Virulome-resistome convergence in swine-associated multidrug-resistant Escherichia coli from Hungary: virulence marker profiles and zoonotic potential.

BACKGROUND

Swine production can sustain dense reservoirs of multidrug-resistant (MDR) Escherichia coli in which mobilizable resistance may co-occur with virulence modules relevant to animal health and One Health risk. We profiled the virulome of Hungarian swine-associated MDR E. coli and assessed whether key virulence signatures co-segregate with selected high-impact resistance flags.

METHODS

In late 2023, E. coli isolates were obtained from four large-scale Hungarian pig farms through routine veterinary diagnostic/surveillance sampling. Phenotypic antimicrobial susceptibility testing was performed for the full collection (n = 203), including extended-spectrum β-lactamase (ESBL) confirmation (n = 127). A whole-genome sequencing (WGS) subset (n = 116) underwent Illumina sequencing and de novo assembly. Virulence genes were identified in silico using curated virulence databases with harmonized identity/coverage thresholds and summarized as gene prevalence and functional modules. Marker-based definitions were applied for extraintestinal pathogenic E. coli (ExPEC)-like and diarrheagenic E. coli (DEC)-like signatures. Acquired antimicrobial resistance genes were annotated using CARD/RGI, and focused analyses considered CTX-M extended-spectrum beta-lactamase (ESBL) genes and rare, high-consequence determinants (mcr-1, qnrB5). Associations between virulence markers and resistance flags were tested using Fisher's exact test. Rather than performing a comprehensive resistome analysis, we focused on selected high-impact resistance determinants (CTX-M, mcr-1, qnrB5) and their co-carriage with virulence markers.

RESULTS

Across 116 genomes, 208 distinct virulence-associated genes were detected; virulence gene load per genome was heterogeneous. Extraintestinal-associated iron acquisition modules were common, including aerobactin (iucABCD/iutA) in 31/116 (26.7%), yersiniabactin (fyuA/irp1/irp2) in 27/116 (23.3%), and salmochelin (iroB/iroN) in 17/116 (14.7%). Toxin-associated determinants were frequent, with hlyA in 33/116 (28.4%). DEC markers occurred in 33/116 (28.4%), including eae in 23/116 (19.8%), stx2 in 6/116 (5.2%), predominantly enterohemorrhagic E. coli (EHEC)-like (stx2 and eae) profiles (5/116; 4.3%) and estIa in 10/116 (8.6%); astA was present in 27/116 (23.3%). Stringent ExPEC-like criteria were met by 6/116 (5.2%), while 15/116 (12.9%) showed convergent profiles co-carrying aerobactin with at least one DEC marker. CTX-M genes were detected in 24/116 (20.7%) and were enriched among aerobactin-positive isolates and DEC-marker-positive isolates; all EHEC-like isolates carried CTX-M. Rare but critical determinants included mcr-1 (3/116; 2.6%) and qnrB5 (2/116; 1.7%).

CONCLUSION

Hungarian swine-associated MDR E. coli show virulome heterogeneity with frequent aerobactin/toxin modules and DEC markers. CTX-M enrichment in these profiles indicates virulome-resistome convergence, supporting integrated One Health surveillance.