Short-term genome stability of serial Clostridium difficile ribotype 027 isolates in an experimental gut model and recurrent human disease

Copyright: © 2013 Eyre et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are creditedClostridium difficile whole genome sequencing has the potential to identify related isolates, even among otherwise indistinguishable strains, but interpretation depends on understanding genomic variation within isolates and individuals.Serial isolates from two scenarios were whole genome sequenced. Firstly, 62 isolates from 29 timepoints from three in vitro gut models, inoculated with a NAP1/027 strain. Secondly, 122 isolates from 44 patients (2–8 samples/patient) with mostly recurrent/on-going symptomatic NAP-1/027 C. difficile infection. Reference-based mapping was used to identify single nucleotide variants (SNVs).Across three gut model inductions, two with antibiotic treatment, total 137 days, only two new SNVs became established. Pre-existing minority SNVs became dominant in two models. Several SNVs were detected, only present in the minority of colonies at one/two timepoints. The median (inter-quartile range) [range] time between patients’ first and last samples was 60 (29.5–118.5) [0–561] days. Within-patient C. difficile evolution was 0.45 SNVs/called genome/year (95%CI 0.00–1.28) and within-host diversity was 0.28 SNVs/called genome (0.05–0.53). 26/28 gut model and patient SNVs were non-synonymous, affecting a range of gene targets.The consistency of whole genome sequencing data from gut model C. difficile isolates, and the high stability of genomic sequences in isolates from patients, supports the use of whole genome sequencing in detailed transmission investigations.Peer reviewe

Evidence for Pervasive Adaptive Protein Evolution in Wild Mice

The relative contributions of neutral and adaptive substitutions to molecular evolution has been one of the most controversial issues in evolutionary biology for more than 40 years. The analysis of within-species nucleotide polymorphism and between-species divergence data supports a widespread role for adaptive protein evolution in certain taxa. For example, estimates of the proportion of adaptive amino acid substitutions (alpha) are 50% or more in enteric bacteria and Drosophila. In contrast, recent estimates of alpha for hominids have been at most 13%. Here, we estimate alpha for protein sequences of murid rodents based on nucleotide polymorphism data from multiple genes in a population of the house mouse subspecies Mus musculus castaneus, which inhabits the ancestral range of the Mus species complex and nucleotide divergence between M. m. castaneus and M. famulus or the rat. We estimate that 57% of amino acid substitutions in murids have been driven by positive selection. Hominids, therefore, are exceptional in having low apparent levels of adaptive protein evolution. The high frequency of adaptive amino acid substitutions in wild mice is consistent with their large effective population size, leading to effective natural selection at the molecular level. Effective natural selection also manifests itself as a paucity of effectively neutral nonsynonymous mutations in M. m. castaneus compared to humans

Comparison of Control of Clostridium difficile Infection in Six English Hospitals Using Whole-Genome Sequencing

Background: Variation in Clostridium difficile infection (CDI) rates between healthcare institutions suggests overall incidence could be reduced if the lowest rates could be achieved more widely. Methods: We investigated whether whole-genome sequencing (WGS) of consecutive C. difficile isolates from six English hospitals over one year (2013-14) could be used to assess infection control performance. Fecal samples with a positive initial screen for C. difficile (GDH or toxin-PCR) were cultured and sequenced. Within each hospital, we estimated the proportion of cases plausibly acquired from previous cases, defined by an isolate ≤2 single nucleotide polymorphisms different from a previous isolate in the last 90-days. Results: 851/971(87.6%) sequenced culture-positive samples were toxigenic, and 451(46.4%) were fecal-toxin-positive. 128/652(20%,95%CI 17-23%) toxigenic isolates >90-days after the study started were genetically-linked to a prior patient’s isolate from the previous 90-days. Hospital-2 had the fewest linked isolates, 7/105(7%,3-13%), hospital-1 an intermediate proportion, 9/70(13%,6-23%), while hospitals 3-6 had similar proportions of linked isolates (22-26%) (p≤0.002 comparing hospital-2 vs 3-6). Results were similar adjusting for locally-circulating ribotypes. Adjusting for hospital, ribotype-027 had the highest proportion of linked isolates (57%, 95%CI 29-81%). Fecal-toxin-positive and toxin-negative patients were similarly infectious in terms of being a potential transmission donor, OR=1.01(0.68-1.49,p=0.97). There was no association between the estimated proportion of cases linked to a previous case within 90-days and testing rates (p=0.60). Conclusions: WGS can be used to identify varying rates of C. difficile transmission in different locations, and offers the potential to allow targeted efforts to reduce CDI incidence

