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J Med Microbiol 55 (2006), 677-688; DOI: 10.1099/jmm.0.46461-0
© 2006 Society for General Microbiology
ISSN 1473-5644

Use of suppression subtractive hybridization to examine the accessory genome of the Liverpool cystic fibrosis epidemic strain of Pseudomonas aeruginosa

Catherine H. M. Smart1, Martin J. Walshaw2, C. Anthony Hart1 and Craig Winstanley1

1 Division of Medical Microbiology and Genitourinary Medicine, University of Liverpool, Liverpool L69 3GA, UK

2 Regional Adult Cystic Fibrosis Unit, Cardiothoracic Centre, Liverpool L14 3PE, UK

Correspondence
Craig Winstanley
C.Winstanley{at}liv.ac.uk

Received 9 December 2005
Accepted 20 February 2006

The Liverpool epidemic strain (LES) of Pseudomonas aeruginosa has been highly successful at colonizing cystic fibrosis (CF) patients throughout the UK, has replaced previously established strains in CF patients, has caused infections of non-CF parents of CF patients, and can cause greater morbidity in CF than other strains of P. aeruginosa. Using suppression subtractive hybridization (SSH) to identify strain-specific sequences, a diagnostic test for the LES based on PCR amplification of SSH sequence PS21 had previously been developed. In this study, the SSH sequence database of LES was substantially increased, using both extension of previous sequences and new rounds of subtraction. Of 92 SSH sequences identified as present in the LES but absent from strain PAO1, 25 were assessed for prevalence amongst a strain panel consisting mainly of LES and non-LES CF isolates. Preliminary analysis of genome sequence data indicated that all SSH sequences that were LES specific or found only rarely in other strains of P. aeruginosa were present on one of three contigs. All of the SSH sequences screened were either unstable amongst LES isolates or were not completely LES specific. Rare false positives were found with the PS21 test. The authors suggest that a second PCR assay designed to detect SSH sequence LESF9 can be used to confirm the identity of the most prevalent CF epidemic lineage in the UK.

Abbreviations: CF, cystic fibrosis; FHA, filamentous haemagglutinin; LES, Liverpool epidemic strain; SSH, suppression subtractive hybridization.




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