Journal of Applied Microbiology最新文献

{"title":"Global diffusion of IncC plasmid harboring blaNDM-1 in the high-risk Escherichia coli ST131 clone.","authors":"Khouloud Bouhalouene, Anis Raddaoui, Yosra Chebbi, Wafa Achour","doi":"10.1093/jambio/lxag229","DOIUrl":"10.1093/jambio/lxag229","url":null,"abstract":"<p><strong>Aims: </strong>The global expansion of quinolone-resistant Escherichia coli (QR-EC) is increasingly associated with β-lactam resistance and mobile genetic elements that facilitate resistance dissemination. This study investigated the molecular mechanisms underlying fluoroquinolone and β-lactam resistance in clinical QR-EC isolates and explored the plasmid type associated.</p><p><strong>Methods and results: </strong>A total of 123 non-duplicate QR-EC clinical isolates responsible mainly for gastrointestinal colonization were collected between 2019 and 2021. Plasmid-mediated quinolone resistance (PMQR), extended-spectrum β-lactamase (ESBL), and carbapenemase genes were screened by PCR. Mutations in the quinolone resistance-determining regions (QRDR) of gyrA and parC were analyzed using sequencing and mismatch amplification mutation assay PCR (MAMA-PCR). Selected isolates underwent multilocus sequence typing (MLST). Whole-genome sequencing (WGS) of a representative extensively drug-resistant strain carrying multiple quinolone resistance determinants, ESBL genes, and carbapenemase genes, was performed. PMQR genes were prevalent among QR-EC, dominated by aac(6')-Ib-cr (60.9% of isolates). ESBL genes were identified in 93.5% of isolates, predominantly blaCTX-M (95.7%). Among ertapenem-resistant isolates (QCR-EC) (n = 18), blaNDM-1 and blaOXA-48 were detected in 13 and 11 isolates, respectively. QRDR mutations were highly frequent, particularly gyrA83 (98.4%) and parC80 (30.9%). Major QCR-EC genotypes belonged to sequence types ST167 (n = 2), ST1196, ST469, and ST410. High-risk E. coli ST131 clone harboring IncC plasmid encoding blaNDM-1 was described for the first time in Africa, following its emergence, in two continents, Asia and America. Despite the very rare description of these strains worldwide, their description in three continents sign their global diffusion.</p><p><strong>Conclusions: </strong>This finding highlights the ongoing spread of carbapenem resistance and underscores the urgent need for strengthened genomic surveillance.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148919854","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A 16S rRNA reverse transcriptase-PCR method for rapid detection of intestinal enterococci in distributed drinking water.","authors":"Leo Heijnen, Eline Stroobach, Adrie Atsma, Gabi van Pelt, Hendrik Jan de Vries, Katrien De Maeyer, Bea Timmermans, Liesbeth Visser, Maja Taucer-Kapteijn, Gertjan Medema","doi":"10.1093/jambio/lxag234","DOIUrl":"https://doi.org/10.1093/jambio/lxag234","url":null,"abstract":"<p><strong>Aim: </strong>To develop and validate a real-time reverse transcription PCR (RT-PCR) method for the rapid and sensitive detection of intestinal enterococci in drinking water. Conventional culture methods require 24-48 h, limiting rapid response to faecal contamination events.</p><p><strong>Methods: </strong>An RT-PCR assay targeting 16S rRNA was developed to detect ten Enterococcus species, selected based on their prevalence in human and animal faeces and drinking water samples. Drinking water samples (100 ml) were filtered, treated with lysozyme, followed by RNA extraction and one-step real-time RT-PCR. Validation followed ISO16140-2:2016 with ISO7899-2:2000 as reference method and an interlaboratory study, using artificially contaminated samples and uncontaminated drinking water samples.</p><p><strong>Results: </strong>The RT-PCR method demonstrated a sensitivity of 94.6%, slightly higher than the reference culture method (89.2%). The combined RLOD value (0.851) indicated comparable detection capability between methods, with slightly improved sensitivity of the RT-PCR for Enterococcus faecium. Inclusivity testing confirmed detection of all targeted species, while exclusivity testing showed no cross-reactivity with non-target bacteria. Interlaboratory validation confirmed consistent performance across laboratories. Application in drinking water monitoring showed 91.8% agreement between RT-PCR and culture results in samples collected after distribution network repairs, with RT-PCR detecting enterococci more frequently than the reference method.