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Paul W. O'toole - One of the best experts on this subject based on the ideXlab platform.
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High-resolution structures of Lactobacillus salivarius transketolase in the presence and absence of thiamine pyrophosphate
Acta crystallographica. Section F Structural biology communications, 2015Co-Authors: Petra Lukacik, Mario Bumann, Carina M C Lobley, Raymond J. Owens, Paul W. O'toole, Victoria Arena De Souza, Martin A. WalshAbstract:Probiotic bacterial strains have been shown to enhance the health of the host through a range of mechanisms including colonization, resistance against pathogens, secretion of antimicrobial compounds and modulation of the activity of the innate immune system. Lactobacillus salivarius UCC118 is a well characterized probiotic strain which survives intestinal transit and has many desirable host-interaction properties. Probiotic bacteria display a wide range of catabolic activities, which determine their competitiveness in vivo. Some lactobacilli are heterofermentative and can metabolize pentoses, using a pathway in which transketolase and transaldolase are key enzymes. L. salivarius UCC118 is capable of pentose utilization because it encodes the key enzymes on a megaplasmid. The crystal structures of the megaplasmid-encoded transketolase with and without the enzyme cofactor thiamine pyrophosphate have been determined. Comparisons with other known transketolase structures reveal a high degree of structural conservation in both the catalytic site and the overall conformation. This work extends structural knowledge of the transketolases to the industrially and commercially important Lactobacillus genus
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Structure of ribose 5-phosphate isomerase from the probiotic bacterium Lactobacillus salivarius UCC118.
Acta crystallographica. Section F Structural biology and crystallization communications, 2012Co-Authors: Carina M C Lobley, Mario Bumann, Pierre Aller, Alice Douangamath, Yamini Reddivari, Louise E Bird, Joanne E Nettleship, Jose Brandao-neto, Raymond J. Owens, Paul W. O'tooleAbstract:The structure of ribose 5-phosphate isomerase from the probiotic bacterium Lactobacillus salivarius UCC188 has been determined at 1.72 Å resolution. The structure was solved by molecular replacement, which identified the functional homodimer in the asymmetric unit. Despite only showing 57% sequence identity to its closest homologue, the structure adopted the typical α and β D-ribose 5-phosphate isomerase fold. Comparison to other related structures revealed high homology in the active site, allowing a model of the substrate-bound protein to be proposed. The determination of the structure was expedited by the use of in situ crystallization-plate screening on beamline I04-1 at Diamond Light Source to identify well diffracting protein crystals prior to routine cryocrystallography.
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Genomic Diversity of Lactobacillus salivarius
Applied and environmental microbiology, 2010Co-Authors: Emma J. Raftis, Elisa Salvetti, Sandra Torriani, Giovanna E. Felis, Paul W. O'tooleAbstract:Strains of Lactobacillus salivarius are increasingly employed as probiotic agents for humans or animals. Despite the diversity of environmental sources from which they have been isolated, the genomic diversity of L. salivarius has been poorly characterized, and the implications of this diversity for strain selection have not been examined. To tackle this, we applied comparative genomic hybridization (CGH) and multilocus sequence typing (MLST) to 33 strains derived from humans, animals, or food. The CGH, based on total genome content, including small plasmids, identified 18 major regions of genomic variation, or hot spots for variation. Three major divisions were thus identified, with only a subset of the human isolates constituting an ecologically discernible group. Omission of the small plasmids from the CGH or analysis by MLST provided broadly concordant fine divisions and separated human-derived and animal-derived strains more clearly. The two gene clusters for exopolysaccharide (EPS) biosynthesis corresponded to regions of significant genomic diversity. The CGH-based groupings of these regions did not correlate with levels of production of bound or released EPS. Furthermore, EPS production was significantly modulated by available carbohydrate. In addition to proving difficult to predict from the gene content, EPS production levels correlated inversely with production of biofilms, a trait considered desirable in probiotic commensals. L. salivarius displays a high level of genomic diversity, and while selection of L. salivarius strains for probiotic use can be informed by CGH or MLST, it also requires pragmatic experimental validation of desired phenotypic traits.
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Probiotic properties of Lactobacillus salivarius and closely related Lactobacillus species.
Future microbiology, 2010Co-Authors: B A Neville, Paul W. O'tooleAbstract:Lactobacillus salivarius has been frequently isolated from the mammalian digestive tract and has been studied as a candidate probiotic. Research to date has described the immunomodulatory properties of the species in cell-lines, mice, rats and humans for the alleviation of intestinal disease and the promotion of host well-being. The ability of L. salivarius to inhibit pathogens and tolerate host antimicrobial defenses demonstrates the adaptation of this species to the gastrointestinal niche. L. salivarius is the best characterized of 25 species in the L. salivarius clade of the genus Lactobacillus. Several other species of this clade are candidate probiotics; however, their probiotic potential has not yet been exploited. This review summarizes the research defining the probiotic nature of L. salivarius, by focusing in particular on L. salivarius UCC118 as a representative strain. The emergent research detailing the probiotic potential of other species in this phylogenetic clade will also be discussed.