Comparison of Multilocus Variable-Number Tandem-Repeat Analysis and Whole-Genome Sequencing for Investigation of Clostridium difficile Transmission

No study to date has compared multilocus variable-number tandem-repeat analysis (MLVA) and whole-genome sequencing (WGS) in an investigation of the transmission of Clostridium difficile infection. Isolates from 61 adults with ongoing and/or recurrent C. difficile infections and 17 asymptomatic carriage episodes in children (201 samples), as well as from 61 suspected outbreaks affecting 2 to 41 patients in 31 hospitals in the United Kingdom (300 samples), underwent 7-locus MLVA and WGS in parallel. When the first and last samples from the same individual taken for a median (interquartile range [IQR]) of 63 days (43 to 105 days) apart were compared, the estimated rates of the evolution of single nucleotide variants (SNVs), summed tandem-repeat differences (STRDs), and locus variants (LVs) were 0.79 (95% confidence interval [CI], 0.00 to 1.75), 1.63 (95% CI, 0.00 to 3.59), and 1.21 (95% CI, 0.00 to 2.67)/called genome/year, respectively. Differences of >2 SNVs and >10 STRDs have been used to exclude direct case-to-case transmission. With the first serial sample per individual being used to assess discriminatory power, across all pairs of samples sharing a PCR ribotype, 192/283 (68%) differed by >10 STRDs and 217/283 (77%) by >2 SNVs. Among all pairs of cases from the same suspected outbreak, 1,190/1,488 (80%) pairs had concordant results using >2 SNVs and >10 STRDs to exclude transmission. For the discordant pairs, 229 (15%) had ≥2 SNVs but ≤10 STRDs, and 69 (5%) had ≤2 SNVs but ≥10 STRDs. Discordant pairs had higher numbers of LVs than concordant pairs, supporting the more diverse measure in each type of discordant pair. Conclusions on whether the potential outbreaks were confirmed were concordant in 58/61 (95%) investigations. Overall findings using MLVA and WGS were very similar despite the fact that they analyzed different parts of the bacterial genome. With improvements in WGS technology, it is likely that MLVA locus data will be available from WGS in the near future

Carbon amendment stimulates benthic nitrogen cycling during the bioremediation of particulate aquaculture waste

The treatment of organic wastes remains one of the key sustainability challenges facing the growing global aquaculture industry. Bioremediation systems based on coupled bioturbation-microbial processing offer a promising route for waste management. We present, for the first time, a combined biogeochemical-molecular analysis of the short-term performance of one such system that is designed to receive nitrogen-rich particulate aquaculture wastes. Using sea cucumbers (Holothuria scabra) as a model bioturbator we provide evidence that adjusting the waste CgN from 5g1 to 20g1 promoted a shift in nitrogen cycling pathways towards the dissimilatory nitrate reduction to ammonium (DNRA), resulting in net NH4+ efflux from the sediment. The carbon amended treatment exhibited an overall net N2 uptake, whereas the control receiving only aquaculture waste exhibited net N2 production, suggesting that carbon supplementation enhanced nitrogen fixation. The higher NH4+ efflux and N2 uptake was further supported by meta-genome predictions that indicate that organic-carbon addition stimulated DNRA over denitrification. These findings indicate that carbon addition may potentially result in greater retention of nitrogen within the system; however, longer-term trials are necessary to determine whether this nitrogen retention is translated into improved sea cucumber biomass yields. Whether this truly constitutes a remediation process is open for debate as there remains the risk that any increased nitrogen retention may be temporary, with any subsequent release potentially raising the eutrophication risk. Longer and larger-scale trials are required before this approach may be validated with the complexities of the in-system nitrogen cycle being fully understood

Extracting science from surveys of our Galaxy

Our knowledge of the Galaxy is being revolutionised by a series of photometric, spectroscopic and astrometric surveys. Already an enormous body of data is available from completed surveys, and data of ever increasing quality and richness will accrue at least until the end of this decade. To extract science from these surveys we need a class of models that can give probability density functions in the space of the observables of a survey -- we should not attempt to "invert" the data from the space of observables into the physical space of the Galaxy. Currently just one class of model has the required capability, so-called "torus models". A pilot application of torus models to understanding the structure of the Galaxy's thin and thick discs has already produced two significant results: a major revision of our best estimate of the Sun's velocity with respect to the Local Standard of Rest, and a successful prediction of the way in which the vertical velocity dispersion in the disc varies with distance from the Galactic plane.Comment: 13 pages. Invited review to appear in Pramana - journal of physics (Indian Academy of Sciences

