MIC) of licheniocin 50.2 inhibited biofilm formation by all CoNS isolates (p < 0.05, respectively), while BGBU1-4 crude extract inhibited biofilm formation by all L. monocytogenes isolates (p < 0.01 and p < 0.05, respectively). Both bacteriocins in concentrations of 100 AU/mL and 200 AU/mL reduced the amount of 24 h old CoNS and L. monocytogenes biofilms (p < 0.05, p < 0.01, p < 0.001). This study suggests that novel bacteriocins have potential to be used for genital application, to prevent biofilm formation and/or to eradicate formed biofilms, and consequently reduce genital and neonatal infections by CoNS and L. monocytogenes.

Antibacterial and Synergistic Activity Against β-Lactamase-Producing Nosocomial Bacteria by Bacteriocin of LAB Isolated From Lesser Known Traditionally Fermented Products of India

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Koel Biswas

2017-04-01

Full Text Available There is an ever-growing need to control antibiotic-resistance owing to alarming resistance to commonly available antimicrobial agents for which contemporary and alternative approaches are being explored. The present study assessed the antibacterial activity of bacteriocins from lactic acid bacteria (LAB from lesser known traditionally fermented products of India for their synergistic potential with common antibiotics against clinical β-lactamases producing pathogens. A total of 84 isolates of LAB were screened for their antibacterial efficacy against Streptococcus pyogenes, Enterococcus faecalis, Escherichia coli, Klebsiella pneumoniae and Bacillus cereus as well as against clinical pathogens harbouring β-lactamase genes such as blaCTX-M, blaVIM, blaIMP, blaSHV and blaNDM. Synergistic activity of bacteriocins were determined in combination with antibiotics namely, cefotaxime, polymyxin B, imipenem and tigecycline. Purified bacteriocins from Lactobacillus, Pediococcus and Enterococcus inhibited the growth of β-lactamase harbouring clinical pathogens which significantly higher inhibitions when compared with antibiotics alone. Minimum inhibitory concentration of the extracts ranged from 6.66 to 26.66 mg/ml and 10 to 33.33 mg/ml for Pediococcus pentosaceus LU11 and Lactobacillus plantarum LS6. The bacteriocinogenic activity of LAB opens scope for bioprospection of antibacterial components in the current struggle against increasing pandrug resistance and slowing down the expansion of multi-drug resistance.

A Sequential Statistical Approach towards an Optimized Production of a Broad Spectrum Bacteriocin Substance from a Soil Bacterium Bacillus sp. YAS 1 Strain

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Amira M. Embaby

2014-01-01

Full Text Available Bacteriocins, ribosomally synthesized antimicrobial peptides, display potential applications in agriculture, medicine, and industry. The present study highlights integral statistical optimization and partial characterization of a bacteriocin substance from a soil bacterium taxonomically affiliated as Bacillus sp. YAS 1 after biochemical and molecular identifications. A sequential statistical approach (Plackett-Burman and Box-Behnken was employed to optimize bacteriocin (BAC YAS 1 production. Using optimal levels of three key determinants (yeast extract (0.48% (w/v, incubation time (62 hrs, and agitation speed (207 rpm in peptone yeast beef based production medium resulted in 1.6-fold enhancement in BAC YAS 1 level (470 AU/mL arbitrary units against Erwinia amylovora. BAC YAS 1 showed activity over a wide range of pH (1–13 and temperature (45–80°C. A wide spectrum antimicrobial activity of BAC YAS 1 against the human pathogens (Clostridium perfringens, Staphylococcus epidermidis, Campylobacter jejuni, Enterobacter aerogenes, Enterococcus sp., Proteus sp., Klebsiella sp., and Salmonella typhimurium, the plant pathogen (E. amylovora, and the food spoiler (Listeria innocua was demonstrated. On top and above, BAC YAS 1 showed no antimicrobial activity towards lactic acid bacteria (Lactobacillus bulgaricus, L. casei, L. lactis, and L. reuteri. Promising characteristics of BAC YAS 1 prompt its commercialization for efficient utilization in several industries.

Structure and properties of the metastable bacteriocin Lcn972 from Lactococcus lactis

Science.gov (United States)

Turner, David L.; Lamosa, Pedro; Rodríguez, Ana; Martínez, Beatriz

2013-01-01

Lactococcus lactis subsp. lactis IPLA 972 produces a polypeptide bacteriocin of 7.5 kDa which has a bactericidal effect on sensitive lactococci, inhibiting septum formation in dividing cells. The active form is a monomer that is metastable under normal conditions but is stabilised by glycerol. The NMR structure of Lcn972 shows a β-sandwich comprising two three-stranded antiparallel β-sheets. Detaching the final strand could allow the sandwich to open, and the irreversible unfolding leads to a loss of antibacterial activity. Covalent linkage of the final strand should increase the stability of Lcn972 and facilitate the study of its interaction with lipid II.

Effect of bacteriocin and exopolysaccharides isolated from probiotic on P. aeruginosa PAO1 biofilm.

Science.gov (United States)

Sharma, Vivek; Harjai, Kusum; Shukla, Geeta

2018-03-01

Microorganisms develop biofilms on indwelling medical devices and are associated with biofilm-related infections, resulting in substantial morbidity and mortality. Therefore, to prevent and control biofilm-associated infections, the present study was designed to assess the anti-biofilm potential of postbiotics derived from probiotic organisms against most prevalent biofilm-forming Pseudomonas aeruginosa PAO1. Eighty lactic acid bacteria isolated from eight neonatal fecal samples possessed antibacterial activity against P. aeruginosa PAO1. Among these, only four lactic acid bacteria produced both bacteriocin and exopolysaccharides but only one isolate was found to maximally attenuate the P. aeruginosa PAO1 biofilm. More specifically, the phenotypic and probiotic characterization showed that the isolated lactic acid bacteria were gram positive, non-motile, and catalase and oxidase negative; tolerated acidic and alkaline pH; has bile salt concentration; showed 53% hydrophobicity; and was found to be non-hemolytic. Phylogenetically, the organism was found to be probiotic Lactobacillus fermentum with accession no. KT998657. Interestingly, pre-coating of a microtiter plate either with bacteriocin or with exopolysaccharides as well as their combination significantly (p < 0.05) reduced the number of viable cells forming biofilms to 41.7% compared with simultaneous coating of postbiotics that had 72.4% biofilm-forming viable cells as observed by flow cytometry and confocal laser scanning microscopy. Therefore, it can be anticipated that postbiotics as the natural biointerventions can be employed as the prophylactic agents for medical devices used to treat gastrointestinal and urinary tract infections.

Effects of the presence of the curing agent sodium nitrite, used in the production of fermented sausages, on bacteriocin production by Weissella paramesenteroides DX grown in meat simulation medium.

Science.gov (United States)

Papagianni, M; Sergelidis, D

2013-06-10

Weissellin A is a listericidal bacteriocin produced by the sausage-isolated strain of Weissella paramesenteroides DX. The response of the strain to various concentrations of the added curing agent NaNO2 (0.0025, 0.005 and 0.01g/L) was evaluated in bioreactor fermentations using a meat simulation medium. The presence of nitrite suppressed bacteriocin production - the effect being more pronounced with increasing concentrations. Weissellin A was produced as a growth-associated metabolite in the absence of nitrite or its presence in the low concentration of 0.005g/L under aerobic conditions. The suppressive effect of nitrite was apparent under conditions supporting increased specific production rates, e.g. 50% and 100% dissolved oxygen tension, but no effect was observed under anaerobic conditions. As the latter prevail in the microenvironment of fermented meat products, the absence of any influence of nitrite on bacteriocin production is an important finding that enlightens the role of this species of lactic acid bacteria in its common substrates. Copyright © 2013 Elsevier Inc. All rights reserved.

Complete genome sequence of bacteriocin-producing Lactobacillus plantarum KLDS1.0391, a probiotic strain with gastrointestinal tract resistance and adhesion to the intestinal epithelial cells.

Science.gov (United States)

Jia, Fang-Fang; Zhang, Lu-Ji; Pang, Xue-Hui; Gu, Xin-Xi; Abdelazez, Amro; Liang, Yu; Sun, Si-Rui; Meng, Xiang-Chen

2017-10-01

Lactobacillus plantarum KLDS1.0391 is a probiotic strain isolated from the traditional fermented dairy products and identified to produce bacteriocin against Gram-positive and Gram-negative bacteria. Previous studies showed that the strain has a high resistance to gastrointestinal stress and has a high adhesion ability to the intestinal epithelial cells (Caco-2). We reported the entire genome sequence of this strain, which contains a circular 2,886,607-bp chromosome and three circular plasmids. Genes, which are related to the biosynthesis of bacteriocins, the stress resistance to gastrointestinal tract environment and adhesive performance, were identified. Whole genome sequence of Lactobacillus plantarum KLDS1.0391 will be helpful for its applications in food industry. Copyright © 2017 Elsevier Inc. All rights reserved.

Kinetic studies of the action of Lactacin F, a bacteriocin produced by Lactobacillus johnsonii that forms poration complexes in the cytoplasmic membrane.

NARCIS (Netherlands)

Abee, T.; Klaenhammer, T.R.; Letellier, L.

1994-01-01

The bacteriocin lactacin F is bactericidal against Lactobacillus delbrueckii, Lactobacillus helveticus, and Enterococcus faecalis. Activity against L. delbrueckii was recently shown to be dependent on two peptides, LafA and LafX, which are encoded within the lactacin F operon (T. R. Klaenhammer,

Bacteriocin- producing strain of Enterococcus faecium EK 13 with probiotic character and its application in the digestive tract of rabbits

Czech Academy of Sciences Publication Activity Database

Lauková, A.; Strompfová, V.; Skřivanová, V.; Volek, Z.; Jindřichová, E.; Marounek, Milan

2006-01-01

Roč. 61, č. 6 (2006), s. 779-782 ISSN 0006-3088. [ Probiotic Conference /2./. Košice, 15.09.2006-19.09.2006] Grant - others:VEGA 2/5139/25 Institutional research plan: CEZ:AV0Z50450515 Keywords : probiotic * bacteriocin * enterocin Subject RIV: GH - Livestock Nutrition Impact factor: 0.213, year: 2006

ABC transporter content diversity in Streptococcus pneumoniae impacts competence regulation and bacteriocin production.

Science.gov (United States)

Wang, Charles Y; Patel, Nisha; Wholey, Wei-Yun; Dawid, Suzanne

2018-06-19

The opportunistic pathogen Streptococcus pneumoniae (pneumococcus) uses natural genetic competence to increase its adaptability through horizontal gene transfer. One method of acquiring DNA is through predation of neighboring strains with antimicrobial peptides called "bacteriocins." Competence and production of the major family of pneumococcal bacteriocins, pneumocins, are regulated by the quorum-sensing systems com and blp , respectively. In the classical paradigm, the ABC transporters ComAB and BlpAB each secretes its own system's signaling pheromone and in the case of BlpAB also secretes the pneumocins. While ComAB is found in all pneumococci, only 25% of strains encode an intact version of BlpAB [BlpAB(+)] while the rest do not [BlpAB(-)]. Contrary to the classical paradigm, it was previously shown that BlpAB(-) strains can activate blp through ComAB-mediated secretion of the blp pheromone during brief periods of competence. To better understand the full extent of com - blp crosstalk, we examined the contribution of each transporter to competence development and pneumocin secretion. We found that BlpAB(+) strains have a greater capacity for competence activation through BlpAB-mediated secretion of the com pheromone. Similarly, we show that ComAB and BlpAB are promiscuous and both can secrete pneumocins. Consequently, differences in pneumocin secretion between BlpAB(+) and BlpAB(-) strains derive from the regulation and kinetics of transporter expression rather than substrate specificity. We speculate that BlpAB(-) strains (opportunists) use pneumocins mainly in a narrowly tailored role for DNA acquisition and defense during competence while BlpAB(+) strains (aggressors) expand their use for the general inhibition of rival strains. Copyright © 2018 the Author(s). Published by PNAS.

Medical and Personal Care Applications of Bacteriocins Produced by Lactic Acid Bacteria

Science.gov (United States)

Dicks, L. M. T.; Heunis, T. D. J.; van Staden, D. A.; Brand, A.; Noll, K. Sutyak; Chikindas, M. L.

The frequent use of antibiotics has led to a crisis in the antibiotic ­resistance of pathogens associated with humans and animals. Antibiotic resistance and the emergence of multiresistant bacterial pathogens have led to the investigation of alternative antimicrobial agents to treat and prevent infections in both humans and animals. Research on antimicrobial peptides, with a special interest on bacteriocins of lactic acid bacteria, is entering a new era with novel applications other than food preservation. Many scientists are now focusing on the application of these peptides in medicinal and personal care products. However, it is difficult to assess the success of such ventures due to the dearth of information that has been published and the lack of clinical trials.

Garvicin A, a Novel Class IId Bacteriocin from Lactococcus garvieae That Inhibits Septum Formation in L. garvieae Strains

Science.gov (United States)

Cárdenas, Nivia; Martínez, Beatriz; Ruiz-Barba, José Luis; Fernández-Garayzábal, José F.; Rodríguez, Juan M.; Gibello, Alicia

2013-01-01

Lactococcus garvieae 21881, isolated in a human clinical case, produces a novel class IId bacteriocin, garvicin A (GarA), which is specifically active against other L. garvieae strains, including fish- and bovine-pathogenic isolates. Purification from active supernatants, sequence analyses, and plasmid-curing experiments identified pGL5, one of the five plasmids found in L. garvieae [M. Aguado-Urda et al., PLoS One 7(6):e40119, 2012], as the coding plasmid for the structural gene of GarA (lgnA), its putative immunity protein (lgnI), and the ABC transporter and its accessory protein (lgnC and lgnD). Interestingly, pGL5-cured strains were still resistant to GarA. Other putative bacteriocins encoded by the remaining plasmids were not detected during purification, pointing to GarA as the main inhibitor secreted by L. garvieae 21881. Mode-of-action studies revealed a potent bactericidal activity of GarA. Moreover, transmission microscopy showed that GarA seems to act by inhibiting septum formation in L. garvieae cells. This potent and species-specific inhibition by GarA holds promise for applications in the prevention or treatment of infections caused by pathogenic strains of L. garvieae in both veterinary and clinical settings. PMID:23666326

Effect of supplementation of yeast with bacteriocin and culture on growth performance, cecal fermentation, microbiota composition, and blood characteristics in broiler chickens

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C. Y. Chen

2017-02-01

Full Text Available Objective The aim of the present study was to investigate the effect of yeast with bacteriocin and Lactobacillus cultures (mixture of Lactobacillus agilis BCRC 10436 and Lactobacillus reuteri BCRC 17476 supplements, alone or in combination, on broiler chicken performance. Methods A total of 300, 1-d-old healthy broiler chickens were randomly divided into five treatment groups: i basal diet (control, ii basal diet+0.25% yeast (Saccharomyces cerevisiae (YC, iii basal diet+0.25% yeast with bacteriocin (BA, iv basal diet+Lactobacillus cultures (LAB, and v basal diet +0.25% yeast with bacteriocin+Lactobacillus cultures (BA+LAB. Growth performance, cecal microbiota, cecal fermentation products, and blood biochemistry parameters were determined when chickens were 21 and 35 d old. Results The supplementation of YC, BA, and BA+LAB resulted in a significantly better feed conversion rate (FCR than that of the control group during 1 to 21 d (p<0.05. The LAB supplementation had a significant effect on the presence of Lactobacillus in the ceca at 35 d. None of the supplements had an effect on relative numbers of L. agilis and L. reuter at 21 d, but the BA supplementation resulted in the decrease of both Lactobacillus strains at 35 d. The BA+LAB supplementation resulted in higher short chain fatty acid (SCFA in the ceca, but LAB supplementation significantly decreased the SCFA at 35 d (p<0.05. All treatments tended to decrease ammonia concentration in the ceca at 21 d, especially in the LAB treatment group. The BA supplementation alone decreased the triacylglycerol (TG concentration significantly at 21 d (p<0.05, but the synergistic effect of BA and LAB supplementation was required to reduce the TG concentration at 35 d. The YC supplementation tended to increase the plasma cholesterol at 21 d and 35 d. However, the BA supplementation significantly decreased the cholesterol and low density lipoprotein cholesterol level at 35 d. In conclusion, the BA

Bacteriocin-like substances of Lactobacillus curvatus P99: characterization and application in biodegradable films for control of Listeria monocytogenes in cheese.

Science.gov (United States)

Marques, Juliana de Lima; Funck, Graciele Daiana; Dannenberg, Guilherme da Silva; Cruxen, Claudio Eduardo Dos Santos; Halal, Shanise Lisie Mello El; Dias, Alvaro Renato Guerra; Fiorentini, Ângela Maria; Silva, Wladimir Padilha da

2017-05-01

The aim of this study was to evaluate the effectiveness of a biodegradable film, with antimicrobial metabolites produced by Lactobacillus curvatus P99 incorporated, targeting the control of Listeria monocytogenes in sliced "Prato" cheese. Tests were performed to evaluate the spectrum of action of cell-free supernatant (CFS) of P99 against different microorganisms, as well as to detect the minimum inhibitory (MIC) and bactericidal (MBC) concentrations against L. monocytogenes Scott A. The detection of genes that encode for the production of bacteriocins and evaluation of their expression were performed. Antimicrobial films were prepared, followed by in vitro and in situ analysis. The MIC and MBC of CFS against L. monocytogenes Scott A was 15.6 μL/mL and 62.5 μL/mL, respectively. Lactobacillus curvatus P99 presented two genes coding for the bacteriocins, which were expressed. Films with added MBC showed activity against different indicator microorganisms and were able to control L. monocytogenes Scott A when used in sliced "Prato" cheese. Copyright © 2016 Elsevier Ltd. All rights reserved.

Detection and characterization of bacteriocin-producing Lactococcus lactis strains Detecção e caracterização de Lactococcus lactis produtores de bacteriocinas

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Izildinha Moreno

1999-04-01

Full Text Available One hundred sixty seven strains of Lactococcus lactis were screened for bacteriocin production by well diffusion assay of GM17 agar. Fourteen (8.4% produced antimicrobial activity other than organic acids, bacteriophages or hydrogen peroxide. The frequency of bacteriocin production ranged from 2% in L. lactis subsp. cremoris up to 12% in L. lactis subsp. lactis. Antimicrobial activities were not observed in any strain of L. lactis subsp. lactis var. diacetylactis. Among thirteen bacteriocin-producing strains and two nisin-producing strains (L. lactis subsp. lactis ATCC 11454 and L. lactis subsp. lactis CNRZ 150, eight (53% were characterized as lactose-positive (Lac+ and proteinase-negative (Prt-. The bacteriocin-producing cultures were also characterized on the basis of plasmid content. All strains had 2 to 7 plasmids with molecular weights varying from 0.5 to 28.1 Mdal. Four strains (ITAL 435, ITAL 436, ITAL 437 and ITAL 438 showed identical profiles and the other were quite distinct.Um total de 167 linhagens de L. lactis foi selecionado para os testes de produção de bacteriocinas pelo método de difusão em poços em agar GM17. Desse total, 14 (8.4% produziram substâncias inibidoras que não foram associadas com ácidos orgânicos, peróxido de hidrogênio e bacteriófagos. A frequência de produção de bacteriocinas variou de 2% em L. lactis subsp. cremoris a 12% em L. lactis subsp. lactis. Nenhuma das linhagens de L. lactis subsp. lactis var. diacetylactis produziu substâncias inibidoras. De 13 linhagens produtoras de bacteriocinas e duas de nisina (L. lactis subsp. lactis ATCC 11454 e L. lactis subsp. lactis CNRZ 150, 8 (53% foram caracterizadas como lactose-positivas (Lac+ e proteinase-negativas (Prt-. As linhagens produtoras de bacteriocinas também foram caracterizadas no seu conteúdo de plasmídios. Elas apresentaram de 2 a 7 plasmídios, com pesos moleculares aproximados de 0.5 a 28.1 Mdal. Quatro linhagens (ITAL 435, ITAL 436

Characterization of a bacteriocin-like substance produced from a novel isolated strain of Bacillus subtilis SLYY-3

Science.gov (United States)

Li, Junfeng; Li, Hongfang; Zhang, Yuanyuan; Duan, Xiaohui; Liu, Jie

2014-12-01

In the present research, the strain SLYY-3 was isolated from sediments of Jiaozhou Bay, Qingdao, China. The strain SLYY-3, which produced a bacteriocin-like substance (BLS), was characterized to be a strain of Bacillus subtillis by biochemical profiling and 16S rDNA sequence analysis. It is the first time to report that Bacillus subtilis from Jiaozhou Bay sediments could produce a BLS. The BLS of B. subtillis SLYY-3 exhibited strong inhibitory activity against gram-positive bacteria (including Staphylococcus aureus and B. subtillis) and some fungi (including Penicillium glaucum, Aspergillus niger and Aspergillus flavus). The antimicrobial activity was detected from culture in the exponential growth phase and reached its maximum when culture entered into stationary growth phase. It was thermo-tolerant even when being kept at 100°C for 60 min without losing any activity and stable over a wide pH range from 1.0 to 12.0 while being inactivated by proteolytic enzyme and trypsin, indicating the proteinaceous nature of the BLS. The BLS was purified by precipitation with hydrochloric acid (HCl) and gel filteration (Sephadex G-100). SDS-PAGE analysis of the extracellular peptides of SLYY-3 revealed a bacteriocin-like protein with a molecular mass of 66 kDa. Altogether, these characteristics indicate the potential of the BLS for food industry as a protection against pathogenic and spoilage microorganisms.

Bacteriocin-Producing Enterococcus faecium LCW 44: A High Potential Probiotic Candidate from Raw Camel Milk.

Science.gov (United States)

Vimont, Allison; Fernandez, Benoît; Hammami, Riadh; Ababsa, Ahlem; Daba, Hocine; Fliss, Ismaïl

2017-01-01

Bacterial isolates from raw camel milk were screened for antibacterial activity using the agar diffusion assay. Ten isolates selected for their inhibition of Gram-positive bacteria were identified by 16S sequencing as Enterococcus faecium or durans . An isolate named E. faecium LCW 44 exhibited the broadest antibacterial spectrum with an inhibitory activity against several Gram-positive strains belonging to the genera Clostridium , Listeria , Staphylococcus , and Lactobacillus. E. faecium LCW 44 was shown to produce N-formylated enterocins L50A and L50B, as revealed by mass spectrometry and PCR analyses. This isolate did not harbor any of the virulence factors tested and was shown to be sensitive to all tested antibiotics. It showed high resistance to gastric and intestinal conditions (78 ± 4% survival). Its adhesion index was evaluated at 176 ± 86 and 24 ± 86 on Caco-2 cells and HT-29 cells, respectively, and it significantly reduced adhesion of Listeria monocytogenes by 65 and 49%, respectively. In Macfarlane broth (simulating the nutrient content of the colon), counts of L. monocytogenes were reduced by 2 log 10 cycles after 24 h in co-culture with E. faecium LCW 44, compared to the increase of 4 log 10 cycles when cultured alone. Comparison with a bacteriocin-non-producing mutant of E. faecium LCW 44 strongly suggests that inhibition of L. monocytogenes was due to bacteriocin production. Altogether, E. faecium LCW 44 thus has potential for use as a probiotic for humans and veterinary medicine.

Bacteriocin-Producing Enterococcus faecium LCW 44: A High Potential Probiotic Candidate from Raw Camel Milk

Directory of Open Access Journals (Sweden)

Allison Vimont

2017-05-01

Full Text Available Bacterial isolates from raw camel milk were screened for antibacterial activity using the agar diffusion assay. Ten isolates selected for their inhibition of Gram-positive bacteria were identified by 16S sequencing as Enterococcus faecium or durans. An isolate named E. faecium LCW 44 exhibited the broadest antibacterial spectrum with an inhibitory activity against several Gram-positive strains belonging to the genera Clostridium, Listeria, Staphylococcus, and Lactobacillus. E. faecium LCW 44 was shown to produce N-formylated enterocins L50A and L50B, as revealed by mass spectrometry and PCR analyses. This isolate did not harbor any of the virulence factors tested and was shown to be sensitive to all tested antibiotics. It showed high resistance to gastric and intestinal conditions (78 ± 4% survival. Its adhesion index was evaluated at 176 ± 86 and 24 ± 86 on Caco-2 cells and HT-29 cells, respectively, and it significantly reduced adhesion of Listeria monocytogenes by 65 and 49%, respectively. In Macfarlane broth (simulating the nutrient content of the colon, counts of L. monocytogenes were reduced by 2 log10 cycles after 24 h in co-culture with E. faecium LCW 44, compared to the increase of 4 log10 cycles when cultured alone. Comparison with a bacteriocin-non-producing mutant of E. faecium LCW 44 strongly suggests that inhibition of L. monocytogenes was due to bacteriocin production. Altogether, E. faecium LCW 44 thus has potential for use as a probiotic for humans and veterinary medicine.

Different bacteriocin activities of Streptococcus mutans reflect distinct phylogenetic lineages

DEFF Research Database (Denmark)

Balakrishnan, M; Simmonds, RS; Kilian, Mogens

2002-01-01

Bacteriocins produced by mutans streptococci are known as mutacins. In this study 16 broadly active mutacin-producing Streptococcus mutans strains from New Zealand, North America and Europe were classified into four groups (A-D) on the basis of differences in their activity in deferred antagonism...... described by Caufield and co-workers. One of the New Zealand isolates of group A (S. mutans strain N) appeared to produce inhibitory activity similar to that of the group I prototype strain UA140. Four other New Zealand isolates of group B (S. mutans strains M19, M34, B34 and D14) had mutacin II......-like activity. The group B mutacin producers differed from the group A mutacin producers in their additional activity against Staph. aureus 46. Seven S. mutans strains (M46, B46, B57, M12, M28, B28 and 13M) were distinguished from the group A and group B mutacin producers in that they inhibited E. faecium TE1...

Enterocin C, a class IIb bacteriocin produced by E. faecalis C901, a strain isolated from human colostrum.

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Maldonado-Barragán, Antonio; Caballero-Guerrero, Belén; Jiménez, Esther; Jiménez-Díaz, Rufino; Ruiz-Barba, José L; Rodríguez, Juan M

2009-07-31

Enterocin C (EntC), a class IIb bacteriocin was purified from culture supernatants of Enterococcus faecalis C901, a strain isolated from human colostrum. Enterocin C consists of two distinct peptides, named EntC1 and EntC2, whose complementary action is required for full antimicrobial activity. The structural genes entC1 and entC2 encoding enterocins EntC1 and EntC2, respectively, and that encoding the putative immunity protein (EntCI) are located in the 9-kb plasmid pEntC, harboured by E. faecalis C901. The N-terminal sequence of both antimicrobial peptides revealed that EntC1 (4284 Da) is identical to Ent1071A, one of the two peptides that form enterocin 1071 (Ent1071), a bacteriocin produced by E. faecalis BFE 1071. In contrast, EntC2 (3867 Da) presents the non-polar alanine residue at position 17 (Ala(17)) instead of the polar threonine residue (Thr(17)) in Ent1071B, the second peptide constituting Ent1071. In spite of peptide similarities, EntC differs from Ent1071 in major aspects, including the complementary activity among its constitutive peptides and its wider inhibitory spectrum of activity. Different amphiphilic alpha-helical conformations between EntC2 and Ent1071B could explain both, acquired complementary activity and increased antimicrobial spectrum.

An isolate of Haemophilus haemolyticus produces a bacteriocin-like substance that inhibits the growth of nontypeable Haemophilus influenzae.

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Latham, Roger D; Gell, David A; Fairbairn, Rory L; Lyons, A Bruce; Shukla, Shakti D; Cho, Kum Yin; Jones, David A; Harkness, Nick M; Tristram, Stephen G

2017-04-01

Nontypeable Haemophilus influenzae (NTHi) frequently colonises the upper respiratory tract and is an important cause of respiratory infections. Resistance to antibiotics is an emerging trend in NTHi and alternative prevention or treatment strategies are required. Haemophilus haemolyticus is a common commensal occupying the same niche as NTHi and, if able to produce substances that inhibit NTHi growth, may have a role as a probiotic. In this study, ammonium sulphate extracts from broth culture of 100 H. haemolyticus isolates were tested for the presence of substances inhibitory to NTHi using a well diffusion assay. One isolate produced a substance that consistently inhibited the growth of NTHi. The substance was inactivated by protease enzymes and had a molecular size of ca. 30 kDa as determined by size exclusion chromatography. When the substance was tested against bacteria from eight Gram-negative and three Gram-positive genera, only Haemophilus spp. were inhibited. Quantitative PCR testing showed the substance to be different to 'haemocin', the previously described bacteriocin of H. influenzae type b. These molecular characteristics, together with narrow-spectrum activity, suggest the substance may be a novel bacteriocin, and there is potential for this H. haemolyticus isolate to function as a probiotic for reduction of colonisation and subsequent infection with NTHi. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

Purification of a Novel Bacteriocin-Like Inhibitory Substance Produced by Enterococcus faecium ICIS 8 and Characterization of Its Mode of Action.

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Vasilchenko, Alexey S; Rogozhin, Eugene A; Valyshev, Alexander V

2017-06-01

The aim of this work was to purify and characterize a bacteriocin-like antimicrobial substance produced by an antagonistic active strain of Enterococcus faecium. A novel bacteriocin-like inhibitory substance (BLIS) produced by the E. faecium ICIS 8 strain was purified and characterized using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and N-terminal amino acid sequencing revealed the following partial sequence: NH 2 -APKEKCFPKYCV. The proteinaceous nature of purified BLIS was assessed by treatment with proteolytic enzyme. Studies of the action of BLIS using bacteriological and bioluminescence assays revealed a dose-dependent inhibition of Listeria monocytogenes 88BK and Escherichia coli K12 TG1 lac::lux viability. The interaction of the BLIS with the bacterial surface led to the compensation of a negative charge value, as shown by zeta-potential measurements. Assessments of membrane integrity using fluorescent probes and atomic force microscopy revealed the permeabilization of the cellular barrier structures in both L. monocytogenes and E. coli. The novel BLIS from E. faecium ICIS 8 was characterized by a unique primary peptide sequence and exerted bactericidal activity against L. monocytogenes and E. coli by disrupting membrane integrity.

Update on the development of a novel dry cow therapy using a bismuth-based intramammary teat seal in combination with the bacteriocin lacticin 3147

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Crispie Fiona

2004-11-01

Full Text Available Public concerns over the widespread prophylactic use of antibiotics have led to a search for alternatives to dry cow therapy for the prevention of intramammary infections. A popular alternative is to infuse a teat seal at drying-off. The teat seal is a viscous non-antibiotic formulation and when it is infused into the teat canal and the teat sinus it forms an internal seal that provides a physical barrier to invasion by mastitis-causing pathogens. Enhancement of teat seal formulations may be achieved using non-antibiotic additives such as bacteriocins, potent proteins produced by some bacteria that have the ability to kill other microorganisms. This paper traces the history of investigations at Moorepark Research Centre into the efficacy of teat seal plus lacticin 3147, a bacteriocin produced by Lactococcus lactis DPC3147, in the prevention of intramammary infections in dry cows. Indications from on-going investigations are that a dry cow formulation combining the two products has considerable potential as a non-antibiotic prophylactic product.

Enterocin F4-9, a Novel O-Linked Glycosylated Bacteriocin.

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Maky, Mohamed Abdelfattah; Ishibashi, Naoki; Zendo, Takeshi; Perez, Rodney Honrada; Doud, Jehan Ragab; Karmi, Mohamed; Sonomoto, Kenji

2015-07-01

Enterococcus faecalis F4-9 isolated from Egyptian salted-fermented fish produces a novel bacteriocin, termed enterocin F4-9. Enterocin F4-9 was purified from the culture supernatant by three steps, and its molecular mass was determined to be 5,516.6 Da by mass spectrometry. Amino acid and DNA sequencing showed that the propeptide consists of 67 amino acid residues, with a leader peptide containing a double glycine cleavage site to produce a 47-amino-acid mature peptide. Enterocin F4-9 is modified by two molecules of N-acetylglucosamine β-O-linked to Ser37 and Thr46. The O-linked N-acetylglucosamine moieties are essential for the antimicrobial activity of enterocin F4-9. Further analysis of the enterocin F4-9 gene cluster identified enfC, which has high sequence similarity to a glycosyltransferase. The antimicrobial activity of enterocin F4-9 covered a limited range of bacteria, including, interestingly, a Gram-negative strain, Escherichia coli JM109. Enterocin F4-9 is sensitive to protease, active at a wide pH range, and moderately resistant to heat. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Influence of adhesion and bacteriocin production by Lactobacillus salivarius on the intestinal epithelial cell transcriptional response.

