title: Quorum sensing and Chromobacterium violaceum: exploitation of violacein production and inhibition for the detection of N-acylhomoserine lactones.
abstract: Quorum sensing relies upon the interaction of a diffusible signal molecule with a transcriptional activator protein to couple gene expression with cell population density. In Gram-negative bacteria, such signal molecules are usually N-acylhomoserine lactones (AHLs) which differ in the structure of their N-acyl side chains. Chromobacterium violaceum, a Gram-negative bacterium commonly found in soil and water, produces the characteristic purple pigment violacein. Previously the authors described a violacein-negative, mini-Tn5 mutant of C. violaceum (CV026) in which pigment production can be restored by incubation with supernatants from the wild-type strain. To develop this mutant as a general biosensor for AHLs, the natural C. violaceum AHL molecule was first chemically characterized. By using solvent extraction, HPLC and mass spectrometry, a single AHL, N-hexanoyl-L-homoserine lactone (HHL), was identified in wild-type C. violaceum culture supernatants which was absent from CV026. Since the production of violacein constitutes a simple assay for the detection of AHLs, we explored the ability of CV026 to respond to a series of synthetic AHL and N-acylhomocysteine thiolactone (AHT) analogues. In CV026, violacein is inducible by all the AHL and AHT compounds evaluated with N-acyl side chains from C4 to C8 in length, with varying degrees of sensitivity. Although AHL compounds with N-acyl side chains from C10 to C14 are unable to induce violacein production, if an activating AHL (e.g. HHL) is incorporated into the agar, these long-chain AHLs can be detected by their ability to inhibit violacein production. The versatility of CV026 in facilitating detection of AHL mixtures extracted from culture supernatants and separated by thin-layer chromatography is also demonstrated. These simple bioassays employing CV026 thus greatly extend the ability to detect a wide spectrum of AHL signal molecules.
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.|Department of Applied Biochemistry and Food Science, University of Nottingham, Sutton Bonington Campus, Loughborough LE12 5RD, UK.
authors.forename: Kay H
authors.initials: KH
authors.lastname: McClean
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.|Department of Applied Biochemistry and Food Science, University of Nottingham, Sutton Bonington Campus, Loughborough LE12 5RD, UK.
authors.forename: Michael K
authors.initials: MK
authors.lastname: Winson
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.|Department of Applied Biochemistry and Food Science, University of Nottingham, Sutton Bonington Campus, Loughborough LE12 5RD, UK.
authors.forename: Leigh
authors.initials: L
authors.lastname: Fish
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.|Department of Applied Biochemistry and Food Science, University of Nottingham, Sutton Bonington Campus, Loughborough LE12 5RD, UK.
authors.forename: Adrian
authors.initials: A
authors.lastname: Taylor
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.
authors.forename: Siri Ram
authors.initials: SR
authors.lastname: Chhabra
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.
authors.forename: Miguel
authors.initials: M
authors.lastname: Camara
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.|Department of Applied Biochemistry and Food Science, University of Nottingham, Sutton Bonington Campus, Loughborough LE12 5RD, UK.
authors.forename: Mavis
authors.initials: M
authors.lastname: Daykin
authors.affiliation: MRC Toxicology Unit, University of Leicester, Leicester LE1 9HN, UK.
authors.forename: John H
authors.initials: JH
authors.lastname: Lamb
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.|Department of Applied Biochemistry and Food Science, University of Nottingham, Sutton Bonington Campus, Loughborough LE12 5RD, UK.
authors.forename: Simon
authors.initials: S
authors.lastname: Swift
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.
authors.forename: Barrie W
authors.initials: BW
authors.lastname: Bycroft
authors.affiliation: Department of Applied Biochemistry and Food Science, University of Nottingham, Sutton Bonington Campus, Loughborough LE12 5RD, UK.
authors.forename: Gordon S A B
authors.initials: GSAB
authors.lastname: Stewart
authors.affiliation: Department of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.
authors.forename: Paul
authors.initials: P
authors.lastname: Williams
mesh_terms: D002861:Chromobacterium / Q000235:genetics / Q000502:physiology*; D015964:Gene Expression Regulation, Bacterial; D006714:Homoserine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism*; D007211:Indoles; D007783:Lactones / Q000737:chemistry / Q000378:metabolism*; D015398:Signal Transduction; D014344:Trypanocidal Agents
chemical_list: D007211:Indoles; D007783:Lactones; D014344:Trypanocidal Agents; D006714:Homoserine; C063155:violacein