Culture-Independent Analyses of Temporal Variation of the Dominant Fecal Microbiota and Targeted Bacterial Subgroups in Crohn's Disease

Abstract

Gut microbiota shows host-specific diversity and temporal stability and significantly contributes to maintenance of a healthy gut. However, in inflammatory bowel disease, this microbiota has been implicated as a contributory factor to the illness. This study compared bacterial dynamics in Crohn's disease patients to those in a control group using a culture-independent method to assess the temporal stability, relative diversity, and similarity of the dominant fecal microbiota, Clostridium spp., Bacteroides spp., Bifidobacterium spp., and lactic acid bacteria spp. (LAB) for all individuals. Fecal samples were collected over several time points from individuals with Crohn's disease who were in remission (n ‫؍‬ 11), from Crohn's disease patients who relapsed into an active Crohn's disease state (n ‫؍‬ 5), and from a control group (n ‫؍‬ 18). Denaturing gradient gel electrophoresis profiles were generated for the different microbial groups by specifically targeting different regions of the 16S rRNA gene and were compared on the basis of similarity and diversity. The temporal stability of dominant species for all Crohn's disease patients was significantly lower (P < 0.005) than that for the control group. Analysis of group-specific profiles for Bifidobacterium spp. found that they were similar in all samples, while the diversity of the LAB varied significantly between the groups, but temporal stability was not significantly altered. We observed significant changes in two functionally important mutualistic groups of bacteria, viz., Clostridium and Bacteroides spp., which may have implications for the host's gut health, since some genera are involved in production of short-chain fatty acid, e.g., butyrate.

Molecular analysis of the gut microbiota of identical twins with Crohn's disease

“…This difference was largely due to lower relative abundances of B. uniformis and higher abundances of B. ovatus and B. vulgatus, in patients with ileal involvement compared with both healthy twins and twins with colonic disease. Several previous reports have also shown an abnormal Bacteroides community in CD patients compared to healthy individuals (Seksik et al, 2003;Ott et al, 2004;Scanlan et al, 2006). For example, Scanlan et al (2006) reported a lower complexity of DGGE (denaturing gradient gel electrophoresis) profiles within the B. fragilis subgroup in CD patients than in healthy individuals and a difficulty in obtaining PCR products from CD patients compared to controls.…”

“…Several previous reports have also shown an abnormal Bacteroides community in CD patients compared to healthy individuals (Seksik et al, 2003;Ott et al, 2004;Scanlan et al, 2006). For example, Scanlan et al (2006) reported a lower complexity of DGGE (denaturing gradient gel electrophoresis) profiles within the B. fragilis subgroup in CD patients than in healthy individuals and a difficulty in obtaining PCR products from CD patients compared to controls. In this study, it was not problematic to obtain PCR products for the Bacteroides group, which could simply reflect differences in Bacteroides abundances within the sample groups of the two studies.…”

“…Previous studies have found that the microbial diversity in the gut is lower in individuals with CD compared to healthy individuals (Seksik et al, 2003;Manichanh et al, 2006;Scanlan et al, 2006). For example, there have been reports of a reduced diversity of Firmicutes (Gophna et al, 2006;Manichanh et al, 2006) and Bacteroides (Ott et al, 2004;Frank et al, 2007) in CD patients.…”

“…are dominant members of the fecal microbiota (Seksik et al, 2003;Ott et al, 2004;Scanlan et al, 2006) and our Bacteroides clone sequences from the same material had the same TRF sizes (see below). Therefore, a group-specific primer set was used during T-RFLP, to focus on the Bacteroides group in the same DNA extracts from the fecal samples that were previously analyzed using general bacterial primers.…”

Molecular-phylogenetic characterization of microbial community imbalances in human inflammatory bowel diseases

“…Indeed, recent studies demonstrate that obesity in humans and ob/ob mice is associated with stereotypical imbalances in the normal gut microbiota (31)(32)(33). Likewise, previous studies of human IBD, using standard culture techniques (34)(35)(36) or molecular analysis (37)(38)(39)(40)(41)(42)(43), have noted alterations in the GI microbiota. However, most of these IBD studies have been limited in statistical power and precision of identification or have examined only the fecal microbiota, which differs substantially from that of the GI mucosa (44).…”