</p><p><strong>Conclusions: </strong>The validated RT-PCR method provides reliable detection of intestinal enterococci in drinking water with sensitivity comparable to the reference method, but with a much shorter time-to-result (3-4 h). This rapid detection capability enables faster responses to faecal contamination events and supports improved public health protection and operational efficiency for drinking water utilities.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":"137 10","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149127563","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Host-microbiome interactions and enteric pathogen infections-from early-life to long-term impacts on gut-brain communication.","authors":"Mélanie G Gareau, Jungjae Park, Kim E Barrett","doi":"10.1093/jambio/lxag237","DOIUrl":"10.1093/jambio/lxag237","url":null,"abstract":"<p><p>Interactions of the host with the microbes that constitute the intestinal microbiota are important for maintaining health and well-being within the gastrointestinal tract and beyond. Disruptions to these interactions, such as those that follow enteric pathogen infection or administration of antibiotics, can lead to persistent pathophysiology, particularly if disruptions occur during early development in neonatal life. Here, we highlight the impacts of altered host-microbe interactions following insults across the life span and discuss emerging mechanisms of action promoting these disruptions (see Graphical Abstract). We also identify potential therapeutics to restore these important pathways and their mechanisms of action.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13662926/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148968905","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterization of hydrolase-associated COD reduction by Bacillus subtilis BY8.","authors":"Zi Yan Jin, Ning Ma, Kai Ru Zhang, Shan Li, Hao Chen Jiang, Bo Wen Zhang, Yan Zhao, Xiang Shan Ji","doi":"10.1093/jambio/lxag243","DOIUrl":"10.1093/jambio/lxag243","url":null,"abstract":"<p><strong>Aims: </strong>This study aimed to evaluate the chemical oxygen demand (COD) reduction ability of BY8 and optimize its fermentation conditions for aquaculture-derived organic matter reduction.</p><p><strong>Methods and results: </strong>BY8 was cultivated in aquaculture feed leachate to assess its growth, COD reduction, and hydrolytic capacity. The genome of BY8 was sequenced and hydrolytic enzyme-related genes were identified. Fermentation conditions were optimized by screening carbon and nitrogen sources and their concentrations. BY8 reduced COD in feed leachate by 45.57% ± 2.47% within 24 h. Clear hydrolysis zones were observed on protease-, amylase-, and cellulase-screening media. Extracellular protease, amylase, and cellulase activities reached 0.68 ± 0.06 U ml-1, 36.31 ± 2.65 U dl-1, and 3.00 ± 0.74 U mg-1 protein at 24 h, respectively. Genome annotation identified 92 non-redundant hydrolase-associated candidate ORFs, including protease/peptidase-, GH13 alpha-glucan/starch-, and cellulose or β-glucan-related candidates. qRT-PCR analysis confirmed transcriptional responses of selected hydrolase-associated genes, with Vpr gene (vpr), the maltogenic α-amylase MdxD gene (mdxD), and the putative FrvX-family M42 peptidase gene (frvX) showing the greatest fold increases at 24 h relative to their respective 0-h levels within the selected functional groups. Fermentation optimization showed that glucose and soybean meal were the most suitable carbon and nitrogen sources for BY8, with optimal concentrations of 5% and 2%, respectively. Under the optimized small-scale fermentation conditions, the viable cell counts reached 1.08 × 10¹¹ CFU ml-1.</p><p><strong>Conclusions: </strong>BY8 showed substantial extracellular hydrolase production and COD reduction in feed leachate, supporting its possible use as a candidate strain for aquaculture organic-load management.