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O'Callaghan, John; Buttó, Ludovica F; MacSharry, John; Nally, Kenneth; O'Toole, Paul W

2012-08-01

Lactobacillus salivarius strain UCC118 is a human intestinal isolate that has been extensively studied for its potential probiotic effects in human and animal models. The objective of this study was to determine the effect of L. salivarius UCC118 on gene expression responses in the Caco-2 cell line to improve understanding of how the strain might modulate intestinal epithelial cell phenotypes. Exposure of Caco-2 cells to UCC118 led to the induction of several human genes (TNFAIP3, NFKBIA, and BIRC3) that are negative regulators of inflammatory signaling pathways. Induction of chemokines (CCL20, CXCL-1, and CXCL-2) with antimicrobial functions was also observed. Disruption of the UCC118 sortase gene srtA causes reduced bacterial adhesion to epithelial cells. Transcription of three mucin genes was reduced significantly when Caco-2 cells were stimulated with the ΔsrtA derivative of UCC118 compared to cells stimulated with the wild type, but there was no significant change in the transcription levels of the anti-inflammatory genes. UCC118 genes that were significantly upregulated upon exposure to Caco-2 cells were identified by bacterial genome microarray and consisted primarily of two groups of genes connected with purine metabolism and the operon for synthesis of the Abp118 bacteriocin. Following incubation with Caco-2 cells, the bacteriocin synthesis genes were transcribed at higher levels in the wild type than in the ΔsrtA derivative. These data indicate that L. salivarius UCC118 influences epithelial cells both through modulation of the inflammatory response and by modulation of intestinal cell mucin production. Sortase-anchored cell surface proteins of L. salivarius UCC118 have a central role in promoting the interaction between the bacterium and epithelial cells.

Molecular Detection and Sensitivity to Antibiotics and Bacteriocins of Pathogens Isolated from Bovine Mastitis in Family Dairy Herds of Central Mexico

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Ma. Fabiola León-Galván

2015-01-01

Full Text Available Thirty-two farms (n=535 cows located in the state of Guanajuato, Mexico, were sampled. Pathogens from bovine subclinical mastitis (SCM and clinical mastitis (CLM were identified by 16S rDNA and the sensitivity to both antibiotics and bacteriocins of Bacillus thuringiensis was tested. Forty-six milk samples were selected for their positive California Mastitis Test (CMT (≥3 and any abnormality in the udder or milk. The frequency of SCM and CLM was 39.1% and 9.3%, respectively. Averages for test day milk yield (MY, lactation number (LN, herd size (HS, and number of days in milk (DM were 20.6 kg, 2.8 lactations, 16.7 animals, and 164.1 days, respectively. MY was dependent on dairy herd (DH, LN, HS, and DM P<0.01, and correlations between udder quarters from the CMT were around 0.49 P<0.01. Coagulase-negative staphylococci were mainly identified, as well as Staphylococcus aureus, Streptococcus uberis, Brevibacterium stationis, B. conglomeratum, and Staphylococcus agnetis. Bacterial isolates were resistant to penicillin, clindamycin, ampicillin, and cefotaxime. Bacteriocins synthesized by Bacillus thuringiensis inhibited the growth of multiantibiotic resistance bacteria such as S. agnetis, S. equorum, Streptococcus uberis, Brevibacterium stationis, and Brachybacterium conglomeratum, but they were not active against S. sciuri, a microorganism that showed an 84% resistance to antibiotics tested in this study.

Production of bioactive substances by intestinal bacteria as a basis for explaining probiotic mechanisms: bacteriocins and conjugated linoleic acid.

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O'Shea, Eileen F; Cotter, Paul D; Stanton, Catherine; Ross, R Paul; Hill, Colin

2012-01-16

The mechanisms by which intestinal bacteria achieve their associated health benefits can be complex and multifaceted. In this respect, the diverse microbial composition of the human gastrointestinal tract (GIT) provides an almost unlimited potential source of bioactive substances (pharmabiotics) which can directly or indirectly affect human health. Bacteriocins and fatty acids are just two examples of pharmabiotic substances which may contribute to probiotic functionality within the mammalian GIT. Bacteriocin production is believed to confer producing strains with a competitive advantage within complex microbial environments as a consequence of their associated antimicrobial activity. This has the potential to enable the establishment and prevalence of producing strains as well as directly inhibiting pathogens within the GIT. Consequently, these antimicrobial peptides and the associated intestinal producing strains may be exploited to beneficially influence microbial populations. Intestinal bacteria are also known to produce a diverse array of health-promoting fatty acids. Indeed, certain strains of intestinal bifidobacteria have been shown to produce conjugated linoleic acid (CLA), a fatty acid which has been associated with a variety of systemic health-promoting effects. Recently, the ability to modulate the fatty acid composition of the liver and adipose tissue of the host upon oral administration of CLA-producing bifidobacteria and lactobacilli was demonstrated in a murine model. Importantly, this implies a potential therapeutic role for probiotics in the treatment of certain metabolic and immunoinflammatory disorders. Such examples serve to highlight the potential contribution of pharmabiotic production to probiotic functionality in relation to human health maintenance. Copyright © 2011 Elsevier B.V. All rights reserved.

Purification and characterization of bacteriocin like substance produced from bacillus lentus with perspective of a new biopreservative for food preservation

International Nuclear Information System (INIS)

Sharma, N.; Gautam, N.

2009-01-01

Molecular weight of bacteriocin like substance (BLIS) of a new strain of Bacillus lentus 121 was found to be approximately 11 kDa. Purification of BLIS was attained by single step gel exclusion chromatography. BLIS was characterized by studying the inhibitory spectrum. It was active at broad pH range, high temperature and high NaCl concentration and showed sensitivity to proteolytic enzymes like trypsin, alpha-chymotrypsin and papain, the characters desirable for food preservation. BLIS extended the shelf stability of milk upto 21 days as a biopreservative. (author)

Lactococcin G is a potassium ion-conducting, two-component bacteriocin.

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Moll, G; Ubbink-Kok, T; Hildeng-Hauge, H; Nissen-Meyer, J; Nes, I F; Konings, W N; Driessen, A J

1996-02-01

Lactococcin G is a novel lactococcal bacteriocin whose activity depends on the complementary action of two peptides, termed alpha and beta. Peptide synthesis of the alpha and beta peptides yielded biologically active lactococcin G, which was used in mode-of-action studies on sensitive cells of Lactococcus lactis. Approximately equivalent amounts of both peptides were required for optimal bactericidal effect. No effect was observed with either the alpha or beta peptide in the absence of the complementary peptide. The combination of alpha and beta peptides (lactococcin G) dissipates the membrane potential (delta omega), and as a consequence cells release alpha-aminoisobutyrate, a non-metabolizable alanine analog that is accumulated through a proton motive-force dependent mechanism. In addition, the cellular ATP level is dramatically reduced, which results in a drastic decrease of the ATP-driven glutamate uptake. Lactococcin G does not form a proton-conducting pore, as it has no effect on the transmembrane pH gradient. Dissipation of the membrane potential by uncouplers causes a slow release of potassium (rubidium) ions. However, rapid release of potassium was observed in the presence of lactococcin G. These data suggest that the bactericidal effect of lactococcin G is due to the formation of potassium-selective channels by the alpha and beta peptides in the target bacterial membrane.

Engineered strains of Streptococcus macedonicus towards an osmotic stress resistant phenotype retain their ability to produce the bacteriocin macedocin under hyperosmotic conditions.

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Anastasiou, Rania; Driessche, Gonzalez Van; Boutou, Effrossyni; Kazou, Maria; Alexandraki, Voula; Vorgias, Constantinos E; Devreese, Bart; Tsakalidou, Effie; Papadimitriou, Konstantinos

2015-10-20

Streptococcus macedonicus ACA-DC 198 produces the bacteriocin macedocin in milk only under low NaCl concentrations (<1.0%w/v). The thermosensitive plasmid pGh9:ISS1 was employed to generate osmotic stress resistant (osmr) mutants of S. macedonicus. Three osmr mutants showing integration of the vector in unique chromosomal sites were identified and the disrupted loci were characterized. Interestingly, the mutants were able to grow and to produce macedocin at considerably higher concentrations of NaCl compared to the wild-type (up to 4.0%w/v). The production of macedocin under hyperosmotic conditions solely by the osmr mutants was validated by the well diffusion assay and by mass spectrometry analysis. RT-PCR experiments demonstrated that the macedocin biosynthetic regulon was transcribed at high salt concentrations only in the mutants. Mutant osmr3, the most robust mutant, was converted in its markerless derivative (osmr3f). Co-culture of S. macedonicus with spores of Clostridium tyrobutyricum in milk demonstrated that only the osmr3f mutant and not the wild-type inhibited the growth of the spores under hyperosmotic conditions (i.e., 2.5%w/v NaCl) due to the production of macedocin. Our study shows how genetic manipulation of a strain towards a stress resistant phenotype could improve bacteriocin production under conditions of the same stress. Copyright © 2015 Elsevier B.V. All rights reserved.

Biochemical and Genetic Characterization of Coagulin, a New Antilisterial Bacteriocin in the Pediocin Family of Bacteriocins, Produced by Bacillus coagulans I4

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Le Marrec, Claire; Hyronimus, Bertrand; Bressollier, Philippe; Verneuil, Bernard; Urdaci, Maria C.

2000-01-01

A plasmid-linked antimicrobial peptide, named coagulin, produced by Bacillus coagulans I4 has recently been reported (B. Hyronimus, C. Le Marrec and M. C. Urdaci, J. Appl. Microbiol. 85:42–50, 1998). In the present study, the complete, unambiguous primary amino acid sequence of the peptide was obtained by a combination of both N-terminal sequencing of purified peptide and the complete sequence deduced from the structural gene harbored by plasmid I4. Data revealed that this peptide of 44 residues has an amino acid sequence similar to that described for pediocins AcH and PA-1, produced by different Pediococcus acidilactici strains and 100% identical. Coagulin and pediocin differed only by a single amino acid at their C terminus. Analysis of the genetic determinants revealed the presence, on the pI4 DNA, of the entire 3.5-kb operon of four genes described for pediocin AcH and PA-1 production. No extended homology was observed between pSMB74 from P. acidilactici and pI4 when analyzing the regions upstream and downstream of the operon. An oppositely oriented gene immediately dowstream of the bacteriocin operon specifies a 474-amino-acid protein which shows homology to Mob-Pre (plasmid recombination enzyme) proteins encoded by several small plasmids extracted from gram-positive bacteria. This is the first report of a pediocin-like peptide appearing naturally in a non-lactic acid bacterium genus. PMID:11097892

Purification of the bacteriocin bavaricin MN and characterization of its mode of action against Listeria monocytogenes Scott A cells and lipid vesicles.

OpenAIRE

Kaiser, A L; Montville, T J

1996-01-01

Bavaricin MN was purified from Lactobacillus sake culture supernatant 135-fold with a final yield of 11%. Sequence analysis revealed bavaricin MN to be a 42-amino-acid peptide having a molecular weight of 4,769 and a calculated pI of 10.0. Computer analysis indicated that the C-terminal region may form an alpha-helical structure with an amphipathic nature deemed important in the interaction of bacteriocins with biological membranes. Bavaricin MN rapidly depleted the membrane potential (delta ...

Purification and characterization of a new bacteriocin active against Campylobacter produced by Lactobacillus salivarius SMXD51.

Science.gov (United States)

Messaoudi, Soumaya; Kergourlay, Gilles; Dalgalarrondo, Michèle; Choiset, Yvan; Ferchichi, Mounir; Prévost, Hervé; Pilet, Marie-France; Chobert, Jean-Marc; Manai, Mohamed; Dousset, Xavier

2012-10-01

Strain SMXD51, isolated from chicken ceca and identified as Lactobacillus salivarius, produced a component that inhibits the growth of Gram-positive and Gram-negative bacteria and especially Campylobacter jejuni. The active peptide from the cell-free supernatant of Lb. salivarius SMXD51 was purified in three steps: (i) precipitation with 80% saturated ammonium sulfate, (ii) elution on a reversed phase SPE UPTI-CLEAN cartridge using different concentrations of acetonitrile, (iii) final purification by reversed phase HPLC on a C(18) column. The mode of action of this peptide of 5383.2 Da was identified as bactericidal, and its amino acid composition was established. This new bacteriocin SMXD51 appears potentially very useful to reduce Campylobacter in poultry prior to processing. Copyright © 2012 Elsevier Ltd. All rights reserved.

[Mode of action of plantaricin L-1, an antilisteria bacteriocin produced by Lactobacillus plantarum].

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Zhou, Wei; Liu, Guo-rong; Li, Ping-lan; Dai, Yun-qing; Zhou, Kang

2007-04-01

Plantaricin L-1, an anti-Listeria bacteriocin, was produced by Lactobacillus plantarum and successfully purified by SP-Sepharose FF cation exchange chromatography. The mechanism on energized cells of Listeria monocytogenes was studied with purified plantaricin L-1. After adding plantaricin L-1 to Listeria monocytogenes at 64 AU/mL, leakage of intercellular K+ ions, inorganic phosphate, lactic dehydrogenase, UV-absorbing materials and the intracellular ATP was observed, and the action resulted in the dissipation of the membrane potential (delta psi) and pH gradient (delta psi), two components of the proton motive force (PMF). All the data suggested that the primary site of action of plantaricin L-1 was the cytoplasmic membrane of sensitive cells. By forming the nonselective pores which leak ions and small organic compounds plantaricin L-1 induced the cells death, this action was similar to membrane corruption caused by peptide effect. Penetrability increased due to the enlarged pore and dysfuction of membrane transporters, which ensured efficient killing of target bacteria.

Isolation of Lactococcus lactis Mutants Simultaneously Resistant to the Cell Wall-Active Bacteriocin Lcn972, Lysozyme, Nisin, and Bacteriophage c2

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Roces, Clara; Courtin, Pascal; Kulakauskas, Saulius; Rodríguez, Ana; Chapot-Chartier, Marie-Pierre

2012-01-01

Lactococcin 972 (Lcn972) is a nonlantibiotic bacteriocin that inhibits cell wall biosynthesis by binding to lipid II. In this work, two mutants resistant to Lcn972, Lactococcus lactis D1 and D1-20, with high (>320 arbitrary units [AU]/ml) and low (80 AU/ml) susceptibilities, respectively, have been isolated. Resistance to Lcn972 did not impose a burden to growth under laboratory conditions, nor did it substantially alter the physicochemical properties of the cell surface. However, the peptidoglycan of the mutants featured a higher content of muropeptides with tripeptide side chains than the wild-type strain, linking for the first time peptidoglycan remodelling to bacteriocin resistance. Moreover, L. lactis lacking a functional d,d-carboxypeptidase DacA (i.e., with a high content of pentapeptide side chain muropeptides) was shown to be more susceptible to Lcn972. Cross-resistance to lysozyme and nisin and enhanced susceptibility to penicillin G and bacitracin was also observed. Intriguingly, the Lcn972-resistant mutants were not infected by the lytic phage c2 and less efficiently infected by phage sk1. Lack of c2 infectivity was linked to a 22.6-kbp chromosomal deletion encompassing the phage receptor protein gene pip. The deletion also included maltose metabolic genes and the two-component system (TCS) F. However, a clear correlation between these genes and resistance to Lcn972 could not be clearly established, pointing to the presence of as-yet-unidentified mutations that account for Lcn972 resistance. PMID:22504807

Characterization and application of enterocin RM6, a bacteriocin from Enterococcus faecalis.

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Huang, En; Zhang, Liwen; Chung, Yoon-Kyung; Zheng, Zuoxing; Yousef, Ahmed E

2013-01-01

Use of bacteriocins in food preservation has received great attention in recent years. The goal of this study is to characterize enterocin RM6 from Enterococcus faecalis OSY-RM6 and investigate its efficacy against Listeria monocytogenes in cottage cheese. Enterocin RM6 was purified from E. faecalis culture supernatant using ion exchange column, multiple C18-silica cartridges, followed by reverse-phase high-performance liquid chromatography. The molecular weight of enterocin RM6 is 7145.0823 as determined by mass spectrometry (MS). Tandem mass spectrometry (MS/MS) analysis revealed that enterocin RM6 is a 70-residue cyclic peptide with a head-to-tail linkage between methionine and tryptophan residues. The peptide sequence of enterocin RM6 was further confirmed by sequencing the structural gene of the peptide. Enterocin RM6 is active against Gram-positive bacteria, including L. monocytogenes, Bacillus cereus, and methicillin-resistant Staphylococcus aureus (MRSA). Enterocin RM6 (final concentration in cottage cheese, 80 AU/mL) caused a 4-log reduction in population of L. monocytogenes inoculated in cottage cheese within 30 min of treatment. Therefore, enterocin RM6 has potential applications as a potent antimicrobial peptide against foodborne pathogens in food.

Caracterização preliminar de bacteriocinas produzidas por seis cepas de bactérias láticas isoladas de produtos cárneos embalados a vácuo Preliminary characterization of bacteriocins produced by six lactic acid bacteria strains isolated from vacuum-packaged meat products

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Elaine C. P. de Martinis

2003-08-01

Full Text Available No presente trabalho, foram estudadas as bacteriocinas produzidas por seis linhagens bacterianas: duas culturas Lactobacillus sake, duas de Lactobacillus curvatus, uma de Leuconostoc mesenteroides, uma de Leuconostoc sp 12. As atividades inibitórias foram quantificadas pelo método da diluição crítica, utilizando-se os indicadores Lactobacillus sake ATCC 15521 e Listeria monocytogenes. As bacteriocinas produzidas foram caracterizadas também quanto à sensibilidade a enzimas, faixa de temperatura na produção, termoestabilidade, estabilidade em diferentes pHs e modo de ação (bactericida ou bacteriostático frente a Listeria monocytogenes. Nenhuma bacteriocina foi destruída pela pepsina, mas todas foram sensíveis à proteinase K, tripsina e alfa-amilase (exceto a bacteriocina produzida por Leuconostoc sp 12, que foi insensível a alfa-amilase. Lactobacillus sake 1, Leuconostoc mesenteroides 11 e Lactobacillus sake 16 apresentaram atividade antilisterial, sendo a maior inibição observada para Lactobacillus sake 1 e Leuconostoc mesenteroides 11 (12.800UA/mL. Lactobacillus sake 1 e Lactobacillus curvatus 5 produziram as bacteriocinas mais termoestáveis. Lactobacillus sake 1 produziu a bacteriocina com maior estabilidade a variações de pH. Todas as bactérias láticas produziram bacteriocina entre 4ºC e 30ºC, sendo esta propriedade muito interessante para futuras aplicações em produtos cárneos refrigerados.In this work, the bacteriocins produced by six bacterial strains were studied (Lactobacillus sake 1, Lactobacillus curvatus 5, Leuconostoc mesenteroides 11, Leuconostoc sp 12, Lactobacillus curvatus 14 and Lactobacillus sake 16. Title of inhibitory activity was determined by critical dilution assay, using Lactobacillus sake ATCC 15521 and Listeria monocytogenes as indicator microorganisms. The inhibitory compounds were also characterized with respect to stability to the action of enzymes, thermostability, stability in several p

Mode of action and in vitro susceptibility of mastitis pathogens to macedocin ST91KM and preparation of a teat seal containing the bacteriocin

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Renee Pieterse

2010-03-01

Full Text Available Mastitis is considered to be the most economically costly disease affecting the dairy industry. Regular dosage of animals with antibiotics, including use of prophylactic concentrations, may select for resistant strains. The purpose of this study was to determine the mode of action of a new bacteriocin (macedocin ST91KM, to evaluate the antimicrobial resistance of mastitis pathogens to antibiotics commonly used in treatment remedies, and to introduce the possible use of an alternative antimicrobial agent. The bacteriocin macedocin ST91KM, produced by Streptococcus gallolyticus subsp. macedonicus ST91KM, is bactericidal to Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus uberis and Staphylococcus aureus associated with mastitis infections, including strains resistant to methicillin and oxacillin. Sensitive cells were deformed and secreted nucleotides, K+ and β-galactosidase when exposed to macedocin ST91KM. Adsorption of the peptide to target cells decreased in the presence of solvents, suggesting that receptors on the cell surfaces have lipid moieties. No adsorption was recorded in the presence of MgCl2, KI and Na2CO3, suggesting that ionic strength plays an important role. A teat seal preparation containing macedocin ST91KM effectively released the peptide and inhibited the growth of S. agalactiae. Macedocin ST91KM could form the basis for alternative dry cow therapy to prevent mastitis infections in dairy cows as it is effective against pathogens that display resistance to conventional antibiotic therapy.

Characterization and Application of Enterocin RM6, a Bacteriocin from Enterococcus faecalis

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En Huang

2013-01-01

Full Text Available Use of bacteriocins in food preservation has received great attention in recent years. The goal of this study is to characterize enterocin RM6 from Enterococcus faecalis OSY-RM6 and investigate its efficacy against Listeria monocytogenes in cottage cheese. Enterocin RM6 was purified from E. faecalis culture supernatant using ion exchange column, multiple C18-silica cartridges, followed by reverse-phase high-performance liquid chromatography. The molecular weight of enterocin RM6 is 7145.0823 as determined by mass spectrometry (MS. Tandem mass spectrometry (MS/MS analysis revealed that enterocin RM6 is a 70-residue cyclic peptide with a head-to-tail linkage between methionine and tryptophan residues. The peptide sequence of enterocin RM6 was further confirmed by sequencing the structural gene of the peptide. Enterocin RM6 is active against Gram-positive bacteria, including L. monocytogenes, Bacillus cereus, and methicillin-resistant Staphylococcus aureus (MRSA. Enterocin RM6 (final concentration in cottage cheese, 80 AU/mL caused a 4-log reduction in population of L. monocytogenes inoculated in cottage cheese within 30 min of treatment. Therefore, enterocin RM6 has potential applications as a potent antimicrobial peptide against foodborne pathogens in food.

Persistence of the oral probiotic Streptococcus salivarius M18 is dose dependent and megaplasmid transfer can augment their bacteriocin production and adhesion characteristics.

Science.gov (United States)

Burton, Jeremy P; Wescombe, Philip A; Macklaim, Jean M; Chai, Melissa H C; Macdonald, Kyle; Hale, John D F; Tagg, John; Reid, Gregor; Gloor, Gregory B; Cadieux, Peter A

2013-01-01

Bacteriocin-producing probiotic Streptococcus salivarius M18 offers beneficial modulatory capabilities within the oral microbiome, apparently through potent inhibitory activity against potentially deleterious bacteria, such as Streptococcus pyogenes. The oral cavity persistence of S. salivarius M18 was investigated in 75 subjects receiving four different doses for 28 days. Sixty per cent of the subjects already had some inhibitor-producing S. salivarius in their saliva prior to probiotic intervention. Strain M18's persistence was dependent upon the dose, but not the period of administration. Culture analysis indicated that in some individuals the introduced strain had almost entirely replaced the indigenous S. salivarius, though the total numbers of the species did not increase. Selected subjects showing either high or low probiotic persistence had their salivary populations profiled using Illumina sequencing of the V6 region of the 16S rRNA gene. Analysis indicated that while certain bacterial phenotypes were markedly modulated, the overall composition of the oral microbiome was not modified by the probiotic treatment. Megaplasmids encoding bacteriocins and adhesion factors were transferred in vitro to generate a transconjugant S. salivarius exhibiting enhanced antimicrobial production and binding capabilities to HEp-2 cells. Since no widespread perturbation of the existing indigenous microbiota was associated with oral instillation and given its antimicrobial activity against potentially pathogenic streptococci, it appears that application of probiotic strain M18 offers potential low impact alternative to classical antibiotic prophylaxis. For candidate probiotic strains having relatively poor antimicrobial or adhesive properties, unique derivatives displaying improved probiotic performance may be engineered in vitro by megaplasmid transfer.

Persistence of the oral probiotic Streptococcus salivarius M18 is dose dependent and megaplasmid transfer can augment their bacteriocin production and adhesion characteristics.

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Jeremy P Burton

Full Text Available Bacteriocin-producing probiotic Streptococcus salivarius M18 offers beneficial modulatory capabilities within the oral microbiome, apparently through potent inhibitory activity against potentially deleterious bacteria, such as Streptococcus pyogenes. The oral cavity persistence of S. salivarius M18 was investigated in 75 subjects receiving four different doses for 28 days. Sixty per cent of the subjects already had some inhibitor-producing S. salivarius in their saliva prior to probiotic intervention. Strain M18's persistence was dependent upon the dose, but not the period of administration. Culture analysis indicated that in some individuals the introduced strain had almost entirely replaced the indigenous S. salivarius, though the total numbers of the species did not increase. Selected subjects showing either high or low probiotic persistence had their salivary populations profiled using Illumina sequencing of the V6 region of the 16S rRNA gene. Analysis indicated that while certain bacterial phenotypes were markedly modulated, the overall composition of the oral microbiome was not modified by the probiotic treatment. Megaplasmids encoding bacteriocins and adhesion factors were transferred in vitro to generate a transconjugant S. salivarius exhibiting enhanced antimicrobial production and binding capabilities to HEp-2 cells. Since no widespread perturbation of the existing indigenous microbiota was associated with oral instillation and given its antimicrobial activity against potentially pathogenic streptococci, it appears that application of probiotic strain M18 offers potential low impact alternative to classical antibiotic prophylaxis. For candidate probiotic strains having relatively poor antimicrobial or adhesive properties, unique derivatives displaying improved probiotic performance may be engineered in vitro by megaplasmid transfer.

Potential of bacteriocins from lab to improve microbial quality of dry-cured and fermented meat products.

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Kęska, Paulina; Stadnik, Joanna; Zielińska, Dorota; Kołożyn-Krajewska, Danuta

2017-01-01

Meat and meat products are an important component of the daily diet. Nevertheless, they are perishable goods and are prone to microbial contamination, which leads to an increased risk to the health of consumers as well as economic losses in the meat industry. Fermentation has been used for thousands of years to preserve meat. As a result of extensive biochemical reactions occurring in meat during fermentation and ripening, the condi- tions inhibiting the growth of pathogenic and spoilage bacteria are formed. These changes are catalyzed by endogenous meat enzymes and exogenous enzymes derived from natural contaminating bacteria or starter cultures applied. In dry-cured and fermented meat products they are represented mainly by lactic acid bacte- ria (LAB) that produce a wide range of compounds, such as bacteriocins, directed against other microorgan- isms. The use of bactericidal peptides does not affect the sensory quality of foodstuffs, so that they attract attention as alternative means of preserving the stability and safety of dry-cured products.

Biochemical and Genetic Evidence that Enterococcus faecium L50 Produces Enterocins L50A and L50B, the sec-Dependent Enterocin P, and a Novel Bacteriocin Secreted without an N-Terminal Extension Termed Enterocin Q

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Cintas, Luis M.; Casaus, Pilar; Herranz, Carmen; Håvarstein, Leiv Sigve; Holo, Helge; Hernández, Pablo E.; Nes, Ingolf F.

2000-01-01

Enterococcus faecium L50 grown at 16 to 32°C produces enterocin L50 (EntL50), consisting of EntL50A and EntL50B, two unmodified non-pediocin-like peptides synthesized without an N-terminal leader sequence or signal peptide. However, the bacteriocin activity found in the cell-free culture supernatants following growth at higher temperatures (37 to 47°C) is not due to EntL50. A purification procedure including cation-exchange, hydrophobic interaction, and reverse-phase liquid chromatography has shown that the antimicrobial activity is due to two different bacteriocins. Amino acid sequences obtained by Edman degradation and DNA sequencing analyses revealed that one is identical to the sec-dependent pediocin-like enterocin P produced by E. faecium P13 (L. M. Cintas, P. Casaus, L. S. Håvarstein, P. E. Hernández, and I. F. Nes, Appl. Environ. Microbiol. 63:4321–4330, 1997) and the other is a novel unmodified non-pediocin-like bacteriocin termed enterocin Q (EntQ), with a molecular mass of 3,980. DNA sequencing analysis of a 963-bp region of E. faecium L50 containing the enterocin P structural gene (entP) and the putative immunity protein gene (entiP) reveals a genetic organization identical to that previously found in E. faecium P13. DNA sequencing analysis of a 1,448-bp region identified two consecutive but diverging open reading frames (ORFs) of which one, termed entQ, encodes a 34-amino-acid protein whose deduced amino acid sequence was identical to that obtained for EntQ by amino acid sequencing, showing that EntQ, similarly to EntL50A and EntL50B, is synthesized without an N-terminal leader sequence or signal peptide. The second ORF, termed orf2, was located immediately upstream of and in opposite orientation to entQ and encodes a putative immunity protein composed of 221 amino acids. Bacteriocin production by E. faecium L50 showed that EntP and EntQ are produced in the temperature range from 16 to 47°C and maximally detected at 47 and 37 to 47�

Bacteriocin-like activity of oral Fusobacterium nucleatum isolated from human and non-human primates Atividade semelhante a bacteriocina de Fusobacterium nucleatum orais isolados de primatas humanos e não-humanos

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Elerson Gaetti-Jardim Júnior

1999-12-01

Full Text Available Fusobacterium nucleatum is indigenous of the human oral cavity and has been involved in different infectious processes. The production of bacteriocin-like substances may be important in regulation of bacterial microbiota in oral cavity. The ability to produce bacteriocin-like substances by 80 oral F. nucleatum isolates obtained from periodontal patients, healthy individuals and Cebus apella monkeys, was examinated. 17.5% of all tested isolates showed auto-antagonism and 78.8% iso- or hetero-antagonism. No isolate from monkey was capable to produce auto-inhibition. In this study, the antagonistic substances production was variable in all tested isolates. Most of the F. nucleatum showed antagonistic activity against tested reference strains. These data suggest a possible participation of these substances on the oral microbial ecology in humans and animals. However, the role of bacteriocins in regulating dental plaque microbiota in vivo is discussed.Fusobacterium nucleatum é indígena da cavidade oral humana e tem sido envolvido em diferentes processos infecciosos. A produção de substâncias semelhantes a bacteriocinas pode ser importante na regulação da microbiota bacteriana da cavidade oral. A capacidade de produzir substâncias tipo bacteriocina de 80 isolados de F. nucleatum orais, obtidos de pacientes com doença periodontal, indivíduos sadios e macaco Cebus apella, foi avaliada. 17,5% de todos os isolados mostrou auto-antagonismo e 78,8% iso- ou hetero-antagonismo. Nenhum isolado de macaco foi capaz de produzir auto-inibição. Neste estudo, a produção de substâncias antagonístas foi variável em todos os isolados testados. A maioria dos F. nucleatum mostrou atividade antagonísta para as cepas de referência testadas. Esses dados sugerem a possível participação dessas substâncias sobre a ecologia microbiana em humanos e animais. Entretanto, o papel das bacteriocinas na regulação da microbiota da placa dental in vivo �

Cloning, production, and functional expression of the bacteriocin enterocin A, produced by Enterococcus faecium T136, by the yeasts Pichia pastoris, Kluyveromyces lactis, Hansenula polymorpha, and Arxula adeninivorans.

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Borrero, Juan; Kunze, Gotthard; Jiménez, Juan J; Böer, Erik; Gútiez, Loreto; Herranz, Carmen; Cintas, Luis M; Hernández, Pablo E

2012-08-01

The bacteriocin enterocin A (EntA) produced by Enterococcus faecium T136 has been successfully cloned and produced by the yeasts Pichia pastoris X-33EA, Kluyveromyces lactis GG799EA, Hansenula polymorpha KL8-1EA, and Arxula adeninivorans G1212EA. Moreover, P. pastoris X-33EA and K. lactis GG799EA produced EntA in larger amounts and with higher antimicrobial and specific antimicrobial activities than the EntA produced by E. faecium T136.

A novel bacteriocin-like substance (BLIS) from a pathogenic strain of Vibrio harveyi.

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Prasad, Sathish; Morris, Peter C; Hansen, Rasmus; Meaden, Philip G; Austin, Brian

2005-09-01

Inter-strain and inter-species inhibition mediated by a bacteriocin-like inhibitory substance (BLIS) from a pathogenic Vibrio harveyi strain VIB 571 was demonstrated against four isolates of the same species, and one culture each of a Vibrio sp., Vibrio fischeri, Vibrio gazogenes and Vibrio parahaemolyticus. The crude BLIS, which was obtained by ammonium-sulphate precipitation of the cell-free supernatant of a 72 h broth culture of strain VIB 571, was inactivated by lipase, proteinase K, pepsin, trypsin, pronase E, SDS and incubation at > or =60 degrees C for 10 min. The activity was stable between pH 2-11 for at least 5 h. Anion-exchange chromatography, gel filtration, SDS-PAGE and two-dimensional gel electrophoresis revealed the presence of a single major peak, comprising a protein with a pI of approximately 5.4 and a molecular mass of approximately 32 kDa. The N-terminal amino acid sequence of the protein comprised Asp-Glu-Tyr-Ile-Ser-X-Asn-Lys-X-Ser-Ser-Ala-Asp-Ile (with X representing cysteine or modified amino acid residues). A similarity search based on the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) generated peptide masses and the N-terminal sequence did not yield any significant matches.

Impairment of the class IIa bacteriocin receptor function and membrane structural changes are associated to enterocin CRL35 high resistance in Listeria monocytogenes.

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Masias, Emilse; Dupuy, Fernando G; da Silva Sanches, Paulo Ricardo; Farizano, Juan Vicente; Cilli, Eduardo; Bellomio, Augusto; Saavedra, Lucila; Minahk, Carlos

2017-07-01

Enterocin CRL35 is a class IIa bacteriocin with anti-Listeria activity. Resistance to these peptides has been associated with either the downregulation of the receptor expression or changes in the membrane and cell walls. The scope of the present work was to characterize enterocin CRL35 resistant Listeria strains with MICs more than 10,000 times higher than the MIC of the WT sensitive strain. Listeria monocytogenes INS7 resistant isolates R2 and R3 were characterized by 16S RNA gene sequencing and rep-PCR. Bacterial growth kinetic was studied in different culture media. Plasma membranes of sensitive and resistant bacteria were characterized by FTIR and Langmuir monolayer techniques. The growth kinetic of the resistant isolates was slower as compared to the parental strain in TSB medium. Moreover, the resistant isolates barely grew in a glucose-based synthetic medium, suggesting that these cells had a major alteration in glucose transport. Resistant bacteria also had alterations in their cell wall and, most importantly, membrane lipids. In fact, even though enterocin CRL35 was able to bind to the membrane-water interface of both resistant and parental sensitive strains, this peptide was only able to get inserted into the latter membranes. These results indicate that bacteriocin receptor is altered in combination with membrane structural modifications in enterocin CRL35-resistant L. monocytogenes strains. Highly enterocin CRL35-resistant isolates derived from Listeria monocytogenes INS7 have not only an impaired glucose transport but also display structural changes in the hydrophobic core of their plasma membranes. Copyright © 2017. Published by Elsevier B.V.

