Genetic Diversity of Carbapenem-Resistant Enterobacteriaceae (CRE) Clinical Isolates From a Tertiary Hospital in Eastern China

Abstract

The prevalence of carbapenem-resistant Enterobacteriaceae (CRE) is increasing globally, with different molecular mechanisms described. Here we studied the molecular mechanisms of carbapenem resistance, including clonal and plasmid dissemination, of 67 CRE isolates collected between 2012 and 2016 from a tertiary hospital in Eastern China, an CRE endemic region. Species identification and susceptibility testing were performed using the BD Phoenix Automated Microbiology System. Isolates were characterized by PCR (for carbapenemases, ESBLs, AmpC and porin genes), multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE), and conjugation transfer experiments. Selected blaKPC-2 -harboring plasmids were subjected to next-generation sequencing using the Illumina Miseq platform. Among the 67 CRE isolates, 42 Klebsiella pneumoniae, 10 Serratia marcescens, 6 Enterobacter cloacae, 2 Raoultella ornithinolytica, 2 K. oxytoca, 1 K. aerogenes, and 4 Escherichia coli isolates were identified. Six different carbapenemases were detected, including blaKPC-2 (n = 45), blaKPC-3 (n = 1), blaNDM-1 (n = 6), blaNDM-5 (n = 1), blaIMP-4 (n = 2), and blaVIM-1 (n = 2); blaOXA-48-like genes were not detected. One E. cloacae strain possessed both blaNDM-1 and blaKPC-3, while two E. cloacae isolates harbored blaNDM-1 and blaVIM-1. ESBLs (CTX-M, SHV, and TEM) and/or AmpC (CMY, DHA, and ACT/MIR) genes were also identified in 59 isolates, including 13 strains that lacked carbapenemases. Several insertions or stop codon mutations were found within porin genes of K. pneumoniae, E. coli and S. marcescens isolates, both with and without carbapenemases. The 42 K. pneumoniae isolates belonged to 12 different sequence types (ST), with ST11 being the most common, while the 6 E. cloacae isolates comprised 4 different STs. The 10 S. marcescens all shared the same PFGE pulsotype, suggestive of clonal spread. Complete plasmid sequencing and PCR screening revealed both intra-strain and inter-species spread of a common blaKPC-2-harboring plasmid in our hospital. Taken together, our study revealed extensive genetic diversity among CRE isolates form a single Chinese hospital. CRE isolates circulating in the hospital differ significantly in their species, STs, porin genes, carbapenemase genes, and their plasmid content, highlighting the complex dissemination of CRE in this endemic region.

A possible link between improper use of carbapenems and prevalence of carbapenem-resistant Klebsiella Pneumoniae in a hospital

“…Six carbapenemase candidate genes were analyzed in the CRKp isolates in this study. The most frequently identi ed gene is bla NDM-1 , which is different from the previous ndings in China that the common epidemic genotype is found to be bla KPC [7,8].…”

Evaluation of MASTDISCS combi Carba plus for the identification of metallo‐β‐lactamases, KPC and OXA‐48 carbapenemase genes in Enterobacteriaceae clinical isolates

“…Currently MBLs, KPC and OXA‐48 carbapenemase genes are considered a major threat in Enterobacteriaceae clinical isolates, representing a potential source of clinical failure in patients treated with almost all β‐lactam regimens (Tängdén and Giske ). Moreover, the production of these enzymes might contribute to their hydrolytic activity and levels of resistance to broad‐spectrum β‐lactams, as well as to the possible migration of these enzymes (Miao et al ).…”

“…Carbapenemase genes were amplified using a panel of primers for detection of all known genes of MBLs ( bla IMP , bla VIM and bla NDM‐1 genes), KPC ( bla KPC gene), OXA‐48 ( bla OXA‐48 gene) carbapenemase genes (Tsakris et al ; Miao et al ). For E. aerogenes isolates, total DNA from logarithmic‐phase‐growth cultures was extracted with the TIANamp Bacteria DNA Kit (TIANGEN Biotech, Beijing, China) according to the manufacturer's protocol.…”

“…Carbapenems are often considered last resort antibiotics in the treatment of infections due to multidrug-resistant Enterobacteriaceae clinical isolates, since they are stable even in response to extended-spectrum b-lactamases (ESBLs) and AmpC enzymes (Tsakris et al 2010;Jean et al 2018;Carrasco-Anabal on et al 2019;Wang et al 2019). However, the carbapenem resistance has been increasingly reported among Enterobacteriaceae in the past few years, which is largely attributed to the production of metallo-b-lactamases (MBLs), KPC and OXA-48 (Yusuf et al 2014;Tijet et al 2016;No€ el et al 2017;Tamma and Simner 2018). Therefore, the dissemination of carbapenemase-producing Enterobacteriaceae (CPE) has been a serious public health problem of global scale, and the early detection of CPE is essential for targeted antimicrobial therapy as well as for effective and timely infection Letters in Applied Microbiology 70, 42--47 © 2019 The Society for Applied Microbiology control.…”

Bloodstream Infections caused by Klebsiella pneumoniae and Serratia marcescens isolates co-harboring NDM-1 and KPC-2.

“…In the last decade, several studies have been reported the emergence of Gram-negative bacteria carrying multiple carbapenemases [1][2][3][4][5][6], isolated from patients with distinct body sites of infection. However, clinical isolates harbouring simultaneously blaKPC-2 and blaNDM-1 are less common.…”