A multitask biosensor for micro-volumetric detection of N-3-oxo-dodecanoyl-homoserine lactone quorum sensing signal
2011
Massai, Francesco | Imperi, Francesco | Quattrucci, Serena | Zennaro, Elisabetta | Visca, Paolo | Leoni, Livia
N-3-oxo-dodecanoyl-homoserine lactone (3OC₁₂-HSL) is the main quorum sensing (QS) signal produced by the human pathogen Pseudomonas aeruginosa, a major cause of hard-to-treat nosocomial infections and years-lasting chronic biofilm infections in the lungs of cystic fibrosis (CF) patients. 3OC₁₂-HSL-dependent QS is considered a promising target for novel anti-pseudomonads drugs. However, the screening systems employed to date for the identification of QS inhibitors (QSI) were aimed at the identification of inhibitors of 3OC₁₂-HSL signaling rather than of the synthesis or the export of this molecule. Moreover, the low concentration of 3OC₁₂-HSL in CF sputum has hampered large scale studies aimed at addressing the role of this molecule in the CF lung infection. Here we describe the construction and characterization of PA14-R3, a new whole-cell biosensor for the quantitative detection of 3OC₁₂-HSL. PA14-R3 provides fast and direct quantification of 3OC₁₂-HSL over a wide range of concentrations (from pM to μM), and proved to be an easy-to-handle, cost-effective and reliable biosensor for high-throughput screening of 3OC₁₂-HSL levels in samples of different origin, including CF sputum. Moreover, the specific features of PA14-R3 made it possible to develop and validate a novel high-throughput screening system for QSI based on the co-cultivation of PA14-R3 with the PA14 wild-type strain. With respect to previous screening systems for QSI, this approach has the advantage of being cost-effective and allowing the identification of compounds targeting, besides 3OC₁₂-HSL signaling, any cellular process critical for QS response, including 3OC₁₂-HSL synthesis and secretion.
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