Protein expression vector and secretion signal peptide optimization to drive the production, secretion, and functional expression of the bacteriocin enterocin A in lactic acid bacteria.

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Borrero, Juan; Jiménez, Juan J; Gútiez, Loreto; Herranz, Carmen; Cintas, Luis M; Hernández, Pablo E

2011-10-20

Replacement of the leader sequence (LS) of the bacteriocin enterocin A (LS(entA)) by the signal peptides (SP) of the protein Usp45 (SP(usp45)), and the bacteriocins enterocin P (SP(entP)), and hiracin JM79 (SP(hirJM79)) permits the production, secretion, and functional expression of EntA by different lactic acid bacteria (LAB). Chimeric genes encoding the SP(usp45), the SP(entP), and the SP(hirJM79) fused to mature EntA plus the EntA immunity genes (entA+entiA) were cloned into the expression vectors pNZ8048 and pMSP3545, under control of the inducible P(nisA) promoter, and in pMG36c, under control of the constitutive P(32) promoter. The amount, antimicrobial activity, and specific antimicrobial activity of the EntA produced by the recombinant Lactococcus lactis, Enterococcus faecium, E. faecalis, Lactobacillus sakei and Pediococcus acidilactici hosts varied depending on the signal peptide, the expression vector, and the host strain. However, the antimicrobial activity and the specific antimicrobial activity of the EntA produced by most of the LAB transformants was lower than expected from their production. The supernatants of the recombinant L. lactis NZ9000 (pNZUAI) and L. lactis NZ9000 (pNZHAI), overproducers of EntA, showed a 1.2- to 5.1-fold higher antimicrobial activity than that of the natural producer E. faecium T136 against different Listeria spp. Copyright © 2011 Elsevier B.V. All rights reserved.

Identification, Characterization, and Recombinant Expression of Epidermicin NI01, a Novel Unmodified Bacteriocin Produced by Staphylococcus epidermidis That Displays Potent Activity against Staphylococci

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Sandiford, Stephanie

2012-01-01

We describe the discovery, purification, characterization, and expression of an antimicrobial peptide, epidermicin NI01, which is an unmodified bacteriocin produced by Staphylococcus epidermidis strain 224. It is a highly cationic, hydrophobic, plasmid-encoded peptide that exhibits potent antimicrobial activity toward a wide range of pathogenic Gram-positive bacteria including methicillin-resistant Staphylococcus aureus (MRSA), enterococci, and biofilm-forming S. epidermidis strains. Purification of the peptide was achieved using a combination of hydrophobic interaction, cation exchange, and high-performance liquid chromatography (HPLC). Matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) analysis yielded a molecular mass of 6,074 Da, and partial sequence data of the peptide were elucidated using a combination of tandem mass spectrometry (MS/MS) and de novo sequencing. The draft genome sequence of the producing strain was obtained using 454 pyrosequencing technology, thus enabling the identification of the structural gene using the de novo peptide sequence data previously obtained. Epidermicin NI01 contains 51 residues with four tryptophan and nine lysine residues, and the sequence showed approximately 50% identity to peptides lacticin Z, lacticin Q, and aureocin A53, all of which belong to a new family of unmodified type II-like bacteriocins. The peptide is active in the nanomolar range against S. epidermidis, MRSA isolates, and vancomycin-resistant enterococci. Other unique features displayed by epidermicin include a high degree of protease stability and the ability to retain antimicrobial activity over a pH range of 2 to 10, and exposure to the peptide does not result in development of resistance in susceptible isolates. In this study we also show the structural gene alone can be cloned into Escherichia coli strain BL21(DE3), and expression yields active peptide. PMID:22155816

Lactic Acid Bacteria and Their Bacteriocins: Classification, Biosynthesis and Applications against Uropathogens: A Mini-Review

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Mduduzi Paul Mokoena

2017-07-01

Full Text Available Several lactic acid bacteria (LAB isolates from the Lactobacillus genera have been applied in food preservation, partly due to their antimicrobial properties. Their application in the control of human pathogens holds promise provided appropriate strains are scientifically chosen and a suitable mode of delivery is utilized. Urinary tract infection (UTI is a global problem, affecting mainly diabetic patients and women. Many uropathogens are developing resistance to commonly used antibiotics. There is a need for more research on the ability of LAB to inhibit uropathogens, with a view to apply them in clinical settings, while adhering to strict selection guidelines in the choice of candidate LAB. While several studies have indicated the ability of LAB to elicit inhibitory activities against uropathogens in vitro, more in vivo and clinical trials are essential to validate the efficacy of LAB in the treatment and prevention of UTI. The emerging applications of LAB such as in adjuvant therapy, oral vaccine development, and as purveyors of bioprotective agents, are relevant in infection prevention and amelioration. Therefore, this review explores the potential of LAB isolates and their bacteriocins to control uropathogens, with a view to limit clinical use of antibiotics.

Antilisterial activity of a broad-spectrum bacteriocin, enterocin LR/6 from Enterococcus faecium LR/6.

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Kumar, Manoj; Srivastava, Sheela

2010-10-01

Enterocin LR/6, a purified bacteriocin, exhibited broad inhibitory spectrum both against related as well as some food-borne pathogens such as Listeria monocytogenes, Yersinia enterocolitica, Aeromonas sp., Shigella sp., and Bacillus licheniformis. In this investigation, we have focused on L. monocytogenes as the target organism, as it is not only an important pathogen but can also survive over a wide range of environmental conditions such as refrigeration temperature, low pH, and high-salt concentration. This allows the pathogen to overcome many food preservation and safety barriers and poses a potential risk to human health. The enterocin LR/6 showed a bactericidal action against L. monocytogenes and completely inhibited the growth on agar plates, supplemented with 200 AU/ml of enterocin LR/6. The effectiveness of enterocin LR/6 in completely killing a population of acid-adapted (pH 5.2, 2 h) L. monocytogenes exposed to different temperatures (4-37 degrees C), pH (2.5-8.0), and osmotic (up to 30% NaCl) stress is reported here. This paper focuses on the key issue of killing of the acid-adapted L. monocytogenes cells under adverse environmental conditions.

Properties of a Bacteriocin Produced by Bacillus subtilis EMD4 Isolated from Ganjang (Soy Sauce).

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Liu, Xiaoming; Lee, Jae Yong; Jeong, Seon-Ju; Cho, Kye Man; Kim, Gyoung Min; Shin, Jung-Hye; Kim, Jong-Sang; Kim, Jeong Hwan

2015-09-01

A Bacillus species, EMD4, with strong antibacterial activity was isolated from ganjang (soy sauce) and identified as B. subtilis. B. subtilis EMD4 strongly inhibited the growth of B. cereus ATCC14579 and B. thuringiensis ATCC33679. The antibacterial activity was stable at pH 3-9 but inactive at pH 10 and above. The activity was fully retained after 15 min at 80°C but reduced by 50% after 15 min at 90°C. The activity was completely destroyed by proteinase K and protease treatment, indicating its proteinaceous nature. The bacteriocin (BacEMD4) was partially purified from culture supernatant by ammonium sulfate precipitation, and QSepharose and Sephadex G-50 column chromatographies. The specific activity was increased from 769.2 AU/mg protein to 8,347.8 AU/mg protein and the final yield was 12.6%. The size of BacEMD4 was determined to be 3.5 kDa by Tricine SDS-PAGE. The N-terminal amino acid sequence was similar with that of Subtilosin A. Nucleotide sequencing of the cloned gene confirmed that BacEMD4 was Subtilosin A. BacEMD4 showed bactericidal activity against B. cereus ATCC14579.

Purification, Characterization, and Mode of Action of Plantaricin GZ1-27, a Novel Bacteriocin against Bacillus cereus.

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Du, Hechao; Yang, Jie; Lu, Xiaohong; Lu, Zhaoxin; Bie, Xiaomei; Zhao, Haizhen; Zhang, Chong; Lu, Fengxia

2018-05-09

Bacillus cereus is an opportunistic pathogen that causes foodborne diseases. We isolated a novel bacteriocin, designated plantaricin GZ1-27, and elucidated its mode of action against B. cereus. Plantaricin GZ1-27 was purified using ammonium sulfate precipitation, gel-filtration chromatography, and RP-HPLC. MALDI-TOF/MS revealed that its molecular mass was 975 Da, and Q-TOF-MS/MS analysis predicted the amino acid sequence as VSGPAGPPGTH. Plantaricin GZ1-27 showed thermostability and pH stability. The antibacterial mechanism was investigated using flow cytometry, confocal laser-scanning microscopy, scanning and transmission electron microscopy, and RT-PCR, which revealed that GZ1-27 increased cell membrane permeability, triggered K + leakage and pore formation, damaged cell membrane integrity, altered cell morphology and intracellular organization, and reduced the expression of genes related to cytotoxin production, peptidoglycan synthesis, and cell division. These results suggest that plantaricin GZ1-27 effectively inhibits B. cereus at both the cellular and the molecular levels and is a potential natural food preservative targeting B. cereus.

Evaluation of bacteriocin-producing Lactobacillus sakei 1 against Listeria monocytogenes 1/2a growth and haemolytic activity Avaliação de Lactobacillus sakei 1 produtor de bacteriocina frente a Listeria monocytogenes 1/2a e sua atividade hemolítica

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Rafael C.R. Martinez

2005-03-01

Full Text Available Bacteriocin-producing Lactobacillus sakei 1 was cultivated in Brain-Heart Infusion broth (24 h at 25ºC. The culture supernatant was neutralized, filter sterilized and used to test the activity of bacteriocin against Listeria monocytogenes 1/2a, at 8ºC and 15ºC. Non-bacteriocinogenic Lactobacillus sakei ATCC 15521 was used as a negative control. L. monocytogenes 1/2a was inoculated in culture supernatant medium from L. sakei 1 and L. sakei ATCC 15521 and the listerial populations were determined after 0, 5 and 10 days. The bacteriocin production was quantified as arbitrary units per mL (AU/mL using agar antagonism test. Additionally, to investigate if L. monocytogenes virulence pattern could be changed after bactericion exposure, the ability of L. monocytogenes to cause haemolysis in sheep red blood cells was determined, before and after exposure to bacteriocin at 8ºC. In the presence of the antimicrobial peptide, at 8ºC, L. monocytogenes population decreased, but growth of resistant cells was observed. At 15ºC, there was no difference between test and control. Furthermore, the haemolytic activity of L. monocytogenes 1/2a was not altered by exposure to L. sakei 1 bacteriocin, which suggests no change in its virulence pattern.Lactobacillus sakei 1 produtor de bacteriocina foi cultivado em caldo Infusão Cérebro-Coração por 24h a 25ºC. O sobrenadante da cultura foi neutralizado, esterilizado por filtração e usado para testar a atividade da bacteriocina frente a Listeria monocytogenes 1/2a, a 8ºC e 15ºC. Lactobacillus sakei ATCC 15521 não bacteriocinogênico, foi utilizado como controle negativo. L. monocytogenes 1/2a foi inoculada no sobrenadante da cultura de L.sakei 1 e L. sakei ATCC 15521 e as populações listeriais foram determinadas após 0, 5 e 10 dias. A produção de bacteriocina foi quantificada como unidades arbitrárias por mL (UA/mL, utilizando-se o teste de antagonismo em ágar. Adicionalmente, para investigar se o padr

Influence of baking enzymes on antimicrobial activity of five bacteriocin-like inhibitory substances produced by lactic acid bacteria isolated from Lithuanian sourdoughs.

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Narbutaite, V; Fernandez, A; Horn, N; Juodeikiene, G; Narbad, A

2008-12-01

To evaluate the effect of four different baking enzymes on the inhibitory activity of five bacteriocin-like inhibitory substances (BLIS) produced by lactic acid bacteria (LAB) isolated from Lithuanian sourdoughs. The overlay assay and the Bioscreen methods revealed that the five BLIS exhibited an inhibitory effect against spore germination and vegetative outgrowth of Bacillus subtilis, the predominant species causing ropiness in bread. The possibility that the observed antibacterial activity of BLIS might be lost after treatment with enzymes used for baking purposes was also examined. The enzymes tested; hemicellulase, lipase, amyloglucosidase and amylase had little or no effect on the majority of the antimicrobial activities associated with the five BLIS studied. This study suggests a potential application in the sourdough baking industry for these antimicrobial producing LAB strains in the control of B. subtilis spore germination and vegetative outgrowth.

Production of bacteriocin-like inhibitory substances (BLIS by Streptococcus salivarius strains isolated from the tongue and throat of children with and without sore throat Produção de substâncias inibidoras semelhantes à bacteriocina por cepas de Streptococcus salivarius, isoladas da língua e garganta de crianças com e sem dor de garganta

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Vera Fantinato

1999-12-01

Full Text Available Streptococcus salivarius strains, isolated from children with and without sore throat, were tested for bacteriocin production against Streptococcus pyogenes. S. salivarius strains producing bacteriocin-like inhibitory substances (BLIS against S. pyogenes were more frequently found in children without sore throat. These results suggest that these children may be protected against sore throat by the presence of BLIS-positive S. salivarius strains.Cepas de Streptococcus salivarius, isoladas de crianças com e sem dor de garganta, foram testadas quanto à produção de bacteriocina contra Streptococcus pyogenes. Os resultados mostraram que as crianças que não tinham dor de garganta possuiam, na boca, cepas de bactérias produtoras de substâncias inibidoras semelhantes à bacteriocina contra S. pyogenes.

Preservation of large yellow croaker (Pseudosciaena crocea) by Coagulin L1208, a novel bacteriocin produced by Bacillus coagulans L1208.

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Fu, Linglin; Wang, Chong; Ruan, Xinming; Li, Gang; Zhao, Yu; Wang, Yanbo

2018-02-02

Large yellow croaker (Pseudosciaena crocea) is a cultivated fish of great economic importance and abundant nutritional value. However, due to its high protein and water contents, it is susceptible to decomposition, leading to considerable economic loss and adverse effects on consumer health. Here, we assessed the function of the bacterial strain Bacillus coagulans L1208 (Bcoa) in preserving large yellow croaker during storage at 4°C and found that Bcoa elongates the shelf-life significantly. Further investigations showed that Bcoa prolongs the storage time mainly by suppressing the growth of spoilage bacteria. Moreover, a novel bacteriocin, designated as Coagulin L1208 and produced by Bcoa, was purified and identified by N-terminal sequencing. Finally, the activity of Coagulin L1208 for suppressing spoilage bacteria during the preservation of large yellow croaker was assessed. Our results reveal the mechanism by which Bcoa aids the preservation of large yellow croaker and identify Coagulin L1208 as a potential novel antiseptic. Copyright © 2017 Elsevier B.V. All rights reserved.

A modified R-type bacteriocin specifically targeting Clostridium difficile prevents colonization of mice without affecting gut microbiota diversity.

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Gebhart, Dana; Lok, Stephen; Clare, Simon; Tomas, Myreen; Stares, Mark; Scholl, Dean; Donskey, Curtis J; Lawley, Trevor D; Govoni, Gregory R

2015-03-24

Clostridium difficile is a leading cause of nosocomial infections worldwide and has become an urgent public health threat requiring immediate attention. Epidemic lineages of the BI/NAP1/027 strain type have emerged and spread through health care systems across the globe over the past decade. Limiting person-to-person transmission and eradicating C. difficile, especially the BI/NAP1/027 strain type, from health care facilities are difficult due to the abundant shedding of spores that are impervious to most interventions. Effective prophylaxis for C. difficile infection (CDI) is lacking. We have genetically modified a contractile R-type bacteriocin ("diffocin") from C. difficile strain CD4 to kill BI/NAP1/027-type strains for this purpose. The natural receptor binding protein (RBP) responsible for diffocin targeting was replaced with a newly discovered RBP identified within a prophage of a BI/NAP1/027-type target strain by genome mining. The resulting modified diffocins (a.k.a. Avidocin-CDs), Av-CD291.1 and Av-CD291.2, were stable and killed all 16 tested BI/NAP1/027-type strains. Av-CD291.2 administered in drinking water survived passage through the mouse gastrointestinal (GI) tract, did not detectably alter the mouse gut microbiota or disrupt natural colonization resistance to C. difficile or the vancomycin-resistant Enterococcus faecium (VREF), and prevented antibiotic-induced colonization of mice inoculated with BI/NAP1/027-type spores. Given the high incidence and virulence of the pathogen, preventing colonization by BI/NAP1/027-type strains and limiting their transmission could significantly reduce the occurrence of the most severe CDIs. This modified diffocin represents a prototype of an Avidocin-CD platform capable of producing targetable, precision anti-C. difficile agents that can prevent and potentially treat CDIs without disrupting protective indigenous microbiota. Treatment and prevention strategies for bacterial diseases rely heavily on traditional

Effect of autochthonous bacteriocin-producing Lactococcus lactis on bacterial population dynamics and growth of halotolerant bacteria in Brazilian charqui.

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Biscola, Vanessa; Abriouel, Hikmate; Todorov, Svetoslav Dimitrov; Capuano, Verena Sant'Anna Cabral; Gálvez, Antonio; Franco, Bernadette Dora Gombossy de Melo

2014-12-01

Charqui is a fermented, salted and sun-dried meat product, widely consumed in Brazil and exported to several countries. Growth of microorganisms in this product is unlikely due to reduced Aw, but halophilic and halotolerant bacteria may grow and cause spoilage. Charqui is a good source of lactic acid bacteria able to produce antimicrobial bacteriocins. In this study, an autochthonous bacteriocinogenic strain (Lactococcus lactis subsp. lactis 69), isolated from charqui, was added to the meat used for charqui manufacture and evaluated for its capability to prevent the growth of spoilage bacteria during storage up to 45 days. The influence of L. lactis 69 on the bacterial diversity during the manufacturing of the product was also studied, using denaturing gradient gel electrophoresis (DGGE). L. lactis 69 did not affect the counts and diversity of lactic acid bacteria during manufacturing and storage, but influenced negatively the populations of halotolerant microorganisms, reducing the spoilage potential. The majority of tested virulence genes was absent, evidencing the safety and potential technological application of this strain as an additional hurdle to inhibit undesirable microbial growth in this and similar fermented meat products. Copyright © 2014 Elsevier Ltd. All rights reserved.

Purification and characterization of multiple bacteriocins and an inducing peptide produced by Enterococcus faecium NKR-5-3 from Thai fermented fish.

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Ishibashi, Naoki; Himeno, Kohei; Fujita, Koji; Masuda, Yoshimitsu; Perez, Rodney Honrada; Zendo, Takeshi; Wilaipun, Pongtep; Leelawatcharamas, Vichien; Nakayama, Jiro; Sonomoto, Kenji

2012-01-01

Enterocins NKR-5-3A, B, C, and D were purified from the culture supernatant of Enterococcus faecium NKR-5-3 and characterized. Among the four purified peptides, enterocin NKR-5-3A (5242.3 Da) was identical to brochocin A, produced by Brochothrix campestris ATCC 43754, in mature peptides, and its putative synergistic peptide, enterocin NKR-5-3Z, was found to be encoded in ent53Z downstream of ent53A, encoding enterocin NKR-5-3A. Enterocin NKR-5-3B (6316.4 Da) showed a broad antimicrobial spectrum, and enterocin NKR-5-3C (4512.8 Da) showed high activity against Listeria. Enterocin NKR-5-3D (2843.5 Da), showing high homology to an inducing peptide produced by Lactobacillus sakei 5, induced the production of the enterocins. The enterocins showed different antimicrobial spectra and intensities. E. faecium NKR-5-3 concomitantly produced enterocins NKR-5-3A, B, C, and D which probably belong to different classes of bacteriocins. Furthermore, NKR-5-3 production was induced by enterocin NKR-5-3D.

Molecular Occurrence of Enterocin A Gene among Enterococcus faecium Strains Isolated from Gastro-Intestinal Tract and Antimicrobial Effect of this Bacteriocin Against Clinical Pathogens

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Mitra Salehi

2014-06-01

Materials and Methods: In this study occurrence of class II enterocin structural gene (enterocin A in a target of 42 Enterococcus faecium strains, isolated from gastrointestinal tract of animal have been surveyed. E. faecium identification and occurrence of enterocin A gene was performed by PCR method. Cell-free neutralized supernatant of gene positive strains was used to test bacteriocin production and antimicrobial spectrum of supernatant was assayed by wall diffusion method on the gram-positive and negative indicators bacteriaResults: Based on our results, 73.8% of isolated strains had enterocin A gene that they inhibited growth of indicator bacteria such as clinical strain of Pseudomonas aeruginosa, Salmonella enteric PTCC1709, Listeria monocytogenes, Bacillus cereus and Bacillus subtilis.Conclusions: Studied enterocins have growth inhibitory spectrum on Gram-positive and Gram-negative bacteria especially against pathogenic bacteria in the gastrointestinal tract. Therefore, these strains have the potential to explore and use as, alternative antimicrobial compound and bio-preservatives in food or feed or as probiotics.

Purification and mass spectrometry based characterization of a pediocin produced by Pediococcus acidilactici 13.

Science.gov (United States)

Altuntaş, Evrim Güneş; Ayhan, Kamuran; Peker, Selen; Ayhan, Beycan; Demiralp, Duygu Ozel

2014-10-01

Bacteriocins are antimicrobial peptides produced by several bacterial species. Among the bacteriocins pediocin-like bacteriocins have a significant inhibitory activity on the foodborne pathogens especially on Listeria monocytogenes. This study aims to select a simple and usable purification method to purify/concentrate the antimicrobial peptide and characterization of the bacteriocin produced by Pediococcus acidilactici 13 by using proteomic approaches which is a recent omic technology. For purification dialysis, ultrafiltration method was used, and as a result of this study the bacteriocin activity reached 819,200 AU/mL from 102,400 AU/mL initially. Two dimensional gel electrophoresis and then matrix-assisted laser desorption ionization/time of flight mass spectrometry (MALDI-TOF MS) analysis were carried out to identify the current bacteriocin and related proteins. Obtained data revealed similarity to pediocin PA-1 transport/processing ATP-binding protein PedD (accession number: P36497), pediocin operon PedC (accession number: Q68GC4) and bacteriocin pediocin PA-1 (accession number: P29430) from UniProtKB/Swiss-Prot databank, thus the bacteriocin produced by P. acidilactici 13 is considered similar to pediocin PA-1.

Common occurrence of antibacterial agents in human intestinal microbiota

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Fatima eDrissi

2015-05-01

Full Text Available Laboratory experiments have revealed many active mechanisms by which bacteria can inhibit the growth of other organisms. Bacteriocins are a diverse group of natural ribosomally-synthesized antimicrobial peptides produced by a wide range of bacteria and which seem to play an important role in mediating competition within bacterial communities. In this study, we have identified and established the structural classification of putative bacteriocins encoded by 317 microbial genomes in the human intestine. On the basis of homologies to available bacteriocin sequences, mainly from lactic acid bacteria, we report the widespread occurrence of bacteriocins across the gut microbiota: 175 bacteriocins were found to be encoded in Firmicutes, 79 in Proteobacteria, 34 in Bacteroidetes and 25 in Actinobacteria. Bacteriocins from gut bacteria displayed wide differences among phyla with regard to class distribution, net positive charge, hydrophobicity and secondary structure, but the α-helix was the most abundant structure. The peptide structures and physiochemical properties of bacteriocins produced by the most abundant bacteria in the gut, the Firmicutes and the Bacteroidetes, seem to ensure low antibiotic activity and participate in permanent intestinal host defence against the proliferation of harmful bacteria. Meanwhile, the potentially harmful bacteria, including the Proteobacteria, displayed highly effective bacteriocins, probably supporting the virulent character of diseases. These findings highlight the eventual role played by bacteriocins in gut microbial competition and their potential place in antibiotic therapy.

Assessing the antimicrobial activities of Ocins

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Shilja eChoyam

2015-09-01

Full Text Available The generation of a zone of inhibition on a solid substrate indicates the bioactivity of antimicrobial peptides such as bacteriocin and enterocin. The indicator strain plays a significant role in bacteriocin assays. Other characteristics of bacteriocins, such as their dispersal ability and the different zymogram components, also affect bacteriocin assays. However, universal well diffusion assays for antimicrobials, irrespective of their ability to diffuse (bacteriocin, enterocin, do not exist. The ability of different zymography components to generate non-specific activities have rarely been explored in the literature. The purpose of the present work was to evaluate the impact of two major factors (diffusion and no diffusion in a solid substrate bioassay, and to document the adverse effects of sodium dodecyl sulfate in zymograms used to estimate the approximate molecular weight of bacteriocins.

Mutacins and bacteriocins like genes in Streptococcus mutans isolated from participants with high, moderate, and low salivary count.

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Soto, Carolina; Padilla, Carlos; Lobos, Olga

2017-02-01

To detect S. mutans producers of mutacins and bacteriocins like substances (BLIS) from saliva of participants with low, moderate, and high salivary counts. 123 strains of S. mutans were obtained from participants with low, moderate, and high salivary counts (age 18 and 20 years old) and their antibacterial capacity analyzed. By using PCR amplification, the expression levels of mutacins and BLIS genes were studied (expressed in arbitrary units/ml) in all three levels. S. mutans strains from participants with low salivary counts show high production of mutacins (63%). In contrast, participants with moderate and high salivary counts depict relatively low levels of mutacins (22 and 15%, respectively). Moreover, participants with low salivary counts showed high expression levels of genes encoding mutacins, a result that correlates with the strong antimicrobial activity of the group. Participants with moderate and high salivary counts however depict low expression levels of mutacin related genes, and little antimicrobial activity. No BLIS were detected in any of the groups studied. S. mutans isolated from the saliva of participants with low bacterial counts have significant antibacterial capacity compared to that of participants with moderate and high salivary counts. The superior lethality of S. mutans in participants with low salivary counts is likely due to the augmented expression of mutacin- related genes. Copyright © 2016 Elsevier Ltd. All rights reserved.

Purification and Characterization of a Novel Anti-Campylobacter Bacteriocin Produced by Lactobacillus curvatus DN317.

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Zommiti, Mohamed; Almohammed, Hamdan; Ferchichi, Mounir

2016-12-01

The lactic acid bacteria (LAB) microbiota of Saudi chicken ceca was determined. From 60 samples, 204 isolates of lactic acid bacteria were obtained. Three isolates produced antimicrobial activities against Campylobacter jejuni, Listeria monocytogenes, and Bacillus subtilis. The isolate DN317, which had the highest activity against Campylobacter jejuni ATCC 33560, was identified as Lactobacillus curvatus (GenBank accession numbers: KX353849 and KX353850). Full inhibitory activity was observed after a 2-h incubation with the supernatant at pH values between 4 and 8. Only 16% of the activity was conserved after a treatment at 121 °C for 15 min. The use of proteinase K, pepsin, chymotrypsin, trypsin, papain, and lysozyme drastically reduced the antimicrobial activity. However, lipase, catalase, and lysozyme had no effect on this activity. The active peptide produced by Lactobacillus curvatus DN317 was purified by precipitation with an 80% saturated ammonium sulfate solution, and two steps of reversed phase HPLC on a C18 column. The molecular weight of this peptide was 4448 Da as determined by MALDI-ToF. N-terminal sequence analysis using Edman degradation revealed 47 amino acid residues (UniProt Knowledgebase accession number C0HK82) revealing homology with the amino acid sequences of sakacin P and curvaticin L442. The antimicrobial activity of the bacteriocin, namely curvaticin DN317, was found to be bacteriostatic against Campylobacter jejuni ATCC 33560. The use of microbial antagonism by LAB is one of the best ways to control microorganisms safely in foods. This result constitutes a reasonable advance in the antimicrobial field because of its potential applications in food technology.

Genomic and Proteomic Characterization of Bacteriocin-Producing Leuconostoc mesenteroides Strains Isolated from Raw Camel Milk in Two Southwest Algerian Arid Zones

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Zineb Benmechernene

2014-01-01

Full Text Available Information on the microbiology of camel milk is very limited. In this work, the genetic characterization and proteomic identification of 13 putative producing bacteriocin Leuconostoc strains exhibiting antilisterial activity and isolated from camel milk were performed. DNA sequencing of the 13 selected strains revealed high homology among the 16S rRNA genes for all strains. In addition, 99% homology with Leuconostoc mesenteroides was observed when these sequences were analysed by the BLAST tool against other sequences from reference strains deposited in the Genbank. Furthermore, the isolates were characterized by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDITOF MS which allowed for the identification of 2 mass peaks 6242 m/z and 5118 m/z that resulted to be specific to the species L. mesenteroides. Remarkably, the phyloproteomic tree provided more intraspecific information of L. mesenteroides than phylogenetic analysis. Accordingly, phyloproteomic analysis grouped L. mesenteroides strains into different subbranches, while all L. mesenteroides isolates were grouped in the same branch according to phylogenetic analysis. This study represents, to our knowledge, the first report on the use of MALDI-TOF MS on the identification of LAB isolated from camel milk.

Genomic and proteomic characterization of bacteriocin-producing Leuconostoc mesenteroides strains isolated from raw camel milk in two southwest Algerian arid zones.

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Benmechernene, Zineb; Fernández-No, Inmaculada; Quintela-Baluja, Marcos; Böhme, Karola; Kihal, Mebrouk; Calo-Mata, Pilar; Barros-Velázquez, Jorge

2014-01-01

Information on the microbiology of camel milk is very limited. In this work, the genetic characterization and proteomic identification of 13 putative producing bacteriocin Leuconostoc strains exhibiting antilisterial activity and isolated from camel milk were performed. DNA sequencing of the 13 selected strains revealed high homology among the 16S rRNA genes for all strains. In addition, 99% homology with Leuconostoc mesenteroides was observed when these sequences were analysed by the BLAST tool against other sequences from reference strains deposited in the Genbank. Furthermore, the isolates were characterized by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDITOF MS) which allowed for the identification of 2 mass peaks 6242 m/z and 5118 m/z that resulted to be specific to the species L. mesenteroides. Remarkably, the phyloproteomic tree provided more intraspecific information of L. mesenteroides than phylogenetic analysis. Accordingly, phyloproteomic analysis grouped L. mesenteroides strains into different subbranches, while all L. mesenteroides isolates were grouped in the same branch according to phylogenetic analysis. This study represents, to our knowledge, the first report on the use of MALDI-TOF MS on the identification of LAB isolated from camel milk.

The safe enterocin DD14 is a leaderless two-peptide bacteriocin with anti-Clostridium perfringens activity.

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Caly, Delphine L; Chevalier, Mickaël; Flahaut, Christophe; Cudennec, Benoit; Al Atya, Ahmed Khassaf; Chataigné, Gabrielle; D'Inca, Romain; Auclair, Eric; Drider, Djamel

2017-03-01

Enterococcus faecalis 14, a strain previously isolated from meconium, displayed activity against four Clostridium perfringens isolates when co-cultured on agar plates. The anti-Clostridium activity was ascribed to the production of enterocin DD14, which was subsequently purified. The minimum inhibitory concentration (MIC) of enterocin DD14 against one collection strain and one clinical C. perfringens strain was determined at 50 µg/mL. Furthermore, using the intestinal epithelial cell line IPEC-1, it was shown that E. faecalis 14 was not cytotoxic after 24 h of contact, and no cytotoxicity was observed when IPEC-1 cells were incubated with pure enterocin DD14 for 4 h. Enterocin DD14 was characterised using mass spectrometry and was shown to consist of two small proteins of 5200.74 Da and 5206.41 Da, respectively. The two peptides (DD14A and DD14B) have highly similar amino acid sequences and no signal peptide, which classifies enterocin DD14 as a class IIb leaderless two-peptide bacteriocin. The genes encoding DD14A and DD14B were sequenced and were shown to be 100% identical to other previously described enterocins MR10A and MR10B, in contrast to the producing strains, which are different. Consequently, the present in vitro study supports the potential of this E. faecalis 14 strain and/or its purified enterocin DD14 as putative anti-C. perfringens compounds in chickens. Copyright © 2017. Published by Elsevier B.V.

Assessment of the Bacteriocinogenic Potential of Marine Bacteria Reveals Lichenicidin Production by Seaweed-Derived Bacillus spp.

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Gillian E. Gardiner

2012-10-01

Full Text Available The objectives of this study were (1 to assess the bacteriocinogenic potential of bacteria derived mainly from seaweed, but also sand and seawater, (2 to identify at least some of the bacteriocins produced, if any and (3 to determine if they are unique to the marine environment and/or novel. Fifteen Bacillus licheniformis or pumilus isolates with antimicrobial activity against at least one of the indicator bacteria used were recovered. Some, at least, of the antimicrobials produced were bacteriocins, as they were proteinaceous and the producers displayed immunity. Screening with PCR primers for known Bacillus bacteriocins revealed that three seaweed-derived Bacillus licheniformis harbored the bli04127 gene which encodes one of the peptides of the two-peptide lantibiotic lichenicidin. Production of both lichenicidin peptides was then confirmed by mass spectrometry. This is the first definitive proof of bacteriocin production by seaweed-derived bacteria. The authors acknowledge that the bacteriocin produced has previously been discovered and is not unique to the marine environment. However, the other marine isolates likely produce novel bacteriocins, as none harboured genes for known Bacillus bacteriocins.

