Characterization of an Enterobacter cloacae Strain Producing both KPC and NDM Carbapenemases by Whole-Genome Sequencing

ABSTRACTA carbapenem-resistantEnterobacter cloacaestrain, WCHECl-14653, causing a fatal bloodstream infection, was characterized by genome sequencing and conjugation experiments. The strain carried two carbapenemase genes,blaNDM-1andblaKPC-2, on separate IncF plasmids. The coexistence ofblaNDM-1andblaKPC-2conferred slightly higher-level carbapenem resistance compared with that ofblaNDM-1orblaKPC-2alone, and the coexistence of two IncF plasmids may generate new platforms for spreading carbapenemase genes.

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Genomically Informed Surveillance for Carbapenem-Resistant Enterobacteriaceae in a Health Care System

mBio ◽

10.1128/mbio.01030-15 ◽

2015 ◽

Vol 6 (4) ◽

Cited By ~ 60

Author(s):

Nicole D. Pecora ◽

Ning Li ◽

Marc Allard ◽

Cong Li ◽

Esperanza Albano ◽

...

Keyword(s):

Health Care ◽

Drug Resistance ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Resistance Genes ◽

Enterobacter Cloacae ◽

Mobile Elements ◽

Whole Genome ◽

Content Type ◽

Carbapenem Resistant

ABSTRACT Carbapenem-resistant Enterobacteriaceae (CRE) are an urgent public health concern. Rapid identification of the resistance genes, their mobilization capacity, and strains carrying them is essential to direct hospital resources to prevent spread and improve patient outcomes. Whole-genome sequencing allows refined tracking of both chromosomal traits and associated mobile genetic elements that harbor resistance genes. To enhance surveillance of CREs, clinical isolates with phenotypic resistance to carbapenem antibiotics underwent whole-genome sequencing. Analysis of 41 isolates of Klebsiella pneumoniae and Enterobacter cloacae, collected over a 3-year period, identified K. pneumoniae carbapenemase (KPC) genes encoding KPC-2, −3, and −4 and OXA-48 carbapenemases. All occurred within transposons, including multiple Tn4401 transposon isoforms, embedded within more than 10 distinct plasmids representing incompatibility (Inc) groups IncR, -N, -A/C, -H, and -X. Using short-read sequencing, draft maps were generated of new KPC-carrying vectors, several of which were derivatives of the IncN plasmid pBK31551. Two strains also had Tn4401 chromosomal insertions. Integrated analyses of plasmid profiles and chromosomal single-nucleotide polymorphism (SNP) profiles refined the strain patterns and provided a baseline hospital mobilome to facilitate analysis of new isolates. When incorporated with patient epidemiological data, the findings identified limited outbreaks against a broader 3-year period of sporadic external entry of many different strains and resistance vectors into the hospital. These findings highlight the utility of genomic analyses in internal and external surveillance efforts to stem the transmission of drug-resistant strains within and across health care institutions. IMPORTANCE We demonstrate how detection of resistance genes within mobile elements and resistance-carrying strains furthers active surveillance efforts for drug resistance. Whole-genome sequencing is increasingly available in hospital laboratories and provides a powerful and nuanced means to define the local landscape of drug resistance. In this study, isolates of Klebsiella pneumoniae and Enterobacter cloacae with resistance to carbapenem antibiotics were sequenced. Multiple carbapenemase genes were identified that resided in distinct transposons and plasmids. This mobilome, or population of mobile elements capable of mobilizing drug resistance, further highlighted the degree of strain heterogeneity while providing a detailed timeline of carbapenemase entry into the hospital over a 3-year period. These surveillance efforts support effective targeting of infection control resources and the development of institution-specific repositories of resistance genes and the mobile elements that carry them.

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Putative Integrative Mobile Elements That Exploit the Xer Recombination Machinery Carrying blaIMI-Type Carbapenemase Genes in Enterobacter cloacae Complex Isolates in Singapore

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01542-17 ◽

2017 ◽

Vol 62 (1) ◽

Cited By ~ 6

Author(s):

Tse H. Koh ◽

Nurdyana Binte Abdul Rahman ◽

Jeanette W. P. Teo ◽

My-Van La ◽

Balamurugan Periaswamy ◽

...

Keyword(s):

Phylogenetic Analysis ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Multilocus Sequence Typing ◽

Enterobacter Cloacae ◽

Mobile Elements ◽

Whole Genome ◽

Content Type ◽

Flanking Regions ◽

Enterobacter Cloacae Complex

ABSTRACT Whole-genome sequencing was performed on 16 isolates of the carbapenemase-producing Enterobacter cloacae complex to determine the flanking regions of bla IMI-type genes. Phylogenetic analysis of multilocus sequence typing (MLST) targets separated the isolates into 4 clusters. The bla IMI-type genes were all found on Xer-dependent integrative mobile elements (IMEX). The IMEX elements of 5 isolates were similar to those described in Canada, while the remainder were novel. Five isolates had IMEX elements lacking a resolvase and recombinase.

