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Functional characterization of probiotic surface layer protein-carrying Lactobacillus amylovorus strains.

ABSTRACT: BACKGROUND: Adhesiveness to intestinal epithelium, beneficial immunomodulating effects and the production of pathogen-inhibitory compounds are generally considered as beneficial characteristics of probiotic organisms. We showed the potential health-promoting properties and the mechanisms of probiotic action of seven swine intestinal Lactobacillus amylovorus isolates plus the type strain (DSM 20531T) by investigating their adherence to porcine intestinal epithelial cells (IPEC-1) and mucus as well as the capacities of the strains to i) inhibit the adherence of Escherichia coli to IPEC-1 cells, ii) to produce soluble inhibitors against intestinal pathogens and iii) to induce immune signaling in dendritic cells (DCs). Moreover, the role of the L. amylovorus surface (S) -layers - symmetric, porous arrays of identical protein subunits present as the outermost layer of the cell envelope - in adherence to IPEC-1 cells was assessed using a novel approach which utilized purified cell wall fragments of the strains as carriers for the recombinantly produced S-layer proteins. RESULTS: Three of the L. amylovorus strains studied adhered to IPEC-1 cells, while four strains inhibited the adherence of E. coli, indicating additional mechanisms other than competition for binding sites being involved in the inhibition. None of the strains bound to porcine mucus. The culture supernatants of all of the strains exerted inhibitory effects on the growth of E. coli, Salmonella, Listeria and Yersinia, and a variable, strain-dependent induction was observed of both pro- and anti-inflammatory cytokines in human DCs. L. amylovorus DSM 16698 was shown to carry two S-layer-like proteins on its surface in addition to the major S-layer protein SlpA. In contrast to expectations, none of the major S-layer proteins of the IPEC-1 -adhering strains mediated bacterial adherence. CONCLUSIONS: We demonstrated adhesive and significant pathogen inhibitory efficacies among the swine intestinal L. amylovorus strains studied, pointing to their potential use as probiotic feed supplements, but no independent role could be demonstrated for the major S-layer proteins in adherence to epithelial cells. The results indicate that many intestinal bacteria may coexist with and confer benefits to the host by mechanisms not attributable to adhesion to epithelial cells or mucus.

SUBMITTER: Hynonen U

PROVIDER: S-EPMC4236617 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Functional characterization of probiotic surface layer protein-carrying Lactobacillus amylovorus strains.

Hynönen Ulla U Kant Ravi R Lähteinen Tanja T Pietilä Taija E TE Beganović Jasna J Smidt Hauke H Uroić Ksenija K Avall-Jääskeläinen Silja S Palva Airi A

BMC microbiology 20140728

<h4>Background</h4>Adhesiveness to intestinal epithelium, beneficial immunomodulating effects and the production of pathogen-inhibitory compounds are generally considered as beneficial characteristics of probiotic organisms. We showed the potential health-promoting properties and the mechanisms of probiotic action of seven swine intestinal Lactobacillus amylovorus isolates plus the type strain (DSM 20531T) by investigating their adherence to porcine intestinal epithelial cells (IPEC-1) and mucus ...[more]

