Biology subjects

Evans, R.

Publications and source records attributed to Evans, R..

At least 19 recordsLinked to original sources

bioRxiv2026

CA19-9 induces microenvironment remodeling in pancreatic ductal adenocarcinoma

Durable therapeutic efficacy remains a major barrier to improving outcomes for patients with pancreatic ductal adenocarcinoma (PDAC). An immunosuppressive tumor microenvironment (TME) is a hallmark of PDAC and has been demonstrated to be a dominant driver of therapeutic resistance. The aberrant glycan CA19-9 is prevalent in PDAC and drives tumor progression, but the paracrine mechanisms by which it contributes to TME remodeling are unknown. To address this, we mapped TME changes and performed functional analyses using a genetically engineered mouse model (GEMM) harboring KrasG12D mutation and inducible CA19-9 expression. Elevation of CA19-9 led to expansion of antigen-presenting cancer associated fibroblasts (apCAFs) and regulatory T cells (Tregs), which can drive immunosuppression. Antibody blockade of CA19 -9 resulted in significant restoration of normal histology and decreased apCAF and Treg populations. We dissected the paracrine signaling mechanisms that drive this TME remodeling in vitro using mouse and human organoid mono- and co-culture models as well as in vivo using GEMMs and syngeneic orthotopic transplantation models. CA19-9 induced IL1a and TGFb expression, reprogramming pancreatic mesothelial cells into apCAFs in vitro, which in turn directly ligated naive Cd4+ T cells resulting in Treg differentiation in co-cultures. Antibody blockade of IL1a and TGFb in mice led to reduced apCAF and Treg differentiation. We previously reported that CA19-9 modification of the secreted Fbln3 protein increased Egfr engagement and now find that the induction of IL1a and TGFb expression by CA19-9 is dependent on Fbln3 hyperactivation of EGFR signaling. Genetic depletion of Fbln3 led to reduced tumor progression and increased Cd8+ T cell infiltration in mice. Together these findings identify a previously unknown signaling axis driving immunosuppressive phenotypes in PDAC, uncovering multiple potential nodes to relieve the immunosuppressive pressures within the PDAC TME.

cancer biology↗

bioRxiv2026

A systematic comparison of tools for predicting antimicrobial resistance from nanopore sequence data

Antimicrobial resistance (AMR) presents a pressing need to ensure that the right antimicrobials are used to target the right microbes at the right time. Ideally, the appropriate antimicrobial is selected after patient samples have been cultured and assessed with antimicrobial sensitivity testing (AST). However, the time needed for culture-based diagnosis leads to immediate empirical treatment, often with broad-spectrum and/or high-tier antimicrobials. Direct nanopore metagenomic whole genome sequencing to identify pathogens and predict their antimicrobial resistance is a rapid and patient-side alternative. A limitation of this approach is potential inconsistencies in in silico predicted AMR phenotypes. Here, we benchmarked the current performance of in silico AMR prediction strategies for nanopore-generated long read data. Using nanopore data paired with AST phenotyping for 201 samples representing 27 bacterial species, we assessed the impact of basecalling mode, data volume, and assembly strategy, and compared the performance of eight in silico AMR prediction tools with seven AMR databases. We found that basecalling accuracy mode does not significantly affect the overall accuracy of in silico AMR predictions, but assembly strategy and data volume both do. Prediction tools using the ResFinder database scored best for balanced accuracy (0.80 {+/-} 0.02 for both ResFinder and ABRicate), whilst DeepARG scored best for sensitivity (0.65 {+/-} 0.03); predictions were more accurate for some antibiotic classes and genera than others. However, even the best performing in silico AMR prediction strategy missed some resistance identified by lab-based AST. We conclude therefore that, currently, in silico AMR prediction can supplement lab-based AST, but cannot yet replace it.

microbiology↗

bioRxiv2026

Ruminosignatures associated with methane emissions and feed efficiency across geographies and cattle breeds

