Whole Genome Sequencing of Carbapenem-Resistant
            <i>Klebsiella Pneumoniae</i>
            : Evolutionary Analysis for Outbreak Investigation

Fontana, Carla; Angeletti, Silvia; Mirandola, Walter; Cella, Eleonora; Lai, Alessia; Zehender, Gianguglielmo; Favaro, Marco; Leoni, Davide; Rose, D. Delle; Gherardi, Giovanni; Florio, Lucía; Salemi, Marco; Andreoni, Massimo; Sarmati, Loredana; Ciccozzi, Massimo

doi:10.2217/fmb-2019-0074

Cited by 10 publications

(7 citation statements)

References 50 publications

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“…Many studies of CPE outbreak investigations have focused on clonal transmission, relying upon single-nucleotide polymorphism distances indicative of clonal bacterial spread to determine genomically-linked transmission 10 , 18 , 19 . As plasmid-mediated transmission accounts for a significant prevalence of CPE transmission at a population-level, plasmid identification and linkage would be necessary for accurate CPE outbreak investigation 20 .…”

Section: Discussionmentioning

confidence: 99%

Whole genome sequencing reveals hidden transmission of carbapenemase-producing Enterobacterales

Marimuthu

Koh²,

Harbarth³

et al. 2022

Nat Commun

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Carbapenemase-producing Enterobacterales (CPE) infection control practices are based on the paradigm that detected carriers in the hospital transmit to other patients who stay in the same ward. The role of plasmid-mediated transmission at population level remains largely unknown. In this retrospective cohort study over 4.7 years involving all multi-disciplinary public hospitals in Singapore, we analysed 779 patients who acquired CPE (1215 CPE isolates) detected by clinical or surveillance cultures. 42.0% met putative clonal transmission criteria, 44.8% met putative plasmid-mediated transmission criteria and 13.2% were unlinked. Only putative clonal transmissions associated with direct ward contact decreased in the second half of the study. Both putative clonal and plasmid-mediated transmission associated with indirect (no temporal overlap in patients’ admission period) ward and hospital contact did not decrease during the study period. Indirect ward and hospital contact were identified as independent risk factors associated with clonal transmission. In conclusion, undetected CPE reservoirs continue to evade hospital infection prevention measures. New measures are needed to address plasmid-mediated transmission, which accounted for 50% of CPE dissemination.

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Section: Discussionmentioning

confidence: 99%

Whole genome sequencing reveals hidden transmission of carbapenemase-producing Enterobacterales

Marimuthu

Koh²,

Harbarth³

et al. 2022

Nat Commun

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“…The spread of CR-KP has gradually increased and even led to outbreaks (Kizilay et al, 2020;Pellicé et al, 2021). Molecular epidemiology plays an important role in tracking the spread of MDR bacteria to provide data for controlling their dissemination (Fontana et al, 2020). Methods for studying the molecular epidemiology of CR-KP include multilocus sequence typing (MLST), pulsed-field gel electrophoresis, enterobacterial repetitive intergenic consensus-polymerase chain reaction, and metagenomics next-generation sequencing.…”

Section: Introductionmentioning

confidence: 99%

Four Types of ST11 Novel Mutations From Increasing Carbapenem-Resistant Klebsiella pneumoniae in Guangdong, 2016–2020

et al. 2021

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Objectives: This study aimed to explore changes in carbapenem-resistant Klebsiella pneumoniae (CR-KP) isolates collected in Guangdong over the period of 2016–2020.Methods: Antibacterial susceptibility was quantified through VITEK 2 compact and K-B method. Carbapenemase phenotypes and genotypes were characterized by modified carbapenem inactivation method (mCIM), EDTA-carbapenem inactivation method (eCIM), and polymerase chain reaction (PCR). Molecular characteristics and evolutionary trends were analyzed by multilocus sequence typing and evolutionary tree.Results: Isolates (2,847) of K. pneumoniae were separated in 2016–2020, and the separate rate of CR-KP increased from 5.65 to 9.90% (p = 0.009). The top 3 wards were intensive care unit (ICU) (21.92%), neonatal wards (13.70%), and respiratory wards (12.33%). In 146 CR-KP strains, serine carbapenemase was the main phenotype, and KPC was the main genotype, and 57 contained two resistant genes, and 1 contained three resistant genes. Two polygenic strains were first found: IMP + GES and KPC + NDM + VIM, but all the phenotypes were metalloenzyme, which indicated that metalloenzyme was usually the first choice for CR-KP resistance. In addition, all the ST54 of metalloenzyme type contained IMP, and all the ST45, ST37, and ST76 contained OXA. ST11 was the most prevalent (42.47%); ST11 and its mutants proved the predominant sequence type making up 51.1% of the carbapenemase-producing isolates. A novel type of ST11 mutation, the rpoB was mutated from sequence 1 to sequence 146, was in an independent separate branch on the evolutionary tree and was resistant to all antibacterial agents. The other three mutants, rpoB 1–15, infB 3–148, and infB 3–80, are also resistant to all antibacteria. Of note, all the four mutants produced serine carbapenemase and contained KPC, and indicated that the prevalent strain in China, ST11, has serious consequences and potential outbreaks.Conclusion: The infection rate of CR-KP has increased, and ICU and neonatal wards have become the key infection areas. Producing serine enzyme, the KPC genotype, and ST11 are the predominant CR-KP. Polygenic strains and ST11 mutation made clinical treatment difficult and may become a potential threat.

