Dissemination of OXA-23 carbapenemase-producing Proteus mirabilis and Escherichia coli is driven by transposon-carrying lineages in the UK

Carbapenem-resistant Enterobacterales are a significant threat to global public health. Here, we characterize blaOXA-23-positive Proteus mirabilis (n=8) and Escherichia coli (n=3) isolates from human clinical samples collected between 2021-2024 in the UK. Whole genome sequencing (WGS) was used to generate data, and a core gene SNP-based phylogenetic tree was constructed to assess the genomic relatedness among the isolates. To provide an international context, we included publicly available genomes. Short-read mapping to a reference genome enabled reconstruction of the genomic neighborhood around blaOXA-23. Minimum inhibitory concentration (MIC) determination was performed using broth microdilution and results interpreted using EUCAST guidelines. UK P. mirabilis isolates belonged to ST142 and were closely related (2-13 SNPs) to French isolates from 2017-2019. E. coli ST38 isolates harboured blaOXA-23 and showed high genetic relatedness (5-9 SNPs) among themselves. In P. mirabilis, blaOXA-23 was associated with transposon Tn6703, while E. coli harboured a novel composite transposon, designated Tn7816, bordered by two copies of IS15DIV and with three copies of blaOXA-23. blaOXA-23 was integrated into the chromosome in all isolates. All isolates were resistant to amoxicillin/clavulanic acid (>32 mg/L) and with meropenem MICs above the EUCAST screening cut-off (0.5-1 mg/L). In conclusion, UK blaOXA-23-positive P. mirabilis isolates belong to the same clonal lineage (ST142) previously reported in Belgium, Germany, Switzerland and France, suggesting introduction of this lineage into the UK. This is the first report of an E. coli ST38 lineage with chromosomally-encoded blaOXA-23 located within a novel transposon Tn7816. WGS plays an important role in identifying the mechanism(s) of transmission of emerging carbapenemase genes. IMPACT STATEMENTMost diagnostic assays primarily focus on detection of the big 5 carbapenemase gene families (KPC, OXA-48-like, NDM, VIM and IMP), which are globally dominant in Enterobacterales. OXA-23-like carbapenemase genes are predominantly identified in Acinetobacter baumannii meaning that their presence in Enterobacterales is likely underestimated. In this study, we report the identification of blaOXA-23 in Proteus mirabilis and Escherichia coli isolated from UK clinical samples following incorporation of blaOXA-23-like as a target in the multiplex-PCR assay used to screen all Gram-negative bacteria referred to the UKs national reference laboratory for investigation of carbapenem resistance. To enhance our understanding of the genomic epidemiology of blaOXA-23 we utilized short-read sequencing to characterize all isolates, and long-read assembly polished with short-read to determine the genomic context of blaOXA-23 in E. coli. WGS analysis provided invaluable insights into the genomic relatedness among the isolates, identifying that blaOXA-23-positive P. mirabilis isolates were closely related to those previously reported in Europe and uncovered a novel transposon associated with blaOXA-23 in E. coli. In addition, blaOXA-23 was chromosomally located in all isolates, with the potential for stable vertical inheritance. The results of this study highlight the need to further characterize Enterobacterales isolates suspected of carbapenemase production but negative for the big 5 carbapenemase gene families and further demonstrates the role of WGS in characterizing bacterial strains and mobile genetic elements associated with the emergence and transmission of antimicrobial resistance mechanisms. DATA SUMMARYIllumina short-read, contigs and MIC (if available) data for P. mirabilis (n=8) and E. coli (n=3) blaOXA-23-positive isolates from the UK are available in the ENA database under Bioproject PRJEB80458. The Illumina polished version of the long-read-only (Nanopore) assembly for 1697008 (ES1) E. coli isolate is accessible under the assembly accession number GCA_964341145. The accession number for each genome and the metadata generated in this study are provided in the Table 1. O_TBL View this table: org.highwire.dtl.DTLVardef@1245c4forg.highwire.dtl.DTLVardef@28785corg.highwire.dtl.DTLVardef@f5ed6eorg.highwire.dtl.DTLVardef@1b6fd5corg.highwire.dtl.DTLVardef@b3b96f_HPS_FORMAT_FIGEXP M_TBL O_FLOATNOTable 1.C_FLOATNO O_TABLECAPTIONAccession numbers, metadata and antimicrobial susceptibility testing (AST) data for P. mirabilis (n=8) and E. coli (n=3) isolates obtained from human clinical samples in the UK. The table displays the minimum inhibitory concentration (MIC) values (in mg/L) for 12 antibiotics. MIC values were categorized according to the EUCAST13 guidelines using clinical breakpoints: 1) susceptible, indicated by the white box, 2) susceptible, increased exposure, indicated by the diagonal pattern, and 3) resistant, indicated by the grey box. C_TABLECAPTION C_TBL