PMID: 25070625

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Project description:BACKGROUND:Probiotics are defined as adequate amount of live microorganisms able to confer health benefits on the host. Currently, most commercially available probiotic products in the market belong to genera Lactobacillus. Traditional dairy products are usually rich source of Lactobacillus strains with significant health benefits. In order to evaluate the probiotic potential of these bacteria, it is essential to assess their health benefits, efficacy, and safety. OBJECTIVES:The probiotic efficacy of two Lactobacillus strains namely Lactobacillus pentosus LP05 and L. brevis LB32 was evaluated. They were previously isolated from ewes' milk in a rural area in East Azerbaijan, Iran. MATERIALS AND METHODS:The selected isolates were tested for certain phenotypic characters and identified to genus and species level by 16S rRNA gene sequencing and species specific primers. Further analysis included acid and bile resistance, antagonistic activity, cholesterol removing ability, survival in simulated gastric and upper intestine contents, aggregation and coaggregation properties. Finally, the adhering ability of the selected Lactobacillus strains to epithelial cells was tested using Caco-2 cell lines. RESULTS:The selected isolates tolerated bile salt concentrations ranging from 0.5% to 3%, however their coefficient of inhibition were varied. Both isolates hydrolyzed bile and grew at pH values of 3, 4, and 5, while isolate LP05 was not able to hydrolyze arginine. Based on 16s rRNA gene sequencing and species-specific primers, the isolates were identified as L. brevis LB32 and L. pentosus LP05. In contrast to simulated gastric conditions, the growth rate of the isolates in alkaline conditions of upper intestine increased significantly with the passage of time reaching its maximum in 24 hours. These 2 isolates inhibited the growth of Listeria monocytogenes, Salmonella enteritidis, Shigella dysenteriae, Staphylococcus aureus, and Streptococcus pneumonia. Furthermore, L. brevis LB32 was able to reduce approximately 86% of cholesterol compared to L. pentosus LP05, which showed only 69% of reduction. Higher aggregation and coaggregation percentage and adherence to Caco-2 cell line was observed in L. pentosus LP05 compared to L. brevis LB32. CONCLUSIONS:This research study proved the presence of viable probiotic LAB microflora in the ewe milk with enhanced health benefits. The 2 selected Lactobacillus strains could be exploited in dairy or pharmaceutical industry in future.

Project description:BACKGROUND:We evaluated the functional capacity of plantaricin-producing Lactobacillus plantarum SF9C and S-layer-carrying Lactobacillus brevis SF9B to withstand gastrointestinal transit and to compete among the gut microbiota in vivo. Considering the probiotic potential of Lb. brevis SF9B, this study aims to investigate the antibacterial activity of Lb. plantarum SF9C and their potential for in vivo colonisation in rats, which could be the basis for the investigation of their synergistic functionality. RESULTS:A plantaricin-encoding cluster was identified in Lb. plantarum SF9C, a strain which efficiently inhibited the growth of Listeria monocytogenes ATCC® 19111™ and Staphylococcus aureus 3048. Homology-based three-dimensional (3D) structures of SF9C plantaricins PlnJK and PlnEF were predicted using SWISS-MODEL workspace and the helical wheel representations of the plantaricin peptide helices were generated by HELIQUEST. Contrary to the plantaricin-producing SF9C strain, the S-layer-carrying SF9B strain excluded Escherichia coli 3014 and Salmonella enterica serovar Typhimurium FP1 from the adhesion to Caco-2 cells. Finally, PCR-DGGE analysis of the V2-V3 regions of the 16S rRNA gene confirmed the transit of the two selected lactobacilli through the gastrointestinal tract (GIT). Microbiome profiling via the Illumina MiSeq platform revealed the prevalence of Lactobacillus spp. in the gut microbiota of the Lactobacillus-treated rats, even on the 10th day after the Lactobacillus application, compared to the microbiota of the healthy and AlCl3-exposed rats before Lactobacillus treatment. CONCLUSION:The combined application of Lb. plantarum SF9C and Lb. brevis SF9B was able to influence the intestinal microbiota composition in rats, which was reflected in the increased abundance of Lactobacillus genus, but also in the altered abundances of other bacterial genera, either in the model of healthy or aberrant gut microbiota of rats. The antibacterial activity and capacity to withstand in GIT conditions contributed to the functional aspects of SF9C and SF9B strains that could be incorporated in the probiotic-containing functional foods with a possibility to positively modulate the gut microbiota composition.

Dataset Information

Functional characterization of probiotic surface layer protein-carrying Lactobacillus amylovorus strains.

Publications

Functional characterization of probiotic surface layer protein-carrying Lactobacillus amylovorus strains.

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