Quorum Sensing

Chapter 16

ImagingN-Acyl Homoserine Lactone Quorum

Sensing In Vivo

Louise Dahl Hultqvist, Maria Alhede, Tim Holm Jakobsen,

Michael Givskov, and Thomas Bjarnsholt

Abstract

In order to studyN-acyl homoserine lactone (AHL)-based quorum sensing in vivo, we present a protocol
using anEscherichia colistrain equipped with aluxR-based monitor system, which in the presence of
exogenous AHL molecules expresses a green fluorescent protein (GFP). Lungs from mice challenged
intratracheally with alginate beads containing both aPseudomonas aeruginosastrain together with the
E. colimonitor strain can be investigated at different time points postinfection. Epifluorescent or confocal
scanning laser microscopy (CSLM) is used to detect the GFP-expressingE. colimonitor strain in the lung
tissues, indicating production and excretion of AHLs in vivo by the infectingP. aeruginosa.

Key wordsQuorum sensing,Pseudomonas aeruginosa, Pulmonary infection model, Epifluorescence

microscopy, CSLM, In vivo

1 Introduction

The biofilm mode of growth is known to protectPseudomonas

aeruginosaagainst the host defense and antibiotics [1–6]. Due to

the high density of cells in biofilms,P. aeruginosais able to make

use of its cell–cell communication (quorum sensing (QS)) systems.

P. aeruginosaemploys twoN-acyl-L-homoserine lactone (AHL)

signal molecule based QS sensors, encoded by thelasandrhl

systems. Both systems are organized with a LuxR-type transcrip-

tional regulator, LasR and RhlR, and a cognate LuxI-type synthe-

tase, LasI and RhlI, that mainly synthesize the signal moleculesN-

(3-oxododecanoyl)-L-homoserine lactone (3-oxo-C 12 -HSL) and

N-butanoyl-L-homoserine lactone (C 4 -HSL), respectively [7, 8].

In addition to the AHL signal molecule based QS sensors a third

QS system, thePseudomonasquinolone signal (PQS) based system,

has been identified [9].

In simple terms, AHL regulation comprises anIgene encoding

the AHL synthetase and an R gene encoding the receptor

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_16,©Springer Science+Business Media LLC 2018

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