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Degradation and utilization of resistant starch by microbiota in the human large intestineZe, Xiaolei January 2013 (has links)
Resistant starch (RS) is the major energy source for microbial growth in the human large intestine and the fermentation of RS in the gut has a potential role in maintaining host health. Molecular profiling of the complex microbial communities in the human gut has revealed a remarkable inter-individual variation of their composition, which could be linked to variations in substrate fermentation in different individuals. In this project, significant inter-individual variation was seen in the composition of the human gut microbial communities and in the response to changes in dietary intake, using a range of qPCR primer sets. The range of qPCR primer sets was expanded in this study, for the assessment of an uncultured bacterial group related to Oscillibacter and Oscillospira spp. This group of bacteria showed a significant increase in obese subjects on a high RS diet and weight loss diet in samples from a previous human dietary intervention study. The group showing the greatest stimulation by RS, however, was related to Ruminococcus bromii. Furthermore, two out of 14 volunteers who showed very low R. bromii numbers also showed incomplete RS3 fermentation in the study of Walker et al. 2011. It was therefore decided to investigate the activity of human colonic bacteria against RS in vitro. Anaerobic pure cultures and defined co-incubations were performed to compare the abilities to degrade RS by four of the most abundant amylolytic species present in the human colon, which were Ruminococcus bromii, Eubacterium rectale, Bifidobacterium adolescentis and Bacteroides thetaiotaomicron. The four species showed the highest utilization of RS2 and RS3 (47-77%) when the RS was autoclaved in the medium, however, the utilization by E. rectale and B. thetaiotaomicron was limited for boiled RS (7-40%) and for raw RS (0.25-18%). Pairwise co-incubation of the four species revealed that utilization of boiled RS2 and RS3 by the other three amylolytic species was significantly increased in the presence of R. bromii, even in a medium that does not permit growth of R. bromii itself. The growth of Anaerostipes hadrus, which is a nonstarch utilizer, was also promoted in a co-incubation with R. bromii. Assays of soluble reducing sugar revealed that the bacterial growth stimulated by R. bromii relies on utilization of breakdown products released from RS by R. bromii amylases. Moreover, addition of R. bromii, but not other three amylolytic species, restored RS3 fermentation in vitro by the faecal bacterial community from one of the volunteers who had previously XVIII shown low starch fermentability in vivo. These results therefore suggest that R. bromii is a „keystone‟ species for the degradation of RS in the human large intestine. Finally, the starch-degrading enzyme system of R. bromii was investigated. Genome mining revealed 15 GH13 (glycoside hydrolases family 13) amylases and 8 SBDs (starchbinding domains), as well as many structural modules of cohesins and dockerins that are known to be involved in protein-protein interactions in related Ruminococcus species. The major extracellular amylases active against RS were identified by zymogram analysis together with peptide sequencing of the excised active bands. The gene products responsible were then identified from draft genome sequence information and found to include three large proteins with dockerin modules, one of which also contained a cohesin. An additional gene product encoding a cohesin domain was identified from the genome sequence that was considered a candidate for cell-surface attachement. The two cohesin domains were therefore over-expressed in order to investigate protein-protein interactions. Western-blotting using the over-expressed cohesins as probes demonstrated cohesindockerin interactions between certain proteins. Possible models for the organization of a R. bromii multienzyme amylase system were then proposed based on these computerbased analysis and on the results generated from protein-protein interaction studies.
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Structural and functional diversity of flagellins expressed by gut bacteriaMonnais, Edouard January 2013 (has links)
Flagellins are proteins of microbial origin that confer motility to bacteria. They exhibit a stupendous conservation in their structure, required for assembly into polymeric flagella. The various environmental and evolutionary pressures encountered by motile bacteria have however generated a multiplicity of primary protein sequences. In mammals, specific receptors, namely TLR5 and Ipaf, are devoted to the immunosurveillance of flagellins. We sought to explore the diversity of flagellin structures expressed by the intestinal microbiota and how these proteins may influence the host innate immune response. In this context, recombinant flagellins derived from Gram-negative and Gram-positive, pathogenic and commensal bacteria from the γ-Proteobacteria and the Clostridium sub-phyla were expressed in E.coli and then purified. Protein sequence alignment and phylogenetic analysis of twelve flagellins revealed clustering dependent on three criteria: the phylum, the genus and the commensal or pathogenic nature of the bacteria. We next characterized these structures functionally using in vitro cell systems including epithelial and dendritic cells. In a TLR5-dependent manner, flagellin induced the secretion of pro-inflammatory chemokines in intestinal epithelial cells, which are considered crucial in triggering the recruitment of immune cell effectors. Of importance, the secretion levels were directly dependent of the flagellin structure, as confirmed by the various agonistic potentials determined in dose-response experiments. We observed that commensal flagellins were in general less potent than pathogenic flagellins. Similarly, the effects on dendritic cells following flagellin exposure were ligand-dependent. Overall, TLR5 activation constitutes a ligand-dependent TLR5-flagellin complex formation with sequential activation of MAPK and/or NF-kB and signalling, resulting in differential modulation of the intracellular signalling cascade downstream. The results presented in this thesis suggest innate signalling mediated by flagellated pathogens and commensals may affect the intestinal immune status in a number of distinct ways, either by promoting active immune response or by promoting tolerance response.
