Clostridium difficile is a Gram-positive spore-forming anaerobe and a major cause of antibiotic-associated diarrhoea. Disruption of the commensal microbiota, such as through treatment with ...broad-spectrum antibiotics, is a critical precursor for colonisation by C. difficile and subsequent disease. Furthermore, failure of the gut microbiota to recover colonisation resistance can result in recurrence of infection. An unusual characteristic of C. difficile among gut bacteria is its ability to produce the bacteriostatic compound para-cresol (p-cresol) through fermentation of tyrosine. Here, we demonstrate that the ability of C. difficile to produce p-cresol in vitro provides a competitive advantage over gut bacteria including Escherichia coli, Klebsiella oxytoca and Bacteroides thetaiotaomicron. Metabolic profiling of competitive co-cultures revealed that acetate, alanine, butyrate, isobutyrate, p-cresol and p-hydroxyphenylacetate were the main metabolites responsible for differentiating the parent strain C. difficile (630Δerm) from a defined mutant deficient in p-cresol production. Moreover, we show that the p-cresol mutant displays a fitness defect in a mouse relapse model of C. difficile infection (CDI). Analysis of the microbiome from this mouse model of CDI demonstrates that colonisation by the p-cresol mutant results in a distinctly altered intestinal microbiota, and metabolic profile, with a greater representation of Gammaproteobacteria, including the Pseudomonales and Enterobacteriales. We demonstrate that Gammaproteobacteria are susceptible to exogenous p-cresol in vitro and that there is a clear divide between bacterial Phyla and their susceptibility to p-cresol. In general, Gram-negative species were relatively sensitive to p-cresol, whereas Gram-positive species were more tolerant. This study demonstrates that production of p-cresol by C. difficile has an effect on the viability of intestinal bacteria as well as the major metabolites produced in vitro. These observations are upheld in a mouse model of CDI, in which p-cresol production affects the biodiversity of gut microbiota and faecal metabolite profiles, suggesting that p-cresol production contributes to C. difficile survival and pathogenesis.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Autism spectrum disorder (ASD) is a neurodevelopmental condition with hallmark behavioral manifestations including impaired social communication and restricted repetitive behavior. In addition, many ...affected individuals display metabolic imbalances, immune dysregulation, gastrointestinal dysfunction, and altered gut microbiome compositions.
We sought to better understand nonbehavioral features of ASD by determining molecular signatures in peripheral tissues through mass spectrometry methods (ultrahigh performance liquid chromatography–tandem mass spectrometry) with broad panels of identified metabolites. Herein, we compared the global metabolome of 231 plasma and 97 fecal samples from a large cohort of children with ASD and typically developing control children.
Differences in amino acid, lipid, and xenobiotic metabolism distinguished ASD and typically developing samples. Our results implicated oxidative stress and mitochondrial dysfunction, hormone level elevations, lipid profile changes, and altered levels of phenolic microbial metabolites. We also revealed correlations between specific metabolite profiles and clinical behavior scores. Furthermore, a summary of metabolites modestly associated with gastrointestinal dysfunction in ASD is provided, and a pilot study of metabolites that can be transferred via fecal microbial transplant into mice is identified.
These findings support a connection between metabolism, gastrointestinal physiology, and complex behavioral traits and may advance discovery and development of molecular biomarkers for ASD.
Mycobacterium tuberculosis drug resistance (DR) challenges effective tuberculosis disease control. Current molecular tests examine limited numbers of mutations, and although whole genome sequencing ...approaches could fully characterise DR, data complexity has restricted their clinical application. A library (1,325 mutations) predictive of DR for 15 anti-tuberculosis drugs was compiled and validated for 11 of them using genomic-phenotypic data from 792 strains. A rapid online 'TB-Profiler' tool was developed to report DR and strain-type profiles directly from raw sequences. Using our DR mutation library, in silico diagnostic accuracy was superior to some commercial diagnostics and alternative databases. The library will facilitate sequence-based drug-susceptibility testing.
Pooideae grasses contain some of the world's most important crop and forage species. Although much work has been conducted on understanding the genetic basis of trait diversification within a few ...annual Pooideae, comparative studies at the subfamily level are limited by a lack of perennial models outside 'core' Pooideae. We argue for development of the perennial non-core genus Melica as an additional model for Pooideae, and provide foundational data regarding the group's biogeography and history of character evolution.