The cattle rumen microbiota represents a highly complex and dynamic ecosystem, whose organization and connection to host phenotypes are of the highest importance to food security and the environment. In this study, we analyzed the rumen microbiota, from 2,492 cattle belonging to five different breeds and production systems across five countries, categorizing them into microbial co-abundance groups referred to as Ruminosignatures. We identified twelve distinct Ruminosignatures, including two that were consistently observed across all populations and were dominated by the genus Prevotella and UBA2810. Additional Ruminosignatures showed breed-and diet-specific patterns and collectively explained 96-99% of the variance in rumen microbial composition. The abundances of several Ruminosignatures were associated with methane emissions and feed efficiency, and were influenced by host genetics, with heritability estimates ranging from 0.09 to 0.51. The Ruminosignature dominated by UAB2810 was negatively associated with methane emissions across all datasets and positively linked to feed efficiency in Holstein from Italy and crossbred from Ireland. Additionally, the type of production system affects both the occurrence of Ruminosignatures and their impact on host phenotypes, emphasizing the need for context-specific approaches to modulate the rumen microbiome. Overall, our results offer new perspectives on the assembly of ruminal microbes and underscore the potential of the Ruminosignatures framework for microbiome-informed precision agriculture and breeding initiatives aimed at enhancing feed efficiency and minimizing the environmental impact of cattle farming.

microbiology↗

bioRxiv2026

The Demographic and GDP Impacts of Slowing Biological Aging

Biological aging imposes significant socio-economic costs, increasing health expenses, reducing productivity, stalling population growth and straining social systems, culminating in reduced economic activity. We draw insights from interviews with 102 scientists working on aging biology and develop four macroeconomic simulations: slowing brain aging, slowing reproductive aging, and an overall delay in biological aging (including the novel concept of "replacing aging"). Our model is calibrated to represent how slowing biological aging manifests in the US economy and population through the channels of mortality, fertility, and productivity rates by age. We simulate the economic and demographic impacts of near-future advancements in aging science. We find that a one-year delay in brain aging alone could add $201 billion annually to US GDP. A one-year delay in overall biological aging could boost GDP by $408 billion annually, yielding $27.1 trillion in net present value in the long run.

scientific communication and education↗

bioRxiv2025

Beyond species means - the intraspecific contribution to global wood density variation

Wood density is central for estimating vegetation carbon storage and a plant functional trait of great ecological and evolutionary importance. However, the global extent of wood density variation is unclear, especially at the intraspecific level. We assembled the most comprehensive wood density collection to date (GWDD v.2), including 109,626 records from 16,829 plant species across woody life forms and biomes. Using the GWDD v.2, we explored the sources of variation in wood density within individuals, within species, and across environmental gradients. Intraspecific variation accounted for up to 15% of overall wood density variation (sd = 0.068 g cm-3). Sapwood densities varied 50% less than heartwood densities, and branchwood densities varied 30% less than trunkwood densities. Individuals in extreme environments (dry, hot, acidic soils) had higher wood density than conspecifics elsewhere (+0.02 g cm-3, [~]4% of the mean). Intraspecific environmental effects strongly tracked interspecific patterns (r = 0.83) but were only 20-30% as large and varied considerably among taxa. Individual plant wood density was difficult to predict (RMSE > 0.08 g cm-3; single-measurement R2 = 0.59). We recommend (i) systematic within-species sampling for local applications, and (ii) expanded taxonomic coverage combined with integrative models for robust estimates across ecological scales.

plant biology↗

bioRxiv2025

A global map of wood density

Wood density influences how quickly woody plants grow, how long they live and how much carbon they store, yet its global variation remains poorly mapped. Here we combined 109,626 wood density measurements from 16,829 species with 300,949 vegetation plots to produce a km-scale map of community-weighted wood density for every woody biome. Our model led to a prediction accuracy 32-51 % higher than previous global products, and a 1.8-3.7-fold wider wood density range (0.28-1.00 g cm-3; global mean: 0.57 g cm-3) than previously assumed. Spatial cross-validation showed low bias ({+/-}2.5 % of the mean), and uncertainties decreased from 20% in poorly sampled drylands and boreal regions to 5% in data-rich temperate forests. Mean annual temperature was the best predictor of community-weighted mean wood density, increasing by 0.01 g cm-3 for every 1{degrees}C change. We deliver a low-bias, high-resolution wood density layer for Earth system models, together with spatially explicit error maps. This study represents a major step forward for carbon accounting and trait-based forecasts of vegetation change.

plant biology↗

bioRxiv2024

Platelet-specific P2Y1 receptor deficient mice have suppressed leukocyte recruitment in response to lipopolysaccharide.

