Emergence of dual β-lactam and colistin resistance via blaFRI-8 and mcr-10.2 co-carriage on an IncFII-family plasmid in Enterobacter vonholyi

Enterobacter vonholyi isolate E323169 represents a rare case of co-carriage of the antimicrobial resistance genes (ARGs) bla FRI-8, mcr-10. 2, isolated from a clinical rectal swab. E323169 represents one of only 11 known E. vonholyi genomes. To date, within the NCBI Pathogen Detection resource, bla FRI-8 was identified in three assemblies and mcr-10. 2 in two. In that dataset, bla FRI-8 co-occurred with an mcr-10 hit only once (Enterobacter asburiae). This study analyzes the genomic, phenotypic and epidemiological importance of this rare co-occurrence. Species identification for E323169 was initially assigned by MALDI-TOF and subsequently confirmed using a multifactorial genomic workflow. Antimicrobial susceptibilities were determined by MIC assay. The genome of E323169 was sequenced on an Illumina-NextSeq-1000, assembled, and annotated for ARGs, virulence factors, and plasmid replicons detection. Comparative phylogenomics used all canonical E. vonholyi RefSeq assemblies, and NCBI metadata were analysed for plasmid distributions of bla FRI-8 and mcr-10. 2. E323169 carried six ARGs: four chromosomally encoded (blaACT-91, fosA, oqxA10, oqxB9) and two plasmid-borne (bla FRI-8 and mcr-10. 2) co-located on IncFII (p14) ₁ₚ14 replicon. Additional plasmid replicons: Col (MG828) ₁ and ColRNAI₁ were also identified. By mining the NCBI Pathogen Detection pipeline, we identified bla FRI-8 on IncFII replicon in E. asburiae JBIWA002, and mcr-10. 2 on a multi-replicon (IncFIB/IncFII) plasmid in E. kobei 11, 778-yvys. This report, to our knowledge, represents the first E. vonholyi isolate co-harboring bla FRI-8 and mcr-10. 2 on a single IncFII family plasmid in a non Enterobacter cloacae complex species, showing the widening host range of plasmid-mediated resistance to carbapenems/colistin. These findings suggest IncFII-family plasmids as recurrent scaffolds for the accumulation of high-impact resistance determinants in Enterobacter and suggest that monitoring IncFII backbones may provide an early warning signal for future convergence events. The detection of Enterobacter vonholyi across human, animal, plant, and environmental sources underscore its relevance within a One Health framework, highlighting the potential for cross-sectoral circulation of antimicrobial resistance. The convergence of bla FRI-8 and mcr-10. 2 on an IncFII plasmid in E. vonholyi exposes an unrecognized reservoir of last-line resistance in a species prone to misidentification. Together, these findings emphasize the importance of integrating genomic surveillance into routine diagnostics to identify hidden reservoirs of carbapenem and colistin resistance and to strengthen infection prevention strategies before wider dissemination occurs. • All available E. vonholyi RefSeq genomes (n = 11) compared species-wide. • First clinical E. vonholyi with blaFRI-8 + mcr-10. 2 on one IncFII plasmid. • Irish isolate E323169 shows carbapenem and colistin resistance. • ARGs mostly chromosomal; plasmid co-carriage unique to E323169. • MALDI-TOF misID risk underscores need for genomic confirmation.