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Megaplasmid pMP118 of Lactobacillus salivarius
Microbial Megaplasmids, 2009Co-Authors: Paul W. O'toole, Emma J. RaftisAbstract:Lactobacillus salivarius UCC118 habours a multiple-replicon genome, including a 242 kb megaplasmid designated pMP118. pMP118 carries a number of contingency genes, which work in conjunction with chromosomally encoded genes and pathways to broaden the metabolic flexibility of this strain. This increases the potential viability of UCC118 in the competitive environment of the gastrointestinal tract. Annotation and functional studies have indicated that pMP118 contributes significantly to the probiotic properties of UCC118, encoding a bile salt hydrolase gene and a potent broad-spectrum bacteriocin. pMP118-related megaplasmids have been established as a general feature of the species Lactobacillus salivarius. Megaplasmids have also been identified in six other Lactobacillus species of intestinal origin. Dissemination of the pMP118-related megaplasmids may have occurred by a conjugation apparatus, which is now non-functional in pMP118. The analysis of pMP118 highlights the contribution of this replicon to the biology and ecology of this commensal species.
Luis M. Quiros - One of the best experts on this subject based on the ideXlab platform.
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Surface glycosaminoglycans mediate adherence between HeLa cells and Lactobacillus salivarius Lv72
BMC microbiology, 2013Co-Authors: Rebeca Martín, Carla Martin, Susana Escobedo, Juan E. Suárez, Luis M. QuirosAbstract:Background The adhesion of lactobacilli to the vaginal surface is of paramount importance to develop their probiotic functions. For this reason, the role of HeLa cell surface proteoglycans in the attachment of Lactobacillus salivarius Lv72, a mutualistic strain of vaginal origin, was investigated.
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Surface glycosaminoglycans mediate adherence between HeLa cells and Lactobacillus salivarius Lv72
BMC Microbiology, 2013Co-Authors: Rebeca Martín, Carla Martin, Susana Escobedo, Juan E. Suárez, Luis M. QuirosAbstract:Background: The adhesion of lactobacilli to the vaginal surface is of paramount importance to develop their probiotic functions. For this reason, the role of HeLa cell surface proteoglycans in the attachment of Lactobacillus salivarius Lv72, a mutualistic strain of vaginal origin, was investigated. Results: Incubation of cultures with a variety of glycosaminoglycans (chondroitin sulfate A and C, heparin and heparan sulfate) resulted in marked binding interference. However, no single glycosaminoglycan was able to completely abolish cell binding, the sum of all having an additive effect that suggests cooperation between them and recognition of specific adhesins on the bacterial surface. In contrast, chondroitin sulfate B enhanced cell to cell attachment, showing the relevance of the stereochemistry of the uronic acid and the sulfation pattern on binding. Elimination of the HeLa surface glycosaminoglycans with lyases also resulted in severe adherence impairment. Advantage was taken of the Lactobacillus-glycosaminoglycans interaction to identify an adhesin from the bacterial surface. This protein, identify as a soluble binding protein of an ABC transporter system (OppA) by MALDI-TOF/(MS), was overproduced in Escherichia coli, purified and shown to interfere with L. salivarius Lv72 adhesion to HeLa cells. Conclusions: These data suggest that glycosaminoglycans play a fundamental role in attachment of mutualistic bacteria to the epithelium that lines the cavities where the normal microbiota thrives, OppA being a bacterial adhesin involved in the process.
Michèle Dalgalarrondo - One of the best experts on this subject based on the ideXlab platform.
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Purification and genetic characterisation of the novel bacteriocin LS2 produced by the human oral strain Lactobacillus salivarius BGHO1
International journal of antimicrobial agents, 2012Co-Authors: M. Busarcevic, Michèle DalgalarrondoAbstract:Abstract 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.
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Purification and characterization of a new bacteriocin active against Campylobacter produced by Lactobacillus salivarius SMXD51.
Food microbiology, 2012Co-Authors: Soumaya Messaoudi, Mohamed Manai, Gilles Kergourlay, Hervé Prévost, Jean-marc Chobert, Michèle Dalgalarrondo, Yvan Choiset, Mounir Ferchichi, Marie-france Pilet, Xavier DoussetAbstract: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 UPTICLEAN cartridge using different concentrations of acetonitrile, (iii) final purification by reversed phase HPLC on a C18 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.
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Purification and characterization of a new bacteriocin active against Campylobacter produced by Lactobacillus salivarius SMXD51
Food Microbiology, 2012Co-Authors: Soumaya Messaoudi, Mohamed Manai, Gilles Kergourlay, Hervé Prévost, Jean-marc Chobert, Michèle Dalgalarrondo, Yvan Choiset, Mounir Ferchichi, Marie-france Pilet, Xavier DoussetAbstract: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. (C) 2012 Elsevier Ltd. All rights reserved.
