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We further mapped our genome to the p17-84 OXA plasmid (accession number: CP059478) (Figure 1B), which is located in an ST63 pandrug-resistant I A. pittii i .[15] High nucleotide sequence identity was observed for some of the contigs in the TCM2 genome with p17-84 OXA (Figure 1C). Identified resistance genes were correlated with the actual antimicrobial resistance phenotype of the strain and a I bla i SB OXA-58 sb -bearing plasmid should be harbored by the TCM2 strain. Consequently, we concluded that there was a I bla i SB OXA-58 sb -bearing plasmid in the TCM2 strain, which was probably obtained from other I A. pittii i and I A. nosocomialis i strains collected in China in 2012 and 2014, respectively (Supplementary Table 2). [Extracted from the article]

Hypervirulent carbapenem-resistant Klebsiella pneumoniae (hv-CRKP) has been widely reported and poses a global threat. However, the comprehensive genetic structure of ST11-KL64 hv-CRKP and the possible evolutionary mechanisms from a genetic structure perspective of this high-risk clone remain unclear. Here, a bla KPC-2 - bla NDM-1 -positive ST11-KL64 hv-CRKP isolate was obtained from a human bloodstream infection (BSI). Whole-genome sequencing and bioinformatics analyses revealed that it contained a fusion plasmid, pKPTCM2–1. pKPTCM2–1 is a conjugative plasmid composed of an oriT -positive pLVPK-like virulence plasmid and a type IV secretion system-produced bla NDM-1 -bearing IncX3 plasmid mediated by IS 26 -based co-integration. This progress generated 8-bp target site duplications (TGAAAACC) on both sides. The fusion plasmid possessed self-transferability and could be transferred to bla KPC-2 -harboring ST11-KL64 CRKP to form the ST11-KL64 hv-CRKP clone. The pLVPK-like-positive ST11-KL64 strain exhibited virulence levels similar to those of the typical hypervirulent K. pneumoniae NTUH-2044. The mutation, Tet(A) (A276S), which was believed to lead to tigecycline resistance was observed. Overall, this high-risk clone has emerged as a tremendous threat in fatal BSIs and thus, targeted surveillance is an urgent need to contain the hv-CRKP clones. • One bla KPC-2 - bla NDM-1 -positive ST11-KL64 hv-CRKP in human bloodstream infection. • oriT -positive pLVPK-like virulence plasmid and bla NDM-1 -bearing IncX3 plasmid fusion. • One mutation of Tet(A) (A276S) maybe lead to tigecycline resistance. [ABSTRACT FROM AUTHOR]

Competing Interests: The authors report no conflicts of interest in this work.

Objectives: To characterize one OXA-232-producing wzi93 -KL112-O1 carbapenem-resistant Klebsiella pneumoniae (CRKP) co-harboring chromosomal bla CTX-M-15 and one rmpA2 -associated virulence plasmid., Methods: Minimum inhibitory concentrations (MICs) were measured via broth microdilution method. Conjugation, chemical transformation, string test and Galleria mellonella infection model experiments were also conducted. Whole-genome sequencing (WGS) was performed on the Illumina and Nanopore platforms. Antimicrobial resistance determinants were identified using ABRicate program with ResFinder database. Insertion sequences (ISs) were identified using ISfinder. Bacterial virulence factors were identified using virulence factor database (VFDB). Wzi, capsular polysaccharide (KL) and lipoolygosaccharide (OCL) were analyzed using Kleborate with Kaptive . Phylogenetic analysis of 109 ST15 K. pneumoniae strains was performed using core genome multilocus sequence typing (cgMLST) on the Ridom SeqSphere+ server. MLST, replicons type, SNP strategies and another cgMLST analysis for 45 OXA-232-producing K. pneumoniae strains were further conducted using BacWGSTdb server., Results: K. pneumoniae KPTCM strain belongs to ST15 with wzi93 , KL112 and O1. It possessed a multidrug-resistant (MDR) profile and was resistant to carbapenems (meropenem and ertapenem), ciprofloxacin and amikacin. Virulence assays demonstrated KPTCM strain possesses a low virulence phenotype. WGS revealed it contained one circular chromosome and nine plasmids. The carbapenemase-encoding gene bla OXA-232 was located in a 6141-bp ColKP3-type non-conjugative plasmid and flanked by ΔIS Ecp1 and Δ lysR -Δ ereA . Interestingly, bla CTX-M-15 was located in the chromosome mediated by IS Ecp1- based transposon Tn 2012 . Importantly, it harbored a rmpA2 -associated pLVPK-like virulence plasmid with iutA - iucABCD gene cluster and one IS 26 -mediated MDR fusion plasmid according to 8-bp (AGCTGCAC or GGCCTTTG) target site duplications (TSD). Based on the cgMLST and SNP analysis, data showed OXA-232-producing ST15 K. pneumoniae isolates were mainly isolated from China and have evolved in recent years., Conclusions: Early detection of CRKP strains carrying chromosomal bla CTX-M-15 , OXA-232 carbapenemase and pLVPK-like virulence plasmid is recommended to avoid the extensive spread of this high-risk clone., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Tian, Xing, Zhao, Fan, Bai, Fu and Wang.)