Distinct Microbiotas are Associated with Ileum-Restricted and Colon-Involving Crohn's Disease.

BACKGROUND: The etiology of inflammatory bowel disease is believed to involve a shift in the microbiota toward more proinflammatory species. Crohn's disease (CD) usually manifests as one of three phenotypes, involving inflammation of the terminal ileum, the colon, or both. However, what determines the particular phenotype and the level of disease activity remains unknown. In this study, we aim to characterize the intestinal microbiota associated with different CD phenotypes. METHODS: DNA was extracted from biopsies of 31 patients with ileal, ileocolic, or colon-restricted CD, and also from 5 non-inflammatory bowel disease control subjects, and analyzed by 16S rRNA gene amplicon pyrosequencing. Data were processed using the Quantitative Insights Into Microbial Ecology pipeline and analyzed using linear discriminant analysis with effect size estimation and PICRUSt algorithms. Two additional recently published cohorts were also analyzed in this study. RESULTS: Highly significant separation was observed between bacterial composition of ileal CD compared with CD with colonic involvement (genus level Bray-Curtis P = 0.005, R = 20%). This separation was unaffected by the biopsy's location or its inflammatory state, or by the patients' condition (remission or relapse). Faecalibacterium was strongly reduced in ileal CD compared with CD with colonic involvement, whereas Enterobacteriaceae were more abundant in the former. Fusobacterium relative abundance was strongly correlated with disease activity in patients with ileal-involving, but not in colon-involving, CD. CONCLUSIONS: Ileal and colon-involving CD sustain distinct microbiotas, suggesting that different mechanisms underlie the two major manifestations of CD. The potential contribution of Fusobacterium to inflammation in ileal CD should be further investigated.

Pouch Inflammation Is Associated With a Decrease in Specific Bacterial Taxa.

BACKGROUND & AIMS: Pouchitis is a common long-term complication in patients with ulcerative colitis (UC) undergoing proctocolectomy with ileal pouch-anal anastomosis. Because the inflammation occurs in a previously normal small bowel, studies of this process might provide information about the development of Crohn's disease. Little is known about the intestinal microbiome of patients with pouchitis. We investigated whether specific bacterial populations correlate with the pouch disease phenotype and inflammatory activity. METHODS: We performed a prospective study of patients with UC who underwent pouch surgery (N = 131) from 1981 through 2012 and were followed at Tel Aviv Medical Center. Patients were assigned to groups based on their degree and type of pouch inflammation. Patients with familial adenomatous polyposis after pouch surgery (n = 9), individuals with intact colons undergoing surveillance colonoscopy (n = 10), and patients with UC who did not undergo surgery (n = 9) served as controls. We collected demographic and disease activity data (based on the Pouchitis Disease Activity Index) and measured levels of C-reactive protein. Fecal samples were collected, levels of calprotectin were measured, and microbiota were analyzed by 16S ribosomal RNA gene amplicon pyrosequencing. RESULTS: Increased proportions of the Fusobacteriaceae family correlated with increased disease activity and levels of C-reactive protein in patients with UC who underwent pouch surgery. In contrast, proportions of Faecalibacterium were reduced in patients with pouchitis vs controls; there was a negative correlation between proportion of Faecalibacterium and level of C-reactive protein. There was an association between antibiotic treatment, but not biologic or immunomodulatory therapy, with reduced proportions of 11 genera and with increased proportions of Enterococcus and Enterobacteriaceae. CONCLUSIONS: Reductions in protective bacteria and increases in inflammatory bacteria are associated with pouch inflammation in patients with UC who underwent pouch surgery. The finding that antibiotics exacerbate dysbiosis indicates that these drugs might not provide long-term benefit for patients with pouchitis. Additional studies of this form of dysbiosis could provide information about the pathogenesis of Crohn's disease.

Can Colonoscopy Aspirates be a Substitute for Fecal Samples in Analyses of the Intestinal Microbiota?

There is a growing interest in the study of the human gut microbiota, as correlations between changes in bacterial profiles and diseases are increasingly discovered. Studies in this field generally use fecal samples, but it is often easier to obtain colon content aspirates during colonoscopy. This study used automated ribosomal internal spacer analysis (ARISA) to examine the extent to which the microbiota of colon aspirate samples obtained after bowel cleansing can reflect interindividual differences and serve as a proxy for fecal samples. Pre-bowel preparation fecal samples as well as colonoscopy aspirate samples from the cecum and rectum were obtained from 19 subjects. DNA was extracted from all samples, and comparative analysis was performed, including analysis of similarity (ANOSIM) and nonmetric multidimensional scaling. ANOSIM confirmed that samples from the same individual were well separated from samples from different individuals. Significantly larger differences were found between samples from different individuals than between samples of the same individual (R = 0.7605, p < 0.0001). These findings show that post-bowel preparation aspirates maintain a strong individual signature. Colonoscopy aspirates can therefore serve as a substitute for fecal samples in studies comparing the microbiota of different clinical study groups, especially when fecal samples are unavailable.

