Genomic Characterisation of Pyometra-Associated Escherichia coli in a Lombardy Veterinary Clinic: A Nanopore-Based Case Series

Background/Objectives: Pyometra is a life-threatening uterine infection of intact bitches and queens. Despite growing reports of multidrug-resistant (MDR) Escherichia coli in canine reproductive and urinary infections, no whole-genome data were previously available for pyometra isolates from Italy. This study aimed to characterise, by whole-genome sequencing and comparative genomics, the population structure, resistome and virulome of E. coli causing pyometra in companion animals from northern Italy in the context of European datasets. Methods: Four E. coli isolates (two canine, two feline) from pyometra cases underwent nanopore long-read sequencing. Genomes were compared with Brazilian and Finnish pyometra isolates using core- and accessory-genome analyses, pan-genome partitioning, phylogeny, and gene-based profiling of antimicrobial resistance and virulence determinants. Results: All Italian isolates belonged to phylogroup B2 and to recognised ExPEC sequence types (ST706/O51:H1, ST141/O2:H6, ST372/O75:H31, ST646/O22:H5). Phenotypically, they were uniformly resistant to several penicillins and early/third-generation cephalosporins but remained susceptible to fluoroquinolones, aminoglycosides and trimethoprim–sulphonamide. The combined 57-genome pan-genome was open yet strongly core-dominated; Italian strains shared an efflux- and regulator-centred intrinsic resistome and a rich ExPEC virulence repertoire (P, S, F1C and type 1 fimbriae, multiple siderophores, colibactin, Vat, haemolysin, CNF1) with Brazilian and Finnish isolates. Conclusions: Pyometra-associated E. coli from northern Italian pets belong to globally disseminated high-risk B2 lineages that combine extensive virulence with a largely intrinsic resistome, and currently retain susceptibility to several key drug classes, underscoring an important but vulnerable therapeutic window.