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Enterobacter cloacae Complex Sequence Type 171 Isolates Expressing KPC-4 Carbapenemase Recovered from Canine Patients in Ohio

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01161-18 ◽

2018 ◽

Vol 62 (12) ◽

Cited By ~ 2

Author(s):

Joshua B. Daniels ◽

Liang Chen ◽

Susan V. Grooters ◽

Dixie F. Mollenkopf ◽

Dimitria A. Mathys ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Companion Animals ◽

Enterobacter Cloacae ◽

Sequence Type ◽

Resistant Bacteria ◽

Whole Genome ◽

Content Type ◽

Complex Sequence ◽

Enterobacter Cloacae Complex

ABSTRACT Companion animals are likely relevant in the transmission of antimicrobial-resistant bacteria. Enterobacter xiangfangensis sequence type 171 (ST171), a clone that has been implicated in clusters of infections in humans, was isolated from two dogs with clinical disease in Ohio. The canine isolates contained IncHI2 plasmids encoding blaKPC-4. Whole-genome sequencing was used to put the canine isolates in phylogenetic context with available human ST171 sequences, as well as to characterize their blaKPC-4 plasmids.

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Characterization of SXT/R391 Integrative and Conjugative Elements in Proteus mirabilis Isolates from Food-Producing Animals in China

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02852-15 ◽

2016 ◽

Vol 60 (3) ◽

pp. 1935-1938 ◽

Cited By ~ 25

Author(s):

Chang-Wei Lei ◽

An-Yun Zhang ◽

Hong-Ning Wang ◽

Bi-Hui Liu ◽

Li-Qin Yang ◽

...

Keyword(s):

Escherichia Coli ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Resistance Genes ◽

Proteus Mirabilis ◽

Whole Genome ◽

Content Type ◽

Integrative And Conjugative Elements ◽

Animal Farms

SXT/R391 integrative and conjugative elements (ICEs) were detected in 8 out of 125Proteus mirabilisisolates from food-producing animals in China. Whole-genome sequencing revealed that seven ICEs were identical to ICEPmiJpn1, carrying the cephalosporinase geneblaCMY-2. Another one, designated ICEPmiChn1, carried five resistance genes. All eight ICEs could be transferred toEscherichia colivia conjugation. The results highlight the idea that animal farms are important reservoir of the SXT/R391 ICE-containingP. mirabilis.

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Deciphering the resistome of the widespreadP. aeruginosaST175 international high-risk clone through whole genome sequencing

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01720-16 ◽

2016 ◽

pp. AAC.01720-16 ◽

Cited By ~ 14

Author(s):

Gabriel Cabot ◽

Carla López-Causapé ◽

Alain A. Ocampo-Sosa ◽

Lea M. Sommer ◽

María Ángeles Domínguez ◽

...

Keyword(s):

High Risk ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Carbapenem Resistance ◽

Whole Genome ◽

Resistance Mutations ◽

Online Databases ◽

Genes Encoding ◽

Resistance Traits

Whole genome sequencing (WGS) was used for the characterization of the, frequently extensively-drug resistant (XDR),P. aeruginosahigh-risk clone ST175. A total of eighteen ST175 isolates recovered from 8 different Spanish hospitals were analyzed; four isolates from four different French hospitals were included for comparison. The typical resistance profile of ST175 included penicillins, cephalosporins, monobactams, carbapenems, aminoglycosides, and fluoroquinolones. In the phylogenetic analysis, the four French isolates clustered together with the two isolates from one of the Spanish regions. Sequence variation was analyzed for 146 chromosomal genes related to antimicrobial resistance and horizontally-acquired genes were explored using online databases. The resistome of ST175 was mainly determined by mutational events, with resistance traits common to all or nearly all of the strains, including specificampRmutations leading toampCoverexpression, specific mutations inoprDconferring carbapenem resistance or amexZmutation leading to MexXY overexpression. All isolates additionally harbored anaadBgene conferring gentamicin and tobramycin resistance. Several other resistance traits were specific to certain geographic areas such as a streptomycin resistanceaadA13gene detected in all four isolates from France and in the 2 isolates from the Cantabria region or aglpTmutation conferring fosfomycin resistance detected in all but these six isolates. Finally, several unique resistance mutations were detected in single isolates; particularly interesting among them were those in genes encoding PBPs (PBP1A, PBP3 and PBP4). Thus, these results provide valuable information for understanding the genetic basis of resistance and the dynamics of dissemination and evolution of high-risk clones.