Nurturing the young shoots of talent: Using action research for exploration and theory building

This is an Author's Accepted Manuscript of an article published in European Early Childhood Education Research Journal, 19(4), 433-450, 2011, copyright Taylor & Francis, available online at: http://www.tandfonline.com/10.1080/1350293X.2011.623515.This paper reports the outcomes of a set of action research projects carried out by teacher researchers in 14 local education authorities in England, working collaboratively with university tutors, over a period of three years. The common aim of all the projects was to explore practical ways of nurturing the gifts and talents of children aged four–seven years. The project was funded by the Department of Education and Skills in England as part of the government's gifted and talented programme. The project teachers felt that their understanding of issues relating to nurturing the gifts and talents of younger children was enhanced through their engagement in the project. It was possible to map the findings of the projects to the English government's National Quality Standards for gifted and talented education which include: (1) identification; (2) effective provision in the classroom; (3) enabling curriculum entitlement and choice; (4) assessment for learning; (5) engaging with community, families and beyond. The findings are also analysed within the framework of good practice in educating children in the first years of schooling. Participating practitioners felt that action research offered them a suitable methodology to explore the complexity of the topic of giftedness through cycles of planning, action and reflection and personal theory building

Role of carbonate burial in Blue Carbon budgets

Calcium carbonates (CaCO3) often accumulate in mangrove and seagrass sediments. As CaCO3 production emits CO2, there is concern that this may partially offset the role of Blue Carbon ecosystems as CO2sinks through the burial of organic carbon (Corg). A global collection of data on inorganic carbon burial rates (Cinorg, 12% of CaCO3 mass) revealed global rates of 0.8 TgCinorg yr−1 and 15–62 TgCinorg yr−1 in mangrove and seagrass ecosystems, respectively. In seagrass, CaCO3burial may correspond to an offset of 30% of the net CO2 sequestration. However, a mass balance assessment highlights that the Cinorg burial is mainly supported by inputs from adjacent ecosystems rather than by local calcification, and that Blue Carbon ecosystems are sites of net CaCO3 dissolution. Hence, CaCO3 burial in Blue Carbon ecosystems contribute to seabed elevation and therefore buffers sea-level rise, without undermining their role as CO2 sinks

Longitudinal relationships between caloric expenditure and gray matter in the cardiovascular health study

Background: Physical activity (PA) can be neuroprotective and reduce the risk for Alzheimer’s disease (AD). In assessing physical activity, caloric expenditure is a proxy marker reflecting the sum total of multiple physical activity types conducted by an individual. Objective:To assess caloric expenditure, as a proxy marker of PA, as a predictive measure of gray matter (GM) volumes in the normal and cognitively impaired elderly persons. Methods: All subjects in this study were recruited from the Institutional Review Board approved Cardiovascular Health Study (CHS), a multisite population-based longitudinal study in persons aged 65 and older. We analyzed a sub-sample of CHS participants 876 subjects (mean age 78.3, 57.5% F, 42.5% M) who had i) energy output assessed as kilocalories (kcal) per week using the standardized Minnesota Leisure-Time Activities questionnaire, ii) cognitive assessments for clinical classification of normal cognition, mild cognitive impairment (MCI), and AD, and iii) volumetric MR imaging of the brain. Voxel-based morphometry modeled the relationship between kcal/week and GM volumes while accounting for standard covariates including head size, age, sex, white matter hyperintensity lesions, MCI or AD status, and site. Multiple comparisons were controlled using a False Discovery Rate of 5 percent. Results: Higher energy output, from a variety of physical activity types, was associated with larger GM volumes in frontal, temporal, and parietal lobes, as well as hippocampus, thalamus, and basal ganglia. High levels of caloric expenditure moderated neurodegeneration-associated volume loss in the precuneus, posterior cingulate, and cerebellar vermis. Conclusion:Increasing energy output from a variety of physical activities is related to larger gray matter volumes in the elderly, regardless of cognitive status.Cyrus A. Raji, David A. Merrill, Harris Eyre, Sravya Mallam, Nare Torosyan, Kirk I. Erickson, Oscar L. Lopez , James T. Becker, Owen T. Carmichael, H. Michael Gach, Paul M. Thompson, W.T. Longstreth, Jr. and Lewis H. Kulle