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ObjectiveAn adequate bowel cleansing is essential for a successful colonoscopy. Although purgative consumption is safe for the patient, there is little consensus on how the intestinal microbiota is affected by the procedure, especially regarding the potential long-term consequences.Design23 healthy subjects were randomised into two study groups consuming a bowel preparation (Moviprep), either in two separate doses of 1 L or as a single 2-L dose. Participants donated faecal samples at the baseline, after bowel cleansing, 14 and 28 days after the treatment. The intestinal microbiota composition was determined with phylogenetic microarray as well as quantitative PCR analysis and correlated with the previously quantified faecal serine proteases.ResultsThe lavage introduced an instant and substantial change to the intestinal microbiota. The total microbial load was decreased by 31-fold and 22% of the participants lost the subject-specificity of their microbiota. While the bacterial levels and community composition were essentially restored within 14 days, the rate of recovery was dose dependent: consumption of the purgative in a single dose had a more severe effect on the microbiota composition than that of a double dose, and notably increased the levels of Proteobacteria, Fusobacteria and bacteria related to Dorea formicigenerans. The abundance of the latter also correlated with the amount of faecal serine proteases that were increased after purging.ConclusionsOur results suggest that the bowel cleansing using two separate dosages introduces fewer alterations to the intestinal microbiota than a single dose and hence may be preferred in clinical practice.

Introduction and hypothesis The purpose of this paper is to report our evaluation of changes in intravaginal microbial flora after ring pessary therapy for pelvic organ prolapse (POP) using conventional and our clone library method. Methods Thirteen patients with POP who were fitted with a ring pessary participated in this longitudinal study that incorporates data from before and 1 month after beginning ring pessary therapy. Changes in intravaginal microbial flora were evaluated by conventional methods, i.e., vaginal pH, lactobacillary grade (LAC grade), Nugent score, and culture-based bacterial detection methods. In addition, we performed our clone library method using 16S ribosomal RNA (rRNA) sequencing of vaginal fluid. Results Conventional methods revealed that most patients had abnormal intravaginal microbial flora. Mean numbers of detected bacterial species by the culture-based and our clone library method were 3.1 (1–6) and 11.8 (1–25), respectively. Our clone library method showed that Lactobacillus spp. increased in four and decreased in two cases after ring pessary therapy but reappeared after therapy in two cases; no Lactobacillus spp. were detected in five cases. Conclusions Our study showed that ring pessary therapy did not always disturb intravaginal microbial flora, especially for patients with Lactobacillus spp. prior to ring pessary insertion. Anaerobic circumstances in the vagina after therapy seem to have induced the growth of anaerobic bacteria.

Twelve trachoma-hyperendemic communities were treated with 3 annual mass azithromycin distributions. Children aged 0-9 years were monitored 1 year following the third treatment. An RNA-based test detected ocular chlamydial infection in more children than did a DNA-based test (6.9% vs 4.2%), and in a larger number of communities (8 vs 7).

Assessing the genetic variation of bacteria has become ever more complex as more sequencing data has become available. Here, Maiden and colleagues propose a gene-by-gene approach of analysing whole-genome data; this approach is based on their experience with multilocus sequence typing (MLST) and reflects the functional and evolutionary relationships among bacteria. Multilocus sequence typing (MLST) was proposed in 1998 as a portable sequence-based method for identifying clonal relationships among bacteria. Today, in the whole-genome era of microbiology, the need for systematic, standardized descriptions of bacterial genotypic variation remains a priority. Here, to meet this need, we draw on the successes of MLST and 16S rRNA gene sequencing to propose a hierarchical gene-by-gene approach that reflects functional and evolutionary relationships and catalogues bacteria 'from domain to strain'. Our gene-based typing approach using online platforms such as the Bacterial Isolate Genome Sequence Database (BIGSdb) allows the scalable organization and analysis of whole-genome sequence data.

