In this work, by high-throughput sequencing, antibiotic resistance genes, including class A (bla _CTX-M, bla _Z, bla _TEM, bla _VEB, bla _KLUC, and bla _SFO), class C (bla _SHV, bla _DHA, bla _MIR, bla _AZECL-29, and bla _ACT), and class D (bla _OXA) β-lactamase genes, were identified among the pooled genomic DNA from 212 clinical Enterobacter cloacae isolates. Six bla _MIR-positive E. cloacae strains were identified, and pulsed-field gel electrophoresis (PFGE) showed that these strains were not clonally related. The complete genome of the bla _MIR -positive strain (Y546) consisted of both a chromosome (4.78 Mb) and a large plasmid pY546 (208.74 kb). The extended-spectrum β-lactamases (ESBLs) (bla _SHV-12 and bla _CTX-M-9a) and AmpC (bla _MIR) were encoded on the chromosome, and the pY546 plasmid contained several clusters of genes conferring resistance to metals, such as copper (pco), arsenic (ars), tellurite (ter), and tetrathionate (ttr), and genes encoding many divalent cation transporter proteins. The comparative genomic analyses of the whole plasmid sequence and of the heavy metal resistance gene-encoding regions revealed that the plasmid sequences of Klebsiella pneumoniae (such as pKPN-332, pKPN-3967, and pKPN-262) shared the highest similarity with those of pY546. It may be concluded that a variety of β-lactamase genes present in E. cloacae which confer resistance to β-lactam antibiotics and the emergence of plasmids carrying heavy metal resistance genes in clinical isolates are alarming and need further surveillance.

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