Emergence of an epidemic hypervirulent clone of Enterobacter hormaechei coproducing mcr-9 and carbapenemases

Enterobacter cloacae complex (ECC) is an important nosocomial pathogen encompassing at least 22 species. Some of the species, such as E hormaechei and E asburiae, are clinically prevalent and frequently associated with a multidrug resistance phenotype.1Zhou K Yu W Cao X et al.Characterization of the population structure, drug resistance mechanisms and plasmids of the community-associated Enterobacter cloacae complex in China.J Antimicrob Chemoth. 2018; 73: 66-76Crossref PubMed Scopus (20) Google Scholar ECC is conventionally considered a low-virulence pathogen and often causes infections in immunocompromised patients, although interspecies diversities of pathogenicity have been noted in studies.2Davin-Regli A Lavigne J-P Pagès J-M Enterobacter spp: update on taxonomy, clinical aspects, and emerging antimicrobial resistance.Clin Microbiol Rev. 2019; 32: e00002-e00019Crossref PubMed Scopus (108) Google Scholar A 2021 cohort study uncovered a hypervirulent species of ECC, E bugandensis, which was responsible for fatal septic shocks of newborn babies in a neonatal intensive care unit in France.3Girlich D Ouzani S Emeraud C et al.Uncovering the novel Enterobacter cloacae complex species responsible for septic shock deaths in newborns: a cohort study.Lancet Microbe. 2021; 2: e536-e544Summary Full Text Full Text PDF PubMed Scopus (2) Google Scholar The pathogenicity of the E bugandensis isolates was higher than that of strains from other species evaluated with a Galleria mellonella infection model. In this Comment we report an epidemic hypervirulent clone of E hormaechei showing high invasiveness in a clinical setting. We collected 230 non-repetitive clinical ECC isolates at a tertiary hospital between 2000 and 2018, and two mcr variants (mcr-9 and mcr-10) were detected in 48 of those isolates by PCR screening. 18 sequence types and seven novel sequence trypes were identified in the mcr-producing isolates (MCR-ECC), and ST133 was predominant (20·8%; 10/48), followed by ST418 (8·3%; 4/48), then ST56 (8·3%; 4/48). ST133 and ST418 belong to E hormaechei, and ST56 belongs to E kobei. Multiple carbapenemase genes (12 blaNDM-1, eight blaIMP-4, and one blaIMP-26) were found in 16 of the 48 MCR-ECC isolates, and eight belong to ST133. The ten ST133 isolates were multidrug-resistant, of which eight carbapenemase-producing isolates were resistant to meropenem and imipenem. However, nine isolates showed susceptibility to polymyxin B and colistin (table). 85 ST133 isolates were detected in 3601 ECC genomes retrieved from the National Center for Biotechnology Information (NCBI) Reference Sequence database on October 1, 2021. These isolates were obtained from at least ten countries across five continents, suggestive of an epidemic clone. Of these isolates, 35 (41·2%) harboured mcr-9 and 11 (12·9%) had carbapenemase genes as well as mcr-9 (appendix 1). Four observational studies have reported that ST133 was an endemic carbapenem-resistant clone in multiple countries.4Kananizadeh P Oshiro S Watanabe S et al.Emergence of carbapenem-resistant and colistin-susceptible Enterobacter cloacae complex co-harboring blaIMP-1 and mcr-9 in Japan.BMC Infect Dis. 2020; 20: 282Crossref PubMed Scopus (17) Google Scholar, 5Guillard T Cholley P Limelette A et al.Fluoroquinolone resistance mechanisms and population structure of Enterobacter cloacae non-susceptible to ertapenem in north-eastern France.Front Microbiol. 2015; 61186Crossref PubMed Scopus (21) Google Scholar, 6Kraftova L Finianos M Studentova V et al.Evidence of an epidemic spread of KPC-producing Enterobacterales in Czech hospitals.Sci Rep. 2021; 1115732Crossref PubMed Scopus (3) Google Scholar, 7Sidjabat HE Townell N Nimmo GR et al.Dominance of IMP-4-producing Enterobacter cloacae among carbapenemase-producing Enterobacteriaceae in Australia.Antimicrob Agents Chemother. 