Qualitative and Quantitative Determination of Quorum Sensing Inhibition In Vitro

Tim Holm Jakobsen, Maria Alhede, Louise Dahl Hultqvist, Thomas Bjarnsholt, Michael Givskov

Abstract

The formation of biofilms in conjunction with quorum sensing (QS) regulated expression of virulence by opportunistic pathogens contributes significantly to immune evasion and tolerance to a variety of antimicrobial treatments. The present protocol describes methods to determine the in vitro efficacy of potential QS inhibitors (QSIs). Work on Pseudomonas aeruginosa has shown that chemical blockage of QS is a promising new antimicrobial strategy. Several live bacterial reporter systems have been developed to screen extracts and pure compounds for QSI activity. Here we describe the usage of reporter strains consisting of a lasB-gfp or rhlA-gfp fusion in P. aeruginosa for qualitative and quantitative evaluation of the inhibition of two of the major QS pathways, monitored as reduced expression of green fluorescence. By the use of an in vitro flow cell system it is possible to study the QSI activity by monitoring its ability to interfere with the protective functions of bacterial biofilm. For evaluation of the global effects of QSI compounds, we present a protocol for the DNA microarray based transcriptomics. Using these in vitro methods it is possible to evaluate the potential of various QSI compounds.

Original languageEnglish
JournalMethods in Molecular Biology
Volume1673
Pages (from-to)275-285
Number of pages11
ISSN1064-3745
DOIs
Publication statusPublished - 2018

Keywords

  • Biofilms/growth & development
  • Biological Assay/methods
  • Green Fluorescent Proteins/metabolism
  • Imaging, Three-Dimensional
  • Oligonucleotide Array Sequence Analysis
  • Pseudomonas aeruginosa/physiology
  • Quorum Sensing
  • Rheology

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