Chapter 21
Qualitative and Quantitative Determination of Quorum
Sensing Inhibition In Vitro
Tim Holm Jakobsen, Maria Alhede, Louise Dahl Hultqvist,
Thomas Bjarnsholt, and 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 antimi-
crobial treatments. The present protocol describes methods to determine the in vitro efficacy of potential
QS inhibitors (QSIs). Work onPseudomonas aeruginosahas 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-gfporrhlA-gfpfusion inP. aeruginosafor 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.
Key wordsQuorum sensing inhibitor, Halogenated furanones, QSI monitor screen, DNA micro-
array, In vitro continuous-culture biofilm flow cell system, Confocal scanning laser microscopy1 Introduction
Treatment of infectious diseases is becoming increasingly more
difficult as bacterial resistance to antibiotics evolves with an alarm-
ing rate. Several opportunistic pathogens rely on biofilm formation
and quorum sensing (QS) controlled expression of virulence factors
in the process of establishing persistent infections in humans and
animals. Both processes help the bacteria against the host defense
and otherwise detrimental effects of antimicrobial drug treatments
[1–4]. However, research has shown that administration of QS
inhibitors (QSIs) can block QS controlled phenotypes including
production of virulence factors [5, 6], thereby functioning as anti-
microbials [6, 7].Livia Leoni and Giordano Rampioni (eds.),Quorum Sensing: Methods and Protocols, Methods in Molecular Biology,
vol. 1673,https://doi.org/10.1007/978-1-4939-7309-5_21,©Springer Science+Business Media LLC 2018
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