Comparison of brush and biopsy sampling methods of the ileal pouch for assessment of mucosa-associated microbiota of human subjects

BACKGROUND: Mucosal biopsy is the most common sampling technique used to assess microbial communities associated with the intestinal mucosa. Biopsies disrupt the epithelium and can be associated with complications such as bleeding. Biopsies sample a limited area of the mucosa, which can lead to potential sampling bias. In contrast to the mucosal biopsy, the mucosal brush technique is less invasive and provides greater mucosal coverage, and if it can provide equivalent microbial community data, it would be preferable to mucosal biopsies. RESULTS: We compared microbial samples collected from the intestinal mucosa using either a cytology brush or mucosal biopsy forceps. We collected paired samples from patients with ulcerative colitis (UC) who had previously undergone colectomy and ileal pouch anal anastomosis (IPAA), and profiled the microbial communities of the samples by sequencing V4-V6 or V4-V5 16S rRNA-encoding gene amplicons. Comparisons of 177 taxa in 16 brush-biopsy sample pairs had a mean R(2) of 0.94. We found no taxa that varied significantly between the brush and biopsy samples after adjusting for multiple comparisons (false discovery rate ≤0.05). We also tested the reproducibility of DNA amplification and sequencing in 25 replicate pairs and found negligible variation (mean R(2) = 0.99). A qPCR analysis of the two methods showed that the relative yields of bacterial DNA to human DNA were several-fold higher in the brush samples than in the biopsies. CONCLUSIONS: Mucosal brushing is preferred to mucosal biopsy for sampling the epithelial-associated microbiota. Although both techniques provide similar assessments of the microbial community composition, the brush sampling method has relatively more bacterial to host DNA, covers a larger surface area, and is less traumatic to the epithelium than the mucosal biopsy

Binding energies of quantum dipole in plane

We propose a numerical algorithm based on a discrete variable representation and shifted inverse iterations and apply it to for the analysis of the bound states of edge dislocation modelled by a quantum dipole in a plane. The good agreement with results of recent papers of Amore et al [J. Phys. B 45, 235004 (2012)] was obtained. The error estimates of the previous results of low-lying states energies of other authors were not known due to limitations of the variational approaches and this paper fills this gap presenting calculated low-lying bound states energies by non-variational technique. The probability densities of low-lying states were calculated

Short-Range Interaction Impact on Two-Dimensional Dipolar Scattering

We report numerical investigation of the short range interaction influence on the two-dimensional quantum scattering of two dipoles. The model simulates two ultracold polar molecules collisions in two spatial dimensions. The used algorithm allows us to quantitatively analyse the scattering of two polarized dipoles with account for strongly anisotropic nature of dipolar interaction. The strong dependence of the scattering total cross section on the short range interaction radius was discovered for threshold collision energies. We also discuss differences of calculated scattering cross section dependencies for different polarisation axis tilt angles

A novel conjugative transposon carrying an autonomously amplified plasmid

ABSTRACTTetracyclines serve as broad-spectrum antibiotics to treat bacterial infections. The discovery of new tetracycline resistance genes has led to new questions about the underlying mechanisms of resistance, gene transfer, and their relevance to human health. We tracked changes in the abundance of a 55-kbp conjugative transposon (CTn214) carrying tetQ, a tetracycline resistance gene, within a Bacteroides fragilis metagenome-assembled genome derived from shotgun sequencing of microbial DNA extracted from the ileal pouch of a patient with ulcerative colitis. The mapping of metagenomic reads to CTn214 revealed the multi-copy nature of a 17,044-nt region containing tetQ in samples collected during inflammation and uninflamed visits. B. fragilis cultivars isolated from the same patient during periods of inflammation harbored CTn214 integrated into the chromosome or both a circular, multi-copy, extrachromosomal region of the CTn214 containing tetQ and the corresponding integrated form. The tetracycline-dependent mechanism for the transmission of CTn214 is nearly identical to a common conjugative transposon found in the genome of B. fragilis (CTnDOT), but the autonomously amplified nature of a circular 17,044-nt region of CTn214 that codes for tetQ and the integration of the same sequence in the linear chromosome within the same cell is a novel observation. Genome and transcriptome sequencing of B. fragilis cultivars grown under different concentrations of tetracycline and ciprofloxacin indicates that tetQ in strains containing the circular form remains actively expressed regardless of treatment, while the expression of tetQ in strains containing the linear form increases only in the presence of tetracycline.IMPORTANCEThe exchange of antibiotic production and resistance genes between microorganisms can lead to the emergence of new pathogens. In this study, short-read mapping of metagenomic samples taken over time from the illeal pouch of a patient with ulcerative colitis to a Bacteroides fragilis metagenome-assembled genome revealed two distinct genomic arrangements of a novel conjugative transposon, CTn214, that encodes tetracycline resistance. The autonomous amplification of a plasmid-like circular form from CTn214 that includes tetQ potentially provides consistent ribosome protection against tetracycline. This mode of antibiotic resistance offers a novel mechanism for understanding the emergence of pathobionts like B. fragilis and their persistence for extended periods of time in patients with inflammatory bowel disease