Species classifier choice is a key consideration when analysing low-complexity food microbiome data

peer-reviewedBackground The use of shotgun metagenomics to analyse low-complexity microbial communities in foods has the potential to be of considerable fundamental and applied value. However, there is currently no consensus with respect to choice of species classification tool, platform, or sequencing depth. Here, we benchmarked the performances of three high-throughput short-read sequencing platforms, the Illumina MiSeq, NextSeq 500, and Ion Proton, for shotgun metagenomics of food microbiota. Briefly, we sequenced six kefir DNA samples and a mock community DNA sample, the latter constructed by evenly mixing genomic DNA from 13 food-related bacterial species. A variety of bioinformatic tools were used to analyse the data generated, and the effects of sequencing depth on these analyses were tested by randomly subsampling reads. Results Compositional analysis results were consistent between the platforms at divergent sequencing depths. However, we observed pronounced differences in the predictions from species classification tools. Indeed, PERMANOVA indicated that there was no significant differences between the compositional results generated by the different sequencers (p = 0.693, R2 = 0.011), but there was a significant difference between the results predicted by the species classifiers (p = 0.01, R2 = 0.127). The relative abundances predicted by the classifiers, apart from MetaPhlAn2, were apparently biased by reference genome sizes. Additionally, we observed varying false-positive rates among the classifiers. MetaPhlAn2 had the lowest false-positive rate, whereas SLIMM had the greatest false-positive rate. Strain-level analysis results were also similar across platforms. Each platform correctly identified the strains present in the mock community, but accuracy was improved slightly with greater sequencing depth. Notably, PanPhlAn detected the dominant strains in each kefir sample above 500,000 reads per sample. Again, the outputs from functional profiling analysis using SUPER-FOCUS were generally accordant between the platforms at different sequencing depths. Finally, and expectedly, metagenome assembly completeness was significantly lower on the MiSeq than either on the NextSeq (p = 0.03) or the Proton (p = 0.011), and it improved with increased sequencing depth. Conclusions Our results demonstrate a remarkable similarity in the results generated by the three sequencing platforms at different sequencing depths, and, in fact, the choice of bioinformatics methodology had a more evident impact on results than the choice of sequencer did.This research was funded by Science Foundation Ireland in the form of a centre grant (APC Microbiome Institute grant number SFI/12/RC/2273). Research in the Cotter laboratory is also funded by Science Foundation Ireland through the PI award “Obesibiotics” (11/PI/1137). Orla O’Sullivan is funded by Science Foundation Ireland through a Starting Investigator Research Grant award (13/SIRG/2160)

Men Are from Quartile One, Women Are from? Relative Age Effect in European Soccer and the Influence of Age, Success, and Playing Status

The relative age effect (RAE) is characterised by an overrepresentation of athletes born earlier in the selection year. Whilst an RAE is consistently evident in male soccer, examinations in female players remain limited. The aim of the present study was to examine the influence of sex, as well as age, success, and playing status in European soccer players. The sample consisted of a total of 6546 soccer players from 55 soccer nations that competed in recent European Championship qualification campaigns. Results indicated an evident RAE in male [p = 0.017] but not female [p = 0.765] players. Male players were over-represented by players born in the first quartile for the U17 [p < 0.001] and U19 [p = 0.001] levels, however, this over-representation did not transfer to senior levels. No RAE was observed at any level for female players. Inside each age group, a slight selection bias towards those born in the first quartile for successful squads was observed but did not significantly differentiate between qualification status for either male or female players. Results from this study highlight the disparity in RAE prevalence between male and female players and raise further questions regarding the value of selecting relatively older players to metrics of success, transition, and selection for senior international soccer

Colonic Gene Expression and Fecal Microbiota in Diarrhea-predominant Irritable Bowel Syndrome : Increased Toll-like Receptor 4 but Minimal Inflammation and no Response to Mesalazine

