Structural Diversity, Fitness Cost, and Stability of a Bla_NDM-1-Bearing Cointegrate Plasmid in Klebsiella pneumoniae and Escherichia coli

Cointegrate/hybrid plasmids combine the genetic elements of two or more plasmids and generally carry abundant antimicrobial resistance determinants. Hence, the spread of cointegrate plasmids will accelerate the transmission of AMR genes. To evaluate the transmission risk caused by cointegrate plasmids, we investigated the structural diversity, fitness cost, and stability of a cointegrate plasmid in Klebsiella pneumoniae YZ6 and Escherichia coli EC600. The cointegrate plasmid pSL131_IncA/C_IncX3 was from a clinical Salmonella Lomita strain. After transferring the plasmid into E. coli EC600 by conjugation, we observed plasmids with different structures, including a full-length original plasmid and two truncated versions. By contrast, DNA fragment deletion and ^blaCTX‐ M‐14 gene insertion in the plasmid were detected in a transconjugant derived from K. pneumoniae YZ6. These results suggest that the structure of the plasmid was unstable during conjugation. Furthermore, both the full‐length plasmid in EC600 and the structurally reorganized plasmid in YZ6 imposed a fitness cost on the bacterial host and enhanced biofilm formation ability. Serial passaging in antibiotic‐free medium resulted in a rapid decline of the plasmid in YZ6. However, the stability of the structurally reorganized plasmid in YZ6 was improved via serial passaging in antibiotic‐containing medium. SNP calling revealed that mutations of the outer membrane porin may play an essential role in this process. These findings indicate that structural versatility could contribute to the dissemination of cointegrate plasmids. Although the plasmid incurred a fitness cost in other Enterobacteriaceae species, positive selection could alleviate the adverse effects.