Les bactériocines des bactéries lactiques : caractéristiques et intérêts pour la bioconservation des produits alimentaires

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Dortu C.

2009-01-01

Full Text Available Bacteriocins from lactic acid bacteria: interest for food products biopreservation. Bacteriocins from lactic acid bacteria are low molecular weight antimicrobial peptides. They have inhibitory activity against the bacteria that are closed related to the producer strains and a narrow inhibitory spectrum. Nevertheless, most of them have activity against some food-born pathogenic bacteria as Listeria monocytogenes. The application of bacteriocins or bacteriocin producing lactic acid bacteria in food products to inhibit pathogenic or food-spoilage bacteria has then been suggested. This review focuses on the classification, structure, function, mode of action, biosynthesis and current food applications of bacteriocins from lactic acid bacteria.

Detectionn and activity of plantaricin OL15 a bacteriocin produced by Lactobacillus plantarum OL15 isolated from Algerian fermented olives.

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Nour-Eddine, Karam

2005-09-01

Full Text Available Lactobacillus plantarum OL15 previously isolated from Algerian fermented green olives produces antimicrobial conpounds in its culture medium. Inhibitory action against other strains of Lactobacillus, Lactococcus , and Propionibacterium was observed . Activity was completely or partially inactivated by proteolytic enzymes, stable at pH values ranging from 3.0 to 8.0, and heat stable even after autoclaving at 121 °C for 15 min. The bacteriocin activity was able to pass through cellulose membranes with 100,000 but not through 10,000 molecular weight cut-off.Lactobacillus plantarum OL15, aislado de aceitunas verdes argelinas, produce un compuesto antimicrobiano en el medio de cultivo, observándose un efecto inhibidor frente a otras cepas de Lactobacillus, Lactococcus y Propionibacterium . Dicha actividad desaparece completa o parcialmente después del tratamiento con enzimas proteolíticas, es estable al pH en el rango de 3,0 a 8,0 y es, asimismo estable al calor incluso después de someterla a un proceso térmico de 121 ºC durante 15 minutos. La bacteriocina pasa a través de membranas de celulosa de corte molecular de 1000.000 pero no a través de las de 10.000.

Co-expression of Nisin Z and Leucocin C as a Basis for Effective Protection Againstin Pasteurized Milk

NARCIS (Netherlands)

Fu, Yuxin; Mu, Dongdong; Qiao, Wanjin; Zhu, Duolong; Wang, Xiangxiang; Liu, Fulu; Xu, Haijin; Saris, Per; Kuipers, Oscar P; Qiao, Mingqiang

2018-01-01

Nisin, an important bacteriocin fromLactococcus lactissubsp., is primarily active against various Gram-positive bacteria. Leucocin C, produced byLeuconostoc carnosum4010, is a class IIa bacteriocin used to inhibit the growth ofListeria monocytogenes.Because two bacteriocins have different modes of

Nisin Z produced by Lactococcus lactis from bullfrog hatchery is active against Citrobacter freundii, a red-leg syndrome related pathogen.

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Quintana, Gabriel; Niederle, Maria V; Minahk, Carlos J; Picariello, Gianluca; Nader-Macías, María E F; Pasteris, Sergio E

2017-09-27

Lactococcus lactis subsp. lactis CRL 1584 isolated from a bullfrog hatchery produces a bacteriocin that inhibits both indigenous Citrobacter freundii (a Red-Leg Syndrome related pathogen) and Lactobacillus plantarum, and Listeria monocytogenes as well. Considering that probiotics requires high cell densities and/or bacteriocin concentrations, the effect of the temperature on L. lactis growth and bacteriocin production was evaluated to find the optimal conditions. Thus, the growth rate was maximal at 36 °C, whereas the highest biomass and bacteriocin activity was achieved between 20 and 30 °C and 20-25 °C, respectively. The bacteriocin synthesis was closely growth associated reaching the maximal values at the end of the exponential phase. Since bacteriocins co-production has been evidenced in bacterial genera, a purification of the bacteriocin/s from L. lactis culture supernatants was carried out. The active fraction was purified by cationic-exchange chromatography and then, a RP-HPLC was carried out. The purified sample was a peptide with a 3353.05 Da, a molecular mass that matches nisin Z, which turned out to be the only bacteriocin produced by L. lactis CRL 1584. Nisin Z showed bactericidal effect on C. freundii and L. monocytogenes, which increased in the presence L-lactic acid + H 2 O 2 . This is the first report on nisin Z production by L. lactis from a bullfrog hatchery that resulted active on a Gram-negative pathogen. This peptide has potential probiotic for raniculture and as food biopreservative for bullfrog meat.

Characterization of lactococci isolated from homemade kefir

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Kojić M.

2007-01-01

Full Text Available Five bacteriocin-producing lactococci isolates from traditionally prepared kefir were determined as Lactococcus lactis subsp. lactis. The analyzed isolates showed different plasmid profiles and no cross inhibition between them was detected. Moreover, natural isolate BGKF26 was resistant to the antimicrobial activity of nisin producing strain NP45. Plasmid curing experiments revealed that the genes encoding bacteriocin and proteinase production are located on separate genetic elements, except in BGKF26. Production of the tested bacteriocins depends on the concentration of casitone or triptone in the medium. Higher concentrations of casitone or triptone induce bacteriocin activity. Our DNA-DNA hybridization analyses suggest that the analyzed antimicrobial compounds probably are lactococcin-like bacteriocins.

Purification, Characterization and Antibacterial Mechanism of ...

African Journals Online (AJOL)

Purpose: To carry out the extraction, purification and biological characterization, and assess the antibacterial activity of bacteriocin from Lactobacillus acidophilus XH1. Methods: Chloroform extraction method was used for bacteriocin extraction while characterization of bacteriocin was carried out by flat-dug well agar ...

Protection of honeybee Apis mellifera by its endogenous and exogenous lactic flora against bacterial infections

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Irakli Janashia

2016-09-01

Three exogenous bacteriocin-producing LAB strains were tested against the same pathogens and against 25 endogenous bacterial isolates representing 11 different LAB species. The screening showed that all the tested exogenous bacteriocin-producing strains inhibited the tested P. larvae strains. The endogenous LAB strains exhibited varied sensitivity profiles when treated with bacteriocin-producing strains. This raises similar challenges to those observed in antibiotic applications leading to dysbacteriosis, even though the efficacy of these bacteriocins against P. larvae in an in vitro system is evident.

Bacteriocinogenic potential of a probiotic strain Bacillus coagulans [BDU3] from Ngari.

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Abdhul, Kaja; Ganesh, Mohan; Shanmughapriya, Santhanam; Vanithamani, Shanmugam; Kanagavel, Murugesan; Anbarasu, Kumarasamy; Natarajaseenivasan, Kalimuthusamy

2015-08-01

Bacteriocin producing strain BDU3 was isolated from a traditional fermented fish of Manipur Ngari. The strain BDU3 was identified as Bacillus coagulans by phenotypic and genotypic characterization. The BDU3 produced novel bacteriocin, which showed an antimicrobial spectrum toward a wide spectrum of food borne, and closely related pathogens with a MIC that ranged between 0.5 and 2.5 μg/mL. The isolate was able to tolerate pH as low as 2.0 and up to 0.2% bile salt concentration. Three step purification was employed to increase the specific activity of the antimicrobial compound. The fractions were further chromatographed by Rp-HPLC C-18 column and the purified bacteriocin had a specific activity of ∼8500 AU/mg. However, the potency of bacteriocin was susceptible to digestion with Proteinase K, Pepsin, SDS, EDTA and Urea. Molecular mass of purified bacteriocin was found to be 1.4 kDa using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF). The functional group was revealed by FTIR analysis. The cytotoxicity assay (MTT) using purified bacteriocin showed 2 times lower EC50 values compared to SDS. This is the smaller bacteriocin ever reported before from B. coagulans with greater antimicrobial potency with lower cytotoxicity. This bacteriocin raises the possibilities to be used as a biopreservative in food industries. Copyright © 2015 Elsevier B.V. All rights reserved.

Potential Applications of the Cyclic Peptide Enterocin AS-48 in the Preservation of Vegetable Foods and Beverages.

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Abriouel, Hikmate; Lucas, Rosario; Omar, Nabil Ben; Valdivia, Eva; Gálvez, Antonio

2010-06-01

Bacteriocins are antimicrobial peptides produced by bacteria. Among them, the enterococcal bacteriocin (enterocin) AS-48 stands for its peculiar characteristics and broad-spectrum antimicrobial activity. AS-48 belongs to the class of circular bacteriocins and has been studied in depth in several aspects: peptide structure, genetic determinants, and mode of action. Recently, a wealth of knowledge has accumulated on the antibacterial activity of this bacteriocin against foodborne pathogenic and spoilage bacteria in food systems, especially in vegetable foods and drinks. This work provides a general overview on the results from tests carried out with AS-48 in different vegetable food categories (such as fruit juices, ciders, sport and energy drinks, fresh fruits and vegetables, pre-cooked ready to eat foods, canned vegetables, and bakery products). Depending on the food substrate, the bacteriocin has been tested alone or as part of hurdle technology, in combination with physico-chemical treatments (such as mild heat treatments or high-intensity pulsed electric fields) and other antimicrobial substances (such as essential oils, phenolic compounds, and chemical preservatives). Since the work carried out on bacteriocins in preservation of vegetable foods and drinks is much more limited compared to meat and dairy products, the results reported for AS-48 may open new possibilities in the field of bacteriocin applications.

Commercial ampholytes used for isoelectric focusing may interfere with bioactivity based purification of antimicrobial peptides

OpenAIRE

Riazi, Shadi; Dover, Sara; Turovskiy, Yevgeniy; Chikindas, Michael L.

2007-01-01

BioRad's Rotofor® system has been frequently used for the purification of proteins and smaller peptides such as bacteriocins. In this study, we report that some commercially available ampholytes used with the Rotofor® isoelectric focusing system possess antimicrobial activity, which may interfere with the purification of bacteriocins and bacteriocin-like substances.

Inhibition of Cronobacter sakazakii by heat labile bacteriocins produced by probiotic LAB isolated from healthy infants.

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Awaisheh, Saddam S; Al-Nabulsi, Anas A; Osaili, Tareq M; Ibrahim, Salam; Holley, Richard

2013-09-01

Cronobacter sakazakii is an opportunistic pathogen that can cause bacteremia, meningitis, and necrotizing enterocolitis, most often in neonates with case-fatality rates that may reach 80%. The antimicrobial activity of lactic acid bacteria against a wide range of foodborne pathogens is well-established in different types of food products. The objective of the current study was to investigate the antibacterial activity of Lactobacillus acidophilus and L. casei isolated from feces of healthy infants against different strains of C. sakazakii in agar and a rehydrated infant milk formula (RIMF) model. The inhibition zones of C. sakazakii around L. acidophilus or L. casei ranged from 22 to 32 mm on eMan Rogosa Sharpe (MRS) agar under aerobic conditions, while a slight reduction in antibacterial activity was noted on modified MRS (0.2% glucose) under anaerobic conditions. It was observed that pH-neutralized cell-free supernatant (CFS) of L. acidophilus or L. casei was inhibitory against tested C. sakazakii strains. The inhibition zones of neutralized CFS were lower than the antibacterial activities of live cultures. The antibacterial activity of CFS was abolished when CFS from L. acidophilus or L. casei was heated at 60 or 80 °C for either 10 min or 2 h, or treated with trypsin or pepsin. This was considered strong evidence that the inhibition was due to the production of bacteriocins by L. casei and L. acidophilus. Both the CFS and active growing cells of L. casei and L. acidophilus were able to reduce the viability of C. sakazakii in the RIMF model. The results may extend the use of natural antimicrobials instead of conventional preservation methods to improve the safety of RIMF. © 2013 Institute of Food Technologists®

Food Applications and Regulation

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Gálvez, Antonio; Abriouel, Hikmate; Omar, Nabil Ben; Lucas, Rosario

This chapter deals with food applications of bacteriocins. Regulatory issues on the different possibilities for incorporating bacteriocins as bioprotectants are discussed. Specific applications of bacteriocins or bacteriocin-producing strains are described for main food categories, including milk and dairy products, raw meats, ready-to-eat meat and poultry products, fermented meats, fish and fish products or fermented fish. The last section of the chapter deals with applications in foods and beverages derived from plant materials, such as raw vegetable foods, fruits and fruit juices, cooked food products, fermented vegetable foods and ­fermented beverages. Results obtained for application of bacteriocins in combination with other hurdles are also discussed for each specific case, with a special emphasis on novel food packaging and food-processing technologies, such as irradiation, pulsed electric field treatments or high hydrostatic pressure treatment.

New insights into enterocin CRL35: mechanism of action and immunity revealed by heterologous expression in Escherichia coli.

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Barraza, Daniela E; Ríos Colombo, Natalia S; Galván, Adriana E; Acuña, Leonardo; Minahk, Carlos J; Bellomio, Augusto; Chalón, Miriam C

2017-09-01

The role of the class IIa bacteriocin membrane receptor protein remains unclear, and the following two different mechanisms have been proposed: the bacteriocin could interact with the receptor changing it to an open conformation or the receptor might act as an anchor allowing subsequent bacteriocin insertion and membrane disruption. Bacteriocin-producing cells synthesize an immunity protein that forms an inactive bacteriocin-receptor-immunity complex. To better understand the molecular mechanism of enterocin CRL35, the peptide was expressed as the suicidal probe EtpM-enterocin CRL35 in Escherichia coli, a naturally insensitive microorganism since it does not express the receptor. When the bacteriocin is anchored to the periplasmic face of the plasma membrane through the bitopic membrane protein, EtpM , E. coli cells depolarize and die. Moreover, co-expression of the immunity protein prevents the deleterious effect of EtpM-enterocin CRL35. The binding and anchoring of the bacteriocin to the membrane has demonstrated to be a sufficient condition for its membrane insertion. The final step of membrane disruption by EtpM-enterocin CRL35 is independent from the receptor, which means that the mannose PTS might not be involved in the pore structure. In addition, the immunity protein can protect even in the absence of the receptor. © 2017 John Wiley & Sons Ltd.

Optimization of culture conditions and medium composition for the production of micrococcin GO5 by Micrococcus sp. GO5.

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Kim, Mi-Hee; Kong, Yoon-Jung; Baek, Hong; Hyun, Hyung-Hwan

2006-01-02

To enhance the production of micrococcin GO5, a bacteriocin produced by Micrococcus sp. GO5, cultivation conditions and medium composition were optimized. The optimal initial pH and temperature for bacteriocin production were 7.0-9.0 and 37 degrees C, respectively. Micrococcus sp. GO5 displayed the highest micrococcin GO5 activity when grown in modified MRS medium that contained lactose or sucrose, rather than glucose, as a carbon source. The maximum bacteriocin activity was obtained in modified MRS medium containing 0.5% tryptone and 1.0% yeast extract as nitrogen sources instead of the other nitrogen sources present in MRS medium. Bacteriocin production was greatly affected by the concentration of K(2)HPO(4); strain GO5 produced eight-fold more bacteriocin in medium containing 2.0-2.5% K(2)HPO(4) than in medium containing 0.2% K(2)HPO(4). The optimal concentration of MgSO(4).7H(2)O for bacteriocin production was 0.5%. The production of micrococcin GO5 was increased 32-fold in shake flask culture and 16-fold in a bioreactor using the optimized medium (TY medium), compared with culturing in MRS medium.

Competitive advantage of bacteriocinogenic strains within lactic acid bacteria consortium of raw milk cheese

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Irena Rogelj

2011-03-01

Full Text Available The presence of gene determinants for different bacteriocins has been already demonstrated in traditional Slovenian types of raw milk cheeses ‘Tolminc’ and ‘Kraški’. These genes were present also in the cultivable microbiota. In this research the aim was to establish how the presence of gene determinants for bacteriocins in microbial consortia is reflected in its antimicrobial activity. In addition, one of the goals was to determine whether the strains that carry gene determinants for bacteriocins have any competitive growth advantage in microbial population. Microbial consortium of ‘Tolminc’ cheese was propagated in milk and examined at the end of propagation its antimicrobial activity and the presence of gene determinants for bacteriocins. Comparison of the results obtained before and after propagation leaded to the conclusion that most of the strains possessing gene determinants for bacteriocins were unable to persist during propagation. The strains which did persist during propagation carried gene determinants for enterocins P, L50B and cytolysin. Antimicrobial activity of consortium before and after propagation was not substantially different and cannot be attributed to any of detected bacteriocins.

Bacteriocinogenic LAB Strains for Fermented Meat Preservation: Perspectives, Challenges, and Limitations.

Science.gov (United States)

Favaro, Lorenzo; Todorov, Svetoslav Dimitrov

2017-12-01

Over the last decades, much research has focused on lactic acid bacteria (LAB) bacteriocins because of their potential as biopreservatives and their action against the growth of spoilage microbes. Meat and fermented meat products are prone to microbial contamination, causing health risks, as well as economic losses in the meat industry. The use of bacteriocin-producing LAB starter or protective cultures is suitable for fermented meats. However, although bacteriocins can be produced during meat processing, their levels are usually much lower than those achieved during in vitro fermentations under optimal environmental conditions. Thus, the direct addition of a bacteriocin food additive would be desirable. Moreover, safety and technological characteristics of the bacteriocinogenic LAB must be considered before their widespread applications. This review describes the perspectives and challenges toward the complete disclosure of new bacteriocins as effective preservatives in the production of safe and "healthy" fermented meat products.

Antibacterial Activity of Some Lactic Acid Bacteria Isolated from an Algerian Dairy Product

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Abdelkader Mezaini

2009-01-01

Full Text Available In the present study, the antibacterial effect of 20 lactic acid bacteria isolates from a traditional cheese was investigated. 6 isolates showed antibacterial effect against Gram positive bacteria. Streptococcus thermophilus T2 strain showed the wide inhibitory spectrum against the Gram positive bacteria. Growth and bacteriocin production profiles showed that the maximal bacteriocin production, by S. thermophilus T2 cells, was measured by the end of the late-log phase (90 AU ml−1 with a bacteriocine production rate of 9.3 (AU ml−1 h−1. In addition, our findings showed that the bacteriocin, produced by S. thermophilus T2, was stable over a wide pH range (4–8; this indicates that such bacteriocin may be useful in acidic as well as nonacidic food. This preliminarily work shows the potential application of autochthonous lactic acid bacteria to improve safety of traditional fermented food.

Heterologous Coproduction of Enterocin A and Pediocin PA-1 by Lactococcus lactis: Detection by Specific Peptide-Directed Antibodies

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Martínez, José M.; Kok, Jan; Sanders, Jan W.; Hernández, Pablo E.

2000-01-01

Antibodies against enterocin A were obtained by immunization of rabbits with synthetic peptides PH4 and PH5 designed, respectively, on the N- and C-terminal amino acid sequences of enterocin A and conjugated to the carrier protein KLH. Anti-PH4-KLH antibodies not only recognized enterocin A but also pediocin PA-1, enterocin P, and sakacin A, three bacteriocins which share the N-terminal class IIa consensus motif (YGNGVXC) that is contained in the sequence of the peptide PH4. In contrast, anti-PH5-KLH antibodies only reacted with enterocin A because the amino acid sequences of the C-terminal parts of class IIa bacteriocins are highly variable. Enterocin A and/or pediocin PA-1 structural and immunity genes were introduced in Lactococcus lactis IL1403 to achieve (co)production of the bacteriocins. The level of production of the two bacteriocins was significantly lower than that obtained by the wild-type producers, a fact that suggests a low efficiency of transport and/or maturation of these bacteriocins by the chromosomally encoded bacteriocin translocation machinery of IL1403. Despite the low production levels, both bacteriocins could be specifically detected and quantified with the anti-PH5-KLH (anti-enterocin A) antibodies isolated in this study and the anti-PH2-KLH (anti-pediocin PA-1) antibodies previously generated (J. M. Martínez, M. I. Martínez, A. M. Suárez, C. Herranz, P. Casaus, L. M. Cintas, J. M. Rodríguez, and P. E. Hernández, Appl. Environ. Microbiol. 64:4536–4545, 1998). In this work, the availability of antibodies for the specific detection and quantification of enterocin A and pediocin PA-1 was crucial to demonstrate coproduction of both bacteriocins by L. lactis IL1403(pJM04), because indicator strains that are selectively inhibited by each bacteriocin are not available. PMID:10919819

Partial purification and characterization of the mode of action of enterocin S37: a bacteriocin produced by Enterococcus faecalis S37 isolated from poultry feces.

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Belguesmia, Y; Choiset, Y; Prévost, H; Dalgalarrondo, M; Chobert, J-M; Drider, D

2010-01-01

The aim of this research was to purify and characterize the mode of action of enterocin S37, a bacteriocin produced by Enterococcus faecalis S37, a strain recently isolated from the chicken feces. Enterocin S37 has a molecular weight comprised between 4 and 5 kDa. It remained active after 1 h at 80(o)C and at pH values ranging from 4.0 to 9.0. Furthermore, cell-free supernatant of Enterococcus faecalis S37 and purified enterocin S37 were active against Gram-positive bacteria including Listeria monocytogenes EGDe, L. innocua F, Enterococcus faecalis JH2-2, and Lactobacillus brevis F145. The purification of enterocin S37 was performed by ammonium sulfate precipitation followed up by hydrophobic-interaction chromatography procedures. Treatment of enterocin S37 with proteinase K, alpha-chymotrypsin, and papain confirmed its proteinaceous nature, while its treatment with lysozyme and lipase resulted in no alteration of activity. Enterocin S37 is hydrophobic, anti-Listeria and likely acting by depletion of intracellular K(+) ions upon action on K(ATP) channels. This study contributed to gain more insights into the mode of action of enterocins.

Partial Purification and Characterization of the Mode of Action of Enterocin S37: A Bacteriocin Produced by Enterococcus faecalis S37 Isolated from Poultry Feces

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Y. Belguesmia

2010-01-01

Full Text Available The aim of this research was to purify and characterize the mode of action of enterocin S37, a bacteriocin produced by Enterococcus faecalis S37, a strain recently isolated from the chicken feces. Enterocin S37 has a molecular weight comprised between 4 and 5 kDa. It remained active after 1 h at 80oC and at pH values ranging from 4.0 to 9.0. Furthermore, cell-free supernatant of Enterococcus faecalis S37 and purified enterocin S37 were active against Gram-positive bacteria including Listeria monocytogenes EGDe, L. innocua F, Enterococcus faecalis JH2-2, and Lactobacillus brevis F145. The purification of enterocin S37 was performed by ammonium sulfate precipitation followed up by hydrophobic-interaction chromatography procedures. Treatment of enterocin S37 with proteinase K, -chymotrypsin, and papain confirmed its proteinaceous nature, while its treatment with lysozyme and lipase resulted in no alteration of activity. Enterocin S37 is hydrophobic, anti-Listeria and likely acting by depletion of intracellular K+ ions upon action on KATP channels. This study contributed to gain more insights into the mode of action of enterocins.

Development of a Chemically Defined Medium for Better Yield and Purification of Enterocin Y31 from Enterococcus faecium Y31

OpenAIRE

Liu, Wenli; Zhang, Lanwei; Yi, Huaxi

2017-01-01

The macro- and micronutrients in traditional medium, such as MRS, used for cultivating lactic acid bacteria, especially for bacteriocin production, have not been defined, preventing the quantitative monitoring of metabolic flux during bacteriocin biosynthesis. To enhance Enterocin Y31 production and simplify steps of separation and purification, we developed a simplified chemically defined medium (SDM) for the growth of Enterococcus faecium Y31 and production of its bacteriocin, Enterocin Y31...

Purification of the bacteriocin bavaricin MN and characterization of its mode of action against Listeria monocytogenes Scott A cells and lipid vesicles.

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Kaiser, A L; Montville, T J

1996-12-01

Bavaricin MN was purified from Lactobacillus sake culture supernatant 135-fold with a final yield of 11%. Sequence analysis revealed bavaricin MN to be a 42-amino-acid peptide having a molecular weight of 4,769 and a calculated pI of 10.0. Computer analysis indicated that the C-terminal region may form an alpha-helical structure with an amphipathic nature deemed important in the interaction of bacteriocins with biological membranes. Bavaricin MN rapidly depleted the membrane potential (delta p) of energized Listeria monocytogenes cells in a concentration-dependent fashion. At a bavaricin MN concentration of 9.0 micrograms/ml, the delta p decreased by 85%. Both the electrical potential (delta psi) and Z delta pH components of the delta p were depleted, and this depletion was not dependent on a threshold level of proton motive force. In addition to studying the effect of bavaricin MN on the delta p of vegetative cells, bavaricin MN-induced carboxyfluorescein (CF) efflux from L. monocytogenes-derived lipid vesicles was also characterized. Bavaricin MN-induced CF leakage was also concentration dependent with an optimum of pH 6.0. The rate of CF efflux was 63% greater in lipid vesicles in which a delta psi was generated compared with that in lipid vesicles in the absence of a delta psi.

Anti-MRSA activities of Enterocins DD28 and DD93 and evidences on their role in the inhibition of biofilm formation

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Ahmed Khassaf eAL ATYA

2016-05-01

Full Text Available Methicillin-resistant Staphylococcus aureus (MRSA, along with other antibiotic resistant bacteria, has become a worrisome superbug worldwide. This work was aimed at studying the efficacies of two class IIb bacteriocins, enterocins DD28 and DD93, against MRSA-S1 grown in planktonic culture and embedded in biofilms. These bacteriocins were purified, from the cultures supernatants of Enterococcus faecalis 28 and 93, using a simplified purification procedure consisting in a cation exchange chromatography and a reversed-phase high-performance liquid chromatography. The anti-Staphylococcal activity of these bacteriocins was shown in-vitro by the assessment of the minimal inhibitory concentration (MIC. Afterwhich, a checkerboard and time-kill kinetics permitted unveiled a synergistic effect of these bacteriocins in combination with erythromycin and kanamycin against the clinical MRSA-S1. These bacteriocins alone or in combination with erythromycin and kanamycin were able to impede the formation of MRSA-S1 biofilms on stainless steel and glace devices as supported by the microbial cell counts, epifluorescence and Scanning Electron Microscope analyses.

Daya Antibakteri Filtrat Asam Laktat dan Bakteriosin Lactobacillus bulgaricus KS1 dalam Menghambat Pertumbuhan Klebsiella pneumoniae Strain ATCC 700603, CT1538, dan S941

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Prima Nanda Fauziah

2015-03-01

Full Text Available Lactobacillus bulgaricus produces lactic acid and bacteriocin which have been reported to have various pharmacologic properties, including their role an antibacterial agent. Klebsiella pneumoniae, as an agent of pneumonia, remains a public health problem in tropical countries. This study was aimed to observe the antibacterial activities of lactic acid filtrate and bacteriocins of L. bulgaricus toward againsts K. pneumoniae strains by in vitro experiment. The experiment took place in Microbiology Laboratory, Teaching Hospital, Padjadjaran University, Bandung, August–October 2012. In vitro laboratory analytic study has been conducted on lactic acid filtrate and bacteriocins of L. bulgaricus against the K. pneumoniae strains. The study used agar pour plate and agar disk diffusion method and analyzed by ANAVA followed by Duncan’s multiple range test (DMRT. The 30% lactic acid filtrate and 20% bacteriocins filtrate concentrations of L. bulgaricus showed bactericidal characteristics againts the growth of K. pneumoniae strains. Greater concentration of lactic acid filtrate and bacteriocins of L. bulgaricus led toincreasing effect of growth inhibition zones of K. pneumoniae strains. Statistical analysis of variance (ANOVA showed that the greatest concentration effect of L. bulgaricus filtratefor inhibiting K. pneumoniae strains was achieved in 90% lactic acid filtrate concentration treatment, whereas the greatest inhibition zones for K. pneumoniae ATCC 700603 was obtaubed in 90% bacteriocins filtrate concentration, amounting 16.667 mm. In conclusion, lactic acid filtrate and bacteriocins L. bulgaricus have antibacterial effects on K. pneumoniae. The level of antibacterial effect of L. bulgaricus against the growth of K. pneumoniae strains depends on the type of filtrate, L. bulgaricus filtrate concentration, and K. pneumoniae strain.

The ColM Family, Polymorphic Toxins Breaching the Bacterial Cell Wall

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Maarten G. K. Ghequire

2018-02-01

Full Text Available Bacteria host an arsenal of antagonism-mediating molecules to combat for ecologic space. Bacteriocins represent a pivotal group of secreted antibacterial peptides and proteins assisting in this fight, mainly eliminating relatives. Colicin M, a model for peptidoglycan-interfering bacteriocins in Gram-negative bacteria, appears to be part of a set of polymorphic toxins equipped with such a catalytic domain (ColM targeting lipid II. Diversifying recombination has enabled parasitism of different receptors and has also given rise to hybrid bacteriocins in which ColM is associated with another toxin module. Remarkably, ColM toxins have recruited a diverse array of immunity partners, comprising cytoplasmic membrane-associated proteins with different topologies. Together, these findings suggest that different immunity mechanisms have evolved for ColM, in contrast to bacteriocins with nuclease activities.

Characterization of an anti-listerial enterocin from wheat silage based Enterococcus faecium.

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Bal, Emel Banu Buyukunal; Isevi, Taner; Bal, Mehmet Ali

2012-10-01

Two Enterococcus faecium and one E. faecalis strains isolated and identified from wheat silage were characterized based on plasmid content, hemolytic activity, antibiotic resistance patterns, bacteriocin production potential, and presence of enterocin structural genes (entA, entB, entP, entL50B). Among the isolates, only the E. faecium U7 strain exhibited bacteriocin activity against Listeria monocytogenes ATCC 7644, and vancomycin resistant Enterococcus spp. (VRE). A combination of three structural genes (entA, entB, and entP) was detected in E. faecium U7. A relationship between the presence of enterocin structural genes, and bacteriocin activity was detected in E. faecium U7; therefore partially purified enterocin (PPE) was further investigated from the isolate. Several bands of different molecular weights were expressed from PPE extracts following tricine SDS-PAGE analysis. However, the only band showing bacteriocin activity was in an approximate 4-kDa region. PPE treatment with proteinase K, lysozyme, and α -amylase caused complete loss of bacteriocin activity. PPE heat treatment at various temperatures resulted in a notable reduction in bacteriocin expression. Enterocin U7 was relatively heat stable, and presumably exhibits a glucoprotein nature with distinct inhibitory properties. Specific bacterial inhibitory activity of enterocin U7, and the producer strain absence of β -hemolysis and vancomycin susceptibility features deserves further investigation to evaluate its potential application in silage inoculation and food preservation. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mutations near the cleavage site of enterocin NKR-5-3B prepeptide reveal new insights into its biosynthesis.

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Perez, Rodney H; Sugino, Haruki; Ishibashi, Naoki; Zendo, Takeshi; Wilaipun, Pongtep; Leelawatcharamas, Vichien; Nakayama, Jiro; Sonomoto, Kenji

2017-04-01

Enterocin NKR-5-3B (Ent53B) is a 64-residue novel circular bacteriocin synthesized from an 87-residue prepeptide. Albeit through a still unknown mechanism, the EnkB1234 biosynthetic enzyme complex processes the prepeptide to yield its mature active, circular form. To gain insights into the key region/residue that plays a role in Ent53 maturation, several mutations near the cleavage site on the precursor peptide were generated. The interaction of the precursor peptide and EnkB1234 appeared to be hydrophobic in nature. At the Leu1 position, only mutations with helix structure-promoting hydrophobic residues (Ala, Ile, Val or Phe) were able to yield the mature Ent53B derivative. In this study, we also highlight the possible conformation-stabilizing role of the Ent53B leader peptide on the precursor peptide for its interaction with its biosynthetic enzyme complex. Any truncations of the leader peptide moiety interfered in the processing of the prepeptide. However, when propeptides of other circular bacteriocins (circularin A, leucocyclicin Q or lactocyclicin Q) were cloned at the C-terminus of the leader peptide, EnkB1234 could not process them to yield a mature bacteriocin. Taken together, these findings offer new perspectives in our understanding of the possible molecular mechanism of the biosynthesis of this circular bacteriocin. These new perspectives will help advance our current understanding to eventually elucidate circular bacteriocin biosynthesis. Understanding the biosynthetic mechanism of circular bacteriocins will materialize their application potential.

Inactivation of exopolysaccharide and 3-hydroxypropionaldehyde-producing lactic acid bacteria in apple juice and apple cider by enterocin AS-48.

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Martínez-Viedma, Pilar; Abriouel, Hikmate; Omar, Nabil Ben; Valdivia, Eva; López, Rosario Lucas; Gálvez, Antonio

2008-03-01

The bacteriocin enterocin AS-48 was tested against exopolysaccharide producing lactic acid bacteria (LAB) strains of Lactobacillus collinoides, Lactobacillus dioliovorans and Pediococcus parvulus as well as two 3-hydroxypropionaldehyde (3-HPA)-producing Lb. collinoides strains causing apple cider spoilage. In fresh-made apple juice, a bacteriocin concentration of 2.5 microg/ml reduced the LAB viable cell counts below detection levels during the course of incubation at 10 and 22 degrees C for most strains tested, except for Lb. collinoides 5 and Lb. dioliovorans 29. These two strains were significantly inhibited at 10 degrees C by 5 microg/ml AS-48 or completely inactivated at 22 degrees C. In a commercial Basque apple cider, the added bacteriocin (2.5 microg/ml for Lb. collinoides strains 9 and 10, and 5 microg/ml for the rest of strains) completely inactivated all LAB strains tested during storage at 10 as well as 22 degrees C. In the commercial Asturian apple cider tested the LAB strains showed a poor capacity for survival, but the added bacteriocin was equally effective in reducing the numbers of survivors. When a cocktail of the five LAB strains was tested in commercial Basque apple cider, viable cell counts were reduced below detection levels after 2 days for a bacteriocin concentration of 12.5 microg/ml regardless of storage temperature. Comparison of RAPD-PCR profiles revealed that strain Lb. dioliovorans 29 was always the predominant survivor detected in bacteriocin-treated samples.