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149074829","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"AI-empowered viral metagenomics for clinical diagnosis: advances, bottlenecks, and translational pathways.","authors":"Quan Shen, Wenhui Shi, Chaoran Liu, Zheming Cao, Yutong Fu, Alfred Ndjekadom, Xiaochun Wang, Yuwei Liu, Shixing Yang, Likai Ji, Henan Li, Chenglin Zhou, Junfeng Liu, Wen Zhang","doi":"10.1093/jambio/lxag239","DOIUrl":"10.1093/jambio/lxag239","url":null,"abstract":"<p><p>Viral metagenomics, leveraging high-throughput sequencing technologies, provides comprehensive, hypothesis-free characterization of viral communities in clinical specimens, establishing itself as a pivotal tool for clinical diagnosis, pathogen discovery, and epidemiological surveillance of viral infectious diseases. The integration of artificial intelligence (AI) has demonstrated transformative potential in viral metagenomic data analysis, significantly enhancing sequence classification, feature extraction, pattern recognition, and result interpretation, thereby improving analytical efficiency, accuracy, and automation. Machine-learning and deep-learning approaches have shown potential for detecting low-abundance viral signals and mitigating background noise in complex clinical samples, and facilitating pathogen identification in complex clinical samples, substantially augmenting the clinical utility of viral metagenomics. Nevertheless, routine clinical implementation faces persistent challenges, including high host nucleic acid background, low viral titers, contamination control, limited reference databases, absence of standardized analytical pipelines, and limited interpretability and generalizability of some predictive models. Furthermore, clinical validation, result reproducibility, ethical compliance, and data security represent critical translational barriers that impede widespread adoption. This review comprehensively summarizes recent advances and clinical applications of AI-empowered viral metagenomics, highlighting its transformative potential in pathogen detection, diagnosis of challenging infections, infectious disease surveillance, and precision medicine. We comprehensively address major technical bottlenecks and translational barriers that must be overcome to facilitate standardized development and clinical implementation of this emerging paradigm.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149080787","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Windows of opportunity for genotoxic E. coli in colorectal cancer initiation.","authors":"Jarryd Michael Boath, Luigi Lukas Nardella, Jens Puschhof","doi":"10.1093/jambio/lxag192","DOIUrl":"10.1093/jambio/lxag192","url":null,"abstract":"<p><p>Polyketide synthase-positive Escherichia coli (pks+ E. coli) has attracted significant attention following the discovery that its genotoxic metabolite, colibactin, imprints distinct mutational signatures in intestinal epithelial cells that are detectable in ∼12% of colorectal cancer (CRC) cases. Although these signatures are present in established tumors, growing evidence suggests that colibactin acts early in tumorigenesis, positioning pks+ E. coli as a potential contributor to CRC initiation. Mechanistic studies show that colibactin genotoxicity is highly context dependent and requires direct epithelial contact-a condition that is typically restricted in the healthy adult intestine. Here, we propose that colibactin mutagenesis occurs during discrete temporal windows when host barriers and microbial ecology transiently permit pks+ E. coli-epithelial interactions. These windows may arise during inflammatory disease states or early-life gut development, two contexts that share disrupted epithelial defenses, expansion of facultative anaerobes, and dynamic oxygen gradients. Defining such permissive states will be essential for establishing causal links between pks+ E. coli exposure and CRC initiation and may reveal opportunities for prevention by targeting microbial colonization during critical periods of susceptibility.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148619814","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Bioactive chitosan dressings functionalized with indole alkaloids suppress biofilm development and virulence traits in bacterial species relevant to wound infections.","authors":"Pedro Henrique de Jesus Barbosa, Marcos Vinicius de Souza Pereira, Gisele de Vasconcelos Sobreira Braathen, Francisco Witallo Souza do Nascimento, Dayvson Ayala Costa, Henriete da Silva Vieira, Gaspar Diaz-Muñoz, Jemmyson Romário de Jesus, Ciro Cesar Rossi, Marisa Alves Nogueira Diaz","doi":"10.1093/jambio/lxag246","DOIUrl":"https://doi.org/10.1093/jambio/lxag246","url":null,"abstract":"<p><strong>Aims: </strong>Chronic wound infections are frequently associated with biofilm-forming pathogens that impair healing and reduce the effectiveness of antimicrobial therapies. This study evaluated the antimicrobial, antibiofilm, synergistic, and antivirulence activities of two indole alkaloids, 3-(1H-indol-3-yl)propanoic acid (C1) and 1H-indole-2-carboxylic acid (C2), and investigated their incorporation into chitosan-based biodressings for infected wound management.