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Genomic Characterization of Multidrug-Resistant Carbapenemase-Producing Enterobacter cloacae ECL189, Co-producing KPC-2, NDM-1, TEM-1, TEM-95, and SHV-66

Jundishapur Journal of Microbiology ◽

10.5812/jjm.105761 ◽

2021 ◽

Vol 13 (11) ◽

Author(s):

Na Du ◽

Shumin Liu ◽

Jing Yao ◽

Kai Yang ◽

Yun Lin ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Resistance Genes ◽

Virulence Genes ◽

Enterobacter Cloacae ◽

Biological Information ◽

Whole Genome ◽

Carbapenem Resistant ◽

Transmission Mechanisms ◽

Antimicrobial Resistance Genes

Background: Carbapenem-resistant Enterobacteriaceae (CRE) has become a public health threat due to resistance to multiple antibiotics. The production of β-lactamase is the most important resistance mechanism of Enterobacteriaceae. Although isolates producing KPC-2 or NDM-1 enzymes have been reported widely, isolates co-producing KPC-2, NDM-1, TEM-1, TEM-95, SHV-66, and other β-lactamases have rarely been detected in the same strain, especially in Enterobacter cloacae. Objectives: In this study, we identified and sequenced the genome of carbapenem-resistant E. cloacae ECL189 to in-depth analyze the resistance and transmission mechanisms of E. cloacae. Methods: We investigated the antimicrobial susceptibility of ECL189 by a VITEK 2 system, E-test gradient strips, and K-B method. Whole-genome sequencing was used by the PacBio RS II platform and Illumina HiSeq 4000 platform. Antimicrobial resistance genes, virulence genes, non-coding RNA, and repeat sequences were predicted by biological information databases. A PCR was used to further confirm that the blaKPC-2, blaNDM-1, blaTEM-1, blaTEM-95, and blaSHV-66 genes existed in ECL189. A conjugation experiment was performed to determine the transferability of resistance. Molecular typing of ECL189 was done by multilocus sequence typing (MLST). Results: Enterobacter cloacae ECL189 was resistant to 21 out of 23 tested antibiotics, but its transconjugant was resistant to 10 out of 18 tested antibiotics. The genome of ECL189 consisted of a 5,026,406 bp chromosome and four circular plasmids. In total, 26 resistance genes and 58 resistance proteins were identified. In addition, 77 determinants associated with bacterial virulence were identified. A large number of resistance and virulence genes were located in the plasmids. The results of whole-genome sequencing were consistent with the β-lactamase genes. The MLST analysis revealed that this strain belonged to ST74. Conclusions: This study further revealed the resistance, virulence, and transmission mechanisms of carbapenem-resistant E. cloacae. Resistance and virulence genes spread in bacteria by the horizontal transfer of plasmids, which should attract more attention in relevant departments.

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Molecular characterization of rectal isolates of carbapenemase-negative carbapenem-resistant enterobacterales obtained from ICU patients in Kuwait by whole-genome sequencing

Journal of Medical Microbiology ◽

10.1099/jmm.0.001409 ◽

2021 ◽

Vol 70 (9) ◽

Author(s):

Amani Al Fadhli ◽

Wafaa Jamal ◽

Vincent O. Rotimi

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Carbapenem Resistance ◽

Efflux Pumps ◽

Multidrug Resistant ◽

Whole Genome ◽

Genetic Characteristics ◽

Carbapenem Resistant ◽

Genes Encoding ◽

Amoxicillin Clavulanic Acid

Introduction. Carbapenem-resistant enterobacterales (CRE) are listed among the most urgent antibiotic resistance threats. Hypothesis. Previous studies on the mechanisms of CRE in Kuwait have focused on carbapenemases. There have been no studies on non-carbapenemase-producing CRE in Kuwait. Aim/Gap Statement. The aim of this study was to investigate the genetic characteristics of non-carbapenemase-producing carbapenem-resistant enterobacterales (NCPE) isolates using whole-genome sequencing (WGS). Methodology. Fourteen confirmed NCPE isolates that were negative for genes encoding carbapenemase production by polymerase chain reaction (PCR) assays using rectal swabs from intensive care unit patients were characterized using phenotypic, PCR and WGS methods. Susceptibility testing was performed via Etest and clonality via multi-locus sequence typing (MLST). Results. All of the isolates were resistant to ertapenem; 78.6 % were resistant to imipenem, meropenem and trimethoprim–sulfamethoxazole. Resistance to the other antibiotics was variable, ranging from 28.5 (colistin) through 50 (tigecycline) and 64.3 (amikacin) up to 85.7 % against both amoxicillin–clavulanic acid and ciprofloxacin. WGS detected several resistance genes mediating the production of β-lactamases, genes encoding an outer-membrane porin permeability mutation resulting in reduced susceptibility to β-lactams, including carbapenems, and genes for multidrug-resistant (MDR) efflux pumps. The isolates also possessed global activator protein MarA, which mediated reduced permeability to β-lactams. The existence of β-lactamase genes, overexpression of MDR efflux pumps and reduced permeability mediated by the porin genes were responsible for carbapenem resistance. Conclusions. This finding reflects the superior detection capabilities offered by WGS analysis, which can be used to complement traditional methods and overcome their limited resolution in clinical settings.

