Label-free SRM-based relative quantification of antibiotic resistance mechanisms in Pseudomonas aeruginosa clinical isolates
Both acquired and intrinsic mechanisms play a crucial role in Pseudomonas aeruginosa antibiotic resistance. Many clinically relevant resistance mechanisms result from changes in gene expression, namely multidrug efflux pump overproduction, AmpC beta-lactamase induction or derepression, and inactivat...
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doaj-2f7123314d1545039d5fcde1bbd631a72020-11-24T22:54:21ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2015-02-01610.3389/fmicb.2015.00081130033Label-free SRM-based relative quantification of antibiotic resistance mechanisms in Pseudomonas aeruginosa clinical isolatesYannick eCharretier0Thilo eKöhler1Tiphaine eCecchini2Tiphaine eCecchini3Chloé eBardet4Chloé eBardet5Abdessalam eCherkaoui6Catherine eLlanes7Pierre eBogaerts8Sonia eChatellier9Jean-Philippe eCharrier10Jacques eSchrenzel11Jacques eSchrenzel12Geneva University HospitalsUniversity of GenevaInstitute for Analytical SciencesbioMérieux SALimoges UniversitybioMérieux SAGeneva University HospitalsUniversité de Franche-ComtéCHU Dinant-Godinne UCL NamurbioMérieux SAbioMérieux SAGeneva University HospitalsGeneva University HospitalsBoth acquired and intrinsic mechanisms play a crucial role in Pseudomonas aeruginosa antibiotic resistance. Many clinically relevant resistance mechanisms result from changes in gene expression, namely multidrug efflux pump overproduction, AmpC beta-lactamase induction or derepression, and inactivation or repression of the carbapenem-specific porin OprD. Changes in gene expression are usually assessed using reverse-transcription quantitative real-time PCR (RT-qPCR) assays. Here, we evaluated label-free Selected Reaction Monitoring (SRM)-based mass spectrometry to directly quantify proteins involved in antibiotic resistance. We evaluated the label-free SRM using a defined set of P. aeruginosa isolates with known resistance mechanisms and compared it against RT-qPCR. Referring to efflux systems, we found a more robust relative quantification of antibiotic resistance mechanisms by SRM than RT-qPCR. The SRM-based approach was applied to a set of clinical P. aeruginosa isolates to detect antibiotic resistance proteins. This multiplexed SRM-based approach is a rapid and reliable method for the simultaneous detection and quantification of resistance mechanisms and we demonstrate its relevance for antibiotic resistance prediction.http://journal.frontiersin.org/Journal/10.3389/fmicb.2015.00081/fullPseudomonas aeruginosaRT-qPCRSRMCarbapenem resistanceOprD porinmultidrug efflux system |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Yannick eCharretier Thilo eKöhler Tiphaine eCecchini Tiphaine eCecchini Chloé eBardet Chloé eBardet Abdessalam eCherkaoui Catherine eLlanes Pierre eBogaerts Sonia eChatellier Jean-Philippe eCharrier Jacques eSchrenzel Jacques eSchrenzel |
spellingShingle |
Yannick eCharretier Thilo eKöhler Tiphaine eCecchini Tiphaine eCecchini Chloé eBardet Chloé eBardet Abdessalam eCherkaoui Catherine eLlanes Pierre eBogaerts Sonia eChatellier Jean-Philippe eCharrier Jacques eSchrenzel Jacques eSchrenzel Label-free SRM-based relative quantification of antibiotic resistance mechanisms in Pseudomonas aeruginosa clinical isolates Frontiers in Microbiology Pseudomonas aeruginosa RT-qPCR SRM Carbapenem resistance OprD porin multidrug efflux system |
author_facet |
Yannick eCharretier Thilo eKöhler Tiphaine eCecchini Tiphaine eCecchini Chloé eBardet Chloé eBardet Abdessalam eCherkaoui Catherine eLlanes Pierre eBogaerts Sonia eChatellier Jean-Philippe eCharrier Jacques eSchrenzel Jacques eSchrenzel |
author_sort |
Yannick eCharretier |
title |
Label-free SRM-based relative quantification of antibiotic resistance mechanisms in Pseudomonas aeruginosa clinical isolates |
title_short |
Label-free SRM-based relative quantification of antibiotic resistance mechanisms in Pseudomonas aeruginosa clinical isolates |
title_full |
Label-free SRM-based relative quantification of antibiotic resistance mechanisms in Pseudomonas aeruginosa clinical isolates |
title_fullStr |
Label-free SRM-based relative quantification of antibiotic resistance mechanisms in Pseudomonas aeruginosa clinical isolates |
title_full_unstemmed |
Label-free SRM-based relative quantification of antibiotic resistance mechanisms in Pseudomonas aeruginosa clinical isolates |
title_sort |
label-free srm-based relative quantification of antibiotic resistance mechanisms in pseudomonas aeruginosa clinical isolates |
publisher |
Frontiers Media S.A. |
series |
Frontiers in Microbiology |
issn |
1664-302X |
publishDate |
2015-02-01 |
description |
Both acquired and intrinsic mechanisms play a crucial role in Pseudomonas aeruginosa antibiotic resistance. Many clinically relevant resistance mechanisms result from changes in gene expression, namely multidrug efflux pump overproduction, AmpC beta-lactamase induction or derepression, and inactivation or repression of the carbapenem-specific porin OprD. Changes in gene expression are usually assessed using reverse-transcription quantitative real-time PCR (RT-qPCR) assays. Here, we evaluated label-free Selected Reaction Monitoring (SRM)-based mass spectrometry to directly quantify proteins involved in antibiotic resistance. We evaluated the label-free SRM using a defined set of P. aeruginosa isolates with known resistance mechanisms and compared it against RT-qPCR. Referring to efflux systems, we found a more robust relative quantification of antibiotic resistance mechanisms by SRM than RT-qPCR. The SRM-based approach was applied to a set of clinical P. aeruginosa isolates to detect antibiotic resistance proteins. This multiplexed SRM-based approach is a rapid and reliable method for the simultaneous detection and quantification of resistance mechanisms and we demonstrate its relevance for antibiotic resistance prediction. |
topic |
Pseudomonas aeruginosa RT-qPCR SRM Carbapenem resistance OprD porin multidrug efflux system |
url |
http://journal.frontiersin.org/Journal/10.3389/fmicb.2015.00081/full |
work_keys_str_mv |
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