</p><p><strong>Methods and results: </strong>Antimicrobial activity was evaluated by agar diffusion, minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), biofilm inhibition, and checkerboard assays. Cytotoxicity was assessed using RAW 264.7 macrophages and HaCaT keratinocytes. Biodressings were characterized by Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis, porosity, and release kinetics, while antimicrobial activity and gene expression were evaluated by agar diffusion and RT-qPCR. Both compounds exhibited broad-spectrum antibacterial activity, particularly against Staphylococcus aureus and Staphylococcus epidermidis, inhibited biofilm formation at sub-MIC concentrations, and showed synergistic or additive interactions with conventional antibiotics. The alkaloids displayed low cytotoxicity and were successfully incorporated into chitosan matrices, showing a biphasic release profile over the evaluated period while preserving antimicrobial activity. Functionalized biodressings significantly downregulated genes involved in adhesion (fimH), biofilm formation (icaA, icaD, aap, and pslA), quorum sensing (lasR), and virulence regulation (sarA).</p><p><strong>Conclusions: </strong>Indole alkaloid-loaded chitosan biodressings constitute a promising multifunctional platform that combines antibacterial, antibiofilm, antibiotic-potentiating, and antivirulence activities while maintaining favorable cytocompatibility. These findings support further investigation of indole alkaloid-loaded chitosan biodressings as a potential biomaterial-based strategy for managing wound-associated bacterial infections.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149109864","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Perilla seed oil reshapes the rumen microbiome and increases fermentation end-products in vitro.","authors":"Yimeng Cui, Boyan Zhang, Xianzhe Jiang, Halidai Rehemujiang, Guishan Xu, Yongqiang Li, Bing Wang","doi":"10.1093/jambio/lxag233","DOIUrl":"10.1093/jambio/lxag233","url":null,"abstract":"<p><strong>Aims: </strong>The effect of Perilla frutescens seed oil (PSO) on an in vitro rumen microbial ecosystem was evaluated by integrating fermentation measurements, microbiome profiling, metagenomics, and untargeted metabolomics.</p><p><strong>Methods and results: </strong>Rumen inoculum was incubated for 24 h with a control TMR substrate (CK), TMR supplemented with 23.7 mg of Perilla seeds per bottle (PS), or TMR supplemented with 8.5 μL of Perilla seed oil per bottle (PSO), with the PS and PSO treatments providing equivalent amounts of seed oil. Fermentation kinetics and volatile fatty acids were measured, and microbial and metabolic responses were characterized using 16S rRNA gene sequencing, metagenomics, KEGG and CAZy annotation, untargeted metabolomics, and MetOrigin2 source tracing. PSO increased maximum gas production and total volatile fatty acid concentrations while maintaining pH within the physiological range. Community diversity was unchanged, but PSO altered microbial composition, including increases in Firmicutes, Verrucomicrobia, Vagococcus, Clostridium, and Lactobacillus and decreases in Shigella sonnei and Methanosarcina sp. Ant1. PSO also altered microbial functional profiles and increased several lipid- and vitamin-associated metabolites, including linoleic acid, 13-HODE, 9-oxoODE, pantothenic acid, and thiamine, while reducing lactate.</p><p><strong>Conclusions: </strong>PSO changed rumen microbial community structure and functional potential in parallel with increased fermentation end-products and extensive metabolic shifts. These in vitro findings identify microbial and metabolic responses that warrant validation in vivo.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148919795","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"In vitro digestion and fecal fermentation profiles of Cremastra appendiculata polysaccharide and evaluation of its antioxidant and hypoglycemic capacities.","authors":"Xue- Yu Peng, Yan-Bing Hu, Xian-Le Luo, Xia Luan, Yang Yang, Xiao-Jun Li, Xue Zhang","doi":"10.1093/jambio/lxag238","DOIUrl":"10.1093/jambio/lxag238","url":null,"abstract":"<p><strong>Aims: </strong>Cremastra appendiculata is a traditional Chinese herbal medicine for liver cancer treatment. C. appendiculata polysaccharides (CAP) as a biomacromolecular showed significant lipid-lowering activities. However, the gastrointestinal metabolic fate and the bioactivities of the fermented products remain poorly understood.