A role for the P2Y1 receptor in inflammation has been established using a pharmacological approach over an acute 4 hour time span. However, nucleotide-structure P2Y1 receptor antagonists have limited experimental use due to inadequate pharmacokinetics, and inability to decipher global versus cell specific effects in vivo. The creation of a conditional knock out (platelet) P2Y1 transgenic mouse model was designed to overcome these restrictions. A homozygous P2Y1 LoxP mouse colony was created using CRISPR/Cas9 technology, and crossed with a hemizygous P2Y1 LoxP with PF4-cre to provide offspring that are homozygous for P2Y1 LoxP flanked allele, and hemizygous for the PF4cre (platelet P2Y1-/-) and offspring homozygous for P2Y1 LoxP flanked allele, but non-carriers for PF4cre (control mice). Animals were intranasally administered LPS to induce pulmonary inflammation to assess the influence of phenotype on leukocyte recruitment. 24 hours post intranasal LPS administration; pulmonary neutrophil and platelet recruitment were significantly suppressed, despite the fact that neutrophils retained the ability to migrate to fMLP ex vivo. Circulating platelet and leukocyte numbers were not different between control and platelet P2Y1-/- animals. Tail bleeding times revealed the platelet P2Y1-/- mice had a severe bleeding phenotype. This is the first demonstration of a platelet specific P2Y1-/- mouse model to confirm the importance of platelet P2Y1 receptors in the regulation of inflammatory responses with a 60-70% inhibition of leukocyte recruitment over an extended time period compared to previous pharmacological studies. platelet P2Y1-/- mice will help further elucidate the mechanisms by which P2Y1 receptors regulate platelet activation during inflammation. Key PointsO_LIMice selectively deficient in the platelet P2Y1 have suppressed leukocyte and platelet recruitment during inflammation. C_LIO_LIWe provide a methodology to determine mechanistic relevance that is otherwise limited by pharmacological approaches. C_LI

pharmacology and toxicology↗

bioRxiv2024

Identification of molecular and functional subtypes using chronic pancreatitis patient-derived organoid models

Chronic pancreatitis (CP) affects [~]3 million people worldwide, yet altering the course of disease is challenging. We developed a patient-derived organoid (PDO) platform to investigate the molecular pathogenesis of this disease and identify therapeutic strategies. We generated 36 PDOs from patients with idiopathic, hereditary, and alcohol-related CP with high genetic concordance. PDOs retained inflammation-associated transcriptional and proteomic features. Transcriptomic profiling revealed three molecular subtypes of CP independent of etiology. We discovered widespread dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) in half of the CP PDOs, including those with wildtype CFTR. Clinically available CFTR modulators stabilized mutant or wildtype CFTR, restored CFTR function, and decreased mitogenic and inflammatory signaling. This work provides the first comprehensive PDO platform for modeling CP. We demonstrate the utility of this platform for precision therapeutic investigations. Our findings reveal CFTR modulators as a broadly applicable and effective therapeutic strategy.

cell biology↗

bioRxiv2023

The Pvc15 D2-Pnf SP Interaction Mediates AAA ATPase Activity, Payload Stability, and Translocation into the PVC Tube Lumen.

The Photorhabdus virulence cassette (PVC) is an elegant, multi-protein, contractile nanostructure which injects functionally bioactive polypeptides into the eukaryotic cytosol. Signal peptides (SPs) are N-terminal amino acid motifs from native payload proteins which can also associate wide range of heterologous proteins to the PVC tube lumen. In addition, Pvc15 is a classic AAA ATPase encoded within the PVC operon which encodes an N-domain and tandem AAA domains D1 and D2. This work finds that Pvc15s ATPase activity is mediated by the E555 residue situated in the Walker B motif of the sole functional ATPase domain, D2. Pvc15 multimerisation may be required to translocate payload into the PVC tube lumen, which is facilitated by intact N and D1 domains. In addition, ATPase activity requires the presence of other PVC operon components which hints at a PVC-regulated mode of action for loading. Furthermore, D2 is a chaperone for the native Pnf SP, though C-terminal truncations of the Pnf SP confers Pvc15-independent stability to the payload. These findings provide insight into the nuanced roles of the Pvc15-SP interaction for payload-PVC association and the effects that SP length and type has on payload stability and PVC-loading ability.

microbiology↗

bioRxiv2023

Chemotherapeutic regulation of the ROS/MondoA-dependent TXNIP/GDF15 axis; and derivation of a new organoid metric as a predictive biomarker

