Role of Intestinal Microbiota in Ulcerative Colitis

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Discussion and conclusion 160 7. Discussion and perspectives communications (Takaishi et al., 2008;Sokol et al., 2009), and could indicate that the commensal microbiota in remission is unstable and unfavorable events could contribute to microbial compositional changes provoking flare‐ups. However, further longitudinal studies are needed to obtain sufficient information both microbiological and immunological on this area, hence helping elucidate if the microbial compositional changes are causal to or consequence of disease. The ability of the fecal microbiota derived from UC patients in either remission or relapse and healthy subjects to colonize an artificial mucus layer was examined in Paper 2. The results from this study demonstrated that the mucosal microbial community differs from that of the lumen, whether the microbiota was derived from healthy subjects, UC patients in remission or relapse. These findings are in agreement with previous in vivo (Zoetendal et al., 2002;Macfarlane, 2008) and in vitro studies (Macfarlane et al., 2005;Van den Abbeele et al., 2011a). Additionally, the results from Paper 2 revealed that bifidobacteria and lactobacilli from UC patients in relapse were found in significantly lower levels in mucus than when derived from healthy subjects. However, the question is, whether the low levels of adhered lactic acid bacteria from UC patients in relapse are a result of low baseline levels in feces or decreased capacity of the bacteria to adhere? Based on results from Paper 1 (using some of the same individuals as in Paper 2), it was demonstrated that the level of fecal Lactobacillus spp. was significantly lower in UC patients in relapse than in healthy subjects. However, no significant difference was observed for the fecal levels of Bifidobacterium spp.. This could imply that lactic acid bacteria derived from UC patients in relapse have decreased capacity to colonize the mucus independent of baseline levels in feces. The decreased capacity of bifidobacteria and lactobacilli to colonize the mucus could be due to changed expression of adhesion molecules. Adhesion‐promoting molecules have been observed in species of Lactobacillus and Bifidobacterium including fimbriae (Pridmore et al., 2004;Kankainen et al., 2009;Gilad et al., 2011), Msa, mannose‐specific adhesin protein (Pretzer et al., 2005), MucBP domain containing proteins (Kleerebezem et al., 2010), and elongation factor (EF‐Tu) (Granato et al., 2004;Gilad et al., 2011). However, proteomic analyses of bifidobacteria or lactobacilli isolated from UC patients have to my knowledge not previously been conducted to reveal, if changes in bacterial phenotypes occur during mucosal inflammation. The decreased capacity to colonize mucus could also be due to intra‐species variations within the lactic acid bacteria genera. In Paper 1 and 2, different species of Bifidobacterium were quantified
Discussion and conclusion 161 7. Discussion and perspectives using SYBR‐Green based assays with primers targeting either 16S rRNA gene sequences or 16S‐23S rRNA intergenic spacer regions. The primers were selected based on sensitivity and specificity, which led to a limited number of Bifidobacterium species target. However, quantification of numerous species of lactic acid bacteria found in the human gut is needed to draw any conclusions on intragenic compositional changes from individual samples. The design of primers may give some problems when targeting species within a genus, since primer specificity and sensitivity can be difficult to obtain. Even though 16S rRNA gene sequences can be used for species identification and quantification ((Matsuki et al., 1998;Byun et al., 2004;Matsuki et al., 2004) and Paper 1 – 2), they have some limitations in discriminating species of related taxa or within the same genus, due to high 16S rRNA sequence similarities (Fox et al., 1992). Protein‐coding genes or 16‐23S rRNA intergenic spacer regions may be a better choice, since both exhibit higher sequence variations than 16S rRNA gene sequences (Saikaly et al., 2007). Previous studies have used 16S‐23S rRNA intergenic spacer regions (Haarman and Knol, 2005), as well as Paper 1 or recA (Masco et al., 2007) to quantify species within Bifidobacterium showing high specificity. The low levels of lactic acid bacteria derived from UC patients found in the artificial mucus layer in Paper 2 have previously been described for bifidobacteria from mucus layer of biopsy specimens from UC patients (Macfarlane et al., 2004;Mylonaki et al., 2005). As mentioned above, the results from Paper 1 demonstrated low abundances of fecal lactobacilli in UC patients in relapse than in healthy subjects, but no significant difference was observed for the fecal bifidobacteria. Low levels of fecal bifidobacteria in UC patients in relapse have previously been described by Sokol et al. (2009). The discrepancy in results could be due to differences in the amount of participants in the two studies: Paper 1 is based upon six participants in each group whereas the study by Sokol et al. (2009) covers thirteen UC patients in replase and twenty seven healthy controls. The low amount of participants in Paper 1 is a weakness in the study, since it gives a low confidence interval (high margin of error); hence is not as representative for a true population, as when using a high amount of participants. Additionally, due to high inter‐individual variations significant power can be difficult to reach. However, when comparing Paper 1 to previous communications (Takaishi et al., 2008;Qin et al., 2010;Arumugam et al., 2011), several similar tendencies can be found, which justifies the results of the study.
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Role of Intestinal Microbiota in Ul
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Role of Intestinal Microbiota in Ul
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Preface Preface This thesis present
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Summary Summary The microbiota of t
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Dansk sammendrag Dansk sammendrag M
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Introduction and objectives Introdu
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List of Manuscripts Not included in
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List of contents List of Centents P
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List of Centents Methodology append
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1. The intestinal environment Theor
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Theoretical part 5 1. The intestina
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2. The colonic environment Theoreti
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Theoretical part 9 2. The colonic e
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Table 1: The presence of glycoside
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Theoretical part Figure 3: The colo
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3. Inflammatory Bowel disease Theor
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Theoretical part 17 3. Inflammatory
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Theoretical part 19 4. Modulation o
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Table 4: Clinical trials on the pre
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Theoretical part 5. Production of p
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Methodology part
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Methodology part 6. Methodology, co
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Introduction Methodology part 42 Pa
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Abstract Background Detailed knowle
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depending the level of disease acti
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in 1 x TAE at 60 °C for 16 h at 36
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Statistics PCA were generated by SA
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The PCA of the Gram‐positive bact
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layer of UC patients and found that
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Acknowledgements The authors thank
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Table 2 ‐ 16S rRNA gene and 16S
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1. Firmicutes phylum 2. Bacteroidet
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Supplementary Figure S1. Dice clust
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Reference List 1. Ahmed S, Macfarla
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32. Matsuki T, Watanabe K, Fujimoto
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Methodology part Paper 2 Fecal lact
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Fecal lactobacilli and bifidobacter
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Introduction The mucus layer lining
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efore enrolment and there was no si
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(Bio‐Rad Labs, Hercules, Californ
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Microbial community analysis using
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difference from the luminal microbi
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that C. coccoides group and C. lept
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Table 1 ‐ 16S rRNA gene of phylum
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Table 2 ‐ Preference of bacterial
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Figure 1. A) Schematic overview of
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A. B. Figure 3. Principal component
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15. Fooks LJ, Gibson GR. (2002) In
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47. Ouwehand AC, Suomalainen T, Tol
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Methodology part Paper 3 Paper 3 In
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APPLIED AND ENVIRONMENTAL MICROBIOL
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8338 VIGSNÆS ET AL. APPL. ENVIRON.
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Page 128 and 129: Methodology part Introduction The a
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Page 197 and 198: References Tannock,G.W. (2010). Ana
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Role of Intestinal Microbiota in Ulcerative Colitis

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