Molecular Characterization of IMP-1-Producing Enterobacter cloacae Complex Isolates in Tokyo

Abstract

Although KPC enzymes are most common among carbapenemases produced by Enterobacter cloacae complex globally, the epidemiology varies from one country to another. While previous studies have suggested that IMP enzymes are most common in Japan, detailed analysis has been scarce thus far. Here, we carried out a molecular epidemiological study and plasmid analysis of IMP-1-producing E. cloacae complex isolates collected from three hospitals in central Tokyo using whole-genome sequencing. Seventy-one isolates were classified into several sequence types (STs), and 49 isolates were identified as Enterobacter hormaechei ST78. Isolates of ST78 were divided into three clades by core-genome single nucleotide polymorphism (SNP)-based phylogenetic analysis. Whereas isolates of clade 3 were isolated from only one hospital, isolates of clade 1 and 2 were identified from multiple hospitals. Ten of 12 clade 1 isolates and 1 of 4 clade 2 isolates carried bla_IMP-1 on IncHI2 plasmids, with high similarity of genetic structures. In addition, these plasmids shared backbone structures with IncHI2 plasmids carrying bla_IMP reported from other countries of the Asia-Pacific region. All isolates of clade 3 except one carried bla_IMP-1 in In1426 on IncW plasmids. An isolate of clade 3, which lacked IncW plasmids, carried bla_IMP-1 in In1426 on an IncFIB plasmid. These observations suggest that IMP-producing E. cloacae complex isolates with a diversity of host genomic backgrounds have spread in central Tokyo, and they indicate the possible contribution of IncHI2 plasmids toward this phenomenon.

Keywords: Enterobacter cloacae complex; IMP-1; carbapenemase-producing Enterobacteriaceae; metallo-β-lactamase; plasmid; whole-genome sequencing.

FIG 1

Phylogenetic tree of IMP-1-producing Enterobacter hormaechei ST78 isolates (n = 49; TUM prefixes have been removed for clarity) constructed with maximum-likelihood phylogenetic analysis based on single nucleotide polymorphisms (SNPs) in the core genome and excluding homologous recombination sequences. The core-genome region was 86.9% (4,027,037/4,633,407 bp) of the genome of the reference strain, E. cloacae ECNIH3 ST97. The scale distance corresponds to the number of substitutions per site.

FIG 2

Comparison of pMTY11043_IncHI2 (GenBank accession number AP018352.1) carrying bla_IMP-1 with pEC-IMP (GenBank accession number EU855787) and pIMP4-SEM (KX810825) drawn with EasyFig, version 2.1. These IncHI2 plasmids belong to pMLST-ST1. Block arrows indicate confirmed or putative open reading frames (ORFs) and their orientations. Arrow size is proportional to the predicted ORF length. The color code is as follows: green, replication initiation protein genes; blue, conjugal transfer genes; cyan, transposase genes; yellow, heavy metal resistance genes; red, integrase genes; magenta, antibiotic resistance genes; orange, toxin-antitoxin system genes. Putative, hypothetical, and unknown genes are represented by gray arrows.

FIG 3

Comparison of the nucleotide sequences of pMTY10660_IncW harboring bla_IMP-1 (GenBank accession number AP018350.1) and pR388 (BR000038). The color code is the same as that described in the legend of Fig. 2.

FIG 4

Structural features of In1426, a class 1 integron containing bla_IMP-1, and surrounding nucleotide sequences in pMTY10695_IncFIB (GenBank accession number AP018351.1). The insertion sequence IS6100 was located downstream of In1426. The central part of the transposase gene of ISPa38 (2,154 bp/2,967 bp) was lacking. The missing region is in orange. IRR, inverted repeat, right; IRL, inverted repeat, left.