AUTHOR=Cai Yimei , Chen Cha , Zhao Mei , Yu Xuegao , Lan Kai , Liao Kang , Guo Penghao , Zhang Weizheng , Ma Xingyan , He Yuting , Zeng Jianming , Chen Liang , Jia Wei , Tang Yi-Wei , Huang Bin 

TITLE=High Prevalence of Metallo-β-Lactamase-Producing Enterobacter cloacae From Three Tertiary Hospitals in China

JOURNAL=Frontiers in Microbiology

VOLUME=10

YEAR=2019

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2019.01610

DOI=10.3389/fmicb.2019.01610

ISSN=1664-302X

ABSTRACT=<p><italic>Enterobacter cloacae</italic> has recently emerged as one of the most common carbapenem-resistant <italic>Enterobacteriaceae</italic>. The emergence and spread of metallo-β-lactamase-producing <italic>E. cloacae</italic> have posed an immediate threat globally. Here, we investigated the molecular characteristics of 84 carbapenem-resistant <italic>Enterobacter cloacae</italic> (CREL) collected from three tertiary hospitals in China between 2012 and 2016. Species identification and antimicrobial susceptibility testing were performed using a VITEK-2 system. Carbapenems, polymyxins B, and tigecycline were tested by broth microdilution method. The carbapenem in activation method (CIM) and cefoxitin three-dimensional test were used to detect carbapenemase and AmpC β-lactamase, respectively. Isolates were screened for β-lactam resistance genes by PCR, and expression of <italic>ompC</italic> and <italic>ompF</italic> was determined by qRT-PCR. Genetic relatedness was performed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST), while selected isolates were subjected to whole-genome sequencing. Among the 84 CREL isolates, 50 (59.5%) were detected as carbapenemase producers. NDM-1 was the dominant carbapenemase (80.0%), followed by IMP-26 (8.0%) and IMP-4 (6.0%). Notably, we identified the first NDM-1 and IMP-1 co-producing <italic>E. cloacae</italic>, carrying plasmids of several incompatibility (Inc) groups, including IncHI2, IncHI2A, and IncN. Most isolates showed decreased expression of <italic>ompC</italic> and/or <italic>ompF</italic>, and contained a broad distribution of ESBLs and AmpC β-lactamases. These findings suggested that different molecular mechanisms, including carbapenemase, ESBL and/or AmpC plus loss of porins, have contributed to carbapenem resistance. The <italic>bla</italic><sub>NDM−1</sub>-harboring plasmids contained highly conserved gene environment around <italic>bla</italic><sub>NDM−1</sub> (<italic>bla</italic><sub>NDM−1</sub>-<italic>ble</italic><sub>MBL</sub>-<italic>trpF</italic>-<italic>dsbD</italic>-<italic>cutA1</italic>-<italic>groES</italic>-<italic>groEL</italic>), which could be associated with the potential dissemination of <italic>bla</italic><sub>NDM−1</sub>. IMP-type MBL was located within a variety of integrons and usually contained various gene cassettes encoding multidrug resistance. These isolates produced 54 different pulsotypes, and were classified into 42 STs by MLST. Nineteen <italic>bla</italic><sub>NDM−1</sub>-positive <italic>E. cloacae</italic> isolates obtained from Ningxia had the same pulsotype (PFGE type 1), belonging to ST78 within clonal complex 74 (CC74). The plasmid-based replicon typing indicated that IncX3 plasmids mediated the dissemination of <italic>bla</italic><sub>NDM−1</sub> among these homologous strains. This is the first report on the outbreak of NDM-1-producing <italic>E. cloacae</italic> ST78 with contribution of IncX3 plasmids in Northwestern China. There's an immediate need to intensify surveillance attentively to prevent and control the further spread of NDM-1 in China.</p>