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      Spatial Localization and Binding of the Probiotic Lactobacillus farciminis to the Rat Intestinal Mucosa: Influence of Chronic Stress

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          Abstract

          The present study aimed at detecting the exogenously applied probiotic Lactobacillus farciminis in rats, after exposure to IBS-like chronic stress, based on 4-day Water Avoidance Stress (WAS). The presence of L. farciminis in both ileal and colonic mucosal tissues was demonstrated by FISH and qPCR, with ileum as the preferential niche, as for the SFB population. A different spatial distribution of the probiotic was observed: in the ileum, bacteria were organized in micro-colonies more or less close to the epithelium whereas, in the colon, they were mainly visualized far away from the epithelium. When rats were submitted to WAS, the L. farciminis population substantially decreased in both intestinal regions, due to a stress-induced increase in colonic motility and defecation, rather than a modification of bacterial binding to the intestinal mucin Muc2.

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          The two mucus layers of colon are organized by the MUC2 mucin, whereas the outer layer is a legislator of host-microbial interactions.

          Malin Johansson, Jessica M Holmén Larsson, Gunnar Hansson (2011)
          The normal intestinal microbiota inhabits the colon mucus without triggering an inflammatory response. The reason for this and how the intestinal mucus of the colon is organized have begun to be unraveled. The mucus is organized in two layers: an inner, stratified mucus layer that is firmly adherent to the epithelial cells and approximately 50 μm thick; and an outer, nonattached layer that is usually approximately 100 μm thick as measured in mouse. These mucus layers are organized around the highly glycosylated MUC2 mucin, forming a large, net-like polymer that is secreted by the goblet cells. The inner mucus layer is dense and does not allow bacteria to penetrate, thus keeping the epithelial cell surface free from bacteria. The inner mucus layer is converted into the outer layer, which is the habitat of the commensal flora. The outer mucus layer has an expanded volume due to proteolytic activities provided by the host but probably also caused by commensal bacterial proteases and glycosidases. The numerous O-glycans on the MUC2 mucin not only serve as nutrients for the bacteria but also as attachment sites and, as such, probably contribute to the selection of the species-specific colon flora. This observation that normal human individuals carry a uniform MUC2 mucin glycan array in colon may indicate such a specific selection.
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            Probiotics and the gut microbiota in intestinal health and disease.

            Melanie Gareau, Philip Sherman, W Allan Walker (2010)
            The use of probiotics is increasing in popularity for both the prevention and treatment of a variety of diseases. While a growing number of well-conducted, prospective, randomized, controlled, clinical trials are emerging and investigations of underlying mechanisms of action are being undertaken, questions remain with respect to the specific immune and physiological effects of probiotics in health and disease. This Review considers recent advances in clinical trials of probiotics for intestinal disorders in both adult and pediatric populations. An overview of recent in vitro and in vivo research related to potential mechanisms of action of various probiotic formulations is also considered.
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              Determination of bacterial load by real-time PCR using a broad-range (universal) probe and primers set.

              Mangala Nadkarni, F. Elizabeth Martin, Nicholas A Jacques … (2002)
              The design and evaluation of a set of universal primers and probe for the amplification of 16S rDNA from the Domain Bacteria to estimate total bacterial load by real-time PCR is reported. Broad specificity of the universal detection system was confirmed by testing DNA isolated from 34 bacterial species encompassing most of the groups of bacteria outlined in Bergey's Manual of Determinative Bacteriology. However, the nature of the chromosomal DNA used as a standard was critical. A DNA standard representing those bacteria most likely to predominate in a given habitat was important for a more accurate determination of total bacterial load due to variations in 16S rDNA copy number and the effect of generation time of the bacteria on this number, since rapid growth could result in multiple replication forks and hence, in effect, more than one copy of portions of the chromosome. The validity of applying these caveats to estimating bacterial load was confirmed by enumerating the number of bacteria in an artificial sample mixed in vitro and in clinical carious dentine samples. Taking these parameters into account, the number of anaerobic bacteria estimated by the universal probe and primers set in carious dentine was 40-fold greater than the total bacterial load detected by culture methods, demonstrating the utility of real-time PCR in the analysis of this environment.
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                Author and article information

                Contributors
                Wenke Feng: Role: Editor
                Journal
                Journal ID (nlm-ta): PLoS One
                Journal ID (iso-abbrev): PLoS ONE
                Journal ID (publisher-id): plos
                Journal ID (pmc): plosone
                Title: PLoS ONE
                Publisher: Public Library of Science (San Francisco, CA USA )
                ISSN (Electronic): 1932-6203
                Publication date (Electronic): 14 September 2015
                Publication date Collection: 2015
                Volume: 10
                Issue: 9
                Electronic Location Identifier: e0136048
                Affiliations
                [1 ]Université de Toulouse, INSA, UPS, INP, LISBP, 135 Avenue de Rangueil, F-31077 Toulouse, France
                [2 ]INRA, UMR792 Ingénierie des Systèmes Biologiques et des Procédés, F-31400 Toulouse, France
                [3 ]CNRS, UMR5504, F-31400 Toulouse, France
                [4 ]UMR 1331 TOXALIM INRA/INPT/UPS, Equipe de NeuroGastroentérologie et Nutrition, 180 chemin de Tournefeuille, 31027 Toulouse cedex 9, France
                [5 ]Univ Lille Nord de France, USTL, Unité de Glycobiologie Structurale et Fonctionnelle, IFR 147, F-59650 Villeneuve d'Ascq, France
                [6 ]CNRS, UMR 8576, F-59650 Villeneuve d'Ascq, France
                [7 ]Faculté de Médecine Rangueil, Centre de Microscopie Electronique Appliquée à la Biologie (CMEAB), Bât. A5, 133 route de Narbonne, 31062 Toulouse cedex, France
                [8 ]Lallemand SA, 19 Rue des Briquetiers, 31702 Blagnac, France
                University of Louisville School of Medicine, UNITED STATES
                Author notes

                Competing Interests: The funder Lallemand SA provided support in the form of salary for an author (A.A.B.). This does not alter the authors' adherence to PLOS ONE policies on sharing data and materials.

                Conceived and designed the experiments: SDS CRM MML PL VT MMB. Performed the experiments: SDS CRM AR MML CSC BR RL IF AAB MMB. Analyzed the data: SDS CRM AR MML BR RL VT MMB. Contributed reagents/materials/analysis tools: SDS CRM MML CSC BR RL IF AAB. Wrote the paper: SDS CRM VT MMB.

                Article
                Publisher ID: PONE-D-15-19334
                DOI: 10.1371/journal.pone.0136048
                PMC ID: 4569280
                PubMed ID: 26367538
                SO-VID: ab397013-d506-4171-a80a-08daf985d15d
                Copyright statement: Copyright @ 2015
                License:

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited

                History
                Date received : 27 May 2015
                Date accepted : 29 July 2015
                Page count
                Figures: 8, Tables: 0, Pages: 16
                Funding
                This work was supported by INRA (CEPIA Division; http://www.inra.fr) and CNRS (INSIS Institute; www.cnrs.fr) through the doctoral grant of Stéphanie Da Silva (2010-2013). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The funder Lallemand SA provided support in the form of salary for author (Afifa Ait-Belgnaoui), but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section. No additional external funding was received for this study.
                Categories
                Subject: Research Article
                Custom metadata
                Data Availability All relevant data are within the paper and its Supporting Information files.

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