2015; 59: 4059-4066Crossref PubMed Scopus (54) Google Scholar The data suggest that coexistence of mcr-9 and carbapenemase genes was prevalent in ST133. Phylogenetic reconstruction of the 85 ST133 genomes with the ten ST133 genomes obtained in this study suggested that the clone has split into two clades, and our isolates were assigned to one clade alongside those isolates from the UK, USA, Germany, Czech Republic, and South Africa (appendix 1).TableInformation of the ten ST133 isolatesYear of isolateSpecimenAge, yearsGenderMinimum inhibitory concentration (mg/L)mcrCarbapenemase genesCAZCXMFEPIPMMEMCIPAMPGENCSLSXTCHLPMBCSTTGCECC452015Catheter83Female>64*Resistant to the drug.>64*Resistant to the drug.>64*Resistant to the drug.4*Resistant to the drug.4*Resistant to the drug.1*Resistant to the drug.>128*Resistant to the drug.128*Resistant to the drug.>128*Resistant to the drug.>320*Resistant to the drug.>128*Resistant to the drug.224*Resistant to the drug.mcr-9.1blaNDM-1, blaIMP-4ECC462015Urine60Female>64*Resistant to the drug.>64*Resistant to the drug.64*Resistant to the drug.4*Resistant to the drug.4*Resistant to the drug.1*Resistant to the drug.>128*Resistant to the drug.128*Resistant to the drug.128*Resistant to the drug.>320*Resistant to the drug.>128*Resistant to the drug.228*Resistant to the drug.mcr-9.1blaNDM-1, blaIMP-4ECC472016Urine74Female>64*Resistant to the drug.>64*Resistant to the drug.>64*Resistant to the drug.16*Resistant to the drug.16*Resistant to the drug.1*Resistant to the drug.>128*Resistant to the drug.128*Resistant to the drug.>128*Resistant to the drug.>320*Resistant to the drug.>128*Resistant to the drug.222*Resistant to the drug.mcr-9.1blaNDM-1, blaIMP-4ECC482016Abdominal fluid65Male>64*Resistant to the drug.>64*Resistant to the drug.64*Resistant to the drug.4*Resistant to the drug.8*Resistant to the drug.1*Resistant to the drug.>128*Resistant to the drug.128*Resistant to the drug.>128*Resistant to the drug.>320*Resistant to the drug.>128*Resistant to the drug.220·5*Resistant to the drug.mcr-9.1blaNDM-1, blaIMP-4ECC522016Cerebrospinal fluid43Male>64*Resistant to the drug.>64*Resistant to the drug.16*Resistant to the drug.4*Resistant to the drug.4*Resistant to the drug.1*Resistant to the drug.>128*Resistant to the drug.>128*Resistant to the drug.>128*Resistant to the drug.>320*Resistant to the drug.>128*Resistant to the drug.220·5*Resistant to the drug.mcr-9.1blaIMP-4ECC542016Drainage39Male>64*Resistant to the drug.>64*Resistant to the drug.64*Resistant to the drug.2*Resistant to the drug.4*Resistant to the drug.2*Resistant to the drug.>128*Resistant to the drug.>128*Resistant to the drug.>128*Resistant to the drug.>320*Resistant to the drug.>128*Resistant to the drug.220·5mcr-9.1blaIMP-4ECC1472005Pancreas40Male64*Resistant to the drug.>64*Resistant to the drug.20·25<0·06251*Resistant to the drug.>128*Resistant to the drug.128*Resistant to the drug.4>320*Resistant to the drug.>128*Resistant to the drug.24*Resistant to the drug.1*Resistant to the drug.mcr-9.1..ECC1882015Blood40Male>64*Resistant to the drug.>64*Resistant to the drug.>64*Resistant to the drug.4*Resistant to the drug.12*Resistant to the drug.>128*Resistant to the drug.>128*Resistant to the drug.>128*Resistant to the drug.>320*Resistant to the drug.>128*Resistant to the drug.0·250·50·5*Resistant to the drug.mcr-9.1blaIMP-4ECC1892015Urine60Female>64*Resistant to the drug.>64*Resistant to the drug.>64*Resistant to the drug.32*Resistant to the drug.4*Resistant to the drug.>16*Resistant to the drug.>128*Resistant to the drug.>128*Resistant to the drug.>128*Resistant to the drug.>320*Resistant to the drug.>128*Resistant to the drug.