CHARACTERIZATION OF THE PARTIALLY PURIFIED PLANTARCIN SR18 PRODUCED BY LACTOBACILLUS PLANTARUM SR18

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Wagih El-Shouny

2013-04-01

Full Text Available The bacteriocin bound to the cells and that secreted into the culture filtrate of Lactobacillus plantarum SR18 were precipitated by 75% ammomium sulphate, dialysed and further purified by Gel filtration on Sephadex G-100. Bacteriocins were purified from proteins bound to the cell of L. plantarum SR18 (plantarcin SR18 a and culture filtrate proteins (plantarcin SR18 b, respectively. The SDS-PAGE of partially purified Plantarcin SR18a showed a molecular weight of 3.5 KDa. While, plantarcin SR18 b had a molecular weight of 10.3 KDa. The antibacterial activity of the tested plantarcin SR18 preparations suffered no measurable loss after 45 min at 80ºC. Whereas, At 100ºC, significant decrease in the activity of bacteriocin preparations (60- 80 % took place by the end of 45 min. At pH ranged from 5-8, the activity of the plantarcin SR18 preparations suffered no measurable loss. Dissociating agents significantly affected the bacteriocin activity. Thus, tween 80 and mercaptoethanol increased the activity of bacteriocin preparations to 1.2-1.4 fold. Sodium dodecyl sulphate (SDS increased the activity of the tested bacteriocin preparations by about 20%.The lowest residual activity (60% was recorded after treatment with Triton X100 for 45 min. Protease completely inhibited the activities of all forms of plantarcin SR18 after 45 min at 37ºC.

Recent approaches in food bio-preservation - a review.

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Singh, Veer Pal

2018-01-01

Bio-preservation is a technique of extending the shelf life of food by using natural or controlled microbiota or antimicrobials. The fermentation products as well as beneficial bacteria are generally selected in this process to control spoilage and render pathogen inactive. The special interest organism or central organism used for this purpose is lactic acid bacteria (LAB) and their metabolites. They are capable to exhibit antimicrobial properties and helpful in imparting unique flavour and texture to the food products. The major compounds produced by LAB are bacteriocin, organic acids and hydrogen peroxide. Bacteriocin is peptides or proteins with antimicrobial activity. On the basis of size, structure and post-translational modification, bacteriocin is divided into four different classes. Due to non-toxic, non-immunogenic, thermo-resistance characteristics and broad bactericidal activity, LAB bacteriocins are considered good bio-preservative agents. The most common LAB bactriocin is nisin which has wider applications in food industry and has been Food and Drug Administration (FDA) approved. Nisin and other bacteriocin are being used in vegetables products, dairy and meat industries. Apart from LAB metabolites, bacteriophages and endolysins has promising role in food processing, preservation and safety. Bacteriocins and endolysins are more suitable for DNA shuffling and protein engineering to generate highly potent variants with expanded activity spectrum. Genetically modified bacteriophages may also be helpful in bio-preservation, however; their safety issues must be addressed properly before selection as bio-preservative agent.

Recent approaches in food bio-preservation - a review

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Veer Pal Singh

2018-03-01

Full Text Available Bio-preservation is a technique of extending the shelf life of food by using natural or controlled microbiota or antimicrobials. The fermentation products as well as beneficial bacteria are generally selected in this process to control spoilage and render pathogen inactive. The special interest organism or central organism used for this purpose is lactic acid bacteria (LAB and their metabolites. They are capable to exhibit antimicrobial properties and helpful in imparting unique flavour and texture to the food products. The major compounds produced by LAB are bacteriocin, organic acids and hydrogen peroxide. Bacteriocin is peptides or proteins with antimicrobial activity. On the basis of size, structure and post-translational modification, bacteriocin is divided into four different classes. Due to non-toxic, non-immunogenic, thermo-resistance characteristics and broad bactericidal activity, LAB bacteriocins are considered good bio-preservative agents. The most common LAB bactriocin is nisin which has wider applications in food industry and has been Food and Drug Administration (FDA approved. Nisin and other bacteriocin are being used in vegetables products, dairy and meat industries. Apart from LAB metabolites, bacteriophages and endolysins has promising role in food processing, preservation and safety. Bacteriocins and endolysins are more suitable for DNA shuffling and protein engineering to generate highly potent variants with expanded activity spectrum. Genetically modified bacteriophages may also be helpful in bio-preservation, however; their safety issues must be addressed properly before selection as bio-preservative agent.

Co-expression and characterization of enterocin CRL35 and its mutant in Escherichia coli Rosetta

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Masías Emilse

2014-01-01

Full Text Available Even though many sequences and structures of bacteriocins from lactic acid bacteria have been fully characterized so far, little information is currently available about bacteriocins heterologously produced by Escherichia coli. For this purpose, the structural gene of enterocin CRL35, munA, was PCR-amplified using specific primers and cloned downstream of PelB sequence in the pET22b (+ expression vector. E. coli Rosetta (DE3 pLysS was chosen as the host for production and enterocin was purified by an easy two-step protocol. The bacteriocin was correctly expressed with the expected intramolecular disulfide bond. Nevertheless, it was found that a variant of the enterocin, differing by 12 Da from the native polypeptide, was co-expressed by E. coli Rosetta in comparable amount. Indeed, the mutant bacteriocin contained two amino acid substitutions that were characterized by matrix assisted laser desorption ionization-time of flight (MALDI-TOF and HPLCelectrospray (ESI-Q-TOF tandem mass spectrometry (MS/ MS sequencing. This is the first report regarding the production of mutants of pediocin-like bacteriocins in the E. coli expression system.

Inhibition of Staphylococcus aureus in vitro by bacteriocinogenic Lactococcus lactis KTH0-1S isolated from Thai fermented shrimp (Kung-som) and safety evaluation.

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Saelao, Sutanate; Maneerat, Suppasil; Kaewsuwan, Sireewan; Rabesona, Hanitra; Choiset, Yvan; Haertlé, Thomas; Chobert, Jean-Marc

2017-05-01

Lactococcus lactis KTH0-1S isolated from Thai traditional fermented shrimp (Kung-som) is able to produce heat-stable bacteriocin and inhibits food spoilage bacteria and food-borne pathogens. The inhibitory effect of bacteriocin remained intact after treatment with different pHs and after heating, but was sensitive to some proteolytic enzymes. Addition of bacteriocin KTH0-1S to Staphylococcus aureus cultures decreased viable cell counts by 2.8 log CFU/ml, demonstrating a bactericidal mode of action. Furthermore, the growth of S. aureus decreased significantly after 12-h co-cultivation with bacteriocinogenic strain. The molecular mass of bacteriocin KTH0-1S was found to be 3.346 kDa after ammonium sulfate precipitation, reversed phase (C 8 Sep-Pak), cation-exchange chromatography, RP-HPLC on C 8 column and mass spectrometry (MS/MS) analysis. Bacteriocin KTH0-1S was identified as nisin Z using PCR amplification and sequencing. The majority of tested virulence factors were absent, confirming the safety. Evidenced inhibitory effect of this strain, the absence of virulence factors creates the possibility for its application as protective culture to inhibit pathogenic bacteria in the several fermented seafood products.

High Frequency and Diversity of Antimicrobial Activities Produced by Nasal Staphylococcus Strains against Bacterial Competitors.

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Daniela Janek

2016-08-01

Full Text Available The human nasal microbiota is highly variable and dynamic often enclosing major pathogens such as Staphylococcus aureus. The potential roles of bacteriocins or other mechanisms allowing certain bacterial clones to prevail in this nutrient-poor habitat have hardly been studied. Of 89 nasal Staphylococcus isolates, unexpectedly, the vast majority (84% was found to produce antimicrobial substances in particular under habitat-specific stress conditions, such as iron limitation or exposure to hydrogen peroxide. Activity spectra were generally narrow but highly variable with activities against certain nasal members of the Actinobacteria, Proteobacteria, Firmicutes, or several groups of bacteria. Staphylococcus species and many other Firmicutes were insusceptible to most of the compounds. A representative bacteriocin was identified as a nukacin-related peptide whose inactivation reduced the capacity of the producer Staphylococcus epidermidis IVK45 to limit growth of other nasal bacteria. Of note, the bacteriocin genes were found on mobile genetic elements exhibiting signs of extensive horizontal gene transfer and rearrangements. Thus, continuously evolving bacteriocins appear to govern bacterial competition in the human nose and specific bacteriocins may become important agents for eradication of notorious opportunistic pathogens from human microbiota.

Heterologous expression of enterocin A, a bacteriocin from Enterococcus faecium, fused to a cellulose-binding domain in Escherichia coli results in a functional protein with inhibitory activity against Listeria.

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Klocke, Michael; Mundt, Kerstin; Idler, Frank; Jung, Sabrina; Backhausen, Jan E

2005-06-01

The genes for the bacteriocins enterocin A and B were isolated from Enterococcus faecium ATB 197a. Using the pET37b(+) vector, the enterocin genes were fused to an Escherichia coli specific export signal sequence, a cellulose-binding domain (CBD(cenA)) and a S-tag under the control of a T7lac promotor. The constructs were subsequently cloned into E. coli host cells. The expression of the recombinant enterocins had different effects on both the host cells and other Gram-positive bacteria. The expression of entA in Esc. coli led to the synthesis and secretion of functional active enterocin A fusion proteins, which were active against some Gram-positive indicator bacteria, but did not influence the viability of the host cells. In contrast, the expression of enterocin B fusion proteins led to a reduced viability of the host cells, indicating a misfolding of the protein or interference with the cellular metabolism of Esc. coli. Indicator strains of Gram-positive bacteria were not inhibited by purified enterocin B fusion proteins. However, recombinant enterocin B displayed inhibitory activity after the proteolytic cleavage of the fused peptides.

Expression of the mucus adhesion genes Mub and MapA, adhesion-like factor EF-Tu and bacteriocin gene plaA of Lactobacillus plantarum 423, monitored with real-time PCR.

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Ramiah, K; van Reenen, C A; Dicks, L M T

2007-05-30

Expression of the mucus adhesion genes Mub and MapA, adhesion-like factor EF-Tu and bacteriocin gene plaA by Lactobacillus plantarum 423, grown in the presence of bile, pancreatin and at low pH, was studied by real-time PCR. Mub, MapA and EF-Tu were up-regulated in the presence of mucus, proportional to increasing concentrations. Expression of MapA was up-regulated in the presence of 3.0 g/l bile and 3.0 g/l pancreatin at pH 6.5. Similar results were recorded in the presence of 10.0 g/l bile and 10.0 g/l pancreatin at pH 6.5. Expression of Mub was down-regulated in the presence of bile and pancreatin, whilst the expression of EF-Tu and plaA remained unchanged. Expression of Mub and MapA remained unchanged at pH 4.0, whilst expression of EF-Tu and plaA were up-regulated. Expression of MapA was down-regulated in the presence of 1.0 g/l l-cysteine HCl, suggesting that the gene is regulated by transcription attenuation that involves cysteine.

Inactivation of Geobacillus stearothermophilus in canned food and coconut milk samples by addition of enterocin AS-48.

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Viedma, Pilar Martínez; Abriouel, Hikmate; Ben Omar, Nabil; López, Rosario Lucas; Valdivia, Eva; Gálvez, Antonio

2009-05-01

The cyclic bacteriocin enterocin AS-48 was tested on a cocktail of two Geobacillus stearothermophilus strains in canned food samples (corn and peas), and in coconut milk. AS-48 (7 microg/g) reduced viable cell counts below detection levels in samples from canned corn and peas stored at 45 degrees C for 30 days. In coconut milk, bacterial inactivation by AS-48 (1.75 microg/ml) was even faster. In all canned food and drink samples inoculated with intact G. stearothermophilus endospores, bacteriocin addition (1.75 microg per g or ml of food sample) rapidly reduced viable cell counts below detection levels and avoided regrowth during storage. After a short-time bacteriocin treatment of endospores, trypsin addition markedly increased G. stearothermophilus survival, supporting the effect of residual bacteriocin on the observed loss of viability for endospores. Results from this study support the potential of enterocin AS-48 as a biopreservative against G. stearothermophilus.

Inhibitory activity of Lactobacillus curvatus CWBI-B28 against ...

African Journals Online (AJOL)

user

2006-11-16

Nov 16, 2006 ... A bacteriocin-producing strain of Lactobacillus curvatus CWBI-B28 isolated from raw meat was shown ... Key words: Lactobacillus curvatus, Bacteriocin, hydrogen peroxide, ... Lactic acid bacteria (LAB) have long been used in food .... Pronase and catalase solutions were added to two of these tubes to.

Mechanism of Nisin, Pediocin 34, and Enterocin FH99 Resistance in Listeria monocytogenes.

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Kaur, Gurpreet; Singh, Tejinder Pal; Malik, Ravinder Kumar; Bhardwaj, Arun

2012-03-01

Nisin-, pediocin 34-, and enterocin FH99-resistant variants of Listeria monocytogenes ATCC 53135 were developed. In an attempt to clarify the possible mechanisms underlying bacteriocin resistance in L. monocytogenes ATCC 53135, sensitivity of the resistant strains of L. monocytogenes ATCC 53135 to nisin, pediocin 34, and enterocin FH99 in the absence and presence of different divalent cations was assessed, and the results showed that the addition of divalent cations significantly reduced the inhibitory activity of nisin, pediocin 34, and enterocin FH99 against resistant variants of L. monocytogenes ATCC 53135. The addition of EDTA, however, restored this activity suggesting that the divalent cations seem to affect the initial electrostatic interaction between the positively charged bacteriocin and the negatively charged phospholipids of the membrane. Nisin-, pediocin 34-, and enterocin-resistant variants of L. monocytogenes ATCC 53135 were more resistant to lysozyme as compared to the wild-type strain both in the presence as well as absence of nisin, pediocin 34, and enterocin FH99. Ultra structural profiles of bacteriocin-sensitive L. monocytogenes and its bacteriocin-resistant counterparts revealed that the cells of wild-type strain of L. monocytogenes were maximally in pairs or short chains, whereas, its nisin-, pediocin 34-, and enterocin FH99-resistant variants tend to form aggregates. Results indicated that without a cell wall, the acquired nisin, pediocin 34, and enterocin FH99 resistance of the variants was lost. Although the bacteriocin-resistant variants appeared to lose their acquired resistance toward nisin, pediocin 34, and enterocin FH99, the protoplasts of the resistant variants appeared to be more resistant to bacteriocins than the protoplasts of their wild-type counterparts.

High-salt brines compromise autoinducer-mediated bacteriocinogenic Lactobacillus plantarum survival in Spanish-style green olive fermentations.

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Caballero-Guerrero, Belén; Lucena-Padrós, Helena; Maldonado-Barragán, Antonio; Ruiz-Barba, José Luis

2013-02-01

The effect of NaCl on plantaricin production by five Lactobacillus plantarum strains was investigated. Plantaricin production in these strains was found to be regulated by three-component regulatory systems ruled by two different autoinducer peptides (AIPs), either PLNC8IF or Plantaricin A. Bacteriocin activity exhibited by these strains came to a halt in liquid medium containing NaCl concentrations of or above 2%. In contrast, bacteriocin activity was still observed when the producing strains were growing on solid medium containing up to 4% NaCl. Bacteriocin activity in liquid medium containing up to 2% NaCl could be restored by coculturing the producing strains with a selected plantaricin-production inducing strain of Lactococcus lactis. Growth of these bacteriocinogenic L. plantarum strains was monitored in traditional Spanish-style green olive fermentations. Survival of these strains could not be enhanced when provided with a range of plantaricin-production inducing mechanisms previously described, such as constitutive AIP production or coinoculation with a specific bacteriocin-production inducing strain of L. lactis. Our results suggest that it is advisable the use of constitutive bacteriocin producers, or at least non-AIP-dependant ones, as starters for olive fermentations due to the intrinsic physical characteristics of this food fermentation, especially the high salt concentration of the brines currently used. Copyright © 2012 Elsevier Ltd. All rights reserved.

Antibacterial activities of coagulase-negative staphylococci from bovine teat apex skin and their inhibitory effect on mastitis-related pathogens.

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Braem, G; Stijlemans, B; Van Haken, W; De Vliegher, S; De Vuyst, L; Leroy, F

2014-05-01

To explore antibacterial activities of coagulase-negative staphylococci (CoNS) from teat apices of dairy cows towards mastitis-causing pathogens. Of 254 CoNS, 38 displayed bacteriocin-like activity after a first screening. Seven of these strains displayed activity against at least one mastitis-related pathogen (Streptococcus uberis, Streptococcus dysgalactiae and Staphylococcus aureus). Staphylococcus chromogenes L217 displayed the strongest inhibitory effect, being active against all tested mastitis-related pathogens and most tested CoNS. Based on cation exchange and reversed-phase chromatography, in addition to N-terminal Edman degradation and PCR, the antibacterial peptide was identified as a nukacin-type bacteriocin and named nukacin L217. Although staphylococcal bacteriocins are generally found in the cell-free supernatants of liquid cultures, Staph. chromogenes L217 only led to detectable activity when grown on agar medium. Bacteriocin-like activities are not uncommon among CoNS from teat apices and may inhibit mastitis-causing pathogens, as found for nukacin L217 production by Staph. chromogenes L217. Nukacin L217 is the first identified bacteriocin of the species Staph. chromogenes and displays unusual production kinetics, that is, requiring surface growth of its producer. The fact that nukacins are produced by different CoNS species suggests a role in the teat skin ecosystem. © 2014 The Society for Applied Microbiology.

Characterization of antimicrobial substance from Lactobacillus salivarius KL-D4 and its application as biopreservative for creamy filling.

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Therdtatha, Phatthanaphong; Tandumrongpong, Chanabhorn; Pilasombut, Komkhae; Matsusaki, Hiromi; Keawsompong, Suttipun; Nitisinprasert, Sunee

2016-01-01

Lactobacillus salivarius KL-D4 isolated from duck intestine produced bacteriocin which was stable at high temperature and a wide pH range of 3-10. Its cell free supernatant at pH 5.5 exhibited wide inhibitory spectrum against both G+ and G- bacteria. The highest bacteriocin production was obtained in MRS broth supplemented with 0.5 % (w/v) CaCO3 at 6 h by gentle shaking. PCR walking using specific primers at the conserved region of class-II bacteriocin resulted in 4 known genes of kld1, kld2, kld3 and kld4 with 100 % similarity to genes encoding for salivaricin α, β, induction peptide and histidine protein kinase of Lb. salivarius GJ-24 which did not previously report for bacteriocin characterization, while showing 94, 93, 59 and 62 % to other salivaricin gene cluster, respectively. The high activities of 25,600 AU/ml indicated a strong induction peptide expressed by kld3 which has low similarity to previous inducer reported. Based on operon analysis, only kld1, kld3 and kld4 could be expressed and subsequently elucidated that only salivaricin α like bacteriocin was produced and secreted out of the cells. Using protein purification, only a single peptide band obtained showed that this strain produced one bacteriocin which could be salivaricin α namely salivaricin KLD showing about 4.3 kDa on SDS-PAGE. Partial purification by 20 % ammonium sulfate precipitation of the product was tested on the artificial contamination of creamy filling by Bacillus cereus, Enterococcus faecalis, Pseudomonas stutzeri, Staphylococcus sp. and Stenotrophomonas sp. resulting the growth inhibitory efficiency of 4.45-66.9, 11.5-100, 100, 0-28.1 and 5-100 % respectively. Therefore, salivaricin KLD can be a tentative biopreservative for food industry in the future.

Bio-preservative and therapeutic potential of pediocin: recent trends and future perspectives.

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Mehta, Ridhi; Arya, Ridhima; Goyal, Karan; Singh, Mahipal; Sharma, Anil K

2013-12-01

Bacteriocins produced by lactic acid bacteria are of keen interest to the food industry for their bio-preservative potential and antimicrobial properties. The increasing demand for high quality 'safe' foods which are not extensively processed has created a niche for natural food preservatives. The bacteriocins (produced by bacteria) derived their name after the genera that produce them, hence bacteriocins produced by genus Pediococcus are known as Pediocin. Pediocins are antimicrobial peptides which show a strong activity against food spoilage and pathogenic bacteria, also are thermostable in nature as well as stable over a wide range of pH. Pediocin produced by Pediococcus acidilactici, has been generally recognized as safe (GRAS). The current review summarizes about the progress made on the Pediocin research from patent perspective along with the immense potential of these Pediococcus derived bacteriocins not only as antimicrobial, biopreservative and probiotic agents but also for the treatment of cancer, body odors and other health promoting actions. The relevant patents have been listed and briefly analyzed to upgrade and benefit food industries by prolonging the shelf life of various products.

Optimization of nutritional and non-nutritional factors involved for production of antimicrobial compounds from Lactobacillus pentosus SJ65 using response surface methodology

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Appukuttan Saraniya

2014-01-01

Full Text Available Bacteriocins from lactic acid bacteria are ribosomal synthesized antibacterial proteins/ peptides having wide range of applications. Lactobacillus pentosus SJ65, isolated from fermented Uttapam batter (used to prepare south Indian pan cake, produces bacteriocin having a broad spectrum of activity against pathogens. Optimization studies are of utmost important to understand the source of utilization and the conditions to enhance the production of metabolites. In the present study, an attempt was made to identify the parameters involved for maximal production of antimicrobial compounds especially bacteriocin from the isolate L. pentosus SJ65. Initially, optimal conditions, such as incubation period, pH, and temperature were evaluated. Initial screening was done using methodology onevariable-at-a-time (OVAT for various carbon and nitrogen sources. Further evaluation was carried out statistically using Plackett-Burman design and the variables were analyzed using response surface methodology using central composite design. The optimum media using tryptone or soy peptone, yeast extract, glucose, triammonium citrate, MnSO4, dipotassium hydrogen phosphate and tween 80 produced maximum bacteriocin activity.

ANÁLISIS ESTRUCTURAL DE FILETES SAJADOS DE HÍBRIDO DE CACHAMA Piaractus braquypomus x Colossoma macropomum UTILIZANDO BACTERIOCINAS PRODUCIDAS POR Lactobacillus plantarum LPBM10 EMPACADO AL VACÍO

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Héctor Suárez M

2008-08-01

Full Text Available Objective. To determine microstructure, texture and sensory changes in cut cachama hybrid fillets. Materials and methods. Hybrid fillets that were vacuum packed for 30 days at 3°C were analyzed after preservative treatments by crude bacteriocins extract, lactic acid, and control by light microscopy. Results. Space among the muscle fibers increased gradually and the architectural arrangement was altered in all treatments throughout the entire storage period. The lowest alteration in connective tissue degradation and lowest increment in spaces between the muscular fibers was observed with the crude bacteriocin extract treatment. The instrumental textural analysis showed no statistical difference in the stability loss of tissues from fillets among treatments. Sensory analysis indicated that the lancing treatment was effective in diminishing the negative effect of intramuscular bones. The best scores were for cutting fillets with the bacteriocin treatment. Conclusions. The meat texture of fillets was affected during the storage period. Lancing is a procedure that enables the use of fillets without encountering the intramuscular bone problem. The use of crude bacteriocins extract prolongs the shelf-life of fillets.

Potential aquaculture probiont Lactococcus lactis TW34 produces nisin Z and inhibits the fish pathogen Lactococcus garvieae.

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Sequeiros, Cynthia; Garcés, Marisa E; Vallejo, Marisol; Marguet, Emilio R; Olivera, Nelda L

2015-04-01

Bacteriocin-producing Lactococcus lactis TW34 was isolated from marine fish. TW34 bacteriocin inhibited the growth of the fish pathogen Lactococcus garvieae at 5 AU/ml (minimum inhibitory concentration), whereas the minimum bactericidal concentration was 10 AU/ml. Addition of TW34 bacteriocin to L. garvieae cultures resulted in a decrease of six orders of magnitude of viable cells counts demonstrating a bactericidal mode of action. The direct detection of the bacteriocin activity by Tricine-SDS-PAGE showed an active peptide with a molecular mass ca. 4.5 kDa. The analysis by MALDI-TOF-MS detected a strong signal at m/z 2,351.2 that corresponded to the nisin leader peptide mass without the initiating methionine, whose sequence STKDFNLDLVSVSKKDSGASPR was confirmed by MS/MS. Sequence analysis of nisin structural gene confirmed that L. lactis TW34 was a nisin Z producer. This nisin Z-producing strain with probiotic properties might be considered as an alternative in the prevention of lactococcosis, a global disease in aquaculture systems.

Sil: a Streptococcus iniae bacteriocin with dual role as an antimicrobial and an immunomodulator that inhibits innate immune response and promotes S. iniae infection.

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Mo-fei Li

Full Text Available Streptococcus iniae is a Gram-positive bacterium and a severe pathogen to a wide range of economically important fish species. In addition, S. iniae is also a zoonotic pathogen and can cause serious infections in humans. In this study, we identified from a pathogenic S. iniae strain a putative bacteriocin, Sil, and examined its biological activity. Sil is composed of 101 amino acid residues and shares 35.6% overall sequence identity with the lactococcin 972 of Lactococcus lactis. Immunoblot analysis showed that Sil was secreted by S. iniae into the extracellular milieu. Purified recombinant Sil (rSil exhibited a dose-dependent inhibitory effect on the growth of Bacillus subtilis but had no impact on the growths of other 16 Gram-positive bacteria and 10 Gram-negative bacteria representing 23 different bacterial species. Treatment of rSil by heating at 50°C abolished the activity of rSil. rSil bound to the surface of B. subtilis but induced no killing of the target cells. Cellular study revealed that rSil interacted with turbot (Scophthalmus maximus head kidney monocytes and inhibited the innate immune response of the cells, which led to enhanced cellular infection of S. iniae. Antibody blocking of the extracellular Sil produced by S. iniae significantly attenuated the infectivity of S. iniae. Consistent with these in vitro observations, in vivo study showed that administration of turbot with rSil prior to S. iniae infection significantly increased bacterial dissemination and colonization in fish tissues. Taken together, these results indicate that Sil is a novel virulence-associated bacteriostatic and an immunoregulator that promotes S. iniae infection by impairing the immune defense of host fish.

ANTILISTERIAL ACTYVITY OF LACTIC ACID BACTERIA ISOLATED FROM GILTHEAD BREAMS AND SEA BASSES FILLETS PACKAGED MAP AGAINST PRIMITIVE STRAINS OF LISTERIA MONOCYTOGENES

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M. Barile

2011-04-01

Full Text Available Listeria monocytogenes is the causative agent of listeriosis typically caused by ready-to-eat processed food that have a refrigerated shelf-life, but lightly preserved fish products also belong to a high-risk category. Aim of the work was to evaluate antimicrobial activity linked bacteriocin-producing of LAB isolated from gilthead breams and sea basses fillets packaged in modified atmospheres. Fifty-five LAB strains were screened against 21 strains of Listeria monocytogenes, 1 Listeria innocua held in the culture collection of Department of Zootechnical Sciences and Food Ispection (SIA and submitted to antagonistic activity using the spot on lawn and the agar well diffusion assay. Lactococcus lactis sub. lactis Sa31 was able to produce bacteriocin in agar and different broth medium. The bacteriocin man31 showed sensitivity to trypsin, pronase E and papain, inactivation at temperatures ≥ 100°C, bactericidal mode of action and antilisterial act, rapidly. The bacteriocin man31 caused a reduction of L. monocytogenes ½ c growth about log10 > 3 UFC/ml, when was applied on indicator strain at 20,480 AU/ml concentration, in vitro.

Lantibiotics produced by Actinobacteria and their potential applications (a review).

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Gomes, Karen Machado; Duarte, Rafael Silva; de Freire Bastos, Maria do Carmo

2017-02-01

The phylum Actinobacteria, which comprises a great variety of Gram-positive bacteria with a high G+C content in their genomes, is known for its large production of bioactive compounds, including those with antimicrobial activity. Among the antimicrobials, bacteriocins, ribosomally synthesized peptides, represent an important arsenal of potential new drugs to face the increasing prevalence of resistance to antibiotics among microbial pathogens. The actinobacterial bacteriocins form a heterogeneous group of substances that is difficult to adapt to most proposed classification schemes. However, recent updates have accommodated efficiently the diversity of bacteriocins produced by this phylum. Among the bacteriocins, the lantibiotics represent a source of new antimicrobials to control infections caused mainly by Gram-positive bacteria and with a low propensity for resistance development. Moreover, some of these compounds have additional biological properties, exhibiting activity against viruses and tumour cells and having also potential to be used in blood pressure or inflammation control and in pain relief. Thus, lantibiotics already described in Actinobacteria exhibit potential practical applications in medical settings, food industry and agriculture, with examples at different stages of pre-clinical and clinical trials.

Resistência bacteriana e ação das bacteriocinas de Lactobacillus spp em Staphylococcus aureus isolados de mastite bovina

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B.R. Pribul

2011-06-01

Full Text Available Staphylococcus aureus is the major pathogen causing intramammary infections in dairy cattle worldwide. Among the factors that contribute to its spread and infectious potential is the ability to overcome the mechanisms of antimicrobials activity. The present work investigated the antimicrobial resistance pattern and sensibility to bacteriocins produced by strains of Lactobacillus spp of 30 isolates of S. aureus from mastitis. From this, 29 are beta-lactamase producers. Eight isolates (26.6% showed resistance to at least four antibiotics being considered multiresistent. All of them were mecA-positive. Otherwise, all isolates tested showed sensibility to at least one of the four bacteriocin producer strains. Due to the significant depletion of the efficacy of antimicrobials, pathogen growth inhibition by bacteriocins seems an alternative of biological control in infectious processes.

Generation of Food-Grade Lactococcal Starters Which Produce the Lantibiotics Lacticin 3147 and Lacticin 481

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O'Sullivan, Lisa; Ryan, Maire P.; Ross, R. Paul; Hill, Colin

2003-01-01

Transconjugant lactococcal starters which produce both lantibiotics lacticin 3147 and lacticin 481 were generated via conjugation of large bacteriocin-encoding plasmids. A representative of one of the resultant strains proved more effective at killing Lactobacillus fermentum and inhibiting the growth of Listeria monocytogenes LO28H than either of the single bacteriocin-producing parental strains, demonstrating the potential of these transconjugants as protection cultures for food safety applications. PMID:12788782

POTENTIAL OF Lactococcus lactis subsp. lactis MTCC 3041 AS A BIOPRESERVATIVE

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Neha Sharma

2013-10-01

Full Text Available Lactic acid bacteria especially in developing countries can be exploited against frequently occurring spoilage organisms of fresh fruits and vegetables in addition to pathogens. Keeping in views this antagonism imparted by bacteria Lactococci, the present study was taken and effectiveness of bacteriocin of Lactococci was also studied in preservatives and enzymes. Lactic acid bacteria Lactococcus lactis subs. Lactis MTCC 3041 was used as bacteriocin producer strain. Isolation of most frequently occurring spoilage organisms from spoiled Mango and Kinnow was done by microbiological procedures and were identified by microscopic studies as Isolate 1 and Isolate 2. It has limited use in processed salted food as no zone of inhibition was observed at and above 5% NaCl (w/v.0.3% (w/v is the minimum concentration of KMS that provides stress to the microorganism for the production of bacteriocin. It is not suitable for food having sodium benzoate as preservative as with increase in concentration growth of Lactococcus lactis decreases. Presence of bacteriocin hinders the growth of the isolate 1 as fresh weight of the mycelium in test sample is 7.09% less than the control. Being non-pathogenic this organism can be safely used against spoilage organisms in addition to food borne pathogens.

Enterocin P Causes Potassium Ion Efflux from Enterococcus faecium T136 Cells

OpenAIRE

Herranz, Carmen; Cintas, Luis M.; Hernández, Pablo E.; Moll, Gert N.; Driessen, Arnold J. M.

2001-01-01

Enterocin P is a bacteriocin produced by Enterococcus faecium P13. We studied the mechanism of its bactericidal action using enterocin-P-sensitive E. faecium T136 cells. The bacteriocin is incapable of dissipating the transmembrane pH gradient. On the other hand, depending on the buffer used, enterocin P dissipates the transmembrane potential. Enterocin P efficiently elicits efflux of potassium ions, but not of intracellularly accumulated anions like phosphate and glutamate. Taken together, t...

The Cyclic Antibacterial Peptide Enterocin AS-48: Isolation, Mode of Action, and Possible Food Applications

OpenAIRE

Grande Burgos, Mar?a Jos?; P?rez Pulido, Rub?n; L?pez Aguayo, Mar?a del Carmen; G?lvez, Antonio; Lucas, Rosario

2014-01-01

Enterocin AS-48 is a circular bacteriocin produced by Enterococcus. It contains a 70 amino acid-residue chain circularized by a head-to-tail peptide bond. The conformation of enterocin AS-48 is arranged into five alpha-helices with a compact globular structure. Enterocin AS-48 has a wide inhibitory spectrum on Gram-positive bacteria. Sensitivity of Gram-negative bacteria increases in combination with outer-membrane permeabilizing treatments. Eukaryotic cells are bacteriocin-resistant. This ...