Supplementing available ITS and ndhF sequence data, we built a preliminary Bayesian-based Melica phylogeny, and used it to understand how the genus has diversified in relation to geography, climate and trait variation surveyed from various floras. We also determine biomass accumulation under controlled conditions for Melica species collected across different latitudes and compare inflorescence development across two taxa for which whole genome data are forthcoming.
Our phylogenetic analyses reveal three strongly supported geographically structured Melica clades that are distinct from previously hypothesized subtribes. Despite less geographical affinity between clades, the two sister 'Ciliata' and 'Imperfecta' clades segregate from the more phylogenetically distant 'Nutans' clade in thermal climate variables and precipitation seasonality, with the 'Imperfecta' clade showing the highest levels of trait variation. Growth rates across Melica are positively correlated with latitude of origin. Variation in inflorescence morphology appears to be explained largely through differences in secondary branch distance, phyllotaxy and number of spikelets per secondary branch.
The data presented here and in previous studies suggest that Melica possesses many of the necessary features to be developed as an additional model for Pooideae grasses, including a relatively fast generation time, perenniality, and interesting variation in physiology and morphology. The next step will be to generate a genome-based phylogeny and transformation tools for functional analyses.
Malaria cases caused by the zoonotic parasite Plasmodium knowlesi are being increasingly reported throughout Southeast Asia and in travelers returning from the region. To test for evidence of ...signatures of selection or unusual population structure in this parasite, we surveyed genome sequence diversity in 48 clinical isolates recently sampled from Malaysian Borneo and in five lines maintained in laboratory rhesus macaques after isolation in the 1960s from Peninsular Malaysia and the Philippines. Overall genomewide nucleotide diversity (π = 6.03 × 10(-3)) was much higher than has been seen in worldwide samples of either of the major endemic malaria parasite species Plasmodium falciparum and Plasmodium vivax. A remarkable substructure is revealed within P. knowlesi, consisting of two major sympatric clusters of the clinical isolates and a third cluster comprising the laboratory isolates. There was deep differentiation between the two clusters of clinical isolates mean genomewide fixation index (FST) = 0.21, with 9,293 SNPs having fixed differences of FST = 1.0. This differentiation showed marked heterogeneity across the genome, with mean FST values of different chromosomes ranging from 0.08 to 0.34 and with further significant variation across regions within several chromosomes. Analysis of the largest cluster (cluster 1, 38 isolates) indicated long-term population growth, with negatively skewed allele frequency distributions (genomewide average Tajima's D = -1.35). Against this background there was evidence of balancing selection on particular genes, including the circumsporozoite protein (csp) gene, which had the top Tajima's D value (1.57), and scans of haplotype homozygosity implicate several genomic regions as being under recent positive selection.
Malaria is a major public health problem that is actively being addressed in a global eradication campaign. Increased population mobility through international air travel has elevated the risk of ...re-introducing parasites to elimination areas and dispersing drug-resistant parasites to new regions. A simple genetic marker that quickly and accurately identifies the geographic origin of infections would be a valuable public health tool for locating the source of imported outbreaks. Here we analyse the mitochondrion and apicoplast genomes of 711 Plasmodium falciparum isolates from 14 countries, and find evidence that they are non-recombining and co-inherited. The high degree of linkage produces a panel of relatively few single-nucleotide polymorphisms (SNPs) that is geographically informative. We design a 23-SNP barcode that is highly predictive (~92%) and easily adapted to aid case management in the field and survey parasite migration worldwide.
Correct vascular differentiation requires distinct patterns of gene expression in different subtypes of endothelial cells. Members of the ETS transcription factor family are essential for the ...transcriptional activation of arterial and angiogenesis-specific gene regulatory elements, leading to the hypothesis that they play lineage-defining roles in arterial and angiogenic differentiation directly downstream of VEGFA signalling. However, an alternative explanation is that ETS binding at enhancers and promoters is a general requirement for activation of many endothelial genes regardless of expression pattern, with subtype-specificity provided by additional factors. Here we use analysis of Ephb4 and Coup-TFII (Nr2f2) vein-specific enhancers to demonstrate that ETS factors are equally essential for vein, arterial and angiogenic-specific enhancer activity patterns. Further, we show that ETS factor binding at these vein-specific enhancers is enriched by VEGFA signalling, similar to that seen at arterial and angiogenic enhancers. However, while arterial and angiogenic enhancers can be activated by VEGFA in vivo, the Ephb4 and Coup-TFII venous enhancers are not, suggesting that the specificity of VEGFA-induced arterial and angiogenic enhancer activity occurs via non-ETS transcription factors. These results support a model in which ETS factors are not the primary regulators of specific patterns of gene expression in different endothelial subtypes.
