Dataset Information

Plasmid diversity among genetically related Klebsiella pneumoniae blaKPC-2 and blaKPC-3 isolates collected in the Dutch national surveillance.

ABSTRACT: Carbapenemase-producing Klebsiella pneumoniae emerged as a nosocomial pathogen causing morbidity and mortality in patients. For infection prevention it is important to track the spread of K. pneumoniae and its plasmids between patients. Therefore, the major aim was to recapitulate the contents and diversity of the plasmids of genetically related K. pneumoniae strains harboring the beta-lactamase gene blaKPC-2 or blaKPC-3 to determine their dissemination in the Netherlands and the former Dutch Caribbean islands from 2014 to 2019. Next-generation sequencing was combined with long-read third-generation sequencing to reconstruct 22 plasmids. wgMLST revealed five genetic clusters comprised of K. pneumoniae blaKPC-2 isolates and four clusters consisted of blaKPC-3 isolates. KpnCluster-019 blaKPC-2 isolates were found both in the Netherlands and the Caribbean islands, while blaKPC-3 cluster isolates only in the Netherlands. Each K. pneumoniae blaKPC-2 or blaKPC-3 cluster was characterized by a distinct resistome and plasmidome. However, the large and medium plasmids contained a variety of antibiotic resistance genes, conjugation machinery, cation transport systems, transposons, toxin/antitoxins, insertion sequences and prophage-related elements. The small plasmids carried genes implicated in virulence. Thus, implementing long-read plasmid sequencing analysis for K. pneumoniae surveillance provided important insights in the transmission of a KpnCluster-019 blaKPC-2 strain between the Netherlands and the Caribbean.

SUBMITTER: Hendrickx APA

PROVIDER: S-EPMC7546619 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Plasmid diversity among genetically related Klebsiella pneumoniae blaKPC-2 and blaKPC-3 isolates collected in the Dutch national surveillance.

Hendrickx Antoni P A APA Landman Fabian F de Haan Angela A Borst Dyogo D Witteveen Sandra S van Santen-Verheuvel Marga G MG van der Heide Han G J HGJ Schouls Leo M LM

Scientific reports 20201008 1

Carbapenemase-producing Klebsiella pneumoniae emerged as a nosocomial pathogen causing morbidity and mortality in patients. For infection prevention it is important to track the spread of K. pneumoniae and its plasmids between patients. Therefore, the major aim was to recapitulate the contents and diversity of the plasmids of genetically related K. pneumoniae strains harboring the beta-lactamase gene blaKPC-2 or blaKPC-3 to determine their dissemination in the Netherlands a ...[more]

PMID: 33033293

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Project description:Ceftazidime-avibactam (CZA) therapy has significantly improved survival rates for patients infected by carbapenem-resistant bacteria, including KPC producers. However, resistance to CZA is a growing concern, attributed to multiple mechanisms. In this study, we characterized four clinical CZA-resistant Klebsiella pneumoniae isolates obtained between July 2019 and December 2020. These isolates expressed novel allelic variants of blaKPC-2 resulting from changes in hotspots of the mature protein, particularly in loops surrounding the active site of KPC. Notably, KPC-80 had an K269_D270insPNK mutation near the Lys270-loop, KPC-81 had a del_I173 mutation within the Ω-loop, KPC-96 showed a Y241N substitution within the Val240-loop and KPC-97 had an V277_I278insNSEAV mutation within the Lys270-loop. Three of the four isolates exhibited low-level resistance to imipenem (4 µg/mL), while all remained susceptible to meropenem. Avibactam and relebactam effectively restored carbapenem susceptibility in resistant isolates. Cloning mutant blaKPC genes into pMBLe increased imipenem MICs in recipient Escherichia coli TOP10 for blaKPC-80, blaKPC-96, and blaKPC-97 by two dilutions; again, these MICs were restored by avibactam and relebactam. Frameshift mutations disrupted ompK35 in three isolates. Additional resistance genes, including blaTEM-1, blaOXA-18 and blaOXA-1, were also identified. Interestingly, three isolates belonged to clonal complex 11 (ST258 and ST11) and one to ST629. This study highlights the emergence of CZA resistance including unique allelic variants of blaKPC-2 and impermeability. Comprehensive epidemiological surveillance and in-depth molecular studies are imperative for understanding and monitoring these complex resistance mechanisms, crucial for effective antimicrobial treatment strategies.ImportanceThe emergence of ceftazidime-avibactam (CZA) resistance poses a significant threat to the efficacy of this life-saving therapy against carbapenem-resistant bacteria, particularly Klebsiella pneumoniae-producing KPC enzymes. This study investigates four clinical isolates exhibiting resistance to CZA, revealing novel allelic variants of the key resistance gene, blaKPC-2. The mutations identified in hotspots surrounding the active site of KPC, such as K269_D270insPNK, del_I173, Y241N and V277_I278insNSEAV, prove the adaptability of these pathogens. Intriguingly, low-level resistance to imipenem and disruptions in porin genes were observed, emphasizing the complexity of the resistance mechanisms. Interestingly, three of four isolates belonged to clonal complex 11. This research not only sheds light on the clinical significance of CZA resistance but also shows the urgency for comprehensive surveillance and molecular studies to inform effective antimicrobial treatment strategies in the face of evolving bacterial resistance.