</p><p><strong>Methods and results: </strong>Simulated salivary, gastric, intestinal digestion models, together with an in vitro human fecal fermentation system, were employed to evaluate the physicochemical profiles of CAP. Variations in molecular weight, reducing sugars, free monosaccharides, short-chain fatty acids (SCFAs), and pH were determined. 16S rRNA gene sequencing was performed to characterize alterations in fecal fermentation microbial communities, and the in vitro α-amylase and α-glucosidase inhibitory activities and antioxidant activities were assessed. As a result, during the simulated fermentation process, CAP remained structurally stable throughout simulated gastrointestinal digestion. During fecal fermentation, CAP was degraded by the fecal microbiota as the molecular weight decreased and the reducing sugars increased. Moreover, CAP significantly enhanced SCFAs production, particularly acetic acid and propionic acid, with a reduction in fermentation pH. CAP selectively enriched beneficial bacteria, including Bifidobacterium, Bacteroides, Lactobacillus, and Odoribacter, while suppressing potentially harmful microorganisms, such as Proteobacteria, Bilophila, and Eggerthella. Furthermore, fecal fermentation of CAP enhanced inhibitory effects against α-amylase and α-glucosidase and antioxidant activities.</p><p><strong>Conclusions: </strong>These findings provide mechanistic insights into the health-promoting properties of CAP and support its future development as a functional food ingredient and gut microbiota regulator.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148976921","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Pathogenic Escherichia coli isolated from recreational waters in northwestern Wyoming display multidrug resistance.","authors":"Anahita Ghorbani Tajani, Aniket Sharma, Kelsey C Ruehling, Bledar Bisha","doi":"10.1093/jambio/lxag240","DOIUrl":"10.1093/jambio/lxag240","url":null,"abstract":"<p><strong>Aim: </strong>To determine the prevalence, antimicrobial resistance, and genomic characteristics of antimicrobial-resistant Escherichia coli isolated from recreational freshwater in the Greys-Hoback watershed, Wyoming, USA.</p><p><strong>Methods and results: </strong>A total of 263 presumptive E. coli isolates were recovered using membrane filtration (EPA Method 1603), of which 218 (83%) were confirmed as E. coli by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Antimicrobial susceptibility testing by broth microdilution identified 20 (∼9%) isolates with clinically relevant resistance phenotypes, including six multidrug-resistant (MDR; ≥3 antimicrobial classes). Whole-genome sequencing of the resistant subset identified blaEC alleles in all genomes, along with additional resistance determinants, including sul2, tet, aph variants, and floR genes. Plasmid replicons were detected in most isolates, with IncF-type plasmids predominating and associated with MDR gene clusters. Contig-level analysis identified plasmid-associated MDR regions in selected isolates, although plasmid reconstruction was limited by short-read assemblies. Phylogenetic analyses revealed multiple sequence types and O: H serotypes, demonstrating substantial genetic diversity and suggesting that the resistant E. coli population originated from multiple sources rather than from clonal expansion. All sequenced isolates carried multiple virulence-associated genes, indicating co-occurrence of resistance and virulence determinants.</p><p><strong>Conclusions: </strong>By integrating antimicrobial susceptibility testing, whole-genome sequencing, and plasmid reconstruction, we show that recreational freshwater in northwestern Wyoming harbors genetically diverse antimicrobial-resistant pathogenic E. coli, including MDR isolates associated with putative mobile resistance elements. These findings support the inclusion of recreational water systems in environmental AMR surveillance frameworks and highlight the value of integrating phenotypic and genomic approaches for isolate characterization.</p>","PeriodicalId":15036,"journal":{"name":"Journal of Applied Microbiology","volume":" ","pages":""},"PeriodicalIF":2.8,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13637474/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149012410","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}