title: Metabolic in Vivo Labeling Highlights Differences of Metabolically Active Microbes from the Mucosal Gastrointestinal Microbiome between High-Fat and Normal Chow Diet.
abstract: The gastrointestinal microbiota in the gut interacts metabolically and immunologically with the host tissue in the contact zone of the mucus layer. For understanding the details of these interactions and especially their dynamics it is crucial to identify the metabolically active subset of the microbiome. This became possible by the development of stable isotope probing techniques, which have only sparsely been applied to microbiome research. We applied the in vivo stable isotope approach using15N-labeled diet with subsequent identification of metabolically active bacterial species. Four-week old male Sprague-Dawley rats were randomly assigned to chow diet (CD, n =15) and high-fat diet (HFD, n =15). After 11 weeks, three animals from each group were sacrificed for baseline characterization of anthropometric and metabolic obesity. The remaining animals were exposed to either a15N-labeled (n =9) or a14N-unlabeled experimental diet (n =3). Three rats from each cohort (HFD and CD) were sacrificed at 12, 24, and 72 h. The remaining three animals from each cohort, which received the14N-unlabeled diet, were sacrificed after 72 h. The colon was harvested and divided into three equal sections (proximal, medial, and distal), and the mucus layer of each specimen was sampled by scraping. We identified the active subset in an HFD model of obesity in comparison with lean controls rats using metaproteomics. In addition, all samples were investigated by 16S rRNA amplicon gene sequencing. The active microbiome of the HFD group showed an increase in bacterial taxa for Verrucomicrobia and Desulfovibrionaceae. In contrast with no significant changes in alpha diversity, time- and localization-dependent effects in beta-diversity were clearly observed. In terms of enzymatic functions the HFD group showed strong affected metabolic pathways such as energy production and carbohydrate metabolism. In vivo isotope labeling combined with metaproteomics provides a valuable method to distinguish the active from the non-active bacterial phylogenetic groups that are relevant for microbiota-host interaction. For morbid obesity such analysis may provide potentially new strategies for targeted pre- or probiotic treatments.
authors.affiliation: Department of Cardiac Surgery, University of Leipzig, Heart Center , 04289 Leipzig, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Oberbach
authors.affiliation: Department of Molecular Systems Biology, UFZ-Helmholtz Centre for Environmental Research , 04318 Leipzig, Germany.
authors.forename: Sven-Bastiaan
authors.initials: SB
authors.lastname: Haange
authors.affiliation: Department of Cardiac Surgery, University of Leipzig, Heart Center , 04289 Leipzig, Germany.|Department of Pediatric Surgery, University of Leipzig , 04103 Leipzig, Germany.
authors.forename: Nadine
authors.initials: N
authors.lastname: Schlichting
authors.affiliation: Department of Cardiac Surgery, University of Leipzig, Heart Center , 04289 Leipzig, Germany.|Integrated Research and Treatment Center (IFB) Adiposity Diseases, University of Leipzig , 04103 Leipzig, Germany.
authors.forename: Marco
authors.initials: M
authors.lastname: Heinrich
authors.affiliation: Integrated Research and Treatment Center (IFB) Adiposity Diseases, University of Leipzig , 04103 Leipzig, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Lehmann
authors.affiliation: Department of Paediatric and Adolescent Surgery, Medical University of Graz , 8036 Graz, Austria.
authors.forename: Holger
authors.initials: H
authors.lastname: Till
authors.affiliation: Laboratory of Microbiology, Wageningen University , 6708 PB Wageningen, The Netherlands.
authors.forename: Floor
authors.initials: F
authors.lastname: Hugenholtz
authors.affiliation: Department of Cardiac Surgery, University of Leipzig, Heart Center , 04289 Leipzig, Germany.|Integrated Research and Treatment Center (IFB) Adiposity Diseases, University of Leipzig , 04103 Leipzig, Germany.
authors.forename: Yvonne
authors.initials: Y
authors.lastname: Kullnick
authors.affiliation: Laboratory of Microbiology, Wageningen University , 6708 PB Wageningen, The Netherlands.
authors.forename: Hauke
authors.initials: H
authors.lastname: Smidt
authors.affiliation: Department of Ecological Modelling, UFZ-Helmholtz Centre for Environmental Research , 04318 Leipzig, Germany.
authors.forename: Karin
authors.initials: K
authors.lastname: Frank
authors.affiliation: Department of Molecular Systems Biology, UFZ-Helmholtz Centre for Environmental Research , 04318 Leipzig, Germany.
authors.forename: Jana
authors.initials: J
authors.lastname: Seifert
authors.affiliation: Department of Molecular Systems Biology, UFZ-Helmholtz Centre for Environmental Research , 04318 Leipzig, Germany.
authors.forename: Nico
authors.initials: N
authors.lastname: Jehmlich
authors.affiliation: Department of Molecular Systems Biology, UFZ-Helmholtz Centre for Environmental Research , 04318 Leipzig, Germany.|Institute of Biochemistry, Faculty of Biosciences, Pharmacy and Psychology, University of Leipzig , 04103 Leipzig, Germany.|Department of Life Sciences and Chemistry, Centre for Microbial Communities, University of Aalborg , 9220 Aalborg, Denmark.
authors.forename: Martin
authors.initials: M
authors.lastname: von Bergen
mesh_terms: D000818:Animals; D059305:Diet, High-Fat; D000069196:Gastrointestinal Microbiome / Q000235:genetics*; D041981:Gastrointestinal Tract / Q000382:microbiology*; D054884:Host-Pathogen Interactions / Q000235:genetics; D007553:Isotope Labeling; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D009092:Mucous Membrane / Q000382:microbiology; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D061327:Verrucomicrobia / Q000235:genetics* / Q000302:isolation & purification
chemical_list: D012336:RNA, Ribosomal, 16S
keywords: 16S rRNA gene sequencing; gut microbiota; metaproteomics; mucus layer; protein-based stable isotope probing