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“…Multilocus sequence typing (MLST) and acquired antimicrobial resistance genes were identified by using the online CGE tool ( https://cge.cbs.dtu.dk ). Phylogenetic relationships were evaluated by core genome multilocus sequence typing (cgMLST) analysis, followed by generation of a minimum-spanning tree by using SeqSphere+ software (Ridom GmbH, Muenster, Germany) [ 29 ]. Virulence genes were predicted by using the online tools VFanalyzer ( http://www.mgc.ac.cn/cgi-bin/VFs/v5/main.cgi?func=VFanalyzer ) and Kaptive Web ( http://kaptive.holtlab.net/ ).…”

Section: Methodsmentioning

confidence: 99%

Resistance evolution of hypervirulent carbapenem-resistant Klebsiella pneumoniae ST11 during treatment with tigecycline and polymyxin

Jin

Chen

Shen

et al. 2021

Emerging Microbes & Infections

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Hypervirulent carbapenem-resistant Klebsiella pneumoniae (hv-CRKP) has recently aroused increasing attention, especially ST11, the predominant CRKP clone in China.Here, we report a case of hv-CRKP-associated infection and reveal the in-host evolution of its mechanism of resistance to tigecycline and polymyxin under clinical therapy. A total of 11 K. pneumoniae carbapenemase (KPC)-producing CRKP strains were consecutively isolated from a male patient who suffered from continuous and multisite infections and successively received antibiotic treatment, including tigecycline and polymyxin. String and antimicrobial susceptibility tests identified seven hypermucoviscous strains and three tigecycline-resistant and four colistin-resistant strains. Galleria mellonella larvae infection model confirmed the hypervirulence. Pulsed-field gel electrophoresis (PFGE) separated three PFGE clusters among all strains, and further Southern blotting detected that blaKPC-2 was located on the same-sized plasmid. Whole-genome sequencing based on Illumina showed that all strains belonged to the hv-CRKP ST11-KL64 clone. Diverse hypervirulence factors and resistance genes were identified. Further sequencing with the Nanopore platform was performed on the CRKP-Urine1 strain, which contained one virulence plasmid (pVi-CRKP-Urine1) and two resistance plasmids (pKPC-CRKP-Urine1 and pqnrS1-CRKP-Urine1). The gene mutations responsible for tigecycline or colistin resistance were then amplified with PCR followed by sequencing, which indicated that mutations of ramR and lon were the potential loci for tigecycline resistance and that the pmrB, phoQ and mgrB genes for colistin resistance. A novel frameshift mutation of lon was identified in the high-level tigecycline-resistant strain (MIC, 128 mg/L). The results indicate that the hypervirulent ST11-KL64 clone is a potential threat to antiinfection treatment and is capable of rapid and diverse evolution of resistance during tigecycline and polymyxin treatment.

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Whole Genome Sequencing of Carbapenem-Resistant Klebsiella Pneumoniae : Evolutionary Analysis for Outbreak Investigation

Cited by 10 publications

References 50 publications

Whole genome sequencing reveals hidden transmission of carbapenemase-producing Enterobacterales

Whole genome sequencing reveals hidden transmission of carbapenemase-producing Enterobacterales

Four Types of ST11 Novel Mutations From Increasing Carbapenem-Resistant Klebsiella pneumoniae in Guangdong, 2016–2020

Resistance evolution of hypervirulent carbapenem-resistant Klebsiella pneumoniae ST11 during treatment with tigecycline and polymyxin

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