Chemotherapy, the standard of care treatment for cancer patients with advanced disease, has been increasingly recognised to activate host immune responses to produce durable outcomes. Here, in colorectal adenocarcinoma (CRC) we identify chemotherapy-induced Thioredoxin Interacting Protein (TXNIP), a MondoA-dependent tumor suppressor gene, as a negative regulator of Growth/Differentiation Factor 15 (GDF15). GDF15 is a negative prognostic factor in CRC and promotes the differentiation of regulatory T cells (Tregs), through CD48 ligation. Intriguingly, multiple models including patient-derived tumor organoids demonstrate that loss of TXNIP/GDF15 axis functionality is associated with advanced disease or chemotherapeutic resistance, with transcriptomic or proteomic GDF15/TXNIP ratios showing potential as a prognostic biomarker. These findings illustrate a potentially common pathway where chemotherapy-induced epithelial stress drives local immune remodelling for patient benefit, with disruption of this pathway seen in refractory or advanced cases.

cancer biology↗

bioRxiv2023

Accelerated computations for iterative-solver techniques in single-step BLUP models

In the last decade, a number of methods have been suggested to deal with large amounts of genetic data in genomic predictions. Yet, steadily growing population sizes and the suboptimal use of computational resources are pushing the practical application of these approaches to their limits. As an extension to the C/CUDA library miraculix, we have developed tailored solutions for the computation of genotype matrix multiplications which is a critical bottleneck in the empirical evaluation of many statistical models. We demonstrate the benefits of our solutions at the example of single-step models which make repeated use of this kind of multiplication. Targeting modern Nvidia(R) GPUs as well as a broad range of CPU architectures, our implementation significantly reduces the time required for the estimation of breeding values in large population sizes. miraculix is released under the Apache 2.0 license and is freely available at https://github.com/alexfreudenberg/miraculix.

bioinformatics↗

bioRxiv2023

Altered neutrophil extracellular traps in response to Mycobacterium tuberculosis in persons living with HIV with no previous TB and negative TST and IGRA

Persons living with HIV (PLWH) have an increased risk for tuberculosis (TB). After prolonged and repeated exposure, some PLWH never develop TB and test persistently negative in tests of immune sensitization tuberculin skin test (TST) and interferon gamma release assays (IGRA) for Mycobacterium tuberculosis (Mtb). This group has been identified and defined as HIV+ persistently TB, tuberculin and IGRA negative (HITTIN). To investigate potential innate mechanisms unique to individuals with the HITTIN phenotype we compared their neutrophil Mtb infection response to that of PLWH, with no TB history, but who test persistently IGRA positive, and tuberculin positive (HIT). Neutrophil samples from 17 HITTIN (PMNHITTIN) and 11 HIT (PMNHIT) were isolated and infected with Mtb H37Rv for 1h and 6h. RNA was extracted and used for RNAseq analysis. At 1h of Mtb infection, PMNHITTIN displayed 151 significantly upregulated and 40 significantly downregulated differentially expressed genes (DEGs) and PMNHIT 98 significantly upregulated and 11 significantly downregulated DEGs. At the 6h timepoint, PMNHITTIN displayed 3106 significantly upregulated and 3548 significantly downregulated DEGs while PMNHIT had 3816 significantly up- and 3794 significantly downregulated DEGs. There was no significant differential transcriptional response at 1h between infected PMNHITTIN and PMNHIT. However, when contrasting the log2FC 6h infection response to Mtb from PMNHITTIN against PMNHIT, 2285 genes showed significant differential response between the two groups. Apoptosis and NETosis were key pathways linked to the enrichment of genes in PMNHITTIN when contrasted to PMNHIT after 6h infection with Mtb. Fluorescence microscopy revealed relatively lower neutrophil extracellular trap formation and cell loss in PMNHITTIN compared to PMNHIT, showing that PMNHITTIN have a distinct response to Mtb.

genetics↗

bioRxiv2023

A hybrid and poly-polish workflow for the complete and accurate assembly of phage genomes: a case study of ten przondoviruses

