Identification of N-acylhomoserine lactones in
mucopurulent respiratory secretions from cystic fibrosis patients
Catherine E. Chambersa,1, Michelle B. Vissera,1, Ute Schwabb, Pamela A. Sokola,*
aDepartment of Microbiology and Infectious Diseases, University of Calgary Health Sciences Centre, Calgary, Alta., Canada T2N 4N1
bDepartment of Food Science, Cornell University, Ithaca, NY 14853, USA
Received 18 September 2004; received in revised form 24 December 2004; accepted 31 January 2005
First published online 9 February 2005
Edited by C.W. Penn
Pseudomonas aeruginosa and species of the Burkholderia cepacia complex are the primary bacterial pathogens contributing to
lung disease in patients with cystic fibrosis. Quorum sensing systems using N-acyl homoserine lactone (AHL) signal molecules
are involved in the regulation of a number of virulence factors in these species. Extracts of mucopurulent respiratory secretions from
13 cystic fibrosis patients infected with P. aeruginosa and/or strains of the B. cepacia complex were fractionated using reverse-phase
fast pressure liquid chromatography and analyzed for the presence of AHLs using a traI–luxCDABE-based reporter that responds
to AHLs with acyl chains ranging between 4 and 12 carbons. Using this assay system, a broad range of AHLs were detected and
identified despite being present at low concentrations in limited sample volumes. N-(3-oxo-dodecanoyl)-L-homoserine lactone, N-(3-
oxo-decanoyl)-L-homoserine lactone and N-octanoyl-L-homoserine lactone (OHL) were the AHLs most frequently identified. OHL
and N-decanoyl-L-homoserine lactone were detected in nanomolar concentrations compared to picomolar amounts of the 3-oxo-
derivatives of the AHLs identified.
? 2005 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.
Keywords: N-Acylhomoserine lactone; Burkholderia cepacia; Pseudomonas aeruginosa; Quorum sensing; Cystic fibrosis
Cystic fibrosis (CF) is due to a mutation in the cystic
fibrosis transmembrane regulator (CFTR) which leads
to lung disease characterized by altered fluid transport,
increased mucus viscosity, and reduced mucociliary
clearance resulting in persistent microbial colonization
(reviewed in [1,2]). Pseudomonas aeruginosa and strains
of the Burkholderia cepacia complex (Bcc) are the pri-
mary pathogens responsible for morbidity and mortality
in CF patients [1,3,4].
Quorum sensing is a cell density-dependent mecha-
nism used by bacteria to coordinate gene expression
via diffusible signal molecules termed N-acylhomoserine
lactones (AHLs). These systems consist of an AHL syn-
thase belonging to the LuxI protein family as well as a
transcriptional regulator belonging to the LuxR family
. P. aeruginosa has two inter-related AHL mediated
signaling systems, lasIR and rhlIR, as well as a non-
AHL signal 2-heptyl-3-hydroxy-4-quinolone (PQS for
Pseudomonas quinolone signal) [6–9]. LasI directs the
synthesis of N-(3-oxododecanoyl)-L-HSL (3-oxo-C12-
HSL) and N-(3-oxooctanoyl)-L-HSL (3-oxo-C8-HSL)
0378-1097/$22.00 ? 2005 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.
*Corresponding author. Tel.: +1 403 220 6037; fax: +1 403 270 2772.
E-mail address: firstname.lastname@example.org (P.A. Sokol).
1C.E.C. and M.B.V. contributed equally to this work.
FEMS Microbiology Letters 244 (2005) 297–304
 Bjarnason, J., Southward, C.M. and Surette, M.G. (2003)
Genomic profiling of iron-responsive genes in Salmonella enterica
serovar typhimurium by high-throughput screening of a random
promoter library. J. Bacteriol. 185, 4973–4982.
 Shaw, P.D., Ping, G., Daly, S.L., Cha, C., Cronan, Jr., J.E.,
Rinehart, K.L. and Farrand, S.K. (1997) Detecting and charac-
terizing N-acyl-homoserine lactone signal molecules by thin-layer
chromatography. Proc. Natl. Acad. Sci. USA 94, 6036–6041.
 Singh, P.K., Schaefer, A.L., Parsek, M.R., Moninger, T.O.,
Welsh, M.J. and Greenberg, E.P. (2000) Quorum-sensing signals
indicate that cystic fibrosis lungs are infected with bacterial
biofilms. Nature 407, 762–764.
 Chun, C.K., Ozer, E.A., Welsh, M.J., Zabner, J. and Greenberg,
E.P. (2004) Inactivation of a Pseudomonas aeruginosa quorum-
sensing signal by human airway epithelia. Proc. Natl. Acad. Sci.
USA 101, 3587–3590.
 Isles, A., Maclusky, I., Corey, M., Gold, R., Prober, C., Fleming,
P. and Levison, H. (1984) Pseudomonas cepacia infection in cystic
fibrosis: an emerging problem. J. Pediatr. 104, 206–210.
 Cieri, M.V., Mayer-Hamblett, N., Griffith, A. and Burns, J.L.
(2002) Correlation between an in vitro invasion assay and a
murine model of Burkholderia cepacia lung infection. Infect.
Immun. 70, 1081–1086.
 Sajjan, U., Corey, M., Humar, A., Tullis, E., Cutz, E., Ackerley,
C. and Forstner, J. (2001) Immunolocalisation of Burkholderia
cepacia in the lungs of cystic fibrosis patients. J. Med. Microbiol.
 Riedel, K., Hentzer, M., Geisenberger, O., Huber, B., Steidle, A.,
Wu, H., Hoiby, N., Givskov, M., Molin, S. and Eberl, L. (2001)
Pseudomonas aeruginosa and Burkholderia cepacia in mixed
biofilms. Microbiology 147, 3249–3262.
C.E. Chambers et al. / FEMS Microbiology Letters 244 (2005) 297–304