Project description:The emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP) isolates in Egyptian hospitals has been reported. However, the genetic basis and analysis of the plasmids associated with carbapenem-resistant hypervirulent K. pneumoniae (CR-HvKP) in Egypt have not been presented. Therefore, we attempted to decipher the plasmid sequences that are responsible for transferring the determinants of carbapenem resistance, particularly bla NDM-1 and bla KPC-2 Out of 34 K. pneumoniae isolates collected from two tertiary hospitals in Egypt, 31 were CRKP. Whole-genome sequencing revealed that our isolates were related to 13 different sequence types (STs). The most prevalent ST was ST101, followed by ST383 and ST11. Among the CRKP isolates, one isolate named EBSI036 has been reassessed by Nanopore sequencing. Genetic environment analysis showed that EBSI036 carried 20 antibiotic resistance genes and was identified as a CR-HvKP strain: it harbored four plasmids, namely, pEBSI036-1-NDM-VIR, pEBSI036-2-KPC, pEBSI036-3, and pEBSI036-4. The two carbapenemase genes bla NDM-1 and bla KPC-2 were located on plasmids pEBSI036-1-NDM-VIR and pEBSI036-2-KPC, respectively. The IncFIB:IncHI1B hybrid plasmid pEBSI036-1-NDM-VIR also carried some virulence factors, including the regulator of the mucoid phenotype (rmpA), the regulator of mucoid phenotype 2 (rmpA2), and aerobactin (iucABCD and iutA). Thus, we set out in this study to analyze in depth the genetic basis of the pEBSI036-1-NDM-VIR and pEBSI036-2-KPC plasmids. We report a high-risk clone ST11 KL47 serotype of a CR-HvKP strain isolated from the blood of a 60-year-old hospitalized female patient from the intensive care unit (ICU) in a tertiary care hospital in Egypt, which showed the cohabitation of a novel hybrid plasmid coharboring the bla NDM-1 and virulence genes and a bla KPC-2-carrying plasmid.IMPORTANCE CRKP has been registered in the critical priority tier by the World Health Organization and has become a significant menace to public health. The emergence of CR-HvKP is of great concern in terms of both disease and treatment. In-depth analysis of the carbapenemase-encoding and virulence plasmids may provide insight into ongoing recombination and evolution of virulence and multidrug resistance in K. pneumoniae Thus, this study serves to alert contagious disease clinicians to the presence of hypervirulence in CRKP isolates in Egyptian hospitals.

Dataset Information

Plasmid diversity among genetically related Klebsiella pneumoniae blaKPC-2 and blaKPC-3 isolates collected in the Dutch national surveillance.

Publications

Plasmid diversity among genetically related Klebsiella pneumoniae bla<sub>KPC-2</sub> and bla<sub>KPC-3</sub> isolates collected in the Dutch national surveillance.

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