Bacteriophages (phages) within the Przondovirus genus are T7-like podoviruses belonging to the Studiervirinae subfamily, within the Autographiviridae family and have a highly conserved genome organisation. The genome size of these phages ranges from 37 kb to 42 kb, encode 50-60 genes and are characterised by the presence of direct terminal repeats (DTRs) flanking the linear chromosome. These DTRs are often deleted during short-read-only and hybrid assemblies. Moreover, long-read-only assemblies are often littered with sequencing and/or assembly errors and require additional curation. Here, we present the isolation and characterisation of ten novel przondoviruses targeting Klebsiella spp. We describe HYPPA - a HYbrid and Poly-polish Phage Assembly workflow, which utilises long-read assemblies in combination with short-read sequencing to resolve phage DTRs and correcting errors, negating the need for laborious primer walking and Sanger sequencing validation. Our data demonstrate the importance of careful curation of phage assemblies before publication, and prior to using them for comparative genomics. IMPACT STATEMENTThe current workflows employed for phage genome assembly are often error-prone and can lead to many incomplete phage genomes being deposited within databases. This can create challenges when performing comparative genomics, and may also lead to incorrect taxonomic assignment. To overcome these challenges we proposed HYPPA, a workflow that can produce complete and high-quality phage genomes without the need for laborious lab-based validation. DATA SUMMARYPhage raw reads are available from the National Centre for Biotechnology Information Sequence Read Archive (NCBI-SRA) under the BioProject number PRJNA914245. Phage annotated genomes have been deposited at GenBank under the accessions OQ579023-OQ579032 (Table 1). Bacterial WGS data for clinical preterm infant samples have been deposited at GenBank under BioProject accession PRJNA471164 (Table S1). Bacterial raw reads for food samples are available from NCBI-SRA with individual accessions (SAMN33593347-SAMN33593351), and can be found under the BioProject number PRJNA941224 (Table S1). Strain-specific details for bacteria and publicly-available phages used in these analyses, along with accessions for the latter can be found in Table S1 and Table S6, respectively. The CL1-CL8 clinical Klebsiella strains (Table S1) were under a Materials Transfer Agreement, for which sequencing data and strain information is not available. O_TBL View this table: org.highwire.dtl.DTLVardef@8cae53org.highwire.dtl.DTLVardef@8e512org.highwire.dtl.DTLVardef@1f71a67org.highwire.dtl.DTLVardef@1e6947dorg.highwire.dtl.DTLVardef@11ebc58_HPS_FORMAT_FIGEXP M_TBL O_FLOATNOTable 1.C_FLOATNO O_TABLECAPTIONPrzondoviruses within the collection to date and data relating to the closest database relative. C_TABLECAPTION C_TBL

microbiology↗

bioRxiv2023

An efficient method for high molecular weight bacterial DNA extraction suitable for shotgun metagenomics from skin swabs

The human skin microbiome represents a variety of complex microbial ecosystems that play a key role in host health. Molecular methods to study these communities have been developed but have been largely limited to low-throughput quantification and short amplicon sequencing, providing limited functional information about the communities present. Shotgun metagenomic sequencing has emerged as a preferred method for microbiome studies as it provides more comprehensive information about the species/strains present in a niche and the genes they encode. However, the relatively low bacterial biomass of skin, in comparison to other areas such as the gut microbiome, makes obtaining sufficient DNA for shotgun metagenomic sequencing challenging. Here we describe an optimised high-throughput method for extraction of high molecular weight DNA suitable for shotgun metagenomic sequencing. We validated the performance of the extraction method, and analysis pipeline on skin swabs collected from both adults and babies. The pipeline effectively characterised the bacterial skin microbiota with a cost and throughput suitable for larger longitudinal sets of samples. Application of this method will allow greater insights into community compositions and functional capabilities of the skin microbiome. Impact StatementDetermining the functional capabilities of microbial communities within different human microbiomes is important to understand their impacts on health. Extraction of sufficient DNA is challenging, especially from low biomass samples, such as skin swabs suitable for shotgun metagenomics, which is needed for taxonomic resolution and functional information. Here we describe an optimised DNA extraction method that produces enough DNA from skin swabs, suitable for shotgun metagenomics, and demonstrate it can be used to effectively characterise the skin microbiota. This method will allow future studies to identify taxonomic and functional changes in the skin microbiota which is needed to develop interventions to improve and maintain skin health. Data SummaryAll sequence data and codes can be accessed at: NCBI Bio Project ID: PRJNA937622 DOI: https://github.com/quadram-institute-bioscience/coronahit_guppy DOI: https://github.com/ilianaserghiou/Serghiou-et-al.-2023-Codes

genomics↗

bioRxiv2022

Discovery and molecular basis of chloride as an allosteric activator and catalytic inhibitor for Class-D lactamases.

