Project description:This SuperSeries is composed of the following subset Series: GSE11944: Mucosal Glycan Foraging Enhances the Fitness and Transmission of a Saccharolytic Human Distal Gut Symbiont GSE11953: Mucosal Glycan Foraging Enhances the Fitness and Transmission of a Saccharolytic Human Distal Gut Symbiont: ECF mutant GSE11962: Growth of B. thetaiotaomicron on purified host mucosal glycans and glycan fragments Refer to individual Series
Project description:Symbiotic bacteria inhabiting the distal human gut have evolved under intense pressure to utilize complex carbohydrates, predominantly plant cell wall glycans abundant in our diets. These substrates are recalcitrant to depolymerization by digestive enzymes encoded in the human genome, but are efficiently targeted by some of the ~103-104 bacterial species that inhabit this niche. These species augment our comparatively narrow carbohydrate digestive capacity by unlocking otherwise unusable sugars and fermenting them into host-absorbable forms, such as short-chain fatty acids. We used phenotype profiling, whole-genome transcriptional analysis and molecular genetic approaches to investigate complex glycan utilization by two fully sequenced and closely related human gut symbionts: Bacteroides thetaiotaomicron and Bacteroides ovatus. Together these species target all of the common glycosidic linkages found in the plant cell wall, as well as host polysaccharides, but each species exhibits a unique ‘glycan niche’: in vitro B. thetaiotaomicron targets plant cell wall pectins in addition to linkages contained in host N- and O-glycans; B. ovatus uniquely targets hemicellulosic polysaccharides along with several pectins, but is deficient in host glycan utilization. Growth of Bacteroides thetaiotaomicron in vitro in minimal medium plus different purified complex glycans. Observation of increased gene expression was used to determine genes that are involved in metabolism of each glycan. Two biological replicates each.
Project description:Symbiotic bacteria inhabiting the distal human gut have evolved under intense pressure to utilize complex carbohydrates, predominantly plant cell wall glycans abundant in our diets. These substrates are recalcitrant to depolymerization by digestive enzymes encoded in the human genome, but are efficiently targeted by some of the ~103-104 bacterial species that inhabit this niche. These species augment our comparatively narrow carbohydrate digestive capacity by unlocking otherwise unusable sugars and fermenting them into host-absorbable forms, such as short-chain fatty acids. We used phenotype profiling, whole-genome transcriptional analysis and molecular genetic approaches to investigate complex glycan utilization by two fully sequenced and closely related human gut symbionts: Bacteroides thetaiotaomicron and Bacteroides ovatus. Together these species target all of the common glycosidic linkages found in the plant cell wall, as well as host polysaccharides, but each species exhibits a unique ‘glycan niche’: in vitro B. thetaiotaomicron targets plant cell wall pectins in addition to linkages contained in host N- and O-glycans; B. ovatus uniquely targets hemicellulosic polysaccharides along with several pectins, but is deficient in host glycan utilization.
Project description:Bacteroides thetaiotaomicron was grown and transcriptionally profiled on a number of different host mucosal glycans and their component mono- and disaccharides.
Project description:Symbiotic bacteria inhabiting the distal human gut have evolved under intense pressure to utilize complex carbohydrates, predominantly plant cell wall glycans abundant in our diets. These substrates are recalcitrant to depolymerization by digestive enzymes encoded in the human genome, but are efficiently targeted by some of the ~103-104 bacterial species that inhabit this niche. These species augment our comparatively narrow carbohydrate digestive capacity by unlocking otherwise unusable sugars and fermenting them into host-absorbable forms, such as short-chain fatty acids. We used phenotype profiling, whole-genome transcriptional analysis and molecular genetic approaches to investigate complex glycan utilization by two fully sequenced and closely related human gut symbionts: Bacteroides thetaiotaomicron and Bacteroides ovatus. Together these species target all of the common glycosidic linkages found in the plant cell wall, as well as host polysaccharides, but each species exhibits a unique ‘glycan niche’: in vitro B. thetaiotaomicron targets plant cell wall pectins in addition to linkages contained in host N- and O-glycans; B. ovatus uniquely targets hemicellulosic polysaccharides along with several pectins, but is deficient in host glycan utilization. Bacteroides ovatus bacteria were grown either in vitro on defined complex glycan sources, or in vivo in the intestinal tract of gnotobiotic mice fed variable diets. Increased in vitro gene expression was used to indicate the genes required for metabolism of complex glycans and compared to in vivo transcriptional activity to determine expression in the mouse gut.
