Premium
A dual biosensor for 2‐alkyl‐4‐quinolone quorum‐sensing signal molecules
Author(s) -
Fletcher Matthew P.,
Diggle Stephen P.,
Crusz Shanika A.,
Chhabra Siri Ram,
Cámara Miguel,
Williams Paul
Publication year - 2007
Publication title -
environmental microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.954
H-Index - 188
eISSN - 1462-2920
pISSN - 1462-2912
DOI - 10.1111/j.1462-2920.2007.01380.x
Subject(s) - quorum sensing , bioreporter , biosensor , biology , microbiology and biotechnology , pseudomonas aeruginosa , biofilm , bioluminescence , pseudomonas , bacteria , biochemistry , reporter gene , gene , gene expression , genetics
Summary Pseudomonas , Burkholderia and Alteromonas species produce diverse 2‐alkyl‐4‐quinolones (AHQs) which inhibit the growth of bacteria, algae and phytoplankton, chelate iron, modulate mammalian host immune defences and act as quorum‐sensing (QS) signal molecules. To facilitate the detection, identification and quantification of the major Pseudomonas aeruginosa AHQs 2‐heptyl‐3‐hydroxy‐4‐quinolone (PQS) and 2‐heptyl‐4‐quinolone (HHQ) we developed two different AHQ biosensors. These were constructed by introducing either a lecA :: luxCDABE or a pqsA :: luxCDABE reporter gene fusion into a P. aeruginosa pqsA mutant which cannot synthesize AHQs. While both biosensors responded similarly to PQS (EC 50 18 ± 4 μM), the pqsA :: luxCDABE biosensor was most sensitively activated by HHQ (EC 50 0.44 ± 0.1 μM). This biosensor was also activated albeit less sensitively by (i) PQS analogues with alkyl chains varying from C1 to C11, (ii) HHQ analogues with C9 and C11 alkyl chains and (iii) 2‐heptyl‐4‐hydroxyquinoline‐N‐oxide (HHQNO). The AHQ biosensor also responded differentially to the AHQs present in cell free culture supernatants prepared from PAO1 and isogenic strains carrying mutations in genes ( pqsA , pqsH , lasR , lasI , rhlR , rhlI ) known to influence AHQ production. The AHQ profiles of P. aeruginosa strains was also evaluated by overlaying thin layer chromatogram (TLC) plates with the pqsA :: luxCDABE biosensor. In PAO1, three major bioluminescent spots were observed which correspond to PQS, HHQ and a mixture of 2 nonyl‐4‐quinolone and HHQNO. We also noted that on TLC plates the biosensor not only produced bioluminescence in response to AHQs but also the green pigment, pyocyanin which offers an alternative visual indicator for AHQ production.