Dissemination and Stability of the -Carrying IncX3-Type Plasmid among Multiclonal Klebsiella pneumoniae Isolates.

NDM-5 carbapenemase was mainly identified in , while the rapid transmission of among has raised serious public attention. This study identified 14 NDM-5-producing isolates from 107 carbapenem-resistant isolates, recovered from blood, urine, and normally sterile body fluids of pediatric patients from January 2016 to December 2018. All NDM-5-producing isolates were highly resistant to β-lactams, while tigecycline and polymyxin B exhibited excellent antimicrobial activity. These 14 strains belonged to 9 different sequence types (STs) and displayed various pulsed-field gel electrophoresis (PFGE) patterns, suggesting that they were not clonally related. S1-PFGE followed by Southern blotting showed that the gene was located on an ∼46-kb IncX3 plasmid in all strains. All -carrying plasmids were successfully transferred into recipient J53. PCR-based sequencing demonstrated that all of the -carrying plasmids shared highly similar backbones, with nucleotide sequence identity of >99%. Moreover, this plasmid displayed high sequence similarity to the previously reported epidemic IncX3 -carrying plasmids, with dynamic changes observed only in -surrounding elements. Interestingly, the IncX3 -carrying plasmids showed strong stability in clinical isolates when cultured in antibiotic-free medium. However, after the conjugation inhibitor linoleic acid was added, a gradual increase in the level of IncX3 plasmid loss could be observed. Clinical isolates displayed 10% to 15% -carrying plasmid loss after coculture with linoleic acid for 5 days. These results showed that the IncX3 plasmid facilitated the dissemination of among multiclonal strains in children and that conjugal transfer contributed significantly to IncX3 plasmid stability within The emergence and spread of New Delhi metallo-β-lactamase (NDM)-producing have been a serious challenge to public health, and NDM-5 shows increased resistance to carbapenems compared with other variants. NDM-5 has been identified mostly in but has rarely been described in and other isolates. Here, we present the dissemination of highly similar 46-kb IncX3 -carrying plasmids among multiclonal strains in children, highlighting the horizontal gene transfer of among strains via the IncX3 plasmid. Moreover, the IncX3 -carrying plasmids displayed strong stability in clinical strains when cultured in antibiotic-free medium, and the plasmid maintenance was attributed partly to conjugal transfer. Plasmid conjugation is mediated by the type IV secretion system (T4SS), and T4SS is conserved among all epidemic IncX3 -carrying plasmids. Therefore, combining conjugation inhibition and promotion of plasmid loss would be an effective strategy to limit the conjugation-assisted persistence of IncX3 -carrying plasmids.