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Microbial Community Analysis Reveals High Level Phylogenetic Alterations in the Overall Gastrointestinal Microbiota of Diarrhoea-Predominant Irritable Bowel Syndrome Sufferers

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Microbial Community Analysis Reveals High Level Phylogenetic Alterations in the Overall Gastrointestinal Microbiota of Diarrhoea-Predominant Irritable Bowel Syndrome Sufferers

Lotta Krogius-Kurikka et al. BMC Gastroenterol.

Abstract

Background: A growing amount of scientific evidence suggests that microbes are involved in the aetiology of irritable bowel syndrome (IBS), and the gastrointestinal (GI) microbiota of individuals suffering from diarrhoea-predominant IBS (IBS-D) is distinguishable from other IBS-subtypes. In our study, the GI microbiota of IBS-D patients was evaluated and compared with healthy controls (HC) by using a high-resolution sequencing method. The method allowed microbial community analysis on all levels of microbial genomic guanine plus cytosine (G+C) content, including high G+C bacteria.

Methods: The collective faecal microbiota composition of ten IBS-D patients was analysed by examining sequences obtained using percent G+C (%G+C) -based profiling and fractioning combined with 16S rRNA gene clone library sequencing of 3267 clones. The IBS-D library was compared with an analogous healthy-control library of 23 subjects. Real-time PCR analysis was used to identify phylotypes belonging to the class Gammaproteobacteria and the order Coriobacteriales.

Results: Significant differences were found between clone libraries of IBS-D patients and controls. The microbial communities of IBS-D patients were enriched in Proteobacteria and Firmicutes, but reduced in the number of Actinobacteria and Bacteroidetes compared to control. In particular, 16S rDNA sequences belonging to the family Lachnospiraceae within the phylum Firmicutes were in greater abundance in the IBS-D clone library.

Conclusions: In the microbiota of IBS-D sufferers, notable differences were detected among the prominent bacterial phyla (Firmicutes, Actinobacteria, Bacteroidetes, and Proteobacteria) localized within the GI tract.

Figures

Figure 1
Figure 1
Distribution of sequences and OTUs among IBS-D and HC clone libraries. The unique sequences or OTUs (cut-off-level of 98%) in the IBS-D and HC libraries are indicated in red and blue, respectively. The grey area in the group-wise bars is a mirror-image of shared sequences or OTUs and it is presented on both sides of the y-axis. The number of singleton OTUs is given in parentheses. The roman numerals XIV and IV within the Firmicutes indicate the corresponding Clostridium rRNA clusters, 1) Other Firmicutes, 2) Acidobacteria, Cyanobacteria, TM7 and Verrucomicrobia.
Figure 2
Figure 2
Rank-abundance plots of the IBS-D and HC libraries. The curves for IBS-D and HC libraries are indicated in red and blue, respectively. To make the image more compact, the OTUs with the highest number of sequences are deleted (an OTU of 395 sequences for IBS-D and an OTU of 160 sequences for HC).
Figure 3
Figure 3
Relative abundance of phyla in the IBS-D and HC libraries in the RDP library compare analysis [34]. Significantly differing (p < 0.01) abundances of sequences (s) or OTUs with a cut-off-level of 98% for sequence similarity (o) between the IBS-D and HC libraries are indicated. The "unclassified" phyla have a bootstrap value below 80%.
Figure 4
Figure 4
Percentage of shared and unique OTUs (n = 578) and sequences (n = 6466) in the combined IBS-D and HC libraries. Number of sequences and OTUs (cut-off-level of 98%) are given in parentheses. The number of singletons is indicated after a semicolon. A common alignment was used for the determination of shared OTUs.
Figure 5
Figure 5
Abundance of shared OTUs. Proportion of shared OTUs in the IBS-D and HC libraries according to the SONS analysis [36].
Figure 6
Figure 6
Real-time PCR results from assays for Enterobacteriaceae (A) and Eggerthella lenta -like (B) phylotypes. Samples from IBS-D and HC subjects are in red and blue, respectively. The values are 16S rRNA gene copy numbers per gram of faeces (log10 values), and the detection limit is set to 104. Vertical lines are medians from the Mann-Whitney U-test.

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