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NF-κB signalling pathway regulation in intestinal epithelial cells in response to commensal and pathogenic bacteria and metagenomic clonesFerraria, Vanessa January 2013 (has links)
The study of the human microbiota is of enormous importance because it represents a significant diverse microbial population that co-habits the intestine and influences host physiology, immune homeostasis, gene and protein expression and, importantly, overall health status. The microbiota is crucial in regulation of immune tolerance. Epithelial cells are considered an important cell population involved in host microbial interactions as they are the first intact cell layer in contact with bacteria colonizing the gut. Understanding how bacteria influence the host response in health and in disease may improve the understanding of how inflammation and inflammatory bowel diseases (IBD) develop and also provide insights into which bacterial population promote health and maintain immune homeostasis. The main objective of this thesis was to investigate the response of epithelial cells to commensal and pathogenic bacteria and metagenomic clones (derived from the human gut microbiota). Both commensal and pathogenic bacteria present microbial-associated molecular patterns (MAMPs) that are recognised by innate immune receptors such as Toll-like receptors (TLRs). The mechanisms by which commensal bacteria maintain immune tolerance in the gut, whereas pathogens induce strong immune responses, are not fully understood. We investigated the hypothesis that commensal bacteria control immune homeostasis, in part, by regulating NF-κB mediated signalling. In particular, the effect of specific gut microbes on the expression of the NF-κB repressor A20, a zinc protein that is encoded by TNFAIP3 gene, which induces strong down-regulation of the canonical NF-kB pathway was investigated. This thesis revealed that immune regulation by both pathogenic and commensal bacteria involves both positive and negative regulation of the NF-kB pathway mediated by specific MAMPs and metabolites. The significance of these findings in relation to immune homeostasis and inflammation are discussed
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Neurotensin and its receptors in human isolated intestine in health and disease : focus on motility aspectsAzriel, Yael, Medical Sciences, Faculty of Medicine, UNSW January 2005 (has links)
Neurotensin (NT) is a brain-gut peptide, localised to mucosal endocrine N-cells in the gastrointestinal tract. The most potent stimuli for NT release are ingested fats. NT, in vivo, inhibits the migrating myoelectric complex and stimulates colonic motility. The aim of this thesis was to investigate NT and its receptors in human isolated intestine, focusing on motility aspects. ???Normal??? colon and ileum were obtained from patients undergoing resection for carcinoma, and ???disease??? colon was obtained from patients undergoing resection for ulcerative colitis (UC), Crohn???s disease (CD), diverticular disease (DD) and slow transit constipation (STC). In functional experiments with ???normal??? ascending and sigmoid colon, NT contracted taenia coli strips by direct mechanisms (EC50 ??? 100 nM). In circular muscle (CM) strips, contractions were mediated by NT-induced release of enteric mediators (indirect actions), which were regionally specific, as well as by direct actions on the smooth muscle. In contrast, in ???normal??? terminal ileum, the predominant action of NT was a potent (EC50 ??? 3 nM), directly mediated inhibition of spontaneous contractions in longitudinal muscle strips. NT receptors were characterised using radioligand binding with [125I]-NT and [3H]-NT, and localised using autoradiography and immunohistochemistry and confocal microscopy. In the sigmoid colon, binding sites corresponded to NTS1 receptors and could be visualised on the smooth muscle and the enteric ganglia, supporting both direct and neuronal actions for NT, respectively. In the ileum, a similar distribution of binding sites could be visualised. In gastrointestinal disease, NT contractile responses were reduced in UC, DD and STC colon CM strips, but were unchanged in CD. In contrast, NT responses in all diseases were largely unchanged in TC strips, suggesting disease/treatment-induced alterations in NT mechanisms. In UC, cholinergic mechanisms appeared to be altered, whereas in DD disease, prostaglandin mechanisms were affected. In STC, there was an apparent loss of a normally predominant component in NT-mechanisms, but contractile mechanisms appeared to be retained. In summary, these studies demonstrated regional and muscle layer specific actions and mechanisms of action for NT, and support alterations in enteric signaling in disease.
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The source and action of immunoglobulins in the dog intestine.Heddle, Robert John. January 1978 (has links) (PDF)
Thesis (Ph.D.) -- University of Adelaide, Dept. of Microbiology, 1978.
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La lambliase étude biologique-clinique et thérapeutique.Champsaur, André Félicien, January 1933 (has links)
Issued also as Thèse pour le doctorat en médecine - Paris. / "Bibliographie": p. 123-134.
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Intestinal microflora of chickensJohansson, Karl Richard, January 1948 (has links)
Thesis (Ph. D.)--University of Wisconsin--Madison, 1948. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves [i]-xvii).
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Observations on the transmission and the epizootiology of the virus of mink enteritisBouillant, Alain Marcel Pierre. January 1963 (has links)
Thesis (M.S.)--University of Wisconsin--Madison, 1963. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 95-99).
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Reductions of sterols by intestinal microorganismsNeiderhiser, Dewey Harold. January 1963 (has links)
Thesis (Ph. D.)--University of Wisconsin--Madison, 1963. / Typescript. Abstracted in Dissertation abstracts, v. 23 (1963) no. 9, p. 3109. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaf 92).
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Gluconate metabolism in Lactobacillus and its role in persistence in the human intestineJenkins, Julie Kay, January 2005 (has links)
Thesis (Ph. D.)--Ohio State University, 2005. / Title from first page of PDF file. Document formatted into pages; contains xi, 102 p.; also includes graphics (some col.). Includes bibliographical references (p. 95-102). Available online via OhioLINK's ETD Center
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