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 (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.