Background/Aims Diarrhea-predominant irritable bowel syndrome (IBS-D) has been previously associated with evidence of immune activation and altered microbiota. Our aim is to assess the effect of the anti-inflammatory agent, mesalazine, on inflammatory gene expression and microbiota composition in IBS-D. Methods We studied a subset of patients (n = 43) from a previously published 12-week radomized placebo-controlled trial of mesalazine. Mucosal biopsies were assessed by immunohistochemistry and reverse transcription-polymerase chain reaction for a range of markers of inflammation, altered permeability, and sensory receptors including Toll-like receptors (TLRs) at randomization after treatment. All biopsy data were compared to 21 healthy controls. Patient's stool microbiota composition was analysed through 16S ribosomal RNA sequencing. Results We found no evidence of increased immune activation compared to healthy controls. However, we did find increased expression of receptors in both sensory pathways and innate immune response including TLR4. Higher TLR4 expression was associated with greater urgency. TLR4 expression correlated strongly with the expression of the receptors bradykinin receptor B2, chemerin chemokine-like receptor 1, and transient receptor potential cation channel, subfamily A, member 1 as well as TLR4's downstream adaptor myeloid differentiation factor 88. Mesalazine had minimal effect on either gene expression or microbiota composition. Conclusions Biopsies from a well-characterized IBS-D cohort showed no substantial inflammation. Mesalazine has little effect on gene expression and its previous reported effect on fecal microbiota associated with much greater inflammation found in inflammatory bowel diseases is likely secondary to reduced inflammation. Increased expression of TLR4 and correlated receptors in IBS may mediate a general increase in sensitivity to external stimuli, particularly those that signal via the TLR system.Peer reviewe

Bacterial genomics reveal the complex epidemiology of an emerging pathogen in Arctic and boreal ungulates

Northern ecosystems are currently experiencing unprecedented ecological change, largely driven by a rapidly changing climate. Pathogen range expansion, and emergence and altered patterns of infectious disease, are increasingly reported in wildlife at high latitudes. Understanding the causes and consequences of shifting pathogen diversity and host-pathogen interactions in these ecosystems is important for wildlife conservation, and for indigenous populations that depend on wildlife. Among the key questions are whether disease events are associated with endemic or recently introduced pathogens, and whether emerging strains are spreading throughout the region. In this study, we used a phylogenomic approach to address these questions of pathogen endemicity and spread for Erysipelothrix rhusiopathiae, an opportunistic multi-host bacterial pathogen associated with recent mortalities in arctic and boreal ungulate populations in North America. We isolated E. rhusiopathiae from carcasses associated with large-scale die-offs of muskoxen in the Canadian Arctic Archipelago, and from contemporaneous mortality events and/or population declines among muskoxen in northwestern Alaska and caribou and moose in western Canada. Bacterial genomic diversity differed markedly among these locations; minimal divergence was present among isolates from muskoxen in the Canadian Arctic, while in caribou and moose populations, strains from highly divergent clades were isolated from the same location, or even from within a single carcass. These results indicate that mortalities among northern ungulates are not associated with a single emerging strain of E. rhusiopathiae, and that alternate hypotheses need to be explored. Our study illustrates the value and limitations of bacterial genomic data for discriminating between ecological hypotheses of disease emergence, and highlights the importance of studying emerging pathogens within the broader context of environmental and host factors

Postinfective bowel dysfunction following Campylobacter enteritis is characterised by reduced microbiota diversity and impaired microbiota recovery

Objectives Persistent bowel dysfunction following gastroenteritis (postinfectious (PI)-BD) is well recognised, but the associated changes in microbiota remain unclear. Our aim was to define these changes after gastroenteritis caused by a single organism, Campylobacter jejuni, examining the dynamic changes in the microbiota and the impact of antibiotics. Design A single-centre cohort study of 155 patients infected with Campylobacter jejuni. Features of the initial illness as well as current bowel symptoms and the intestinal microbiota composition were recorded soon after infection (visit 1, 80 days later (visits 2 and 3). Microbiota were assessed using 16S rRNA sequencing. Results PI-BD was found in 22 of the 99 patients who completed the trial. The cases reported significantly looser stools, with more somatic and gastrointestinal symptoms. Microbiota were assessed in 22 cases who had significantly lower diversity and altered microbiota composition compared with the 44 age-matched and sex-matched controls. Moreover 60 days after infection, cases showed a significantly lower abundance of 23 taxa including phylum Firmicutes, particularly in the order Clostridiales and the family Ruminoccocaceae, increased Proteobacteria abundance and increased levels of Fusobacteria and Gammaproteobacteria. The microbiota changes were linked with diet; higher fibre consumption being associated with lower levels of Gammaproteobacteria. Conclusion The microbiota of PI-BD patients appeared more disturbed by the initial infection compared with the microbiota of those who recovered. The prebiotic effect of high fibre diets may inhibit some of the disturbances seen in PI-BD.Peer reviewe

