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.",

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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

U2 - 10.1016/j.diagmicrobio.2020.115076

DO - 10.1016/j.diagmicrobio.2020.115076

M3 - Article

SN - 0732-8893

VL - 97

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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