Antimicrobial properties of indigenous Lactobacillus sakei strain

OpenAIRE

Vesković-Moračanin Slavica; Obradović D.; Velebit B.; Borović Branka; Škrinjar Marija; Turubatović L.

2010-01-01

The strain I 154 of Lactobacillus sakei has been isolated from traditionally fermented sausages in the course of the realization of the international project (INCO PROJECT No ICA4-CT-2002-10037). This strain exhibited the ability for bacteriocin production. Antimicrobial properties of the isolated bacteriocin (sakacine), its sensibility towards proteolytic enzymes, as well as the effect of increased to high temperatures on its stability have been examined in this work. Semi purified bacterioc...

Purification and Characterization of Suicin 65, a Novel Class I Type B Lantibiotic Produced by Streptococcus suis.

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Vaillancourt, Katy; LeBel, Geneviève; Frenette, Michel; Fittipaldi, Nahuel; Gottschalk, Marcelo; Grenier, Daniel

2015-01-01

Bacteriocins are antimicrobial peptides of bacterial origin that are considered as a promising alternative to the use of conventional antibiotics. Recently, our laboratory reported the purification and characterization of two lantibiotics, suicin 90-1330 and suicin 3908, produced by the swine pathogen and zoonotic agent Streptococcus suis (serotype 2). In this study, a novel bacteriocin produced by S. suis has been identified and characterized. The producing strain S. suis 65 (serotype 2) was found to belong to the sequence type 28, that includes strains known to be weakly or avirulent in a mouse model. The bacteriocin, whose production was only possible following growth on solid culture medium, was purified to homogeneity by cationic exchange and reversed-phase high-pressure liquid chromatography. The bacteriocin, named suicin 65, was heat, pH and protease resistant. Suicin 65 was active against all S. suis isolates tested, including antibiotic resistant strains. Amino acid sequencing of the purified bacteriocin by Edman degradation revealed the presence of modified amino acids suggesting a lantibiotic. Using the partial sequence obtained, a blast was performed against published genomes of S. suis and allowed to identify a putative lantibiotic locus in the genome of S. suis 89-1591. From this genome, primers were designed and the gene cluster involved in the production of suicin 65 by S. suis 65 was amplified by PCR. Sequence analysis revealed the presence of ten open reading frames, including a duplicate of the structural gene. The structural genes (sssA and sssA') of suicin 65 encodes a 25-amino acid residue leader peptide and a 26-amino acid residue mature peptide yielding an active bacteriocin with a deducted molecular mass of 3,005 Da. Mature suicin 65 showed a high degree of identity with class I type B lantibiotics (globular structure) produced by Streptococcus pyogenes (streptococcin FF22; 84.6%), Streptococcus macedonicus (macedocin ACA-DC 198; 84

Purification and Characterization of Suicin 65, a Novel Class I Type B Lantibiotic Produced by Streptococcus suis.

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Katy Vaillancourt

Full Text Available Bacteriocins are antimicrobial peptides of bacterial origin that are considered as a promising alternative to the use of conventional antibiotics. Recently, our laboratory reported the purification and characterization of two lantibiotics, suicin 90-1330 and suicin 3908, produced by the swine pathogen and zoonotic agent Streptococcus suis (serotype 2. In this study, a novel bacteriocin produced by S. suis has been identified and characterized. The producing strain S. suis 65 (serotype 2 was found to belong to the sequence type 28, that includes strains known to be weakly or avirulent in a mouse model. The bacteriocin, whose production was only possible following growth on solid culture medium, was purified to homogeneity by cationic exchange and reversed-phase high-pressure liquid chromatography. The bacteriocin, named suicin 65, was heat, pH and protease resistant. Suicin 65 was active against all S. suis isolates tested, including antibiotic resistant strains. Amino acid sequencing of the purified bacteriocin by Edman degradation revealed the presence of modified amino acids suggesting a lantibiotic. Using the partial sequence obtained, a blast was performed against published genomes of S. suis and allowed to identify a putative lantibiotic locus in the genome of S. suis 89-1591. From this genome, primers were designed and the gene cluster involved in the production of suicin 65 by S. suis 65 was amplified by PCR. Sequence analysis revealed the presence of ten open reading frames, including a duplicate of the structural gene. The structural genes (sssA and sssA' of suicin 65 encodes a 25-amino acid residue leader peptide and a 26-amino acid residue mature peptide yielding an active bacteriocin with a deducted molecular mass of 3,005 Da. Mature suicin 65 showed a high degree of identity with class I type B lantibiotics (globular structure produced by Streptococcus pyogenes (streptococcin FF22; 84.6%, Streptococcus macedonicus (macedocin ACA

Production of Enterocins L50A, L50B, and IT, a New Enterocin, by Enterococcus faecium IT62, a Strain Isolated from Italian Ryegrass in Japan▿

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Izquierdo, Esther; Bednarczyk, Audrey; Schaeffer, Christine; Cai, Yimin; Marchioni, Eric; Van Dorsselaer, Alain; Ennahar, Saïd

2008-01-01

Enterococcus faecium IT62, isolated from ryegrass in Japan, was shown to produce three different bacteriocins, two of which had molecular masses and amino acid sequences that corresponded to those of enterocin L50A and enterocin L50B. These peptides existed, however, as chemically modified forms that were either N formylated or N formylated and oxidized at Met24. The third bacteriocin, named enterocin IT, had a molecular mass of 6,390 Da, was made up of 54 amino acids, and did not correspond to any known bacteriocin. However, enterocin IT was identical to the C-terminal part of the 16-amino-acid-longer bacteriocin 32 (T. Inoue, H. Tomita, and Y. Ike, Antimicrob. Agents Chemother., 50:1202-1212, 2006). For the first time, the antimicrobial activity spectra for enterocins L50A and L50B were determined separately and included a wide range of gram-positive bacteria but also a few gram-negative strains that were weakly sensitive. Slight differences in the activities of enterocins L50A and L50B were observed, as gram-positive bacteria showed an overall higher level of sensitivity to L50A than to L50B, as opposed to gram-negative ones. Conversely, enterocin IT showed a very narrow antimicrobial spectrum that was limited to E. faecium strains, one strain of Bacillus subtilis, and one strain of Lactococcus lactis. This study showed that E. faecium IT62, a grass-borne strain, produces bacteriocins with very different activity features and structures that may be found in strains associated with food or those of clinical origin, which demonstrates that a particular enterocin structure may be widespread and not related to the producer's origin. PMID:18391036

Production of enterocins L50A, L50B, and IT, a new enterocin, by Enterococcus faecium IT62, a strain isolated from Italian ryegrass in Japan.

Science.gov (United States)

Izquierdo, Esther; Bednarczyk, Audrey; Schaeffer, Christine; Cai, Yimin; Marchioni, Eric; Van Dorsselaer, Alain; Ennahar, Saïd

2008-06-01

Enterococcus faecium IT62, isolated from ryegrass in Japan, was shown to produce three different bacteriocins, two of which had molecular masses and amino acid sequences that corresponded to those of enterocin L50A and enterocin L50B. These peptides existed, however, as chemically modified forms that were either N formylated or N formylated and oxidized at Met(24). The third bacteriocin, named enterocin IT, had a molecular mass of 6,390 Da, was made up of 54 amino acids, and did not correspond to any known bacteriocin. However, enterocin IT was identical to the C-terminal part of the 16-amino-acid-longer bacteriocin 32 (T. Inoue, H. Tomita, and Y. Ike, Antimicrob. Agents Chemother., 50:1202-1212, 2006). For the first time, the antimicrobial activity spectra for enterocins L50A and L50B were determined separately and included a wide range of gram-positive bacteria but also a few gram-negative strains that were weakly sensitive. Slight differences in the activities of enterocins L50A and L50B were observed, as gram-positive bacteria showed an overall higher level of sensitivity to L50A than to L50B, as opposed to gram-negative ones. Conversely, enterocin IT showed a very narrow antimicrobial spectrum that was limited to E. faecium strains, one strain of Bacillus subtilis, and one strain of Lactococcus lactis. This study showed that E. faecium IT62, a grass-borne strain, produces bacteriocins with very different activity features and structures that may be found in strains associated with food or those of clinical origin, which demonstrates that a particular enterocin structure may be widespread and not related to the producer's origin.

Strategies for Pathogen Biocontrol Using Lactic Acid Bacteria and Their Metabolites: A Focus on Meat Ecosystems and Industrial Environments

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Patricia Castellano

2017-07-01

Full Text Available The globalization of trade and lifestyle ensure that the factors responsible for the emergence of diseases are more present than ever. Despite biotechnology advancements, meat-based foods are still under scrutiny because of the presence of pathogens, which causes a loss of consumer confidence and consequently a fall in demand. In this context, Lactic Acid Bacteria (LAB as GRAS organisms offer an alternative for developing pathogen-free foods, particularly avoiding Listeria monocytogenes, with minimal processing and fewer additives while maintaining the foods’ sensorial characteristics. The use of LAB strains, enabling us to produce antimicrobial peptides (bacteriocins in addition to lactic acid, with an impact on quality and safety during fermentation, processing, and/or storage of meat and ready-to-eat (RTE meat products, constitutes a promising tool. A number of bacteriocin-based strategies including the use of bioprotective cultures, purified and/or semi-purified bacteriocins as well as their inclusion in varied packaging materials under different storage conditions, have been investigated. The application of bacteriocins as part of hurdle technology using non-thermal technologies was explored for the preservation of RTE meat products. Likewise, considering that food contamination with L. monocytogenes is a consequence of the post-processing manipulation of RTE foods, the role of bacteriocinogenic LAB in the control of biofilms formed on industrial surfaces is also discussed.

Strategies for Pathogen Biocontrol Using Lactic Acid Bacteria and Their Metabolites: A Focus on Meat Ecosystems and Industrial Environments

Science.gov (United States)

Castellano, Patricia; Pérez Ibarreche, Mariana; Fontana, Cecilia; Vignolo, Graciela M.

2017-01-01

The globalization of trade and lifestyle ensure that the factors responsible for the emergence of diseases are more present than ever. Despite biotechnology advancements, meat-based foods are still under scrutiny because of the presence of pathogens, which causes a loss of consumer confidence and consequently a fall in demand. In this context, Lactic Acid Bacteria (LAB) as GRAS organisms offer an alternative for developing pathogen-free foods, particularly avoiding Listeria monocytogenes, with minimal processing and fewer additives while maintaining the foods’ sensorial characteristics. The use of LAB strains, enabling us to produce antimicrobial peptides (bacteriocins) in addition to lactic acid, with an impact on quality and safety during fermentation, processing, and/or storage of meat and ready-to-eat (RTE) meat products, constitutes a promising tool. A number of bacteriocin-based strategies including the use of bioprotective cultures, purified and/or semi-purified bacteriocins as well as their inclusion in varied packaging materials under different storage conditions, have been investigated. The application of bacteriocins as part of hurdle technology using non-thermal technologies was explored for the preservation of RTE meat products. Likewise, considering that food contamination with L. monocytogenes is a consequence of the post-processing manipulation of RTE foods, the role of bacteriocinogenic LAB in the control of biofilms formed on industrial surfaces is also discussed. PMID:28696370

Antimicrobial activity of lactic acid bacteria isolated from bekasam against staphylococcus aureus ATCC 25923, escherichia coli ATCC 25922, and salmonella sp

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Sari, Melia; Suryanto, Dwi; Yurnaliza

2018-03-01

Bekasam is an Indonesian fermented food made of fish. As a fermented food, this food may contain some beneficial bacteria like lactic acid bacteria (LAB), which usually have antimicrobial properties such as organic acid, hydrogen peroxide, and a bacteriocin. A study on antimicrobial activity of LAB isolated from bekasam against some pathogenic bacteria has been conducted. The purpose of this study was to know the ability of crude bacteriocin produced LAB of bekasam against Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Salmonella sp. Bekasam sample was taken from South Sumatera. LAB isolation was done using de Man Rogosa and Sharpe agar. A bacterial colony with clear zone was selected and purified to get a single colony. The antagonistic assay of the LAB was conducted in Muller-Hinton agar Selected isolates with higher clearing zone were assayed for antibacterial effect of their crude bacteriocin of different culture incubation time of 6, 9, and 12 hours. The results showed that the crude extract bacteriocin of isolate MS2 of 9 hours culture incubation time inhibited more in Staphylococcus aureus ATCC 25923 with inhibition zone of 13.1 mm, whereas isolate MS9 of 9 hours culture incubation time inhibited more in Escherichia coli ATCC 25922 and Salmonella sp. with inhibition zone of 12.7 and 7.3 mm, respectively.

Action des cultures protectrices : cas des germes lactiques sur la flore alimentaire indésirable

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Privat, K.

2011-01-01

Full Text Available Protective action of cultures: the case of lactic bacteria against undesirable food flora. Protective strains have been used for centuries in the fermentation of a variety of food. The preservative ability of these strains in food is attributed to the production of antimicrobial metabolites including organic acids and bacteriocins. But to secure an extension of shelf life and better food safety, the multiple antimicrobial barriers approach is required. This approach typically uses the principal hurdles temperature (higher or lower, water activity (Aw, pH, redox potentiel (Eh, chemical preservatives, vacuum packaging, modified atmosphere, high hydrostatic pressure (HHP and UV, as additional barriers to protective cultures and/or antimicrobial compounds. This paper reviews the recent literature describing the protective cultures and different ways they use to suppress unwanted flora. Are also briefly presented, classification, biosynthesis, mechanism of action of bacteriocins produced by these cultures and finally the protective benefit of using additional barriers to booster protective cultures and/or their bacteriocins.

Evaluation of Antimicrobial Activity of Probiotic Lactobacillus Strains against Growth and Urease Activity of Proteus spp.

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Leila Goudarzi

2017-10-01

Full Text Available Background:    Nowadays, the use of probiotic bacteria for the prevention and treatment of urinary tract infections is growing. Lactobacillus, as probiotic bacterial genus, is well known for its benefits for the human health.Methods:      The effects of partially purified antimicrobial compounds (bacteriocins and biosurfactants of Lactobacillus strains was assessed and their capacity to in vitro inhibit growth and urease production of various strains of Proteus spp, was studied. Inhibition of the urease production of Proteus spp. at sub-MIC levels was screened using spectrophotometry method.  Results:   Results revealed that semi-purified bacteriocins of L. acidophilus and L. plantarum showed a greater inhibitory activity on the bacterial urease, compared to biosurfactants of L. rhamnosus, L. casei and L. fermentum (P < 0.05.Conclusion:    It can be concluded that bacteriocins may affect Proteus pathogenesis by inhibition of the bacterial urease activity and therefore eliminate the stone formation by these bacteria.

Effects of Different Spices Used in Production of Fermented Sausages on Growth of and Curvacin A Production by Lactobacillus curvatus LTH 1174

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Verluyten, Jurgen; Leroy, Frédéric; de Vuyst, Luc

2004-01-01

Lactobacillus curvatus LTH 1174, a fermented sausage isolate, produces the listericidal bacteriocin curvacin A. The effect of different spices relevant for the production of fermented sausages was investigated in vitro through laboratory fermentations with a meat simulation medium and an imposed pH profile relevant for Belgian-type fermented sausages. The influence on the growth characteristics and especially on the kinetics of curvacin A production with L. curvatus LTH 1174 was evaluated. Pepper, nutmeg, rosemary, mace, and garlic all decreased the maximum specific growth rate, while paprika was the only spice that increased it. The effect on the lag phase was minor except for nutmeg and especially for garlic, which increased it, yet garlic was stimulatory for biomass production. The maximum attainable biomass concentration (Xmax) was severely decreased by the addition of 0.40% (wt/vol) nutmeg, while 0.35% (wt/vol) garlic or 0.80% (wt/vol) white pepper increased Xmax. Nutmeg decreased both growth and bacteriocin production considerably. Garlic was the only spice enhancing specific bacteriocin production, resulting in higher bacteriocin activity in the cell-free culture supernatant. Finally, lactic acid production was stimulated by the addition of pepper, and this was not due to the manganese present because an amount of manganese that was not growth limiting was added to the growth medium. Addition of spices to the sausage mixture is clearly a factor that will influence the effectiveness of bacteriocinogenic starter cultures in fermented-sausage manufacturing. PMID:15294818

Effects of different spices used in production of fermented sausages on growth of and curvacin A production by Lactobacillus curvatus LTH 1174.

Science.gov (United States)

Verluyten, Jurgen; Leroy, Frédéric; De Vuyst, Luc

2004-08-01

Lactobacillus curvatus LTH 1174, a fermented sausage isolate, produces the listericidal bacteriocin curvacin A. The effect of different spices relevant for the production of fermented sausages was investigated in vitro through laboratory fermentations with a meat simulation medium and an imposed pH profile relevant for Belgian-type fermented sausages. The influence on the growth characteristics and especially on the kinetics of curvacin A production with L. curvatus LTH 1174 was evaluated. Pepper, nutmeg, rosemary, mace, and garlic all decreased the maximum specific growth rate, while paprika was the only spice that increased it. The effect on the lag phase was minor except for nutmeg and especially for garlic, which increased it, yet garlic was stimulatory for biomass production. The maximum attainable biomass concentration (X(max)) was severely decreased by the addition of 0.40% (wt/vol) nutmeg, while 0.35% (wt/vol) garlic or 0.80% (wt/vol) white pepper increased X(max). Nutmeg decreased both growth and bacteriocin production considerably. Garlic was the only spice enhancing specific bacteriocin production, resulting in higher bacteriocin activity in the cell-free culture supernatant. Finally, lactic acid production was stimulated by the addition of pepper, and this was not due to the manganese present because an amount of manganese that was not growth limiting was added to the growth medium. Addition of spices to the sausage mixture is clearly a factor that will influence the effectiveness of bacteriocinogenic starter cultures in fermented-sausage manufacturing.

Enterocin P Causes Potassium Ion Efflux from Enterococcus faecium T136 Cells

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Herranz, Carmen; Cintas, Luis M.; Hernández, Pablo E.; Moll, Gert N.; Driessen, Arnold J. M.

2001-01-01

Enterocin P is a bacteriocin produced by Enterococcus faecium P13. We studied the mechanism of its bactericidal action using enterocin-P-sensitive E. faecium T136 cells. The bacteriocin is incapable of dissipating the transmembrane pH gradient. On the other hand, depending on the buffer used, enterocin P dissipates the transmembrane potential. Enterocin P efficiently elicits efflux of potassium ions, but not of intracellularly accumulated anions like phosphate and glutamate. Taken together, these data demonstrate that enterocin P forms specific, potassium ion-conducting pores in the cytoplasmic membrane of target cells. PMID:11181377

Rice Hull Ash and Silicic Acid as Adsorbents for Concentration of Bacteriocins†

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Janes, M. E.; Nannapaneni, R.; Proctor, A.; Johnson, M. G.

1998-01-01

A model procedure has been developed for the rapid extraction of five bacteriocins (nisin, pediocin RS2, leucocin BC2, lactocin GI3, and enterocin CS1) from concentrated freeze-dried crude culture supernatants by adsorption onto acid or alkaline rice hull ash (RHA) or silicic acid (SA). Bacteriocins were adsorbed onto RHA or SA by a pH-dependent method and desorbed by decreasing the pH to 2.5 or 3.0 and heating at 90°C for 5 min. The maximum adsorption and optimal pH range for different bacte...

Efecto de las condiciones de crecimiento y composición del medio de cultivo sobre la producción de bacteriocina de Enterococcus mundtii Tw56

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Marisol Vallejo

2014-07-01

Full Text Available Título en español: Efecto de las condiciones de crecimiento y composición del medio de cultivo sobre la producción de bacteriocina de Enterococcus mundtii Tw56 Título corto: Efecto de las condiciones de crecimiento y composición del medio de cultivo Título en ingles: Effect of growth conditions and culture medium composition on bacteriocin production by Enterococcus mundtii Tw56 Resumen: Enterococcus mundtii Tw56 es una cepa productora de bacteriocina que fue aislada del contenido intestinal de pejerrey (Odontesthes sp.. El objetivo del presente trabajo fue determinar los factores fisicoquímicos y la composición del medio de cultivo para lograr un mayor rendimiento de células viables y producción de bacteriocina. No se observaron cambios en la producción del antimicrobiano cuando la glucosa fue sustituida por fructosa o maltosa en la formulación del medio MRS. Por el contrario, la mayor actividad de las bacteriocinas se obtuvo cuando se utilizó el extracto de carne como fuente única de nitrógeno. Mientras que la máxima biomasa se alcanzó a 35 ºC, las temperaturas óptimas para la producción de bacteriocina se observaron a 25 y30 ºC. El pH inicial óptimo para el crecimiento celular y bioactividad fue 6,5, ambos parámetros disminuyeron cuando la experiencia comenzó a pH 6,0 o 5,5.  La formación de biomasa y la producción de bacteriocina disminuyeron en presencia de cloruro de sodio. La cepa comenzó a producir bacteriocina en la fase exponencial tardía. La actividad aumentó en función de la masa celular y alcanzó el máximo al final de la fase exponencial (12 h. Una disminución de la actividad antimicrobiana se observó en la fase estacionaria (16 h, posiblemente debido a la degradación por enzimas proteolíticas.  Palabras clave: Enterococcus mundtii Tw56, bacteriocina, factores fisicoquímicos, medio de cultivo.  Abstract:  Enterococcus mundtii Tw56 is a bacteriocin-producing strain that was isolated from

Co-expression of Nisin Z and Leucocin C as a Basis for Effective Protection Against Listeria monocytogenes in Pasteurized Milk

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Yuxin Fu

2018-03-01

Full Text Available Nisin, an important bacteriocin from Lactococcus lactis subsp., is primarily active against various Gram-positive bacteria. Leucocin C, produced by Leuconostoc carnosum 4010, is a class IIa bacteriocin used to inhibit the growth of Listeria monocytogenes. Because two bacteriocins have different modes of action, the combined use of them could be a potential strategy for effective inhibition of foodborne pathogens. In this study, L. lactis N8-r-lecCI (N8 harboring lecCI gene coexpressing nisin–leucocin C was constructed based on the food-grade carrier L. lactis N8. Production of both bacteriocins was stably maintained. Antimicrobial measurements showed that the recombinant strain is effectively against Listeria monocytogenes and Staphylococcus aureus and moderately against Salmonella enterica serovar Enteritidis and Escherichia coli because of its stronger antibacterial activity than the parental strain, this result first demonstrated that the co-expression of nisin and leucocin C results in highly efficient antimicrobial activity. The checkerboard assay showed that the antibacterial activity of L. lactis N8-r-lecCI supernatant was enhanced in the presence of low concentration of EDTA. Analysis of the scanning electron microscope image showed the biggest cellular morphology change in L. monocytogenes treated with a mixture of EDTA and L. lactis N8-r-lecCI supernatant. The practical effect was verified in pasteurized milk through time-kill assay. The L. lactis N8-r-lecCI strain expressing both nisin and leucocin C has a promising application prospect in pasteurized milk processing and preservation because of its strong antibacterial activity.

Characterization of a Multipeptide Lantibiotic Locus in Streptococcus pneumoniae

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Natalie Maricic

2016-01-01

Full Text Available Bacterial communities are established through a combination of cooperative and antagonistic interactions between the inhabitants. Competitive interactions often involve the production of antimicrobial substances, including bacteriocins, which are small antimicrobial peptides that target other community members. Despite the nearly ubiquitous presence of bacteriocin-encoding loci, inhibitory activity has been attributed to only a small fraction of gene clusters. In this study, we characterized a novel locus (the pld locus in the pathogen Streptococcus pneumoniae that drives the production of a bacteriocin called pneumolancidin, which has broad antimicrobial activity. The locus encodes an unusual tandem array of four inhibitory peptides, three of which are absolutely required for antibacterial activity. The three peptide sequences are similar but appear to play distinct roles in regulation and inhibition. A modification enzyme typically found in loci encoding a class of highly modified bacteriocins called lantibiotics was required for inhibitory activity. The production of pneumolancidin is controlled by a two-component regulatory system that is activated by the accumulation of modified peptides. The locus is located on a mobile element that has been found in many pneumococcal lineages, although not all elements carry the pld genes. Intriguingly, a minimal region containing only the genes required for pneumolancidin immunity was found in several Streptococcus mitis strains. The pneumolancidin-producing strain can inhibit nearly all pneumococci tested to date and provided a competitive advantage in vivo. These peptides not only represent a unique strategy for bacterial competition but also are an important resource to guide the development of new antimicrobials.

Novel Bacteriocinogenic Lactobacillus plantarum Strains and Their Differentiation by Sequence Analysis of 16S rDNA, 16S-23S and 23S-5S Intergenic Spacer Regions and Randomly Amplified Polymorphic DNA Analysis

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Morteza Shojaei Moghadam

2010-01-01

Full Text Available Six strains of bacteriocinogenic Lactobacillus plantarum (TL1, RG11, RS5, UL4, RG14 and RI11 isolated from Malaysian foods were investigated for their structural bacteriocin genes. A new combination of plantaricin EF and plantaricin W bacteriocin structural genes was successfully amplified from all studied strains, suggesting that they were novel bacteriocin-producing L. plantarum strains. A four-base pair variable region was detected in the short 16S-23S intergenic spacer regions of the studied strains by a comparative analysis with 17 L. plantarum strains deposited in the GenBank, implying they were new genotypes. The studied L. plantarum strains were subsequently differentiated into four groups on the basis of the detected four-base pair variable region of the short 16S-23S intergenic spacer region. Further analysis of the DNA sequence of 23S-5S intergenic spacer region revealed only one type of 23S-5S intergenic spacer region present in the studied strains, indicating it was highly conserved among the studied L. plantarum strains. Three randomly amplified polymorphic DNA experiments using three different combinations of arbitrary primers successfully differentiated the studied L. plantarum strains from each other, confirming they were different strains. In conclusion, the studied L. plantarum strains were shown to be novel bacteriocin producers and high level of strain discrimination could be achieved with a combination of randomly amplified polymorphic DNA analysis and the analysis of the variable region of short 16S-23S intergenic spacer region present in L. plantarum strains.

Antimicrobial Activity – The Most Important Property of Probiotic and Starter Lactic Acid Bacteria

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Blaženka Kos

2010-01-01

Full Text Available The antimicrobial activity of industrially important lactic acid bacteria as starter cultures and probiotic bacteria is the main subject of this review. This activity has been attributed to the production of metabolites such as organic acids (lactic and acetic acid, hydrogen peroxide, ethanol, diacetyl, acetaldehyde, acetoine, carbon dioxide, reuterin, reutericyclin and bacteriocins. The potential of using bacteriocins of lactic acid bacteria, primarily used as biopreservatives, represents a perspective, alternative antimicrobial strategy for continuously increasing problem with antibiotic resistance. Another strategy in resolving this problem is an application of probiotics for different gastrointestinal and urogenital infection therapies.

Enterocin P Selectively Dissipates the Membrane Potential of Enterococcus faecium T136

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Herranz, C.; Chen, Y.; Chung, H.-J.; Cintas, L. M.; Hernández, P. E.; Montville, T. J.; Chikindas, M. L.

2001-01-01

Enterocin P is a pediocin-like, broad-spectrum bacteriocin which displays a strong inhibitory activity against Listeria monocytogenes. The bacteriocin was purified from the culture supernatant of Enterococcus faecium P13, and its molecular mechanism of action against the sensitive strain E. faecium T136 was evaluated. Although enterocin P caused significant reduction of the membrane potential (ΔΨ) and the intracellular ATP pool of the indicator organism, the pH gradient (ΔpH) component of the proton motive force (Δp) was not dissipated. By contrast, enterocin P caused carboxyfluorescein efflux from E. faecium T136-derived liposomes. PMID:11282622

Characterization and Heterologous Expression of the Genes Encoding Enterocin A Production, Immunity, and Regulation in Enterococcus faecium DPC1146

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O’Keeffe, Triona; Hill, Colin; Ross, R. Paul

1999-01-01

Enterocin A is a small, heat-stable, antilisterial bacteriocin produced by Enterococcus faecium DPC1146. The sequence of a 10,879-bp chromosomal region containing at least 12 open reading frames (ORFs), 7 of which are predicted to play a role in enterocin biosynthesis, is presented. The genes entA, entI, and entF encode the enterocin A prepeptide, the putative immunity protein, and the induction factor prepeptide, respectively. The deduced proteins EntK and EntR resemble the histidine kinase and response regulator proteins of two-component signal transducing systems of the AgrC-AgrA type. The predicted proteins EntT and EntD are homologous to ABC (ATP-binding cassette) transporters and accessory factors, respectively, of several other bacteriocin systems and to proteins implicated in the signal-sequence-independent export of Escherichia coli hemolysin A. Immediately downstream of the entT and entD genes are two ORFs, the product of one of which, ORF4, is very similar to the product of the yteI gene of Bacillus subtilis and to E. coli protease IV, a signal peptide peptidase known to be involved in outer membrane lipoprotein export. Another potential bacteriocin is encoded in the opposite direction to the other genes in the enterocin cluster. This putative bacteriocin-like peptide is similar to LafX, one of the components of the lactacin F complex. A deletion which included one of two direct repeats upstream of the entA gene abolished enterocin A activity, immunity, and ability to induce bacteriocin production. Transposon insertion upstream of the entF gene also had the same effect, but this mutant could be complemented by exogenously supplied induction factor. The putative EntI peptide was shown to be involved in the immunity to enterocin A. Cloning of a 10.5-kb amplicon comprising all predicted ORFs and regulatory regions resulted in heterologous production of enterocin A and induction factor in Enterococcus faecalis, while a four-gene construct (entAITD) under the

Characterization of Pediococcus acidilactici strains isolated from rainbow trout (Oncorhynchus mykiss) feed and larvae: safety, DNA fingerprinting, and bacteriocinogenicity.

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Araújo, Carlos; Muñoz-Atienza, Estefanía; Poeta, Patrícia; Igrejas, Gilberto; Hernández, Pablo E; Herranz, Carmen; Cintas, Luis M

2016-05-03

The use of lactic acid bacteria (LAB) as probiotics constitutes an alternative or complementary strategy to chemotherapy and vaccination for disease control in aquaculture. The objectives of this work were (1) the in vitro safety assessment of 8 Pediococcus acidilactici strains isolated from rainbow trout (Oncorhynchus mykiss, Walbaum) feed and larvae; (2) the evaluation of their genetic relatedness; (3) the study of their antimicrobial/bacteriocin activity against fish pathogens; and (4) the biochemical and genetic characterization of the bacteriocin produced by the strain displaying the greatest antimicrobial activity. Concerning the safety assessment, none of the pediococci showed antibiotic resistance nor produced hemolysin or gelatinase, degraded gastric mucin, or deconjugated bile salts. Four strains (50%) produced tyramine or putrescine, but the corresponding genes were not amplified by PCR. Enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR) fingerprinting allowed clustering of the pediococci into 2 well-defined groups (68% similarity). From the 8 pediococci displaying direct antimicrobial activity against at least 3 out of 9 fish pathogens, 6 strains (75%) were identified as bacteriocin producers. The bacteriocin produced by P. acidilactici L-14 was purified, and mass spectrometry and DNA sequencing revealed its identity to pediocin PA-1 (PedPA-1). Altogether, our results allowed the identification of 4 (50%) putatively safe pediococci, including 2 bacteriocinogenic strains. ERIC-PCR fingerprinting was a valuable tool for genetic profiling of P. acidilactici strains. This work reports for the first time the characterization of a PedPA-1-producing P. acidilactici strain isolated from an aquatic environment (rainbow trout larvae), which shows interesting properties related to its potential use as a probiotic in aquaculture.

Bio-physicochemical characterization and applied studies of carotovoricin na5 (crna5) on blb affected rice plants

International Nuclear Information System (INIS)

Jabeen, N.; Rasool, S.A.; Naz, S.A.

2014-01-01

Erwinia carotovora is a common soil borne plant pathogen, which generally infects plants of family Solanacea. In the present study, bacteriocin (CrNA5), produced by an indigenously isolated E. carotovora NA5 has been characterized and its possible anti phytopathogenic potential was shown in the field studies. CrNA5 showed its antimicrobial activity against many gram-positive and gram-negative bacteria including those associated with the plant diseases. The bacteriocin showed substantial stability against wide range of temperatures and pH. Additionally, it was also found resistant to the treatment of metal ions, organic solvents and non-proteolytic enzymes. Conversely, its inactivation by proteinase K and protease suggested its protein nature. Mode of action studies revealed that CrNA5 is bactericidal, particularly against Xanthomonas oryzae oryzae. The electron micrograph of CrNA5 revealed spherical particle (empty head) like structures implicating the vestigial bacteriophage based origin of carotovoricin. In silico analyses were also conducted in order to deduce the plausible ratio of the amino acids present in the protein. The In vivo experiments showed the efficacy of CrNA5 against X. oryzae oryzae (Xoo), the causative agent of bacterial leaf blight (BLB) of rice, both in controlled conditions (green house) as well as in field trials. To the best of our knowledge, the present study is the first of its kind with the bacteriocin of Erwinia origin (tested against the BLB infected plants in the field). It is expected that the present study will help visit new insights of the bacteriocins produced by Erwinia carotovora and their potential (application) as anti phytopathogenic agent. (author)

Antimicrobial properties of lactic acid bacteria isolated from uruguayan artisan cheese

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Martín Fraga Cotelo

2013-12-01

Full Text Available Uruguayan artisan cheese is elaborated with raw milk and non-commercial starters. The associated native microbiota may include lactic acid bacteria and also potentially pathogenic bacteria. Lactic acid bacteria were isolated from artisan cheese, raw milk, and non-commercial starter cultures, and their potential bacteriocin production was assessed. A culture collection of 509 isolates was obtained, and five isolates were bacteriocin-producers and were identified as Enterococcus durans,Lactobacillus casei, and Lactococcus lactis. No evidence of potential virulence factors were found in E. durans strains. These are promising results in terms of using these native strains for cheese manufacture and to obtain safe products.