Project description:Symbiotic bacteria inhabiting the distal human gut have evolved under intense pressure to utilize complex carbohydrates, predominantly plant cell wall glycans abundant in our diets. These substrates are recalcitrant to depolymerization by digestive enzymes encoded in the human genome, but are efficiently targeted by some of the ~103-104 bacterial species that inhabit this niche. These species augment our comparatively narrow carbohydrate digestive capacity by unlocking otherwise unusable sugars and fermenting them into host-absorbable forms, such as short-chain fatty acids. We used phenotype profiling, whole-genome transcriptional analysis and molecular genetic approaches to investigate complex glycan utilization by two fully sequenced and closely related human gut symbionts: Bacteroides thetaiotaomicron and Bacteroides ovatus. Together these species target all of the common glycosidic linkages found in the plant cell wall, as well as host polysaccharides, but each species exhibits a unique ‘glycan niche’: in vitro B. thetaiotaomicron targets plant cell wall pectins in addition to linkages contained in host N- and O-glycans; B. ovatus uniquely targets hemicellulosic polysaccharides along with several pectins, but is deficient in host glycan utilization.
Project description:The large-scale application of genomic and metagenomic sequencing technologies has yielded a number of insights about the metabolic potential of symbiotic human gut microbes. Bacteria that colonize the mucosal layer that overlies the gut epithelium have access to highly-sulfated polysaccharides (i.e., mucin oligosaccharides and glycosaminoglycans), which they could potentially forage as nutrient sources. To be active, sulfatases must undergo a critical post-translational modification catalyzed in anaerobic bacteria by the AdoMet enzyme anSME (anaerobic Sulfatase-Maturating Enzyme). In the present study, we have tested the role of this pathway in the prominent gut symbiont Bacteroides thetaiotaomicron, which possesses more predicted sulfatases (28) than in the human genome and a single predicted anSME. In vitro studies revealed that deletion of its anSME (BT0238) results in loss of sulfatase activity and impaired ability to use sulfated polysaccharides as carbon sources. Co-colonization of germ-free animals with both isogenic strains, or invasion experiments involving the introduction of one then the other strain, established that anSME activity and the sulfatases that are activated via this pathway, are important fitness factors for B. thetaiotaomicron, especially when mice are fed a simple sugar diet that requires this saccharolytic bacterium to adaptively forage on host glycans as nutrients. Whole genome transcriptional profiling of wild-type and the anSME mutant in vivo revealed that loss of this enzyme alters expression of genes involved in mucin utilization and that this disrupted ability to access mucosal glycans likely underlies the observed dramatic colonization defect. Comparative genomic analysis reveals that 100% of 46 fully sequenced human gut Bacteroidetes contain homologs of BT0238 and genes encoding sulfatases, suggesting that this is an important and evolutionarily conserved feature. Three replicate samples from 4 different biological treatment groups: 1. Wild-type B. thetaiotaomicron from the cecum of gnotobiotic mice fed a simple-sugar diet; 2. chuR mutant B. thetaiotaomicron from the cecum of gnotobiotic mice fed a simple-sugar diet; 3. Wild-type B. thetaiotaomicron from the cecum of gnotobiotic mice fed a plant-rich diet; 4. chuR mutant B. thetaiotaomicron from the cecum of gnotobiotic mice fed a plant-rich diet.
