Abstract
Background: Acquired 16S rRNA methyltransferases (16S-RMTases) confer high-level resistance to aminoglycosides and are often associated with β-lactam and quinolone resistance determinants. Methods: Using PCR, whole-genome sequencing and conjugation experiments, we conducted a retrospective genomic surveillance study of 16S-RMTase-producing Enterobacterales, collected between 2006 and 2023, to explore transmission dynamics of methyltransferase and associated antibiotic resistance genes. Results: Among the 10,731 consecutive isolates, 150 (1.4%) from 13 species carried armA (92.7%), rmtB (4.7%), and rmtF + rmtB (2.7%) methyltransferase genes. The coexistence of extended-spectrum β-lactamase (bla(CTX-M-3/15), bla(SHV-12), bla(SFO-1)), carbapenemase (bla(NDM-1/5), bla(VIM-1/4/86), bla(OXA-48)), acquired AmpC (bla(CMY-2/4/99), bla(DHA-1), bla(AAC-1)), and plasmid-mediated quinolone resistance (qnrB, qnrS, aac(6')-Ib-cr) genes within these isolates was also detected. Methyltransferase genes were carried by different plasmids (IncL/M, IncA/C, IncR, IncFIB, and IncFII), suggesting diverse origins and sources of acquisition. armA was co-transferred with bla(CTX-M-3/15), bla(NDM-1), bla(VIM-4/86), bla(OXA-48), bla(CMY-4), aac(6')-Ib-cr, qnrB, and qnrS, while rmtF1 was co-transferred with bla(SFO-1), highlighting the multidrug-resistant nature of these plasmids. Long-read sequencing of ST6260 K. pneumoniae isolates revealed a novel resistance association, with rmtB1 and bla(NDM-5) on the chromosome, bla(OXA-232) on a conjugative ColKP3 plasmid, and rmtF1 with bla(SFO-1) on self-transmissible IncFIB and IncFII plasmids. Conclusions: The genetic plasticity of plasmids carrying methyltransferase genes suggests their potential to acquire additional resistance genes, turning 16S-RMTase-producing Enterobacterales into a persistent public health threat.