The Impact of the Antimicrobial Compounds Produced by Lactic Acid Bacteria on the Growth Performance of Mycobacterium avium subsp. paratuberculosis

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Petr Kralik

2018-04-01

Full Text Available Cell-free supernatants (CFSs extracted from various lactic acid bacteria (LAB cultures were applied to Mycobacterium avium subsp. paratuberculosis (MAP cells to determine their effect on MAP viability. In addition, 5% lactic acid (LA; pH 3 and commercially synthetized nisin bacteriocin were also tested. This procedure was chosen in order to mimic the influence of LAB compounds during the production and storage of fermented milk products, which can be contaminated by MAP. Its presence in milk and milk products is of public concern due to the possible ingestion of MAP by consumers and the discussed role of MAP in Crohn’s disease. Propidium monoazide real-time PCR (PMA qPCR was used for viability determination. Although all CFS showed significant effects on MAP viability, two distinct groups of CFS – effective and less effective – could be distinguished. The effective CFSs were extracted from various lactobacilli cultures, their pH values were mostly lower than 4.5, and their application resulted in >2 log10 reductions in MAP viability. The group of less effective CFS were filtered from Lactococcus and enterococci cultures, their pH values were higher than 4.5, and their effect on MAP viability was <2 log10. LA elicited a reduction in MAP viability that was similar to that of the group of less effective CFS. Almost no effect was found when using commercially synthetized nisin at concentrations of 0.1–1000 μg/ml. A combination of the influence of the type of bacteriocin, the length of its action, bacteriocin production strain, and pH are all probably required for a successful reduction in MAP viability. However, certain bacteriocins and their respective LAB strains (Lactobacillus sp. appear to play a greater role in reducing the viability of MAP than pH.

Combined effect of enterocin and lipase from Enterococcus faecium NCIM5363 against food borne pathogens: mode of action studies.

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Ramakrishnan, Vrinda; Narayan, Bhaskar; Halami, Prakash M

2012-08-01

Food borne diseases have a major impact on public health whose epidemiology is rapidly changing. The whole cells of pathogens involved or their toxins/metabolites affect the human health apart from spoiling sensory properties of the food products finally affecting the food industry as well as consumer health. With pathogens developing mechanisms of antibiotic resistance, there has been an increased need to replace antibiotics as well as chemical additives with naturally occurring bacteriocins. Bacteriocins are known to act mainly against Gram-positive pathogens and with little or no effect towards Gram-negative enteric bacteria. In the present study, combination effect of lipase and bacteriocin produced by Enterococcus faecium NCIM5363, a highly lipolytic lactic acid bacterium against various food pathogens was assessed. The lipase in combination with enterocin exhibited a lethal effect against Gram-negative pathogens. Scanning electron microscopy studies carried out to ascertain the constitutive mode of action of lipase and enterocin revealed that the lipase degrades the cell wall of Gram-negative bacteria and creates a pore through which enterocin enters thereby resulting in cell death. The novelty of this work is the fact that this is the first report revealing the synergistic effect of lipase with enterocin against Gram-negative bacteria.

Purification and characterization of enterocin FH 99 produced by a faecal isolate Enterococcus faecium FH 99.

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Gupta, H; Malik, R K; Bhardwaj, A; Kaur, G; De, S; Kaushik, J K

2010-06-01

Enterococcus faecium FH 99 was isolated from human faeces and selected because of its broad spectrum of inhibitory activity against several Gram-positive foodborne spoilage and pathogenic bacteria. Ent. faecium FH 99 accumulates enterocin in large number in early stationary phase of the growth. The enterocin FH 99 was stable over a wide pH range (2-10) and recovered activity even after treatment at high temperatures (10 min at 100°C). The enterocin was subjected to different purification techniques viz., gel filteration, cation exchange chromatography and reverse-phase high-performance liquid chromatography. The activity was eluted as one individual active fraction. SDSPAGE revealed a molecular weight of less than 6.5 kDa. Studies carried out to identify the genetic determinants for bacteriocin production showed that this trait may be plasmid encoded as loss in both of the plasmids (size>chromosomal DNA) led to loss in bacteriocin production by Ent. faecium FH 99. Ent. faecium strain FH 99 is a newly discovered high bacteriocin producer with Activity Units 1.8 × 10(5) AU ml(-1) and its characteristics indicate that it may have strong potential for application as a protective agent against pathogens and spoilage bacteria in foods.

Assay of enterocin AS-48 for inhibition of foodborne pathogens in desserts.

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Martinez Viedma, Pilar; Abriouel, Hikmate; Ben Omar, Nabil; Lucas López, Rosario; Valdivia, Eva; Gálvez, Antonio

2009-08-01

Enterocin AS-48 was tested against Staphylococcus aureus, Bacillus cereus, and Listeria monocytogenes in different kinds of desserts. The highest activity against S. aureus was detected in baker cream. However, in yogurt-type soy-based desserts and in gelatin pudding, AS-48 (175 arbitrary units [AU]/g) reduced viable cell counts of S. aureus by only 1.5 to 1.8 log units at most. The efficacy of AS-48 in puddings greatly depended on inoculum size, and viable S. aureus counts decreased below detection levels within 24 h for inocula lower than 4 to 5.5 log CFU/g. For L. monocytogenes, bacteriocin concentrations of 52.5 to 87.5 AU/g reduced viable counts below detection levels and avoided regrowth of survivors. The lowest activity was detected in yogurt-type desserts. For B. cereus, viable cell counts were reduced below detection levels for bacteriocin concentrations of 52.5 AU/g in instant pudding without soy or by 175 AU/g in the soy pudding. In gelatin pudding, AS-48 (175 AU/g) reduced viable cell counts of B. cereus below detection levels after 8 h at 10 degrees C or after 48 h at 22 degrees C. Bacteriocin addition also inhibited gelatin liquefaction caused by the proteolytic activity of B. cereus.

[Influence of staphylococcin T on Enterococcus sp. growth].

Science.gov (United States)

Białucha, Agata; Kozuszko, Sylwia; Gospodarek, Eugenia; Bugalski, Roman Marian; Gierlotka, Krzysztof

2007-01-01

Bacteriocins are ribosomally synthesised, extracellular bacterial products. Generally, spectrum of inhibition is limited to the same or closely related species to bacteriocin producer. Staphylococcin T is produced by Staphylococcus cohnii strain. The present study concerns influence of StT to 267 Enterococcus sp. strains growth isolated between 2003 and 2006 in Department of Microbiology University Hospital of dr. A. Jurasz in Bydgoszcz. S. cohnii T antagonistic ability evaluated towards bacteries on Mueller-Hinton Agar (bio Mérieux) in aerobic conditions. After 24 and 48 hours tested enterococci suspensions were plated perpendiculary. Susceptibility to antibiotics was assessed by disc diffusion method according to the guideless of Clinical and Laboratory Standards Institute and National Reference Centre for Antimicrobial Susceptibility. Among Enterococcus sp. strains tested 7.1% were sensitive to StT. The highest percentage of sensitive enterococci isolated from wound swabs, urine, blood and pus. Enterococcus faecium strains dominated (63.2%) among enterococci sensitive to StT. Moderate inhibition degree on S. cohnii T bacteriocin action was observed in majority sensitive enterococci strains. Enterococcus sp. sensitive to StT strains were frequently multidrug resistant (68.4%). According to the study results and increasing resistance to antibiotics, StT could be an alternative agent used to treat infections caused by Enterococcus sp.

Les produits de la mer au Sénégal et le potentiel des bactéries lactiques et des bactériocines pour la conservation

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Diop, MB.

2010-01-01

Full Text Available Fish preservation in Senegal: potential use of lactic acid bacteria and their antibacterial metabolites. In Senegal, fish are first as staple animal protein foods for populations. Socioeconomic constraints hamper the development of industrial preservations. Traditional fish production has some inherent food safety concerns considering the high susceptibility of seafood to bacterial spoilage. Some strategies of seafood products preservation using lactic acid bacteria producing bacteriocin are not very expensive and do not consume energy very much. They can be adapted in the preservation of fish in Senegal particularly in the fermentation in which the catches, generally handled for many hours at ambient temperature, are only added salt (NaCl and sun dried. Lactic acid bacteria producing bacteriocins could be used as starters (with addition of carbohydrates during the preparation of fish for rendering them more resistant to the growth of spoilage and pathogenic bacteria by in situ production of organic acids and bacteriocins. A second alternative of application of these bacteria is to use bactericidal solutions obtained from their cultures as additional barrier to sodium chloride to prevent growth of the flora during fish storage for maturation. These new strategies of preparation combined with drying can be used to enhance microbiological and dietetic qualities of local seafood commodities.

Cloning and Expression of Plantaricin W Produced by Lactobacillus plantarum U10 Isolate from "Tempoyak" Indonesian Fermented Food as Immunity Protein in Lactococcus lactis.

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Lages, Aksar Chair; Mustopa, Apon Zaenal; Sukmarini, Linda; Suharsono

2015-10-01

Plantaricins, one of bacteriocin produced by Lactobacillus plantarum, are already known to have activities against several pathogenic bacterium. L. plantarum U10 isolated from "tempoyak," an Indonesian fermented food, produced one kind of plantaricin designated as plantaricin W (plnW). The plnW is suggested as a putative membrane location of protein and has similar conserved motif which is important as immunity to bacteriocin itself. Thus, due to study about this plantaricin, several constructs have been cloned and protein was analyzed in Lactococcus lactis. In this study, plnW gene was successfully cloned into vector NICE system pNZ8148 and created the transformant named L. lactis NZ3900 pNZ8148-WU10. PlnW protein was 25.3 kDa in size. The concentration of expressed protein was significantly increased by 10 ng/mL nisin induction. Furthermore, PlnW exhibited protease activity with value of 2.22 ± 0.05 U/mL and specific activity about 1.65 ± 0.03 U/mg protein with 50 ng/mL nisin induction. Immunity study showed that the PlnW had immunity activity especially against plantaricin and rendered L. lactis recombinant an immunity broadly to other bacteriocins such as pediocin, fermentcin, and acidocin.

[BIOLOGICAL ACTIVITY OF ANTIMICROBIAL PEPTIDES OF ENTEROCOCCUS FAECIUM].

Science.gov (United States)

Vasilchenko, A S; Rogozhin, E A; Valyshev, A V

2015-01-01

Isolate bacteriocins from Enterococcus faecium metabolites and characterize their effect on cells of Gram positive (Listeria monocytogenes) and Gram negative (Escherichia coli) bacteria. Methods of solid-phase extraction, ion-exchange and reversed phase chromatography were applied for isolation of bacteriocins from cultural medium of bacteria MALDI time-of-flight mass-spectrometry was used for characterization of the obtained preparations. The mechanism of biological effect of peptides was evaluated using DNA-tropic dyes (SYTO 9 and PI) with subsequent registration of fluorescence spectra: Atomic-force microscopy (AFM) was used for characterization of morpho-functional reaction of target cells. Peptide fractions with mass of 1.0 - 3.0 kDa were isolated from enterococci metabolites, that inhibit the growth of indicator microorganisms. E. faecium strain exoproducts were shown to increase membrane permeability during interaction with L. monocytogenes, that results in subsequent detectable disturbance of normal cell morphology of listeria. Alterations of E. coli surface during the effect of purified peptide fraction was detected using AFM. The studies carried out have revealed the effect of bacteriocins of enterococci on microorganisms with various types of cell wall composition and have confirmed the importance of bacterial barrier structure permeability disturbance in the mechanism of antimicrobial effect of enterocins.

Partial characterization of bacitracin like inhibitory substance from bacillus subtilis BS15, a local soil isolate

International Nuclear Information System (INIS)

Alam, S.I.; Kamran, M.; Sohail, M.; Ahmad, A.; Khan, S.A.

2011-01-01

The aim of this study was to investigate the production of bacteriocin/bacteriocin-like inhibitory substances (BLIS) from Bacillus subtilis BS15, isolated from soil. The inhibitory substance was partially purified and characterized as BLIS with a molecular-weight of 3-5 kDa, as determined by SDS-PAGE. Its production was observed during the late exponential phase or at the beginning of stationary-phase. It retained its activity up to 80 deg. C and over a wide range of pH i.e., 3-9. It was found active against several clinically important bacterial species such as Listeria monocytogenes, Staphylococcus aureus, Bacillus cereus, Salmonella typhi and also against the food-spoilage causing microbes, and may be considered as future food preservative. (author)

Nisin and its Antimicrobial Effect in Foods

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Hamparsun Hampikyan

2007-04-01

Full Text Available Nisin is a bacteriocin which is produced by Lactococcus lactis and takes its place in I. class bacteriocins which are known as lantibiotics. Nisin has antimicrobial and bactericidal activity against a broad spectrum of gram positive bacteria and spores of Clostridium spp. and Bacillus spp. According to toxicity studies nisin is considered not toxic to humans. Its first established used was as a preservative in processed cheese products and since than numerous other applications in various foods such as meat and meat products, poultry products, sea products and beverages such as beer, wine have been used safely. In this review, the characteristics of nisin, its usage in food and its antimicrobial effect are considered. [TAF Prev Med Bull 2007; 6(2.000: 142-147

Antimicrobial activities of lactic acid bacteria isolated from akamu ...

African Journals Online (AJOL)

The partially purified inhibitory compounds were screened by agar spot assay method for antagonistic ... The partially purified compounds exhibited strong activity against ... Keywords: Bacteriocins, lactic acid bacteria (LAB), target organisms, ...

Draft Genome Sequence of Leuconostoc mesenteroides 406 Isolated from the Traditional Fermented Mare Milk Airag in Tuv Aimag, Mongolia

OpenAIRE

Morita, Hidetoshi; Toh, Hidehiro; Oshima, Kenshiro; Nakano, Akiyo; Hano, Chihiro; Yoshida, Saki; Nguyen, Tien Thi Thuy; Wulijideligen,; Tashiro, Kosuke; Arakawa, Kensuke; Miyamoto, Taku

2016-01-01

Leuconostoc mesenteroides 406 was isolated from the traditional fermented mare milk airag in Tuv Aimag, Mongolia. This strain produces an antilisterial bacteriocin. Here, we report the draft genome sequence of this organism.

The Cyclic Antibacterial Peptide Enterocin AS-48: Isolation, Mode of Action, and Possible Food Applications

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María José Grande Burgos

2014-12-01

Full Text Available Enterocin AS-48 is a circular bacteriocin produced by Enterococcus. It contains a 70 amino acid-residue chain circularized by a head-to-tail peptide bond. The conformation of enterocin AS-48 is arranged into five alpha-helices with a compact globular structure. Enterocin AS-48 has a wide inhibitory spectrum on Gram-positive bacteria. Sensitivity of Gram-negative bacteria increases in combination with outer-membrane permeabilizing treatments. Eukaryotic cells are bacteriocin-resistant. This cationic peptide inserts into bacterial membranes and causes membrane permeabilization, leading ultimately to cell death. Microarray analysis revealed sets of up-regulated and down-regulated genes in Bacillus cereus cells treated with sublethal bacteriocin concentration. Enterocin AS-48 can be purified in two steps or prepared as lyophilized powder from cultures in whey-based substrates. The potential applications of enterocin AS-48 as a food biopreservative have been corroborated against foodborne pathogens and/or toxigenic bacteria (Listeria monocytogenes, Bacillus cereus, Staphylococcus aureus, Escherichia coli, Salmonella enterica and spoilage bacteria (Alicyclobacillus acidoterrestris, Bacillus spp., Paenibacillus spp., Geobacillus stearothermophilus, Brochothrix thermosphacta, Staphylococcus carnosus, Lactobacillus sakei and other spoilage lactic acid bacteria. The efficacy of enterocin AS-48 in food systems increases greatly in combination with chemical preservatives, essential oils, phenolic compounds, and physico-chemical treatments such as sublethal heat, high-intensity pulsed-electric fields or high hydrostatic pressure.

The Cyclic Antibacterial Peptide Enterocin AS-48: Isolation, Mode of Action, and Possible Food Applications.

Science.gov (United States)

Grande Burgos, María José; Pulido, Rubén Pérez; Del Carmen López Aguayo, María; Gálvez, Antonio; Lucas, Rosario

2014-12-08

Enterocin AS-48 is a circular bacteriocin produced by Enterococcus. It contains a 70 amino acid-residue chain circularized by a head-to-tail peptide bond. The conformation of enterocin AS-48 is arranged into five alpha-helices with a compact globular structure. Enterocin AS-48 has a wide inhibitory spectrum on Gram-positive bacteria. Sensitivity of Gram-negative bacteria increases in combination with outer-membrane permeabilizing treatments. Eukaryotic cells are bacteriocin-resistant. This cationic peptide inserts into bacterial membranes and causes membrane permeabilization, leading ultimately to cell death. Microarray analysis revealed sets of up-regulated and down-regulated genes in Bacillus cereus cells treated with sublethal bacteriocin concentration. Enterocin AS-48 can be purified in two steps or prepared as lyophilized powder from cultures in whey-based substrates. The potential applications of enterocin AS-48 as a food biopreservative have been corroborated against foodborne pathogens and/or toxigenic bacteria (Listeria monocytogenes, Bacillus cereus, Staphylococcus aureus, Escherichia coli, Salmonella enterica) and spoilage bacteria (Alicyclobacillus acidoterrestris, Bacillus spp., Paenibacillus spp., Geobacillus stearothermophilus, Brochothrix thermosphacta, Staphylococcus carnosus, Lactobacillus sakei and other spoilage lactic acid bacteria). The efficacy of enterocin AS-48 in food systems increases greatly in combination with chemical preservatives, essential oils, phenolic compounds, and physico-chemical treatments such as sublethal heat, high-intensity pulsed-electric fields or high hydrostatic pressure.

Development of a Chemically Defined Medium for Better Yield and Purification of Enterocin Y31 from Enterococcus faecium Y31

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Wenli Liu

2017-01-01

Full Text Available The macro- and micronutrients in traditional medium, such as MRS, used for cultivating lactic acid bacteria, especially for bacteriocin production, have not been defined, preventing the quantitative monitoring of metabolic flux during bacteriocin biosynthesis. To enhance Enterocin Y31 production and simplify steps of separation and purification, we developed a simplified chemically defined medium (SDM for the growth of Enterococcus faecium Y31 and production of its bacteriocin, Enterocin Y31. We found that the bacterial growth was unrelated to Enterocin Y31 production in MRS; therefore, both the growth rate and the Enterocin Y31 production were set as the index for investigation. Single omission experiments revealed that 5 g/L NaCl, five vitamins, two nucleic acid bases, MgSO4·7H2O, MnSO4·4H2O, KH2PO4, K2HPO4, CH3COONa, fourteen amino acids, and glucose were essential for the strain’s growth and Enterocin Y31 production. Thus, a novel simplified and defined medium (SDM was formulated with 30 components in total. Consequently, Enterocin Y31 production yield was higher in SDM as compared to either MRS or CDM. SDM improved the Enterocin Y31 production and simplified the steps of purification (only two steps, which has broad potential applications.

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Vol 11, No 11 (2012), Antimicrobial action of purified raspberry flavonoid, Abstract ... solvents extracted samples of Linum usitatissimum by disc diffusion method ... Antimicrobial activities of the bacteriocin-like substances produced by lactic ...

Enhanced biomass production study on probiotic Bacillus subtilis ...

African Journals Online (AJOL)

user

2010-11-22

Nov 22, 2010 ... INTRODUCTION. Probiotic organisms find their potential use in food and ..... complex nutrients, temperature and pH on bacteriocin production by. Bacillus subtilis ... B, Gupta R (2004). Application of statistical experimental.

In vitro study of beneficial properties and safety of lactic acid bacteria isolated from Portuguese fermented meat products.

Science.gov (United States)

Todorov, S D; Franco, B D G M; Wiid, I J

2014-09-01

Many lactic acid bacteria produce bacteriocins with a rather broad spectrum of inhibition, which could offer potential applications in food preservation. Bacteriocin production by starter cultures may bring advantage to these strains in competitive interactions with pathogenic bacteria from the food matrix. The objective of this study was to determine the safety of beneficial strains (Lactobacillus plantarum ST202Ch and ST216Ch, Enterococcus faecium ST211Ch, and Lactobacillus sakei ST22Ch, ST153Ch and ST154Ch) previously isolated from fermented meat products and characterised as bacteriocin producers. Auto-aggregation was strain-specific, and values of 28.97, 27.86 and 28.56% were recorded for L. sakei ST22Ch, ST153Ch and ST154Ch, respectively, 16.95 and 14.58% for L. plantarum ST202Ch and ST216Ch, respectively, and 12.77% for E. faecium ST211Ch. Various degrees of co-aggregation between 28.85 and 44.76% for Listeria monocytogenes 211 and 409, and between 23.60 to 34.96% for E. faecium ATCC 19443 were observed. According to the results of the diffusion method, the studied strains demonstrated susceptibility to penicillin G, ampicillin, amoxicillin, amoxicillin/clavulonic acid, imipenem, linezolid, and tetracycline. In addition, the susceptibility of the six strains to various non-antibiotic commercial drugs was examined. Production of β-galactosidase by L. sakei ST22Ch, ST153Ch and ST154Ch, L. plantarum ST202Ch and ST216Ch, and E. faecium ST211Ch was confirmed by employing sterile filter paper discs impregnated with o-nitrophenyl-β-D-galactopyranose. A statistically significant (P<0.001) inhibition of Mycobacterium tuberculosis growth by bacteriocins produced by L. plantarum ST202Ch (38.3%) and ST216Ch (48.6%), L. sakei ST153Ch (16.2%) and ST154Ch (16.1%), and E. faecium ST211Ch (21.7%) was observed. As determined by the polymerase chain reaction, the tested strains showed a low virulence gene profile.

Determination of antimicrobial activity and production of some ...

African Journals Online (AJOL)

STORAGESEVER

2008-12-17

Dec 17, 2008 ... periods (24, 48 and 72 h) by a spectrophotometric method. The exopolysaccharide .... indicated among treatment means (P≤0.01), means were differentiated using ..... Bacteriocin inhibition of Clostridium botulinum spores by ...

Time series analysis of aerobic bacterial flora during Miso fermentation.

Science.gov (United States)

Onda, T; Yanagida, F; Tsuji, M; Shinohara, T; Yokotsuka, K

2003-01-01

This article reports a microbiological study of aerobic mesophilic bacteria that are present during the fermentation process of Miso. Aerobic bacteria were enumerated and isolated from Miso during fermentation and divided into nine groups using traditional phenotypic tests. The strains were identified by biochemical analysis and 16S rRNA sequence analysis. They were identified as Bacillus subtilis, B. amyloliquefaciens, Kocuria kristinae, Staphylococcus gallinarum and S. kloosii. All strains were sensitive to the bacteriocins produced by the lactic acid bacteria isolated from Miso. The dominant species among the undesirable species throughout the fermentation process were B. subtilis and B. amyloliquefaciens. It is suggested that bacteriocin-producing lactic acid bacteria are effective in the growth prevention of aerobic bacteria in Miso. This study has provided useful information for controlling of bacterial flora during Miso fermentation.

Antibacterial activities of lactic acid bacteria isolated from cow ...

African Journals Online (AJOL)

Method: Escherichia coli, Klebsiella species (spp) and LAB were isolated from thirty different cow faecal samples and the .... The PCR products were purified and sequenced for the ... their ability to produce bacteriocin-like inhibitory sub-.

Effects of agitation speed, temperature, carbon and nitrogen sources ...

African Journals Online (AJOL)

USER

2010-08-16

Aug 16, 2010 ... alive for the treatment of disease. In this study ... virulence factor that was purified from Aeromonas hydro- phila .... salicylic acid (DNS) method (Miller, 1959). Sucrose .... bacteriocin production of recombinant strain L. lactis.

Draft Genome Sequence of Leuconostoc mesenteroides 213M0, Isolated from Traditional Fermented Mare Milk Airag in Bulgan Aimag, Mongolia

OpenAIRE

Morita, Hidetoshi; Toh, Hidehiro; Oshima, Kenshiro; Nakano, Akiyo; Hano, Chihiro; Yoshida, Saki; Bolormaa, Tsognemekh; Burenjargal, Sedkhuu; Nguyen, Co Thi Kim; Tashiro, Kosuke; Arakawa, Kensuke; Miyamoto, Taku

2016-01-01

Leuconostoc mesenteroides 213M0 was isolated from traditional fermented mare milk airag in Bulgan Aimag, Mongolia. This strain produces a listericidal bacteriocin-like inhibitory substance. Here, we report the draft genome sequence of this organism.

Antagonistic effect of brevicin on Gram positive and Gram negative ...

African Journals Online (AJOL)

B. Senthil Kumar

This is stable at high temperature acidic to neutral pH ... applications, though, due to its biochemical properties; it .... All were assessed for the production of bacteriocin ..... coating to control the growth of Listeria monocytogenes on poached.

Production of enterocin A by Enterococcus faecium MMRA isolated from 'Rayeb', a traditional Tunisian dairy beverage.

Science.gov (United States)

Rehaiem, A; Martínez, B; Manai, M; Rodríguez, A

2010-05-01

Characterization and purification of a bacteriocin produced by a wild Enterococcus faecium strain, isolated from a Tunisian traditional fermented milk. Enterococcus faecium MMRA was selected on the basis of its strong anti-Listeria activity. The antibacterial activity was sensitive to proteases, confirming its proteinaceous nature. It was extremely heat stable (15 min at 121 degrees C), remained active over a wide pH range (2-12), and also after treatment with lipase, amylase, organic solvents, detergents, lyophilisation and long-term storage at -20 degrees C. Production of the bacteriocin occurred throughout the logarithmic growth phase, it did not adhere to the surface of the producer cells and the mode of action was bactericidal. After partial purification of the active supernatants, a 4-kDa band with antibacterial activity was revealed by SDS-PAGE electrophoresis and bioassay. Tryptic digestion followed by MALDI-TOF mass spectrometry identified the peptide as enterocin A. The inhibitory activity of Ent. faecium MMRA, a wild strain isolated from the artisan dairy beverage 'Rayeb', is due to the synthesis of an enterocin A. Traditional fresh Tunisian fermented dairy products are generally manufactured with raw milk that can be used as a source of uncharacterized wild lactic acid bacteria strains. To our knowledge, this is the first report on the isolation of an enterocin A producing Ent. faecium from 'Rayeb'. This bacteriocin or the producing strain might have a promising potential in biopreservation to enhance the hygienic quality of this dairy product.

Atypical Genetic Locus Associated with Constitutive Production of Enterocin B by Enterococcus faecium BFE 900

Science.gov (United States)

Franz, Charles M. A. P.; Worobo, Randy W.; Quadri, Luis E. N.; Schillinger, Ulrich; Holzapfel, Wilhelm H.; Vederas, John C.; Stiles, Michael E.

1999-01-01

A purified bacteriocin produced by Enterococcus faecium BFE 900 isolated from black olives was shown by Edman degradation and mass spectrometric analyses to be identical to enterocin B produced by E. faecium T136 from meat (P. Casaus, T. Nilsen, L. M. Cintas, I. F. Nes, P. E. Hernández, and H. Holo, Microbiology 143:2287–2294, 1997). The structural gene was located on a 2.2-kb HindIII fragment and a 12.0-kb EcoRI chromosomal fragment. The genetic characteristics and production of EntB by E. faecium BFE 900 differed from that described so far by the presence of a conserved sequence like a regulatory box upstream of the EntB gene, and its production was constitutive and not regulated. The 2.2-kb chromosomal fragment contained the hitherto undetected immunity gene for EntB in an atypical orientation that is the reverse of that of the structural gene. Typical transport and other genes associated with bacteriocin production were not detected on the 12.0-kb chromosomal fragment containing the EntB structural gene. This makes the EntB genetic system different from most other bacteriocin systems, where transport and possible regulatory genes are clustered. EntB was subcloned and expressed by the dedicated secretion machinery of Carnobacterium piscicola LV17A. The structural gene was amplified by PCR, fused to the divergicin A signal peptide, and expressed by the general secretory pathway in Enterococcus faecalis ATCC 19433. PMID:10224016

Production, purification, sequencing and activity spectra of mutacins D-123.1 and F-59.1.

Science.gov (United States)

Nicolas, Guillaume G; LaPointe, Gisèle; Lavoie, Marc C

2011-04-10

The increase in bacterial resistance to antibiotics impels the development of new anti-bacterial substances. Mutacins (bacteriocins) are small antibacterial peptides produced by Streptococcus mutans showing activity against bacterial pathogens. The objective of the study was to produce and characterise additional mutacins in order to find new useful antibacterial substances. Mutacin F-59.1 was produced in liquid media by S. mutans 59.1 while production of mutacin D-123.1 by S. mutans 123.1 was obtained in semi-solid media. Mutacins were purified by hydrophobic chromatography. The amino acid sequences of the mutacins were obtained by Edman degradation and their molecular mass was determined by mass spectrometry. Mutacin F-59.1 consists of 25 amino acids, containing the YGNGV consensus sequence of pediocin-like bacteriocins with a molecular mass calculated at 2719 Da. Mutacin D-123.1 has an identical molecular mass (2364 Da) with the same first 9 amino acids as mutacin I. Mutacins D-123.1 and F-59.1 have wide activity spectra inhibiting human and food-borne pathogens. The lantibiotic mutacin D-123.1 possesses a broader activity spectrum than mutacin F-59.1 against the bacterial strains tested. Mutacin F-59.1 is the first pediocin-like bacteriocin identified and characterised that is produced by Streptococcus mutans. Mutacin D-123.1 appears to be identical to mutacin I previously identified in different strains of S. mutans.

Production, purification, sequencing and activity spectra of mutacins D-123.1 and F-59.1

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LaPointe Gisèle

2011-04-01

Full Text Available Abstract Background The increase in bacterial resistance to antibiotics impels the development of new anti-bacterial substances. Mutacins (bacteriocins are small antibacterial peptides produced by Streptococcus mutans showing activity against bacterial pathogens. The objective of the study was to produce and characterise additional mutacins in order to find new useful antibacterial substances. Results Mutacin F-59.1 was produced in liquid media by S. mutans 59.1 while production of mutacin D-123.1 by S. mutans 123.1 was obtained in semi-solid media. Mutacins were purified by hydrophobic chromatography. The amino acid sequences of the mutacins were obtained by Edman degradation and their molecular mass was determined by mass spectrometry. Mutacin F-59.1 consists of 25 amino acids, containing the YGNGV consensus sequence of pediocin-like bacteriocins with a molecular mass calculated at 2719 Da. Mutacin D-123.1 has an identical molecular mass (2364 Da with the same first 9 amino acids as mutacin I. Mutacins D-123.1 and F-59.1 have wide activity spectra inhibiting human and food-borne pathogens. The lantibiotic mutacin D-123.1 possesses a broader activity spectrum than mutacin F-59.1 against the bacterial strains tested. Conclusion Mutacin F-59.1 is the first pediocin-like bacteriocin identified and characterised that is produced by Streptococcus mutans. Mutacin D-123.1 appears to be identical to mutacin I previously identified in different strains of S. mutans.

LnqR, a TetR-family transcriptional regulator, positively regulates lacticin Q production in Lactococcus lactis QU 5.

Science.gov (United States)

Iwatani, Shun; Ishibashi, Naoki; Flores, Floirendo P; Zendo, Takeshi; Nakayama, Jiro; Sonomoto, Kenji

2016-09-01

Lacticin Q is an unmodified leaderless bacteriocin produced by Lactococcus lactis QU 5. It has been revealed that the production and self-immunity of lacticin Q are facilitated by a gene cluster lnqQBCDEF The gene for a putative TetR-family transcriptional regulator, termed lnqR, was found nearby the lnqQBCDEF cluster, but its involvement in lacticin Q biosynthesis remained unknown. In this study, we created an LnqR-overexpressing QU 5 recombinant by using lactococcal constitutive promoter P32 The recombinant QU 5 showed enhanced production of and self-immunity to lacticin Q. RT-PCR analysis has revealed that an overexpression of LnqR increases the amounts of lnqQBCDEF transcripts, and these six genes are transcribed as an operon in a single transcriptional unit. Interestingly, LnqR expression and thus lacticin Q production by L. lactis QU 5 was found temperature dependent, while LnzR, an LnqR-homologue, in L. lactis QU 14 was expressed in a similar but not identical manner to LnqR, resulting in dissimilar bacteriocin productivities by these strains. This report demonstrates LnqR as the first TetR-family transcriptional regulator involved in LAB bacteriocin biosynthesis and that, as an exceptional case of TetR-family regulators, LnqR positively regulates the transcription of these biosynthetic genes. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Inhibitory activity of Lactobacillus plantarum LMG P-26358 against Listeria innocua when used as an adjunct starter in the manufacture of cheese

LENUS (Irish Health Repository)

2011-08-30

Abstract Lactobacillus plantarum LMG P-26358 isolated from a soft French artisanal cheese produces a potent class IIa bacteriocin with 100% homology to plantaricin 423 and bacteriocidal activity against Listeria innocua and Listeria monocytogenes. The bacteriocin was found to be highly stable at temperatures as high as 100°C and pH ranges from 1-10. While this relatively narrow spectrum bacteriocin also exhibited antimicrobial activity against species of enterococci, it did not inhibit dairy starters including lactococci and lactobacilli when tested by well diffusion assay (WDA). In order to test the suitability of Lb. plantarum LMG P-26358 as an anti-listerial adjunct with nisin-producing lactococci, laboratory-scale cheeses were manufactured. Results indicated that combining Lb. plantarum LMG P-26358 (at 108 colony forming units (cfu)\\/ml) with a nisin producer is an effective strategy to eliminate the biological indicator strain, L. innocua. Moreover, industrial-scale cheeses also demonstrated that Lb. plantarum LMG P-26358 was much more effective than the nisin producer alone for protection against the indicator. MALDI-TOF mass spectrometry confirmed the presence of plantaricin 423 and nisin in the appropriate cheeses over an 18 week ripening period. A spray-dried fermentate of Lb. plantarum LMG P-26358 also demonstrated potent anti-listerial activity in vitro using L. innocua. Overall, the results suggest that Lb. plantarum LMG P-26358 is a suitable adjunct for use with nisin-producing cultures to improve the safety and quality of dairy products.