Oxacillinase (OXA)-48-like carbapenemases are epidemic class D {beta}-lactamases in Enterobacterales, resulting in high mortality. Though the chemical mechanism has been clearly established, for decades, the link between the biphasic kinetic behaviour of these enzymes, which significantly impacts antibiotic efficacy, and the state of carbamylated lysine has been elusive. Here, substituting N-carbamylated lysine73 with a chemically-stable N-acetyl lysine allows us to prove the origin of catalytic inhibition is not decarbamylation and enables us to capture an unprecedented inactive acyl-intermediate wedged in place by a chloride ion against the conserved residue arginine250. We here identify chloride as a "Janus effector" acting by allosteric activation of the burst phase and inhibition of the steady-state for a series of {beta}-lactam substrates in kinetic assays. Chloride ions are necessarily present in both laboratory and clinical OXA activity assays and their inseparable role is now identified. Our finding suggests a new direction for the discovery of next-generation antibiotics specific for {beta}-lactamases of Class D.

biochemistry↗

bioRxiv2022

A Synthetic ERRα Agonist Induces an Acute AerobicExercise Response and Enhances Exercise Capacity

Repetitive physical exercise induces physiological adaptations in skeletal muscle that improves exercise performance and is effective for the prevention and treatment of several diseases. Here we report the identification of a synthetic agonist for the orphan nuclear receptor ERR (estrogen receptor-related receptor ), SLU-PP-332, that activates an acute aerobic exercise genetic program in skeletal muscle in an ERR-dependent manner. SLU-PP-332 increases mitochondrial function and cellular respiration consistent with induction of this genetic program. When administered to mice, SLU-PP-332 increased the type IIa oxidative skeletal muscle fibers and enhanced exercise endurance. These data indicate the feasibility of targeting ERR for development of compounds that act as exercise mimetics that may be effective in treatment of numerous metabolic disorders and to improve muscle function in the aging.

pharmacology and toxicology↗

bioRxiv2022

Neutrophils infiltrate sensory ganglia and mediate chronic widespread pain in fibromyalgia

Fibromyalgia is a debilitating widespread chronic pain syndrome that occurs in 2-4% of the population. The prevailing view that fibromyalgia results from central nervous system dysfunction has recently been challenged with data showing changes in peripheral nervous system activity. Using a mouse model of chronic widespread pain through hyperalgesic priming of muscle, we show that neutrophils invade sensory ganglia and confer mechanical hypersensitivity on recipient mice, whilst adoptive transfer of immunoglobulin, serum, lymphocytes or monocytes have no effect on pain behaviour. Neutrophil depletion abolishes the establishment of chronic widespread pain in mice. Neutrophils from patients with fibromyalgia also confer pain on mice. A link between neutrophil derived mediators and peripheral nerve sensitisation is already established. These observations suggest new approaches for targeting fibromyalgia pain through an understanding of the mechanisms that cause altered neutrophil activity and interactions with sensory neurons. Significance statementWe used a back-translational model in mice to demonstrate the pro-nociceptive role of neutrophils in fibromyalgia. Adoptive transfer of neutrophils from mice with chronic widespread pain or from patients with fibromyalgia can confer mechanical pain to recipient naive mice, sensitise evoked action potential firing of spinal cord neurons and produce phenotypic changes in cell surface expression of neutrophil proteins that cause infiltration of neutrophils into dorsal root ganglia. These data provide the framework for an immunological basis of chronic widespread pain in fibromyalgia mediated by polymorphonuclear granulocytes.

neuroscience↗

bioRxiv2021

Genetic insights into the management and conservation of Arctic charr in North Wales

Aquatic species throughout the world are threatened by extinction in many parts of their range, particularly in their most southerly distributions. Arctic charr (Salvelinus alpinus) is a Holarctic species with a distribution that includes the glacial lakes of North Wales, towards it southern limit. To date, no genetic studies have been conducted to determine the genetic health of the three remaining native Arctic charr populations in North Wales, despite exposure to stocking and adverse environmental and ecological conditions. We used seven microsatellite loci to determine whether: 1) genetic differentiation existed between native populations; 2) translocated populations from Llyn Peris were genetically similar to the historically connected Llyn Padarn population; and 3) hatchery supplementation negatively impacted genetic diversity in Llyn Padarn. All three native populations retained their genetic integrity, with Llyn Bodlyn showing high levels of divergence (FST = 0.26 {+/-} 0.02SD) as well as low genetic diversity (HO 0.30) compared to remaining populations (HO 0.64 {+/-} 0.14SD). Although evidence suggests that stocking increased the effective population size of Llyn Padarn in the short term without impacting genetic diversity, the long term effects of such practices are yet to be seen. Results provide baseline data for conservation management, and highlight the need for protection of small isolated populations that are being negatively impacted by the processes of genetic drift due to escalating anthropogenic pressures. Continual monitoring of both Arctic charr and their habitats using a combination of methods will increase the likelihood that these threatened and iconic populations will persist in the future.

genetics↗