•Vein-specific enhancers can contain essential ETS motifs.•VEGFA induced an increase in ETS binding at vein, arterial and angiogenic enhancers.•VEGFA stimulation cannot induce vein-specific enhancer activity.
Spoligotyping is a well-established genotyping technique based on the presence of unique DNA sequences in Mycobacterium tuberculosis (Mtb), the causal agent of tuberculosis disease (TB). Although ...advances in sequencing technologies are leading to whole-genome bacterial characterization, tens of thousands of isolates have been spoligotyped, giving a global view of Mtb strain diversity. To bridge the gap, we have developed SpolPred, a software to predict the spoligotype from raw sequence reads. Our approach is compared with experimentally and de novo assembly determined strain types in a set of 44 Mtb isolates. In silico and experimental results are identical for almost all isolates (39/44). However, SpolPred detected five experimentally false spoligotypes and was more accurate and faster than the assembling strategy. Application of SpolPred to an additional seven isolates with no laboratory data led to types that clustered with identical experimental types in a phylogenetic analysis using single-nucleotide polymorphisms. Our results demonstrate the usefulness of the tool and its role in revealing experimental limitations.
SpolPred is written in C and is available from www.pathogenseq.org/spolpred.
francesc.coll@lshtm.ac.uk.
Supplementary data are available at Bioinformatics Online.
Whole-genome sequencing technologies are being increasingly applied to Plasmodium falciparum clinical isolates to identify genetic determinants of malaria pathogenesis. However, genome-wide discovery ...methods, such as haplotype scans for signatures of natural selection, are hindered by missing genotypes in sequence data. Poor correlation between single nucleotide polymorphisms (SNPs) in the P. falciparum genome complicates efforts to apply established missing-genotype imputation methods that leverage off patterns of linkage disequilibrium (LD). The accuracy of state-of-the-art, LD-based imputation methods (IMPUTE, Beagle) was assessed by measuring allelic r2 for 459 P. falciparum samples from malaria patients in 4 countries: Thailand, Cambodia, Gambia, and Malawi. In restricting our analysis to 86 k high-quality SNPs across the populations, we found that the complete-case analysis was restricted to 21k SNPs (24.5%), despite no single SNP having more than 10% missing genotypes. The accuracy of Beagle in filling in missing genotypes was consistently high across all populations (allelic r2, 0.87-0.96), but the performance of IMPUTE was mixed (allelic r2, 0.34-0.99) depending on reference haplotypes and population. Positive selection analysis using Beagle-imputed haplotypes identified loci involved in resistance to chloroquine (crt) in Thailand, Cambodia, and Gambia, sulfadoxine-pyrimethamine (dhfr, dhps) in Cambodia, and artemisinin (kelch13) in Cambodia. Tajima's D-based analysis identified genes under balancing selection that encode well-characterized vaccine candidates: apical merozoite antigen 1 (ama1) and merozoite surface protein 1 (msp1). In contrast, the complete-case analysis failed to identify any well-validated drug resistance or candidate vaccine loci, except kelch13. In a setting of low LD and modest levels of missing genotypes, using Beagle to impute P. falciparum genotypes is a viable strategy for conducting accurate large-scale population genetics and association analyses, and supporting global surveillance for drug resistance markers and candidate vaccine antigens.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Individuals living in endemic areas generally harbour multiple parasite strains. Multiplicity of infection (MOI) can be an indicator of immune status and transmission intensity. It has a potentially ...confounding effect on a number of population genetic analyses, which often assume isolates are clonal. Polymerase chain reaction-based approaches to estimate MOI can lack sensitivity. For example, in the human malaria parasite Plasmodium falciparum, genotyping of the merozoite surface protein (MSP1/2) genes is a standard method for assessing MOI, despite the apparent problem of underestimation. The availability of deep coverage data from massively parallizable sequencing technologies means that MOI can be detected genome wide by considering the abundance of heterozygous genotypes. Here, we present a method to estimate MOI, which considers unique combinations of polymorphisms from sequence reads. The method is implemented within the estMOI software. When applied to clinical P.falciparum isolates from three continents, we find that multiple infections are common, especially in regions with high transmission.
Availability and implementation:
estMOI is freely available from http://pathogenseq.lshtm.ac.uk.
Contact:
samuel.assefa@lshtm.ac.uk
Supplementary information:
Supplementary data are available at Bioinformatics online.
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