Genomic profiling of ESBL/AmpC-producing Escherichia coli from backyard poultry: resistome, virulome, plasmidome, and CRISPR-Cas insights

Introduction Antimicrobial resistance (AMR) is a major One Health concern driven by many factors including unregulated animal farming. This study aimed to perform a comprehensive phenotypic and genotypic characterization of Escherichia coli isolated from backyard poultry reared in households in rural setup in southern India. Methods A total of 48 cloacal samples were collected from poultry birds across 12 epidemiological units using a 30-cluster sampling strategy. E. coli isolation was performed using standard microbiological methods and confirmed by species-specific multiplex PCR. Of the 48 samples, 45 yielded confirmed E. coli isolates, among which 18 ESBL- and/or AmpC-positive isolates were selected for whole genome sequencing (WGS). Genomic analyses included detection of antimicrobial resistance genes, virulence-associated genes, mobile genetic elements, plasmid replicons, biofilm-associated genes, multilocus sequence typing, serotyping, CH typing, Clermont phylogrouping, CRISPR-Cas profiling, and SNP-based phylogenetic analysis. Results A total of 48 cloacal samples yielded 45 confirmed isolates of which 18 detected with ESBL genes including bla SHV (28.8%), bla TEM (26.6%), and bla CTX-M (20%) along with plasmid-mediated AmpC genes (11.11%). Resistome profiling revealed diverse ARGs such as bla CTX-M-15 , bla DHA-1 , qnrS/B, tet(A), sul1/2/3, dfrA var iants, and mphA/B, in addition to intrinsic efflux systems ( acr, emr, mdt, mar ). Virulome analysis showed conserved genes associated with adhesion (fim, csg, ecp), iron acquisition (ent, fep, fes, ybt), and stress response (gad, hlyE, ompT, iss). CRISPR-Cas analysis revealed a high prevalence of Type I-E arrays (83.3%) coexisting with multiple plasmids. MLST and CH typing revealed high genetic diversity across 12 sequence types, including ST10, ST48, ST3107, and ST226. SNP-based phylogeny placed isolates mainly within commensal phylogroups A and B1, with relatedness to global strains. All sequenced isolates were multidrug-resistant, with significant β -lactam resistance enrichment (Z = 3.46, p 0.001). ARG distribution differed significantly among phylogroups ( p = 0.036), and Simpson’s diversity index (D = 0.94) indicated marked clonal heterogeneity. Strong plasmid–ARG associations (Cramér’s V 0.5) suggested plasmid-mediated resistome structuring. Discussion Backyard poultry-derived E. coli showed multidrug resistance, genetic diversity, and virulence-associated traits, highlighting backyard poultry as a potential reservoir for AMR dissemination at the human–animal–environment interface.