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Purification and genetic characterisation of the novel bacteriocin LS2 produced by the human oral strain Lactobacillus salivarius BGHO1
International Journal of Antimicrobial Agents, 2012Co-Authors: M. Busarcevic, Michèle DalgalarrondoAbstract: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 4115.1 Da 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. (C) 2012 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.
Rebeca Martín - One of the best experts on this subject based on the ideXlab platform.
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Surface glycosaminoglycans mediate adherence between HeLa cells and Lactobacillus salivarius Lv72
BMC microbiology, 2013Co-Authors: Rebeca Martín, Carla Martin, Susana Escobedo, Juan E. Suárez, Luis M. QuirosAbstract:Background The adhesion of lactobacilli to the vaginal surface is of paramount importance to develop their probiotic functions. For this reason, the role of HeLa cell surface proteoglycans in the attachment of Lactobacillus salivarius Lv72, a mutualistic strain of vaginal origin, was investigated.
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Surface glycosaminoglycans mediate adherence between HeLa cells and Lactobacillus salivarius Lv72
BMC Microbiology, 2013Co-Authors: Rebeca Martín, Carla Martin, Susana Escobedo, Juan E. Suárez, Luis M. QuirosAbstract:Background: The adhesion of lactobacilli to the vaginal surface is of paramount importance to develop their probiotic functions. For this reason, the role of HeLa cell surface proteoglycans in the attachment of Lactobacillus salivarius Lv72, a mutualistic strain of vaginal origin, was investigated. Results: Incubation of cultures with a variety of glycosaminoglycans (chondroitin sulfate A and C, heparin and heparan sulfate) resulted in marked binding interference. However, no single glycosaminoglycan was able to completely abolish cell binding, the sum of all having an additive effect that suggests cooperation between them and recognition of specific adhesins on the bacterial surface. In contrast, chondroitin sulfate B enhanced cell to cell attachment, showing the relevance of the stereochemistry of the uronic acid and the sulfation pattern on binding. Elimination of the HeLa surface glycosaminoglycans with lyases also resulted in severe adherence impairment. Advantage was taken of the Lactobacillus-glycosaminoglycans interaction to identify an adhesin from the bacterial surface. This protein, identify as a soluble binding protein of an ABC transporter system (OppA) by MALDI-TOF/(MS), was overproduced in Escherichia coli, purified and shown to interfere with L. salivarius Lv72 adhesion to HeLa cells. Conclusions: These data suggest that glycosaminoglycans play a fundamental role in attachment of mutualistic bacteria to the epithelium that lines the cavities where the normal microbiota thrives, OppA being a bacterial adhesin involved in the process.
Marı́a Elena Nader-macı́as - One of the best experts on this subject based on the ideXlab platform.
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Influence of pH, temperature and culture media on the growth and bacteriocin production by vaginal Lactobacillus salivarius CRL 1328.
Journal of applied microbiology, 2002Co-Authors: M.s. Juárez Tomás, Elena Bru, B. Wiese, A. P. De Ruiz Holgado, Marı́a Elena Nader-macı́asAbstract:Aims To study the influence of pH, temperature and culture medium on the growth and bacteriocin production by vaginal Lactobacillus salivarius subsp. salivarius CRL 1328. Methods and results The study was performed using a complete factorial experimental design. Lactobacillus salivarius was cultivated in LAPTg and MRS broths, adjusted to specific initial pH, and at different temperatures of incubation. The growth, which was evaluated by the Gompertz model, was higher in MRS broth than in LAPTg broth. The initial pH of the culture medium and the temperature had a dramatic effect on the production of bacteriocin. The optimal conditions for bacteriocin production were different to those for optimal growth. The decrease in the pH of the culture medium was parallel to the growth; pH had similar final values in both the MRS and the LAPTg broths. Conclusions The optimal growth conditions were recorded in MRS broth, with an initial pH of 6.5 and a temperature of 37 degrees C. The maximum bacteriocin activity was obtained in LAPTg after 6 h at 37 degrees C, and at an initial pH of 6.5 or 8.0. Significance and impact of the study The application of a complete factorial design, and the evaluation of the growth parameters through the Gompertz model, enabled a rapid and simultaneous exploration of the influence of pH, temperature and growth medium on both growth and bacteriocin production by vaginal Lact. salivarius CRL 1328.
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Characterization of a bacteriocin-like substance produced by a vaginal Lactobacillus salivarius strain.
Applied and environmental microbiology, 1999Co-Authors: Virginia S. Ocaña, Aida Pesce De Ruiz Holgado, Marı́a Elena Nader-macı́asAbstract: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.
