Frontiers in Microbiology (Feb 2015)

Label-free SRM-based relative quantification of antibiotic resistance mechanisms in Pseudomonas aeruginosa clinical isolates

  • 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

DOI
https://doi.org/10.3389/fmicb.2015.00081
Journal volume & issue
Vol. 6

Abstract

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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.

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