Intestinal infections and prebiotics: the roles of oligosaccharides in promoting health

Science.gov (United States)

Prebiotic oligosaccharides exert activity against pathogens partly by stimulating the growth and/or activity of commensal bacteria that provide health benefits (lower pH, bacteriocin production, immune system modulation, competitive exclusion). This review describes alternative mechanisms of action...

1 ORIGINAL ARTICLE

African Journals Online (AJOL)

boaz

Bacteriocins produce localized holes in cell wall and cellular membrane ... regeneration of damaged tissues (22). The possibility .... line caudally from the lower margin of the ear. Antisepsis of the ..... the ability to inhibit the growth of the some.

Draft Genome Sequence of Leuconostoc mesenteroides 406 Isolated from the Traditional Fermented Mare Milk Airag in Tuv Aimag, Mongolia.

Science.gov (United States)

Morita, Hidetoshi; Toh, Hidehiro; Oshima, Kenshiro; Nakano, Akiyo; Hano, Chihiro; Yoshida, Saki; Nguyen, Tien Thi Thuy; Wulijideligen; Tashiro, Kosuke; Arakawa, Kensuke; Miyamoto, Taku

2016-03-24

Leuconostoc mesenteroides406 was isolated from the traditional fermented mare milk airag in Tuv Aimag, Mongolia. This strain produces an antilisterial bacteriocin. Here, we report the draft genome sequence of this organism. Copyright © 2016 Morita et al.

Intracellular pH of Mycobacterium avium subsp. paratuberculosis following exposure to antimicrobial compounds monitored at the single cell level

DEFF Research Database (Denmark)

Gaggìa, Francesca; Nielsen, Dennis Sandris; Biavati, Bruno

2010-01-01

for 24h revealed the presence of a subpopulation of cells probably resistant to the antimicrobial compounds tested. Use of nisin and bacteriocin-producing LAB strains could lead to new intervention strategies for the control of MAP based on in vivo application of probiotic cultures as feed additives......Mycobacterium avium subsp. paratuberculosis (MAP) is the etiologic agent of Johne's disease; moreover, it seems to be implicated in the development of Crohn's disease in humans. In the present study, fluorescence ratio imaging microscopy (FRIM) was used to assess changes in intracellular pH (p......H(i)) of one strain of MAP after exposure to nisin and neutralized cell-free supernatants (NCSs) from five bacteriocin-producing lactic acid bacteria (LAB) with known probiotic properties. The evaluation of pH(i) by FRIM provides information about the physiological state of bacterial cells, bypassing the long...

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African Journals Online (AJOL)

Vol 6, No 22 (2007), Bacteriocin and cellulose production by lactic acid bacteria isolated .... Vol 14, No 7 (2015), Bayoud disease of date palm in Algeria: History, ... extract on germination and growth of lettuce and tomato plants, Abstract PDF.

Bacteriocinogenic potential and genotypic characterization of three ...

African Journals Online (AJOL)

Louiza

2015-08-06

Aug 6, 2015 ... Complete inactivation in bacteriocinogenic activity was observed after treatment with .... enterocins, with inhibitory activity against strains closely related to the ... buffered medium; ATCC, American Type Culture Collection; ND, not detected. Phenotypic ..... The thermal stability of bacteriocins produced by.

A probiotic bacterium, Pediococcus pentosaceus OZF, isolated from ...

African Journals Online (AJOL)

Pediococcus pentosaceus OZF, originally isolated from healthy human breast milk, produces antimicrobial activities against many gram-positive bacterial species, including the food borne pathogen, Listeria monocytogenes. A bacteriocin was purified to homogeneity from the supernatant of exponentially growing cells using ...

Influence des substrats carbonés et minéraux sur l'activité des ...

African Journals Online (AJOL)

p < 0.0001). BLIS were stable to heat treatment ranging from 50 °C to 100 °C for 30 min. Conclusion and application of findings: These inhibitory substances produced, could be bacteriocins and could contribute to a better food preservation.

Inhibition of Bacillus cereus and Bacillus weihenstephanensis in raw vegetables by application of washing solutions containing enterocin AS-48 alone and in combination with other antimicrobials.

Science.gov (United States)

Cobo Molinos, Antonio; Abriouel, Hikmate; Lucas López, Rosario; Ben Omar, Nabil; Valdivia, Eva; Gálvez, Antonio

2008-09-01

Enterocin AS-48 is a broad-spectrum cyclic antimicrobial peptide produced by Enterococcus faecalis. In the present study, the bacteriocin was tested alone and in combination with other antimicrobials for decontamination of Bacillus inoculated on alfalfa, soybean sprouts and green asparagus. Washing with enterocin AS-48 solutions reduced viable cell counts of Bacillus cereus and Bacillus weihenstephanensis by 1.0-1.5 and by 1.5-2.38 log units right after application of treatment, respectively. In both cases, the bacteriocin was effective in reducing the remaining viable population below detection levels during further storage of the samples at 6 degrees C, but failed to prevent regrowth in samples stored at 15 or 22 degrees C. Application of washing treatments containing enterocin AS-48 in combination with several other antimicrobials and sanitizers (cinnamic and hydrocinnamic acids, carvacrol, polyphosphoric acid, peracetic acid, hexadecylpyridinium chloride and sodium hypochlorite) greatly enhanced the bactericidal effects. The combinations of AS-48 and sodium hypochlorite, peracetic acid or hexadecylpyridinium chloride provided the best results. After application of the combined treatments on alfalfa sprouts contaminated with B. cereus or with B. weihenstephanensis, viable bacilli were not detected or remained at very low concentrations in the treated samples during a 1-week storage period at 15 degrees C. Inhibition of B. cereus by in situ produced bacteriocin was tested by cocultivation with the AS-48 producer strain E. faecalis A-48-32 inoculated on soybean sprouts. Strain A-48-32 was able to grow and produce bacteriocin on sprouts both at 15 and 22 degrees C. At 15 degrees C, growth of B. cereus was completely inhibited in the cocultures, while a much more limited effect was observed at 22 degrees C. The results obtained for washing treatments are very encouraging for the application of enterocin AS-48 in the decontamination of sprouts. Application of washing

Purification of leucocin A for use on wieners to inhibit Listeria monocytogenes in the presence of spoilage organisms.

Science.gov (United States)

Balay, Danielle R; Dangeti, Ramana V; Kaur, Kamaljit; McMullen, Lynn M

2017-08-16

The aims of this study were to improve the method for purification of leucocin A to increase yield of peptide and to evaluate the efficacy of leucocin A and an analogue of leucocin A (leucocin N17L) to inhibit the growth of Listeria monocytogenes on wieners in the presence of spoilage organisms. Leucocin A was produced by Leuconostoc gelidum UAL187 and purified with a five-fold increase in yield; leucocin N17L was synthesized replacing asparagine at residue 17 with leucine. Five strains of L. monocytogenes associated with foodborne illness were used to assess bacteriocin efficacy in vitro and in situ. Minimum inhibitory concentrations could not be determined in broth; however, on agar the minimum inhibitory concentrations ranged from 11.7-62.5μM and 62.5->500μM for leucocin A and leucocin N17L, respectively. Leucocin N17L was less effective than the native bacteriocin at controlling the growth of L. monocytogenes. The inactivation profiles of L. monocytogenes in broth in the presence of leucocin A suggested each isolate had different levels of resistance to the bacteriocin as determined by the initial bactericidal effect. The formation of spontaneously resistance subpopulations were also observed for each strain of L. monocytogenes. In situ, wieners were inoculated with the spoilage organisms, Carnobacterium divergens and Brochothrix thermosphacta, followed by surface application of purified leucocin A, and inoculated with a cocktail of L. monocytogenes. Wieners were vacuum packaged and stored at 7°C for 16d. Leucocin A reduced the counts L. monocytogenes on wieners during storage, regardless of the presence of C. divergens. B. thermosphacta was unaffected by the presence of leucocin A on wieners over the duration of storage. This study suggests that leucocin A may be beneficial to industry as a surface application on wieners to help reduce L. monocytogenes counts due to post-processing contamination even in the presence of spoilage organisms. However, further

Detection of the pediocin gene pedA in strains from human faeces by real-time PCR and characterization of Pediococcus acidilactici UVA1

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Cereghetti Tania

2007-09-01

Full Text Available Abstract Background Bacteriocin-producing lactic acid bacteria are commonly used as natural protective cultures. Among them, strains of the genus Pediococcus are particularly interesting for their ability to produce pediocin, a broad spectrum antimicrobial peptide with a strong antagonistic activity against the food-borne pathogen Listeria monocytogenes. Furthermore, there is increasing interest in isolating new bacteriocin-producing strains of human intestinal origin that could be developed for probiotic effects and inhibition of pathogenic bacteria in the gut. In this work, we typed a new strain, co-isolated from baby faeces together with a Bifidobacterium thermophilum strain, and characterized its proteinaceous compound with strong antilisterial activity. Results The newly isolated strain UVA1 was identified as a Pediococcus acidilactici by carbohydrate fermentation profile, growth at 50°C and 16S rDNA sequencing. The partially purified bacteriocin was heat resistant up to 100°C, active over a wide range of pH (2 to 9 and susceptible to proteolytic enzymes. The molecular weight, estimated by SDS-PAGE, was similar to that of pediocin AcH/PA-1 (4.5 kDa. P. acidilactici UVA1 harboured a 9.5-kb plasmid that could be cured easily, which resulted in the loss of the antimicrobial activity. Southern hybridization using the DIG-labelled pedA-probe established that the bacteriocin gene was plasmid-borne as for all pediocin described so far. Nucleotide sequence of the whole operon (3.5 kb showed almost 100 % similarity to the pediocin AcH/PA-1 operon. The mRNA transcript for pedA could be detected in P. acidilactici UVA1 but not in the cured derivative, confirming the expression of the pedA-gene in UVA1. Using a new real-time PCR assay, eleven out of seventeen human faecal samples tested were found to contain pedA-DNA. Conclusion We identified and characterised the first pediocin produced by a human intestinal Pediococcus acidilactici isolate and

quorum sensing

African Journals Online (AJOL)

kannappan

2012-10-02

Oct 2, 2012 ... But in the treatment tank the mortality varied from 0.53 to 1.26,. 5.51, 8.17, 10.72 ... Therefore, the quest for alternative methods to control infection ..... weight determination of bacteriocin produced by Lactococcus lactis. D53.

Use of response surface methodology to optimize the drying ...

African Journals Online (AJOL)

CHABI

2016-09-15

Sep 15, 2016 ... ATCC 27844, methicillin resistant S. aureus (MRSA),. Salmonella typhi R ... count (YM), bacteriocin production (BE) and the antimicrobial activity against indicator .... predicted one from the optimized model by calculating the percentage error to ..... A mathematical model considering variable diffusivity ...

Draft Genome Sequence of Leuconostoc mesenteroides 213M0, Isolated from Traditional Fermented Mare Milk Airag in Bulgan Aimag, Mongolia.

Science.gov (United States)

Morita, Hidetoshi; Toh, Hidehiro; Oshima, Kenshiro; Nakano, Akiyo; Hano, Chihiro; Yoshida, Saki; Bolormaa, Tsognemekh; Burenjargal, Sedkhuu; Nguyen, Co Thi Kim; Tashiro, Kosuke; Arakawa, Kensuke; Miyamoto, Taku

2016-03-31

Leuconostoc mesenteroides213M0 was isolated from traditional fermented mare milk airag in Bulgan Aimag, Mongolia. This strain produces a listericidal bacteriocin-like inhibitory substance. Here, we report the draft genome sequence of this organism. Copyright © 2016 Morita et al.

Leucocins 4010 from Leuconostoc carnosum cause a matrix related decrease in intracellular pH of Listeria monocytogenes

DEFF Research Database (Denmark)

Fang, Weihuan; Budde, Birgitte Bjørn; Siegumfeldt, Henrik

2006-01-01

A mixed culture of single cells of Listeria monocytogenes and the bacteriocin producing Leuconostoc carnosum 4010 showed growth inhibition of L. monocytogenes, although the intracellular pH (pHi) of L. monocytogenes followed by fluorescence ratio imaging microscopy was not affected. Furthermore, L...

Recent approaches in food bio-preservation - a review | Singh ...

African Journals Online (AJOL)

Apart from LAB metabolites, bacteriophages and endolysins has promising role in food processing, preservation and safety. Bacteriocins and endolysins are more suitable for DNA shuffling and protein engineering to generate highly potent variants with expanded activity spectrum. Genetically modified bacteriophages may ...

Lactobacillus plantarum inhibits growth of Listeria monocytogenes in an in vitro continuous flow gut model, but promotes invasion of L. monocytogenes in the gut of gnotobiotic rats

DEFF Research Database (Denmark)

Bernbom, Nete; Licht, Tine Rask; Saadbye, Peter

2006-01-01

The ability of the pediocin AcH producing Lactobacillus plantarum DDEN 11007 and its non-producing plasmid-cured isogenic variant, DDEN 12305 to prevent the persistence and growth of Listeria monocytogenes EP2 in two gastrointestinal (GI) tract models was examined. In vitro studies conducted...... in a two-stage continuous flow system showed that L. plantarum DDEN 11007 inhibited L. monocytogenes EP2 under these conditions, while less effect was seen of the non-bacteriocin producing variant. The inhibitory effect was more pronounced at pH 5 than at pH 7. No effect on persistence of L. monocytogenes...... in the GI tract was seen in gnotobiotic rats colonized with either the pediocin AcH producing or the non-bacteriocin producing variant of L. plantarum when compared to rats inoculated with L. monocytogenes EP2 alone. Surprisingly, inoculation of the gnotobiotic animals with either of the L. plantarum...

Purification and characterization of enterocin MC13 produced by a potential aquaculture probiont Enterococcus faecium MC13 isolated from the gut of Mugil cephalus.

Science.gov (United States)

Satish Kumar, R; Kanmani, P; Yuvaraj, N; Paari, K A; Pattukumar, V; Arul, V

2011-12-01

A bacteriocin producer strain MC13 was isolated from the gut of Mugil cephalus (grey mullet) and identified as Enterococcus faecium. The bacteriocin of E. faecium MC13 was purified to homogeneity, as confirmed by Tricine sodium dodecyl sulphate - polyacrylamide gel electrophoresis (SDS-PAGE). Reverse-phase high-performance liquid chromatography (HPLC) analysis showed a single active fraction eluted at 26 min, and matrix-assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry analysis showed the molecular mass to be 2.148 kDa. The clear zone in native PAGE corresponding to enterocin MC13 band further substantiated its molecular mass. A dialyzed sample (semicrude preparation) of enterocin MC13 was broad spectrum in its action and inhibited important seafood-borne pathogens: Listeria monocytogenes , Vibrio parahaemolyticus, and Vibrio vulnificus. This antibacterial substance was sensitive to proteolytic enzymes: trypsin, protease, and chymotrypsin but insensitive to catalase and lipase, confirming that inhibition was due to the proteinaceous molecule, i.e., bacteriocin, and not due to hydrogen peroxide. Enterocin MC13 tolerated heat treatment (up to 90 °C for 20 min). Enterococcus faecium MC13 was effective in bile salt tolerance, acid tolerance, and adhesion to the HT-29 cell line. These properties reveal the potential of E. faecium MC13 to be a probiotic bacterium. Enterococcus faecium MC13 could be used as potential fish probiotic against pathogens such as V. parahaemolyticus, Vibrio harveyi, and Aeromonas hydrophila in fisheries. Also, this could be a valuable seafood biopreservative against L. monocytogenes.

PRODUCTION OF PLANTARCIN BY LACTOBACILLUS PLANTARUM SR18

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Wagih El-Shouny

2012-06-01

Full Text Available Out of 86 lactobacilli previously screened in our laboratory, Lactobacillus plantarum SR18 isolated from yoghurt revealed the largest detected inhibition zone against the selected indicator Streptococcus salivarius 5. The obtained electrophoretic patterns revealed that L. plantarum SR18 was free from plasmids. Exposure of 6 h growing L. plantarum culture to T-8M, 3B ultraviolet B lamp (8w, 220v & 312 nm for 2 h and subsequent growth for further 24 h resulted in an increase of cell-bound bacteriocin titer reached 2 fold at 12 h. Whereas bacteriocin secreted in the culture filtrate was not affected by UV irradiation. Plantarcin SR18 production was maximal (12800 AU/ml between 12 and 18 h by incubation of the culture at 37°C and pH 5-7 in candle jar (CO2. The bacteriocin bound to the cells and that secreted into the culture filtrate of L. plantarum SR18 were precipitated by 75% ammomium sulphate, dialysed and further purified by Gel filtration on Sephadex G-100. The specific activities (AU/mg protein were increased by a factor of about 5.3 and 2.35 for plantarcins purified from proteins bound to the cell of L. plantarum SR18 (plantarcin SR18 a and that secreted into the culture filtrate (plantarcin SR18 b, respectively. Gel filtration of plantarcin SR18a resulted in moderate antibacterial activity (3200 AU/ml and very high activity (25600 AU/ml of plantarcin SR18b.

Quorum-Sensing Regulation of Constitutive Plantaricin by Lactobacillus plantarum Strains under a Model System for Vegetables and Fruits

Science.gov (United States)

Rizzello, Carlo G.; Filannino, Pasquale; Calasso, Maria; Gobbetti, Marco

2014-01-01

This study aimed at investigating the regulatory system of bacteriocin synthesis by Lactobacillus plantarum strains in vegetables and fruits in a model system. Sterile and neutralized cell-free supernatant (CFS) from L. plantarum strains grown in MRS broth showed in vitro antimicrobial activities toward various indicator strains. The highest activity was that of L. plantarum C2. The antimicrobial activity was further assayed on vegetable and fruit agar plates (solid conditions) and in juices (liquid conditions). A regulatory mechanism of bacteriocin synthesis via quorum sensing was hypothesized. The synthesis of antimicrobial compounds seemed to be constitutive under solid conditions of growth on vegetable and fruit agar plates. In contrast, it depended on the size of the inoculum when L. plantarum C2 was grown in carrot juice. Only the inoculum of ca. 9.0 log CFU ml−1 produced detectable activity. The genes plnA, plnEF, plnG, and plnH were found in all L. plantarum strains. The genes plnJK and plnN were detected in only three or four strains. Reverse-phase high-performance liquid chromatography purification and mass spectrometry analysis revealed the presence of a mixture of eight peptides in the most active fraction of the CFS from L. plantarum C2. Active peptides were encrypted into bacteriocin precursors, such as plantaricins PlnJ/K and PlnH and PlnG, which are involved in the ABC transport system. A real-time PCR assay showed an increase in the expression of plnJK and plnG during growth of L. plantarum C2 in carrot juice. PMID:24242246

Characterization of bacterial isolates from the microbiota of mothers' breast milk and their infants.

Science.gov (United States)

Kozak, Kimberly; Charbonneau, Duane; Sanozky-Dawes, Rosemary; Klaenhammer, Todd

2015-01-01

This investigation assessed the potential of isolating novel probiotics from mothers and their infants. A subset of 21 isolates among 126 unique bacteria from breast milk and infant stools from 15 mother-infant pairs were examined for simulated GI transit survival, adherence to Caco-2 cells, bacteriocin production, and lack of antibiotic resistance. Of the 21 selected isolates a Lactobacillus crispatus isolate and 3 Lactobacillus gasseri isolates demonstrated good profiles of in vitro GI transit tolerance and Caco-2 cell adherence. Bacteriocin production was observed only by L. gasseri and Enterococcus faecalis isolates. Antibiotic resistance was widespread, although not universal, among isolates from infants. Highly similar isolates (≥ 97% similarity by barcode match) of Bifidobacterium longum subsp. infantis (1 match), Lactobacillus fermentum (2 matches), Lactobacillus gasseri (6 matches), and Enterococcus faecalis (1 match) were isolated from 5 infant-mother pairs. Antibiotic resistance profiles between these isolate matches were similar, except in one case where the L. gasseri isolate from the infant exhibited resistance to erythromycin and tetracycline, not observed in matching mother isolate. In a second case, L. gasseri isolates differed in resistance to ampicillin, chloramphenicol and vancomycin between the mother and infant. In this study, gram positive bacteria isolated from mothers' breast milk as well as their infants exhibited diversity in GI transit survival and acid inhibition of pathogens, but demonstrated limited ability to produce bacteriocins. Mothers and their infants offer the potential for identification of probiotics; however, even in the early stages of development, healthy infants contain isolates with antibiotic resistance.

Virulence factors and antibiotic susceptibility in enterococci isolated from oral mucosal and deep infections

Directory of Open Access Journals (Sweden)

Gunnar Dahlén

2012-02-01

Full Text Available This study evaluates the presence of virulence factors and antibiotic susceptibility among enterococcal isolates from oral mucosal and deep infections. Forty-three enterococcal strains from oral mucosal lesions and 18 from deep infections were isolated from 830 samples that were sent during 2 years to Oral Microbiology, University of Gothenburg, for analysis. The 61 strains were identified by 16S rDNA, and characterized by the presence of the virulence genes efa A (endocarditis gene, gel E (gelatinase gene, ace (collagen binding antigen gene, asa (aggregation substance gene, cyl A (cytolysin activator gene and esp (surface adhesin gene, tested for the production of bacteriocins and presence of plasmids. MIC determination was performed using the E-test method against the most commonly used antibiotics in dentistry, for example, penicillin V, amoxicillin and clindamycin. Vancomycin was included in order to detect vancomycin-resistant enterococci (VRE strains. Sixty strains were identified as Enterococcus faecalis and one as Enterococcus faecium. All the virulence genes were detected in more than 93.3% (efa A and esp of the E. faecalis strains, while the presence of phenotypic characteristics was much lower (gelatinase 10% and hemolysin 16.7%. Forty-six strains produced bacteriocins and one to six plasmids were detected in half of the isolates. Enterococcal strains from oral infections had a high virulence capacity, showed bacteriocin production and had numerous plasmids. They were generally susceptible to ampicillins but were resistant to clindamycin, commonly used in dentistry, and no VRE-strain was found.

Efficacy of Sakacin on Selected Food Pathogenic Microorganisms ...

African Journals Online (AJOL)

USER

technique and was characterized based on it colony, cell morphology and some ... De Man Rogosa Sharpe (MRS) broth for bacteriocin (sakacin) production. ... tremendous attention as potential bio preservatives in the food and dairy industries. .... positive result,-: Negative result, A : Acid production ,G :Gas production,.

Screening of malting sorghum samples for lactic acid bacteria with ...

African Journals Online (AJOL)

catalase of these inhibitory compounds from the seven bacteria identified them as bacteriocins. Based on standard biochemical and microbiological tests, the isolates were tentatively identified as belonging to Lactococcus spp., Leuconostoc spp., Lactobacillus spp. and Streptococcus spp. However, three isolates (GS3A, ...

Lactococcins : Mode of action, immunity and secretion

NARCIS (Netherlands)

Venema, K; Kok, J; Venema, Gerhardus

1995-01-01

Lactococcus lactis subsp. cremoris 9B4 produces three small (around 5kDa), heat-stable, non-lanthionine containing, membrane active bacteriocins. Amino acid uptake experiments and proton motive force measurements have indicated that these peptides most probably form pores in the cytoplasmic membrane

African Journal of Biotechnology - Vol 8, No 3 (2009)

African Journals Online (AJOL)

Screening for anti-methicillin resistant Staphylococcus aureus (MRSA) bacteriocin producing bacteria · EMAIL FREE FULL TEXT EMAIL FREE FULL TEXT DOWNLOAD ... Seed germination and in vitro propagation of Piliostigma thonningii – an important medicinal plant · EMAIL FREE FULL TEXT EMAIL FREE FULL TEXT

The Centre for Food Innovation -- Research Areas and Potential Projects

Science.gov (United States)

2013-04-01

barley, and rice  frozen red meat (beef, veal, lamb)  processed foods, particularly baby food, wine and beer  fruit juice UNCLASSIFIED 3...support gut health ( probiotics /prebiotics/bacteriocins), development of snack products (bars, FD fruit, HPP fruit, FD yoghurt etc.), novel ingredients

Multiscale Modeling of Microbial Communities

Science.gov (United States)

Blanchard, Andrew

smooth expanding front. More specifically, mutualism promotes spatial homogeneity and population robustness while competition increases spatial segregation and population fluctuations. To examine the generality of these findings, a large set of initial conditions with varying density and species abundance was tested and analyzed. The results and the computational framework presented provide the basis for further explorations of individual based simulations of bacterial communities. For Chapter 4, I consider the role of gene regulation in shaping the outcome of competition between a bacteriocin (i.e. toxin) producing and sensitive strain. In natural systems, bacteriocin production is often conditional, governed by underlying quorum sensing regulatory circuitry. By developing an ordinary differential equation (ODE) model integrating population dynamics with molecular regulation, we find that the ecological contribution of bacteriocin production can be positive or negative, determined by the tradeoff between the benefit of bacteriocins in mediating competition and the fitness cost due to metabolic load. Interestingly, under the naturally occurring scenario where bacteriocin production has a high cost, density-dependent synthesis is more advantageous than constitutive synthesis, which offers a quantitative interpretation for the wide prevalence of density-related bacteriocin production in nature. By incorporating the modeling framework presented in Chapter 3, the results of the ODE model were extended to the spatial setting, providing ecological insights into the costs and benefits of bacteriocin synthesis in competitive environments. For the final research chapter, I consider the impact of growth coupling on protein production at both the single cell and population scales. The same machinery (e.g. ribosomes) and resources (e.g. amino acids and ATP) are used within cells to produce both endogenous (host) and exogenous (circuit) proteins. Thus, the introduction of a gene

2537-IJBCS-Article-Essodolom Taale

African Journals Online (AJOL)

hp

This review focuses on bacteriocins know up to date by highlighting on their particular genetic organization, their mode of ..... sous forme purifiée car leur purification est coûteuse ... C'est la technique d'adsorption qui est la plus utilisée à cause ...

Mode of action of LciA, the lactococcin A immunity protein

NARCIS (Netherlands)

Venema, K.; Haverkort, R.E.; Abee, T.; Haandrikman, A.J.; Leenhouts, K.J.; Leij, L. de; Venema, G.; Kok, J.

Monoclonal antibodies were raised against a fusion between the Escherichia coli maltose-binding protein and LciA, the immunity protein that protects Lactococcus lactis against the effects of the bacteriocin lactococcin A. One of the antibodies directed against the LciA moiety of the fusion protein

In vitro antimicrobial characteristics of bacteriocinproducing ...

African Journals Online (AJOL)

A total of 50 bacteriocin-producing Lactobacillus strains isolated from some Nigerian indigenous fermented foods and beverages (ogi, fufu, garri and nono) and characterized as L. acidophilus, L. casei, L. fermentum, L. lactis and L. plantarum were screened for their inhibitory potentials against food-borne pathogenic ...

Antimicrobials of Bacillus species: mining and engineering

NARCIS (Netherlands)

Zhao, Xin

2016-01-01

Bacillus sp. have been successfully used to suppress various bacterial and fungal pathogens. Due to the wide availability of whole genome sequence data and the development of genome mining tools, novel antimicrobials are being discovered and updated,;not only bacteriocins, but also NRPs and PKs. A

Antibacterial activity and probiotic properties of some lactic acid ...

African Journals Online (AJOL)

Several lactic acid bacteria strains were screened for the production of antibacterial substances active against some pathogenic bacteria. The inhibitory mechanism was investigated and was shown to be dependant of bacteriocin production. The objective was to isolate LAB with antibacterial activity from raib and to select ...

Inhibition of toxicogenic Bacillus cereus in rice-based foods by enterocin AS-48.

Science.gov (United States)

Grande, Maria J; Lucas, Rosario; Abriouel, Hikmate; Valdivia, Eva; Omar, Nabil Ben; Maqueda, Mercedes; Martínez-Bueno, Manuel; Martínez-Cañamero, Magdalena; Gálvez, Antonio

2006-02-01

The antimicrobial effect of the broad-spectrum bacteriocin enterocin AS-48 against the toxicogenic psychrotrophic strain Bacillus cereus LWL1 has been investigated in a model food system consisting of boiled rice and in a commercial infant rice-based gruel dissolved in whole milk stored at temperatures of 37 degrees C, 15 degrees C and 6 degrees C. In food samples supplemented with enterocin AS-48 (in a concentration range of 20-35 mug/ml), viable cell counts decreased rapidly over incubation time, depending on the bacteriocin concentration, the temperature of incubation and the food sample. Enterotoxin production at 37 degrees C was also inhibited. Heat sensitivity of endospores increased markedly in food samples supplemented with enterocin AS-48: inactivation of endospores was achieved by heating for 1 min at 90 degrees C in boiled rice or at 95 degrees C in rice-based gruel. Activity of enterocin AS-48 in rice gruel was potentiated by sodium lactate in a concentration-dependent way.

A novel enterocin T1 with anti-Pseudomonas activity produced by Enterococcus faecium T1 from Chinese Tibet cheese.

Science.gov (United States)

Liu, Hui; Zhang, Lanwei; Yi, Huaxi; Han, Xue; Gao, Wei; Chi, Chunliang; Song, Wei; Li, Haiying; Liu, Chunguang

2016-02-01

An enterocin-producing Enterococcus faecium T1 was isolated from Chinese Tibet cheese. The enterocin was purified by SP-Sepharose and reversed phase HPLC. It was identified as unique from other reported bacteriocins based on molecular weight (4629 Da) and amino acid compositions; therefore it was subsequently named enterocin T1. Enterocin T1 was stable at 80-100 °C and over a wide pH range, pH 3.0-10.0. Protease sensitivity was observed to trypsin, pepsin, papain, proteinase K, and pronase E. Importantly, enterocin T1 was observed to inhibit the growth of numerous Gram-negative and Gram-positive bacteria including Pseudomonas putida, Pseudomonas aeruginosa, Pseudomonas fluorescens, Escherichia coli, Salmonella typhimurium, Shigella flexneri, Shigella sonnei, Staphylococcus aureus, Listeria monocytogenes. Take together, these results suggest that enterocin T1 is a novel bacteriocin with the potential to be used as a bio-preservative to control Pseudomonas spp. in food.

The expression of propionicin PLG-1 gene (plg-1) by lactic starters.

Science.gov (United States)

Mohamed, Sameh E; Tahoun, Mahmoud K

2015-05-01

Propionicin PLG-1 is a bacteriocin produced by Propionibacterium thoenii P127. Such bacteriocin inhibits wide range of food-borne pathogens such as pathogenic Escherichia coli, Pseudomonas aeruginosa, Vibrio parahaemolyticus, Yersinia enterocolitica and a strain of Corynebacterium sp. In the present study, plg-1 gene expressing propionicin PLG-1 was isolated, sequenced for the first time and the resulting sequence was analysed using several web-based bioinformatics programs. The PCR product containing plg-1 gene was transferred to different lactic acid bacterial (LAB) strains using pLEB590 as a cloning vector to give the modified vector pLEBPLG-1. LAB transformants showed an antimicrobial activity against Esch. coli DH5α (most affected strain), Listeria monocytogenes 18116, and Salmonella enterica 25566 as model pathogenic strains. Such LAB transformants can be used in dairy industry to control the food-borne pathogens that are largely distributed worldwide and to feed schoolchildren in the poor countries where dangerous epidemic diseases and diarrhoea prevail.

In Vitro Evaluation of Beneficial Properties of Bacteriocinogenic Lactobacillus plantarum ST8Sh.

Science.gov (United States)

Todorov, Svetoslav Dimitrov; Holzapfel, Wilhelm; Nero, Luis Augusto

2017-06-01

Lactobacillus plantarum ST8Sh, isolated from Bulgarian salami "shpek" and previously characterized as bacteriocin producer, was evaluated for its beneficial properties. Based on the PCR analysis, Lb. plantarum ST8Sh was shown to host a gene related to the production of adhesion proteins such as Mab, Mub, EF, and PrgB. Genetic and physiological tests suggest Lb. plantarum ST8Sh to represent a potential probiotic candidate, including survival in the presence of low levels of pH and high levels of ox bile, production of β-galactosidase, bile salt deconjugation, high level of hydrophobicity, functional auto- and co-aggregation properties, and adhesion to cell lines. Application of semi-purified bacteriocin produced by Lb. plantarum ST8Sh in combination with ciprofloxacin presented synergistic effect on inhibition of Listeria monocytogenes Scott A. Based on observed properties, Lb. plantarum ST8Sh can be considered as a potential probiotic candidate with additional bacteriocinogenic properties.