Purpose: Three NDM-5-producing Enterobacteriaceae (Escherichia coli, Klebsiella pneumoniae , and Citrobacter braakii , one each) were isolated during a screening study for the presence of carbapenemase-producing Enterobacteriaceae (CPE) strains in urban rivers in China. The aim of the present study was to characterize these NDM-5-producing isolates by using whole-genome analysis.
Methods: In vitro susceptibility testing was performed using the broth microdilution method. Conjugation assay was carried out to investigate the transferability of bla _NDM-5-harboring plasmids. Whole-genome sequencing was performed using an Illumina HiSeq combined with the PacBio RSII system. The genetic characteristics of the bla _NDM-5-harboring plasmids were analyzed. Antimicrobial resistance genes and virulence genes were identified from the genome sequences. Phylogenetic analysis was performed based on core genome.
Results: Antimicrobial susceptibility testing showed that all three isolates were resistant to carbapenems, cephalosporins, quinolones, and aminoglycosides, and susceptible to colistin. Whole-genome sequencing showed that each isolate carried multiple antibiotic resistance genes mediating multidrug resistance, and harbored numerous virulence genes, some of which were located on plasmids. In these isolates, bla _NDM-5 was carried by an IncX3 plasmid in K. pneumoniae and C. braakii , and on an IncR/IncX1 plasmid in E. coli . Conjugation experiments showed that these bla _NDM-5-haboring plasmids were successfully transferred to E. coli  J53. Phylogenetic analysis revealed that E. coli  SCLZR49 was present in a cluster with isolates of different origin, K. pneumoniae  SCLZR50 was mainly clustered with clinical isolates, and C. braakii  SCLZR53 had closely genetic relationship with environmental isolates.
Conclusion: This study revealed contamination of the urban river ecosystems by clinically significant carbapenemase gene bla _NDM-5, raising the possibility of plasmid transmission into the environmental from humans and highlighting the need for a constant surveillance of CPE in the environment under the “One-Health” perspective.
Keywords: carbapenemase, antimicrobial resistance, aquatic environment, river water