Protein and messenger RNA (mRNA) copy numbers vary from cell to cell in isogenic bacterial populations. However, these molecules often exist in low copy numbers and are difficult to detect in single cells. We carried out quantitative system-wide analyses of protein and mRNA expression in individual cells with single-molecule sensitivity using a newly constructed yellow fluorescent protein fusion library for Escherichia coli. We found that almost all protein number distributions can be described by the gamma distribution with two fitting parameters which, at low expression levels, have clear physical interpretations as the transcription rate and protein burst size. At high expression levels, the distributions are dominated by extrinsic noise. We found that a single cell's protein and mRNA copy numbers for any given gene are uncorrelated.

Eukaryotic cells spatially organize mRNA processes such as translation and mRNA decay. Much less is clear in bacterial cells where the spatial distribution of mature mRNA remains ambiguous. Using a sensitive method based on quantitative fluorescence in situ hybridization, we show here that in Caulobacter crescentus and Escherichia coli , chromosomally expressed mRNAs largely display limited dispersion from their site of transcription during their lifetime. We estimate apparent diffusion coefficients at least two orders of magnitude lower than expected for freely diffusing mRNA, and provide evidence in C. crescentus that this mRNA localization restricts ribosomal mobility. Furthermore, C. crescentus RNase E appears associated with the DNA independently of its mRNA substrates. Collectively, our findings show that bacteria can spatially organize translation and, potentially, mRNA decay by using the chromosome layout as a template. This chromosome-centric organization has important implications for cellular physiology and for our understanding of gene expression in bacteria. Bacterial organization The lack of compartmentalization in membrane-enclosed compartments in bacteria makes it difficult to determine how mature messenger RNAs are distributed within the cell. Llopis et al . used fluorescence experiments — tricky because bacteria are so small — to follow mRNA dispersal after transcription. They found, surprisingly, that newly transcribed mRNAs showed limited diffusion; they speculate that the packed chromosomal material may itself act as a partition to separate translation from mRNA degradation. In bacteria, the lack of compartmentalization within membrane-enclosed compartments has made it difficult to determine how mature messenger RNAs are spatially distributed. Here the authors use fluorescence experiments in bacteria to follow mRNA dispersal after transcription. They find, surprisingly, that the newly transcribed mRNAs show limited diffusion, and speculate that the packed chromosomal material may itself act as a partition to separate translation from mRNA degradation.

Mycobacterium tuberculosis (Mtb) is a bacterial pathogen that causes tuberculosis (TB), an infectious disease that is responsible for major health and economic costs worldwide 1 . Mtb encounters diverse environments during its life cycle and responds to these changes largely by reprogramming its transcriptional output 2 . However, the mechanisms of Mtb transcription and how they are regulated remain poorly understood. Here we use a sequencing method that simultaneously determines both termini of individual RNA molecules in bacterial cells 3 to profile the Mtb transcriptome at high resolution. Unexpectedly, we find that most Mtb transcripts are incomplete, with their 5′ ends aligned at transcription start sites and 3′ ends located 200–500 nucleotides downstream. We show that these short RNAs are mainly associated with paused RNA polymerases (RNAPs) rather than being products of premature termination. We further show that the high propensity of Mtb RNAP to pause early in transcription relies on the binding of the σ-factor. Finally, we show that a translating ribosome promotes transcription elongation, revealing a potential role for transcription–translation coupling in controlling Mtb gene expression. In sum, our findings depict a mycobacterial transcriptome that prominently features incomplete transcripts resulting from RNAP pausing. We propose that the pausing phase constitutes an important transcriptional checkpoint in Mtb that allows the bacterium to adapt to environmental changes and could be exploited for TB therapeutics. A study reveals that most transcripts in Mycobacterium tuberculosis are incomplete, likely because of the tendency of the transcription machinery in this species to pause on genomic DNA.

