Genomic Characterization of 16S rRNA Methyltransferase-Producing Enterobacterales Reveals the Emergence of Klebsiella pneumoniae ST6260 Harboring rmtF, rmtB, blaNDM-5, blaOXA-232 and blaSFO-1 Genes in a Cancer Hospital in Bulgaria.

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 (blaCTX-M-3/15, blaSHV-12, blaSFO-1), carbapenemase (blaNDM-1/5, blaVIM-1/4/86, blaOXA-48), acquired AmpC (blaCMY-2/4/99, blaDHA-1, blaAAC-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 blaCTX-M-3/15, blaNDM-1, blaVIM-4/86, blaOXA-48, blaCMY-4, aac(6')-Ib-cr, qnrB, and qnrS, while rmtF1 was co-transferred with blaSFO-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 blaNDM-5 on the chromosome, blaOXA-232 on a conjugative ColKP3 plasmid, and rmtF1 with blaSFO-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.