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Sequence variations located at the signal sequence and mid-region within the vacA gene, the 3′-end of the cagA gene, the indel motifs at the 3′-end of the cag pathogenicity island and the regions upstream of the vacA and ribA genes were determined by PCR in 19 paired antral or antrum and corpus Helicobacter pylori isolates obtained at the same endoscopic session, and three antral pairs taken sequentially. Random amplification of polymorphic DNA (RAPD)-PCR and fluorescent amplified fragment length polymorphism (FAFLP)-PCR fingerprinting were applied to these paired clinical isolates. The FAFLP-PCR profiles generated were phylogenetically analysed. For the 22 paired isolates there were no differences within pairs at five of the genetic loci studied. However, six pairs of isolates (27 %), of which four were antrum and corpus pairs, showed differences in the numbers of repeats located at the 3′-end of the cagA gene. RAPD-PCR fingerprinting showed that 16 (73 %) pairs, nine of which were antrum and corpus pairs, possessed identical profiles, while six (27 %) displayed distinctly different profiles, indicating mixed infections. Three of the six pairs showing differences at the 3′-end of the cagA gene yielded identical RAPD-PCR fingerprints. FAFLP-PCR fingerprinting and phylogenetic analysis revealed that all 16 pairs that displayed identical RAPD-PCR profiles had highly similar, but not identical, fingerprints, demonstrating that these pairs were ancestrally related but had undergone minor genomic alterations. Two antrum and corpus pairs of isolates, within the latter group, were isolates obtained from two siblings from the same family. This analysis demonstrated that each sibling was colonized by ancestrally related strains that exhibited differences in vacA genotype characteristics.

Background The intestinal microbiota has been implicated in the pathophysiology of irritable bowel syndrome (IBS). Due to the variable resolutions of techniques used to characterize the intestinal microbiota, and the heterogeneity of IBS, the defined alterations of the IBS intestinal microbiota are inconsistent. We analyzed the composition of the intestinal microbiota in a defined subgroup of IBS patients (diarrhea-predominant IBS, D-IBS) using a technique that provides the deepest characterization available for complex microbial communities. Methods Fecal DNA was isolated from 23 D-IBS patients and 23 healthy controls (HC). Variable regions V1–V3 and V6 of the 16S rRNA gene were amplified from all samples. PCR products were sequenced using 454 high throughput sequencing. The composition, diversity and richness of microbial communities were determined and compared between D-IBS and HC using the quantitative insights into microbial ecology pipeline. Key Results The contribution of bacterial groups to the composition of the intestinal microbiota differed between D-IBS and HC. D-IBS patients had significantly higher levels of Enterobacteriaceae (P = 0.03), and lower levels of Fecalibacterium genera (P = 0.04) compared to HC. β-Diversity values demonstrated significantly lower levels of UniFrac distances in HC compared to D-IBS patients. The richness of 16S rRNA sequences was significantly decreased in D-IBS patients (P

pieces of small and large intestine were cultured for 8 hrs and supernatant collected for measurement of cytokine secretion by Elisa. Splenocytes were isolated and cultured with fecal sonicates obtained from the animal's own stools collected prior to (Day 0) and following PM (Day 7) or vehicle treatment. Results: After 7 days of treatment, WT and IL-10-/mice had increased IL-8 secretion in the small intestine, but no PM-induced changes in colonic secretion. However, following 30 days of treatment, a significant increase in small and large intestinal IL-1β, IL-8, IL-12, and TNFα secretion was seen in WT mice indicating a PMinduced inflammatory response. Compared to WT mice, IL-10-/had increased levels of IL8, IL-12, and TNFα, and responded to PM with enhanced IL-17 secretion in both the small and large intestine. PM-induced alterations in colonic microflora preceded the changes in cytokine secretion, with both WT and IL-10-/mice exhibiting decreased microbial diversity and changes in composition, primarily within the phylum Firmicutes by 7 days of treatment. Alterations in systemic immune function were also observed, in that the splenocytes from the PM-treated mice had decreased proliferation to fecal sonicates as compared with the control group. Conclusions: Oral ingestion of PM collected from urban ambient air appears to elicit an initial acute inflammatory response in the small intestine of mice that is followed by altered mucosal immune function in both the small and large intestine. This altered mucosal immune function is associated with a PM-induced modulation of colonic microflora and provides support for a possible role for airborne pollutants in inflammatory bowel diseases.

we would like to thank Drs. Han and Fioramonti for bringing their recent work to our attention. In particular, it is reassuring that both studies reached a similar conclusion with respect to the beneficial effect of SOD-expressing bacteria on colitis amelioration. Our omission in citing their work

Background Recent studies have suggested a role for an altered intestinal microbiota in the pathophysiology of irritable bowel syndrome (IBS). However, no consensus has been reached regarding the association between specific enteric bacterial groups and IBS. The aim of this study was to investigate the fecal and mucosal-associated microbiota using two independent techniques in intestinal samples from diarrhea-predominant IBS (D-IBS) and healthy controls. Methods Fecal and colonic mucosal biopsy samples were obtained from 10 D-IBS patients and 10 healthy controls. Colonic tissue was collected during a un-sedated un-prepped flexible sigmoidoscopy. Fecal and tissue samples were processed immediately upon collection for culture under aerobic and anaerobic conditions or frozen for further molecular analysis. DNA was extracted from all frozen samples and used to enumerate specific bacterial groups using quantitative real-time PCR (qPCR). Results Culture analysis of intestinal samples demonstrated a significant reduction in the concentration of aerobic bacteria in fecal samples from D-IBS patients when compared to healthy controls (1.4 × 107 vs. 8.4 × 108 CFUs/g feces, P = 0.002). qPCR analysis demonstrated a significant 3.6 fold increase (P = 0.02) in concentrations of fecal Lactobacillus species between D-IBS patients and healthy controls. Conclusions Our culture and molecular data indicate that quantitative differences exist in specific bacterial groups in the microbiota between D-IBS and healthy subjects.