Mapping interactions between the RNA chaperone FinO and its RNA targets

Bacterial conjugation is regulated by two-component repression comprising the antisense RNA FinP, and its protein co-factor FinO. FinO mediates base-pairing of FinP to the 5′-untranslated region (UTR) of traJ mRNA, which leads to translational inhibition of the transcriptional activator TraJ and subsequent down regulation of conjugation genes. Yet, little is known about how FinO binds to its RNA targets or how this interaction facilitates FinP and traJ mRNA pairing. Here, we use solution methods to determine how FinO binds specifically to its minimal high affinity target, FinP stem–loop II (SLII), and its complement SLIIc from traJ mRNA. Ribonuclease footprinting reveals that FinO contacts the base of the stem and the 3′ single-stranded tails of these RNAs. The phosphorylation or oxidation of the 3′-nucleotide blocks FinO binding, suggesting FinO binds the 3′-hydroxyl of its RNA targets. The collective results allow the generation of an energy-minimized model of the FinO–SLII complex, consistent with small-angle X-ray scattering data. The repression complex model was constrained using previously reported cross-linking data and newly developed footprinting results. Together, these data lead us to propose a model of how FinO mediates FinP/traJ mRNA pairing to down regulate bacterial conjugation

The Application of Novel Research Technologies by the Deep Pelagic Nekton Dynamics of the Gulf of Mexico (DEEPEND) Consortium

The deep waters of the open ocean represent a major frontier in exploration and scientific understanding. However, modern technological and computational tools are making the deep ocean more accessible than ever before by facilitating increasingly sophisticated studies of deep ocean ecosystems. Here, we describe some of the cutting-edge technologies that have been employed by the Deep Pelagic Nekton Dynamics of the Gulf of Mexico (DEEPEND; www.deependconsortium.org) Consortium to study the biodiverse fauna and dynamic physical-chemical environment of the offshore Gulf of Mexico (GoM) from 0 to 1,500 m

Socioeconomic Inequality in the Prevalence of Autism Spectrum Disorder: Evidence from a U.S. Cross-Sectional Study

This study was designed to evaluate the hypothesis that the prevalence of autism spectrum disorder (ASD) among children in the United States is positively associated with socioeconomic status (SES).A cross-sectional study was implemented with data from the Autism and Developmental Disabilities Monitoring Network, a multiple source surveillance system that incorporates data from educational and health care sources to determine the number of 8-year-old children with ASD among defined populations. For the years 2002 and 2004, there were 3,680 children with ASD among a population of 557,689 8-year-old children. Area-level census SES indicators were used to compute ASD prevalence by SES tertiles of the population.Prevalence increased with increasing SES in a dose-response manner, with prevalence ratios relative to medium SES of 0.70 (95% confidence interval [CI] 0.64, 0.76) for low SES, and of 1.25 (95% CI 1.16, 1.35) for high SES, (P<0.001). Significant SES gradients were observed for children with and without a pre-existing ASD diagnosis, and in analyses stratified by gender, race/ethnicity, and surveillance data source. The SES gradient was significantly stronger in children with a pre-existing diagnosis than in those meeting criteria for ASD but with no previous record of an ASD diagnosis (p<0.001), and was not present in children with co-occurring ASD and intellectual disability.The stronger SES gradient in ASD prevalence in children with versus without a pre-existing ASD diagnosis points to potential ascertainment or diagnostic bias and to the possibility of SES disparity in access to services for children with autism. Further research is needed to confirm and understand the sources of this disparity so that policy implications can be drawn. Consideration should also be given to the possibility that there may be causal mechanisms or confounding factors associated with both high SES and vulnerability to ASD

Genetic Rearrangements Can Modify Chromatin Features at Epialleles

Analogous to genetically distinct alleles, epialleles represent heritable states of different gene expression from sequence-identical genes. Alleles and epialleles both contribute to phenotypic heterogeneity. While alleles originate from mutation and recombination, the source of epialleles is less well understood. We analyze active and inactive epialleles that were found at a transgenic insert with a selectable marker gene in Arabidopsis. Both converse expression states are stably transmitted to progeny. The silent epiallele was previously shown to change its state upon loss-of-function of trans-acting regulators and drug treatments. We analyzed the composition of the epialleles, their chromatin features, their nuclear localization, transcripts, and homologous small RNA. After mutagenesis by T-DNA transformation of plants carrying the silent epiallele, we found new active alleles. These switches were associated with different, larger or smaller, and non-overlapping deletions or rearrangements in the 3′ regions of the epiallele. These cis-mutations caused different degrees of gene expression stability depending on the nature of the sequence alteration, the consequences for transcription and transcripts, and the resulting chromatin organization upstream. This illustrates a tight dependence of epigenetic regulation on local structures and indicates that sequence alterations can cause epigenetic changes at some distance in regions not directly affected by the mutation. Similar effects may also be involved in gene expression and chromatin changes in the vicinity of transposon insertions or excisions, recombination events, or DNA repair processes and could contribute to the origin of new epialleles