Biology subjects

Leigh, R. J.

Publications and source records attributed to Leigh, R. J..

5 recordsLinked to original sources

bioRxiv2024

Genomic analysis of antimicrobial resistant Escherichia coli isolated from manure and manured agricultural grasslands

Antimicrobial resistance (AMR) is a multifactorial issue involving an intertwining relationship between animals, humans and the environment. The environment can harbour bacteria that are pathogenic to human health, including Escherichia coli, an indicator of environmental faecal contamination. Through culture dependent approaches this study identified 46 E. coli isolates in porcine and bovine manure, non-manured and manured soil, and the phyllosphere of manured grass. The grass isolation highlights grass as an environmental reservoir for E. coli. Whole genome sequencing identified 11 different multi-locus sequence types. We also identified a diverse plasmidome with 23 different plasmid replicon types. The E. coli isolates were phenotypically antibiotic resistance, predominantly multidrug resistant. Additionally, whole genome sequencing identified 31 antibiotic resistance genes, and mutations in the gyrA, parC, and parE genes, conferring fluoroquinolone resistance. The main virulence genes were associated actin mediated locomotion (icsP/sopA), siderophore production and alginate production (algA), which suggest adaptation to survive in the non-gut environment or the UV environment of grass surfaces. These results suggest that E. coli in soils and grasses may adapt to their new environments evolving novel strategies. This study demonstrates grass as an understudied environmental niche of AMR E. coli, which directly links the environment to the grass grazing animal and vice-versa via the circular economy of manure application. Impact statementEscherichia coli is capable of surviving across biomes within One Health. This study sheds light on the genomic elements present in AMR E. coli in the understudied niche of agricultural grassland. Data summaryThe genome sequences have been deposited in Genbank. Bioproject number PRJNA1080214 and SRP491607 in the sequence read archive https://www.ncbi.nlm.nih.gov/sra/?term=SRP491607.

microbiology↗

bioRxiv2024

Plasmids from a complex biome exist as communities

Plasmids play a crucial role in the spread of antimicrobial resistance genes (ARGs) across One Health due to their ability to transfer a wide range of ARGs within and across bacterial species and biomes. We sequenced 173 circularised plasmids transferred from wastewater treatment plant (WWTP) effluent into Escherichia coli and subsequently characterised their genetic content. Multiple multidrug resistant plasmids were identified with a significant number of mega plasmids (>100Kb). Plasmids existing in isolation were rare and almost all existed with other plasmids. Our results suggest that positive epistasis promotes plasmid persistence in WWTP populations in a similar manner to that identified in vitro via infectious transmission, varying properties against plasmid community backgrounds, interactions with a range of other plasmids, source-sink spill-over transmission within the plasmid community rather than the host bacteria and compensatory mutations. We have demonstrated that the plasmid paradox solutions apply to plasmid communities in addition to plasmid host interactions. Our study identified that rather than existing as lone entities plasmids co-exist in small packs, the protection is afforded to the pack not by all members but by one or two and many plasmids coast within this pack as they contain no obvious advantage to the host. Our findings show that we need to enter a new paradigm and study plasmids in packs rather than as single entities in order to understand their transmission across One Health.

microbiology↗

bioRxiv2021

Comparative genomics and pangenomics of vancomycin resistant and susceptible Enterococcus faecium from Irish hospitals across 20 years

Enterococcus faecium has emerged as an important nosocomial pathogen, which is increasingly difficult to treat due to the genetic acquisition of vancomycin resistance. Ireland exhibits a recalcitrant vancomycin resistant bloodstream infection rate compared to other developed countries. A set of 28 vancomycin resistant isolates was sequenced to construct a dataset alongside 61 other publicly available Irish genomes. This dataset was extensively analysed using in-silico methodologies and uncovered distinct evolutionary, coevolutionary, and clinically relevant population trends. These results suggest that a stable (in terms of genome size, GC%, and number of genes), yet genetically diverse population (in terms of gene content) of Enterococcus faecium persist in Ireland with acquired resistance arising via plasmid acquisition (vanA) or to a lesser extent, chromosomal recombination (vanB). Population analysis described five clusters with one cluster partitioned into four clades which transcend isolation dates. Pangenomic and recombination analyses revealed an open (whole genome and chromosomal specific) pangenome illustrating a rampant evolutionary pattern. Comparative resistomics and virulomics uncovered distinct chromosomal and mobilomal propensity for multidrug resistance, widespread chromosomal point-mutation mediated resistance, and chromosomal harboured arsenals of virulence factors. Comparative phagomics revealed a core prophagome of three prophages throughout the dataset. Interestingly, a potential difference in biofilm formation strategies was highlighted by coevolutionary analysis, suggesting differential biofilm genotypes between vanA and vanB isolates. These results highlight the evolutionary history of Irish Enterococcus faecium isolates and may provide an insight into underlying infection dynamics in a clinical setting.

microbiology↗

bioRxiv2021

statSuma: automated selection and performance of statistical comparisons for microbiome studies

There is a reproducibility crisis in scientific studies. Some of these crises arise from incorrect application of statistical tests to data that follow inappropriate distributions, have inconsistent equivariance, or have very small sample sizes. As determining which test is most appropriate for all data in a multicategorical study (such as comparing taxa between sites in microbiome studies), we present statsSuma, an interactive Python notebook (which can be run from any desktop computer using the Google Colaboratory web service) and does not require a user to have any programming experience. This software assesses underlying data structures in a given dataset to advise what pairwise or listwise statistical procedure would be best suited for all data. As some users may be interested in further mining specific trends, statSuma performs 5 different two-tailed pairwise tests (Students t-test, Welchs t-test, Mann-Whitney U-test, Brunner-Munzel test, and a pairwise Kruskal-Wallis H-test) and advises the best test for each comparison. This software also advises whether ANOVA or a multicategorical Kruskal-Wallis H-test is most appropriate for a given dataset and performs both procedures. A data distribution-vs-Gaussian distribution plot is produced for each taxon at each site and a variance plot between all combinations of 2 taxa at each site are produced so Gaussian tests and variance tests can be visually confirmed alongside associated statistical determinants.

microbiology↗

bioRxiv2021

uniForest: an unsupervised machine learning technique to detect outliers and restrict variance in microbiome studies

Isolation Forests is an unsupervised machine learning technique for detecting outliers in continuous datasets that does not require an underlying equivariant or Gaussian distribution and is suitable for use on small datasets. While this procedure is widely used across quantitative fields, to our knowledge, this is the first attempt to solely assess its use for microbiome datasets. Here we present uniForest, an interactive Python notebook (which can be run from any desktop computer using the Google Colaboratory web service) for the processing of microbiome outliers. We used uniForest to apply Isolation Forests to the Healthy Human Microbiome project dataset and imputed outliers with the mean of the remaining inliers to maintain sample size and assessed its prowess in variance reduction in both community structure and derived ecological statistics (-diversity). We also assessed its functionality in anatomical site differentiation (pre- and postprocessing) using principal component analysis, dissimilarity matrices, and ANOSIM. We observed a minimum variance reduction of 81.17% across the entire dataset and in alpha diversity at the Phylum level. Application of Isolation Forests also separated the dataset to an extremely high specificity, reducing variance within taxa samples by a minimum of 81.33%. It is evident that Isolation Forests are a potent tool in restricting the effect of variance in microbiome analysis and has potential for broad application in studies where high levels of microbiome variance is expected. This software allows for clean analyses of otherwise noisy datasets.

microbiology↗