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MCR-1 and OXA-48 In Vivo Acquisition in KPC-Producing Escherichia coli after Colistin Treatment

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02540-16 ◽

2017 ◽

Vol 61 (8) ◽

Cited By ~ 32

Author(s):

Racha Beyrouthy ◽

Frederic Robin ◽

Aude Lessene ◽

Igor Lacombat ◽

Laurent Dortet ◽

...

Keyword(s):

Escherichia Coli ◽

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Whole Genome ◽

Content Type ◽

First Report ◽

E Coli ◽

Carbapenem Resistant

ABSTRACT The spread of mcr-1-encoding plasmids into carbapenem-resistant Enterobacteriaceae raises concerns about the emergence of untreatable bacteria. We report the acquisition of mcr-1 in a carbapenem-resistant Escherichia coli strain after a 3-week course of colistin in a patient repatriated to France from Portugal. Whole-genome sequencing revealed that the Klebsiella pneumoniae carbapenemase-producing E. coli strain acquired two plasmids, an IncL OXA-48-encoding plasmid and an IncX4 mcr-1-encoding plasmid. This is the first report of mcr-1 in carbapenemase-encoding bacteria in France.

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Whole-Genome Sequencing Overrules a Suspected Case of Carbapenem-Resistant Enterobacter cloacae Transmission

Journal of Clinical Microbiology ◽

10.1128/jcm.00915-17 ◽

2017 ◽

Vol 55 (9) ◽

pp. 2868-2870 ◽

Cited By ~ 1

Author(s):

Mark Chen ◽

Sean Conlan ◽

Anna F. Lau ◽

John P. Dekker ◽

Clay Deming ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Enterobacter Cloacae ◽

Suspected Case ◽

Whole Genome ◽

Carbapenem Resistant

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Utility of Whole-Genome Sequencing in Characterizing Acinetobacter Epidemiology and Analyzing Hospital Outbreaks

Journal of Clinical Microbiology ◽

10.1128/jcm.01818-15 ◽

2015 ◽

Vol 54 (3) ◽

pp. 593-612 ◽

Cited By ~ 44

Author(s):

Margaret A. Fitzpatrick ◽

Egon A. Ozer ◽

Alan R. Hauser

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Phylogenetic Trees ◽

Acinetobacter Calcoaceticus ◽

Whole Genome ◽

Strain Typing ◽

Nucleotide Polymorphisms ◽

Content Type ◽

Carbapenem Resistant ◽

Outbreak Investigations

Acinetobacter baumanniifrequently causes nosocomial infections and outbreaks. Whole-genome sequencing (WGS) is a promising technique for strain typing and outbreak investigations. We compared the performance of conventional methods with WGS for strain typing clinicalAcinetobacterisolates and analyzing a carbapenem-resistantA. baumannii(CRAB) outbreak. We performed two band-based typing techniques (pulsed-field gel electrophoresis and repetitive extragenic palindromic-PCR), multilocus sequence type (MLST) analysis, and WGS on 148Acinetobacter calcoaceticus-A. baumanniicomplex bloodstream isolates collected from a single hospital from 2005 to 2012. Phylogenetic trees inferred from core-genome single nucleotide polymorphisms (SNPs) confirmed threeAcinetobacterspecies within this collection. Four majorA. baumanniiclonal lineages (as defined by MLST) circulated during the study, three of which are globally distributed and one of which is novel. WGS indicated that a threshold of 2,500 core SNPs accurately distinguishedA. baumanniiisolates from different clonal lineages. The band-based techniques performed poorly in assigning isolates to clonal lineages and exhibited little agreement with sequence-based techniques. After applying WGS to a CRAB outbreak that occurred during the study, we identified a threshold of 2.5 core SNPs that distinguished nonoutbreak from outbreak strains. WGS was more discriminatory than the band-based techniques and was used to construct a more accurate transmission map that resolved many of the plausible transmission routes suggested by epidemiologic links. Our study demonstrates that WGS is superior to conventional techniques forA. baumanniistrain typing and outbreak analysis. These findings support the incorporation of WGS into health care infection prevention efforts.

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