A highly multiplexed melt-curve assay for detecting the most prevalent carbapenemase, ESBL and AmpC genes

Thomas Edwards; Chris Williams; Y. Teethaisong; J. Sealey; S. Sasaki; G. Hobbs; Luis Cuevas; K. Evans; Emily Adams

doi:10.1016/j.diagmicrobio.2020.115076

A highly multiplexed melt-curve assay for detecting the most prevalent carbapenemase, ESBL and AmpC genes

Thomas Edwards, Chris Williams, Y. Teethaisong, J. Sealey, S. Sasaki, G. Hobbs, Luis Cuevas, K. Evans, Emily Adams

Research output: Contribution to journal › Article › peer-review

19 Citations (Scopus)

Abstract

Resistance to third generation cephalosporins and carbapenems in Gram-negative bacteria is chiefly mediated by beta-lactamases including ESBL, AmpC and carbapenemase enzymes. Routine phenotypic detection methods do not provide timely results, and there is a lack of comprehensive molecular panels covering all important markers.

An ESBL/carbapenemase HRM assay (SHV, TEM, CTX-M ESBL families, and NDM, IMP, KPC, VIM and OXA-48-like carbapenemases) and an AmpC HRM assay (16S rDNA control, FOX, MOX, ACC, EBC, CIT and DHA) were designed, and evaluated on 111 Gram-negative isolates with mixed resistance patterns.

The sensitivity for carbapenemase, ESBL and AmpC genes was 96.7% (95%CI:82.8-99.9%), 93.6% (95%CI:85.7-97.9%) and 93.8% (95%CI:82.8-98.7%), respectively with a specificity of 100% (95%CI:95.6-100%), 93.9% (95%CI:79.8-99.3%) and 93.7% (95%CI:84.5-98.2%).

The HRM assays enable the simultaneous detection of the fourteen most important ESBL, carbapenemase and AmpC genes and could be used as a molecular surveillance tool or to hasten detection of AMR for treatment management.

Original language	English
Article number	115076
Pages (from-to)	115076
Journal	Diagnostic Microbiology and Infectious Disease
Volume	97
Issue number	4
Early online date	8 May 2020
DOIs	https://doi.org/10.1016/j.diagmicrobio.2020.115076
Publication status	Published - 1 Aug 2020

Keywords

Antimicrobial resistance
Carbapenemase
ESBL
High resolution melt analysis

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title = "A highly multiplexed melt-curve assay for detecting the most prevalent carbapenemase, ESBL and AmpC genes",

abstract = "Resistance to third generation cephalosporins and carbapenems in Gram-negative bacteria is chiefly mediated by beta-lactamases including ESBL, AmpC and carbapenemase enzymes. Routine phenotypic detection methods do not provide timely results, and there is a lack of comprehensive molecular panels covering all important markers.An ESBL/carbapenemase HRM assay (SHV, TEM, CTX-M ESBL families, and NDM, IMP, KPC, VIM and OXA-48-like carbapenemases) and an AmpC HRM assay (16S rDNA control, FOX, MOX, ACC, EBC, CIT and DHA) were designed, and evaluated on 111 Gram-negative isolates with mixed resistance patterns.The sensitivity for carbapenemase, ESBL and AmpC genes was 96.7\% (95\%CI:82.8-99.9\%), 93.6\% (95\%CI:85.7-97.9\%) and 93.8\% (95\%CI:82.8-98.7\%), respectively with a specificity of 100\% (95\%CI:95.6-100\%), 93.9\% (95\%CI:79.8-99.3\%) and 93.7\% (95\%CI:84.5-98.2\%).The HRM assays enable the simultaneous detection of the fourteen most important ESBL, carbapenemase and AmpC genes and could be used as a molecular surveillance tool or to hasten detection of AMR for treatment management.",

keywords = "Antimicrobial resistance, Carbapenemase, ESBL, High resolution melt analysis",

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AU - Williams, Chris

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AU - Sasaki, S.

AU - Hobbs, G.

AU - Cuevas, Luis

AU - Evans, K.

AU - Adams, Emily

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AB - Resistance to third generation cephalosporins and carbapenems in Gram-negative bacteria is chiefly mediated by beta-lactamases including ESBL, AmpC and carbapenemase enzymes. Routine phenotypic detection methods do not provide timely results, and there is a lack of comprehensive molecular panels covering all important markers.An ESBL/carbapenemase HRM assay (SHV, TEM, CTX-M ESBL families, and NDM, IMP, KPC, VIM and OXA-48-like carbapenemases) and an AmpC HRM assay (16S rDNA control, FOX, MOX, ACC, EBC, CIT and DHA) were designed, and evaluated on 111 Gram-negative isolates with mixed resistance patterns.The sensitivity for carbapenemase, ESBL and AmpC genes was 96.7% (95%CI:82.8-99.9%), 93.6% (95%CI:85.7-97.9%) and 93.8% (95%CI:82.8-98.7%), respectively with a specificity of 100% (95%CI:95.6-100%), 93.9% (95%CI:79.8-99.3%) and 93.7% (95%CI:84.5-98.2%).The HRM assays enable the simultaneous detection of the fourteen most important ESBL, carbapenemase and AmpC genes and could be used as a molecular surveillance tool or to hasten detection of AMR for treatment management.

KW - Antimicrobial resistance

KW - Carbapenemase

KW - ESBL

KW - High resolution melt analysis

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DO - 10.1016/j.diagmicrobio.2020.115076

M3 - Article

SN - 0732-8893

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SP - 115076

JO - Diagnostic Microbiology and Infectious Disease

JF - Diagnostic Microbiology and Infectious Disease

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M1 - 115076

ER -

A highly multiplexed melt-curve assay for detecting the most prevalent carbapenemase, ESBL and AmpC genes

Abstract

Keywords

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