Whole-Genomic Analysis of NDM-5-Producing Enterobacteriaceae Recovered from an Urban River in China

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.

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.

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.

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.