>32*Resistant to the drug.0·58*Resistant to the drug.mcr-9.1blaNDM-1, blaIMP-4ECC1932016Catheter67Male>64*Resistant to the drug.>64*Resistant to the drug.16*Resistant to the drug.111*Resistant to the drug.>128*Resistant to the drug.128*Resistant to the drug.>128*Resistant to the drug.>320*Resistant to the drug.>128*Resistant to the drug.220·5*Resistant to the drug.mcr-9.1..CAZ=ceftazidime. CXM=cefuroxime. FEP=cefepime. IPM=imipenem. MEM=meropenem. CIP=ciprofloxacin. AMP=ampicillin. GEN=gentamicin. CSL=cefoperazone. SXT=sulfamethoxazole. CHL=chloramphenicol. PMB=polymyxin B. CST=colistin. TGC=tigecycline.* Resistant to the drug. Open table in a new tab CAZ=ceftazidime. CXM=cefuroxime. FEP=cefepime. IPM=imipenem. MEM=meropenem. CIP=ciprofloxacin. AMP=ampicillin. GEN=gentamicin. CSL=cefoperazone. SXT=sulfamethoxazole. CHL=chloramphenicol. PMB=polymyxin B. CST=colistin. TGC=tigecycline. Eight of the ten ST133 isolates were obtained from sterile-site specimens of patients (table), suggestive of an invasive clone. Among the patients positive for ST133, six were men and six were aged 60 years or older. All patients had underlying diseases and received antimicrobial therapy, except for one patient for whom data could not be recovered. A high mortality (40%) was found; four patients died and five patients were discharged. 36 (76·6%) of the 85 ST133 genomes with source information retrieved from NCBI were obtained from sterile-site specimens (appendix 2). In particular, 24 of 39 ST133 isolates in the UK were obtained from blood samples (13 isolates were obtained without source information). Additionally, an outbreak of NDM-1 producing ST133 was reported in a surgical intensive care unit in Croatia, and two of four infected patients died as a result of severe intra-abdominal infections caused by the clone.8Petrosillo N Vranic-Ladavac M Feudi C et al.Spread of Enterobacter cloacae carrying blaNDM-1, blaCTX-M-15, blaSHV-12 and plasmid-mediated quinolone resistance genes in a surgical intensive care unit in Croatia.J Glob Antimicrob Resist. 2016; 4: 44-48Crossref PubMed Scopus (11) Google Scholar Due to the high mortality and invasiveness of ST133, we propose that it could be hypervirulent. The virulence of the ten ST133 isolates collected in this study was further tested with a G mellonella larvae infection model. After being infected for 24 h at an inoculum of 5 × 106 colony forming unit, survival rates of G mellonella were 30% or less for all ST133 isolates (especially 0% for two of them). A survival rate of 0% was recorded for the hypervirulence control strain Klebsiella pneumoniae ATCC43816 (K2 serotype), and a survival rate of 95·8% was recorded for the hypovirulence control strain E cloacae ATCC13047 (appendix 1). These data show that the ST133 clone was hypervirulent. Although multiple epidemic carbapenem-resistant clones of ECC (eg, ST114, ST171, and ST78) have been reported, to our knowledge ST133 is the first carbapenem-resistant hypervirulent clone of ECC frequently causing invasive infections worldwide. The convergence of resistance to last-resort antibiotics and hypervirulence in the species of ECC with clinical significance (ie, E hormaechei) would exacerbate clinical outcomes. We declare no competing interests. This work was supported by the National Key R&D Program of China (2021YFC2300300 and 2022YFE0103200), National Natural Science Foundation of China (82172330 and 81902030), Shenzhen Basic Research Key Project (JCYJ20200109144220704), and Shenzhen Basic Research Project (JCYJ20190807144409307 and JCYJ20210324113608022). The genome sequences have been deposited into GenBank under the bioprojects PRJNA752244 and PRJNA750885. Download .pdf (.72 MB) Help with pdf files Supplementary appendix 1 Download .pdf (.2 MB) Help with pdf files Supplementary appendix 2