Composition and dynamics of the gill microbiota of an invasive Indo-Pacific oyster in the eastern Mediterranean Sea.

Gill bacterial communities of Chama pacifica, an Indo-Pacific invasive oyster to the eastern Mediterranean Sea, were compared with those of Chama savignyi, its northern Red Sea congeneric species. Summer and winter bacterial populations were characterized and compared using 16S rDNA clone libraries, and seasonal population dynamics were monitored by automated ribosomal intergenic spacer analysis (ARISA). Clone libraries revealed a specific clade of bacteria, closely related to marine endosymbionts from the Indo-Pacific, found in both ecosystems, of which one taxon was conserved in oysters from both sites. This taxon was dominant in summer libraries and was weakly present in winter ones, where other members of this group were dominant. ARISA results revealed significant seasonal variation in bacterial populations of Mediterranean Sea oysters, as opposed to Red Sea ones that were stable throughout the year. We suggest that this conserved association between bacteria and oyster reflects either a symbiosis between the oyster host and some of its bacteria, a co-invasion of both parties, or both.

Genotype is a stronger determinant than sex of the mouse gut microbiota.

The mammalian gut microbiota is considered to be determined mostly by diet, while the effect of genotype is still controversial. Here, we examined the effect of genotype on the gut microbiota in normal populations, exhibiting only natural polymorphisms, and evaluated this effect in comparison to the effect of sex. DNA fingerprinting approaches were used to profile the gut microbiota of eight different recombinant inbred mouse lines of the collaborative cross consortium, whose level of genetic diversity mimics that of a natural human population. Analyses based on automated ribosomal internal transcribed spacer analysis demonstrated significant higher similarity of the gut microbiota composition within mouse lines than between them or within same-gender groups. Thus, genetic background significantly impacts the microbiota composition and is a stronger determinant than gender. These findings imply that genetic polymorphisms help shape the intestinal microbiota of mammals and consequently could affect host susceptibility to diseases.

Integration of a foreign gene into a native complex does not impair fitness in an experimental model of lateral gene transfer.

Lateral gene transfer (LGT) is a central force in microbial evolution. The observation that genes encoding subunits of complexes exhibit relatively compatible phylogenies, suggesting vertical descent, can be explained by different evolutionary scenarios. On the one hand, the failure of a new gene product to correctly interact with preexisting protein subunits can make its acquisition neutral-a theory termed the "complexity hypothesis." On the other hand, foreign subunit-encoding genes may reduce the fitness of the new host by disrupting the stoichiometric balance between complex subunits, resulting in purifying selection against gene retention. We previously showed in a model LGT system that overexpression of an orthologous subunit was neutral due to lack of interaction with host subunits. Here, we examine a case where the foreign protein is more similar to its native orthologs, by expressing the RNA polymerase ß subunit (RpoB) of Bacillus subtilis in Escherichia coli. The foreign subunit is shown by coimmunoprecipitation to interact with the host subunits, and to form novel, nonspecific interactions. Nevertheless, the host did not incur any fitness disadvantage, as measured by its growth. We conclude that LGT of complex subunits may be neutral even when the transferred subunit can integrate into the host complex and that this neutrality can be a fertile ground for selective forces once the environment changes.

A systematic assessment of automated ribosomal intergenic spacer analysis (ARISA) as a tool for estimating bacterial richness.

ARISA (automated ribosomal intergenic spacer analysis) is a commonly used method for microbial community analysis that provides estimates of microbial richness and diversity. Here we investigated the potential biases of ARISA in richness estimation by performing computer simulations using 722 complete genomes. Our simulations based on in silico PCR demonstrated that over 8% of bacterial strains represented by complete genomes will never yield a PCR fragment using ARISA primers, usually because their ribosomal RNA genes are not organized in an operon. Despite the tendency of ARISA to overestimate species richness, a strong linear correlation exists between the observed number of fragments, even after binning, and the actual number of species in the sample. This linearity is fairly robust to the taxon sampling in the database as it is also observed on subsets of the 722 genome database using a jackknife approach. However, this linearity disappears when the species richness is high and binned fragment lengths gradually become saturated. We suggest that for ARISA-based richness estimates, where the number of binned lengths observed ranges between 10 and 116, a correction should be used in order to obtain more accurate "species richness" results comparable to 16S rRNA clone-library data.