Comprehensive Analysis of Virulence Determinants and Genomic Islands of blaNDM-1-Producing Enterobacter hormaechei Clinical Isolates from Greece

Abstract

Nosocomial outbreaks of multidrug-resistant (MDR) Enterobacter cloacae complex (ECC) are often reported worldwide, mostly associated with a small number of multilocus-sequence types of E. hormaechei and E. cloacae strains. In Europe, the largest clonal outbreak of blaNDM-1-producing ECC has been recently reported, involving an ST182 E. hormaechei strain in a Greek teaching hospital. In the current study, we aimed to further investigate the genetic make-up of two representative outbreak isolates. Comparative genomics of whole genome sequences (WGS) was performed, including whole genome-based taxonomic analysis and in silico prediction of virulence determinants of the bacterial cell surface, plasmids, antibiotic resistance genes and virulence factors present on genomic islands. The enterobacterial common antigen and the colanic antigen of the cell surface were identified in both isolates, being similar to the gene clusters of the E. hormaechei ATCC 49162 and E. cloacae ATCC 13047 type strains, whereas the two strains possessed different gene clusters encoding lipopolysaccharide O-antigens. Other virulence factors of the bacterial cell surface, such as flagella, fimbriae and pili, were also predicted to be encoded by gene clusters similar to those found in Enterobacter spp. and other Enterobacterales. Secretion systems and toxin–antitoxin systems, which also contribute to pathogenicity, were identified. Both isolates harboured resistance genes to multiple antimicrobial classes, including β-lactams, aminoglycosides, quinolones, chloramphenicol, trimethoprim, sulfonamides and fosfomycin; they carried blaTEM-1, blaOXA-1, blaNDM-1, and one of them also carried blaCTXM-14, blaCTXM-15 and blaLAP-2 plasmidic alleles. Our comprehensive analysis of the WGS assemblies revealed that blaNDM-1-producing outbreak isolates possess components of the bacterial cell surface as well as genomic islands, harbouring resistance genes to several antimicrobial classes and various virulence factors. Differences in the plasmids carrying β-lactamase genes between the two strains have also shown diverse modes of acquisition and an ongoing evolution of these mobile elements.

Comparative Genomics of an Emerging Multidrug-Resistant blaNDM-Carrying ST182 Lineage in Enterobacter cloacae Complex

“…The EC-ML-559 strain from Greece (cluster C) carried a bla NDM-1 -harboring structure ( Tables S3 and S5 ) found in Klebsiella pneumoniae subsp. pneumoniae strain KPX plasmid pKPX-1 from Taiwan [ 15 , 33 ] and Enterobacter hormaechei subsp. xiangfangensis strain ST114 plasmid pLAU_ENM30_NDM1 from Lebanon [ 13 ].…”

“…The conjugative regions ( oriT , relaxase gene, T4CP gene and T4SS gene cluster) of the self-transmissible MGEs were characterized for the ST182 strains that have caused the outbreaks in the Czech Republic [ 11 , 12 ] and Greece [ 14 , 15 ]. In the bla NDM-1 -harboring strain EC-ML559 from Greece (Biosample SAMN33955250), all four conjugative regions were predicted: the oriT (region: 13,705–13,786), the relaxase gene (region: 14,142–16,070), the T4CP gene (region: 19,453–21,645) and the T4SS gene cluster (region: 19,453–44,454).…”

“…Lately, the largest European dissemination of ECC NDM producers has been reported in Greece and the outbreak was caused by an ST182 clonal strain [ 14 ]. Whole-genome taxonomic analysis of two bla NDM-1 -producing strains recovered during the outbreak (EC-ML559 of MLST ST182 and EC-ML621 of ST2143, a single-locus variant of ST182) revealed that both strains were assigned as E. hormaechei [ 15 ]. In silico prediction of components of the bacterial cell surface and genomic islands showed the presence of various virulence factors and resistance genes to several antimicrobial classes, as well as differences in the plasmids carrying β-lactamase genes [ 15 ].…”

Comparative Genomics of an Emerging Multidrug-Resistant blaNDM-Producing ST182 Lineage in Enterobacter cloacae complex

“…The EC-ML-559 strain from Greece (cluster C) carried a blaNDM-1-harbouring structure found in Klebsiellapneumoniae subsp. pneumoniae strain KPX plasmid pKPX-1from Taiwan [14,43] and Enterobacter hormaechei subsp. xiangfangensis strain ST114 plasmid pLAU_ENM30_NDM1 from Lebanon [12].Finally, blaNDM-1-harboring contigs of strain RIVM_C015180 from the Netherlands and strain E472 from Singapore, were similar to plasmids pM214_AC2 (IncA/C) [36] and pJN24NDM (IncN2) [37] types, respectively, which have been previously described in blaNDM-harbouring plasmids of E. coli (Figure S4).…”

“…Lately, the largest European dissemination of ECC NDM-producers has been reported in Greece and the outbreak was caused by a ST182 clonal strain [13]. Whole-genome taxonomic analysis of two blaNDM-1-producing strains recovered during the outbreak (EC-ML559 of MLST ST182 and EC-ML621 of ST2143, a single locus variant of ST182) revealed that both strains were assigned as E. hormaechei [14]. In silico prediction of components of the bacterial cell surface and genomic islands showed the presence of various virulence factors and resistance genes to several antimicrobial classes, as well as differences in the plasmids carrying β-lactamase genes [14].…”