Project description:The adult human gut microbial community is typically dominated by two bacterial phyla (divisions), the Firmicutes and the Bacteroidetes. Little is known about the factors that govern the interactions between their members. Here we examine the niches of representatives of both phyla in vivo. Finished genome sequences were generated from E. rectale and E. eligens, which belong to Clostridium Cluster XIVa, one of the most common gut Firmicute clades. Comparison of these and 25 other gut Firmicutes and Bacteroidetes indicated that the former possess smaller genomes and a disproportionately smaller number of glycan-degrading enzymes. Germ-free mice were then colonized with E. rectale and/or a prominent human gut Bacteroidetes, Bacteroides thetaiotaomicron, followed by whole genome transcriptional profiling of both organisms in their distal gut (cecal) habitat as well as host responses, high resolution proteomic analysis of cecal contents, and biochemical assays of carbohydrate metabolism. B. thetaiotaomicron adapts to E. rectale by upregulating expression of a variety of polysaccharide utilization loci (PULs) encoding numerous glycoside hydrolase gene families, and by signaling the host to produce mucosal glycans that it, but not E. rectale, can access. E. rectale adapts to B. thetaiotaomicron by decreasing production of its glycan-degrading enzymes, increasing expression of selected amino acid and sugar transporters, and facilitating glycolysis by reducing levels of NADH, in part via generation of butyrate from acetate, which in turn is utilized by the gut epithelium. This simplified model of the human gut microbiota illustrates niche specialization and functional redundancy within members of major gut bacterial phyla, and the importance of host glycans as a nutrient foundation that ensures ecosystem stability. Experiment Overall Design: four biological replicates per group. NMRI mice, males (12-14 weeks old).
Project description:The adult human gut microbial community is typically dominated by two bacterial phyla (divisions), the Firmicutes and the Bacteroidetes. Little is known about the factors that govern the interactions between their members. Here we examine the niches of representatives of both phyla in vivo. Finished genome sequences were generated from E. rectale and E. eligens, which belong to Clostridium Cluster XIVa, one of the most common gut Firmicute clades. Comparison of these and 25 other gut Firmicutes and Bacteroidetes indicated that the former possess smaller genomes and a disproportionately smaller number of glycan-degrading enzymes. Germ-free mice were then colonized with E. rectale and/or a prominent human gut Bacteroidetes, Bacteroides thetaiotaomicron, followed by whole genome transcriptional profiling of both organisms in their distal gut (cecal) habitat as well as host responses, high resolution proteomic analysis of cecal contents, and biochemical assays of carbohydrate metabolism. B. thetaiotaomicron adapts to E. rectale by upregulating expression of a variety of polysaccharide utilization loci (PULs) encoding numerous glycoside hydrolase gene families, and by signaling the host to produce mucosal glycans that it, but not E. rectale, can access. E. rectale adapts to B. thetaiotaomicron by decreasing production of its glycan-degrading enzymes, increasing expression of selected amino acid and sugar transporters, and facilitating glycolysis by reducing levels of NADH, in part via generation of butyrate from acetate, which in turn is utilized by the gut epithelium. This simplified model of the human gut microbiota illustrates niche specialization and functional redundancy within members of major gut bacterial phyla, and the importance of host glycans as a nutrient foundation that ensures ecosystem stability.
Project description:Gene expression profiles of Bacteroides thetaiotaomicron in vitro during growth on host mucosal polysaccharides as sole carbon sources. All substrates in this series are derived from porcine gastric mucin and include mucin O-glycans and glycosaminoglycans. Two different culture formats used: 800ml batch-culture bioreactors and 5ml tube cultures (format is indicated within each sample title). Each set of growths was referenced to a minimal medium glucose reference corressponding to the same culture format. Unfractionated porcine mucosal glycan (PMG) growths were compared to previously published in vivo datasets, which were referenced to the 800ml minimal medium glucose reference dataset.