ObjectiveA decade of microbiome studies has linked IBD to an alteration in the gut microbial community of genetically predisposed subjects. However, existing profiles of gut microbiome dysbiosis in adult IBD patients are inconsistent among published studies, and did not allow the identification of microbial signatures for CD and UC. Here, we aimed to compare the faecal microbiome of CD with patients having UC and with non-IBD subjects in a longitudinal study.DesignWe analysed a cohort of 2045 non-IBD and IBD faecal samples from four countries (Spain, Belgium, the UK and Germany), applied a 16S rRNA sequencing approach and analysed a total dataset of 115 million sequences.ResultsIn the Spanish cohort, dysbiosis was found significantly greater in patients with CD than with UC, as shown by a more reduced diversity, a less stable microbial community and eight microbial groups were proposed as a specific microbial signature for CD. Tested against the whole cohort, the signature achieved an overall sensitivity of 80% and a specificity of 94%, 94%, 89% and 91% for the detection of CD versus healthy controls, patients with anorexia, IBS and UC, respectively.ConclusionsAlthough UC and CD share many epidemiologic, immunologic, therapeutic and clinical features, our results showed that they are two distinct subtypes of IBD at the microbiome level. For the first time, we are proposing microbiomarkers to discriminate between CD and non-CD independently of geographical regions.

Objective Antibiotic (AB) usage strongly affects microbial intestinal metabolism and thereby impacts human health. Understanding this process and the underlying mechanisms remains a major research goal. Accordingly, we conducted the first comparative omic investigation of gut microbial communities in faecal samples taken at multiple time points from an individual subjected to β-lactam therapy. Methods The total (16S rDNA) and active (16S rRNA) microbiota, metagenome, metatranscriptome (mRNAs), metametabolome (high-performance liquid chromatography coupled to electrospray ionisation and quadrupole time-of-flight mass spectrometry) and metaproteome (ultra high performing liquid chromatography coupled to an Orbitrap MS2 instrument [UPLC-LTQ Orbitrap-MS/MS]) of a patient undergoing AB therapy for 14 days were evaluated. Results Apparently oscillatory population dynamics were observed, with an early reduction in Gram-negative organisms (day 6) and an overall collapse in diversity and possible further colonisation by ‘presumptive’ naturally resistant bacteria (day 11), followed by the re-growth of Gram-positive species (day 14). During this process, the maximum imbalance in the active microbial fraction occurred later (day 14) than the greatest change in the total microbial fraction, which reached a minimum biodiversity and richness on day 11; additionally, major metabolic changes occurred at day 6. Gut bacteria respond to ABs early by activating systems to avoid the antimicrobial effects of the drugs, while ‘presumptively’ attenuating their overall energetic metabolic status and the capacity to transport and metabolise bile acid, cholesterol, hormones and vitamins; host–microbial interactions significantly improved after treatment cessation. Conclusions This proof-of-concept study provides an extensive description of gut microbiota responses to follow-up β-lactam therapy. The results demonstrate that ABs targeting specific pathogenic infections and diseases may alter gut microbial ecology and interactions with host metabolism at a much higher level than previously assumed.

Background Parkinson’s disease (PD) is a systemic disease clinically defined by the degeneration of dopaminergic neurons in the brain. While alterations in the gut microbiome composition have been reported in PD, their functional consequences remain unclear. Herein, we addressed this question by an analysis of stool samples from the Luxembourg Parkinson’s Study ( n  = 147 typical PD cases, n  = 162 controls). Results All individuals underwent detailed clinical assessment, including neurological examinations and neuropsychological tests followed by self-reporting questionnaires. Stool samples from these individuals were first analysed by 16S rRNA gene sequencing. Second, we predicted the potential secretion for 129 microbial metabolites through personalised metabolic modelling using the microbiome data and genome-scale metabolic reconstructions of human gut microbes. Our key results include the following. Eight genera and seven species changed significantly in their relative abundances between PD patients and healthy controls. PD-associated microbial patterns statistically depended on sex, age, BMI, and constipation. Particularly, the relative abundances of Bilophila and Paraprevotella were significantly associated with the Hoehn and Yahr staging after controlling for the disease duration. Furthermore, personalised metabolic modelling of the gut microbiomes revealed PD-associated metabolic patterns in the predicted secretion potential of nine microbial metabolites in PD, including increased methionine and cysteinylglycine. The predicted microbial pantothenic acid production potential was linked to the presence of specific non-motor symptoms. Conclusion Our results suggest that PD-associated alterations of the gut microbiome can translate into substantial functional differences affecting host metabolism and disease phenotype.