In a previous study of isolates from cystic fibrosis (CF) patients in England and Wales, the Midlands 1 strain of Pseudomonas aeruginosa was identified as the second most common clone, representing 10 % of isolates and found in nearly one-third of all CF centres [Scott, F. W. & Pitt, T. L. (2004). J Med Microbiol53, 609–615]. Using suppression subtractive hybridization, 54 sequences were identified as present in a Midlands 1 strain but were absent from strain PAO1. The distribution of 14 of these sequences amongst representatives of Midlands 1, other CF epidemic strains and unrelated P. aeruginosa CF isolates was determined using PCR assays. Using these data, a PCR-based test was developed that was specific for the Midlands 1 clone, which was confirmed using dot-blot hybridization. By applying the test to CF isolates from a CF centre in Liverpool, a Midlands 1 clone was identified. The identity was confirmed using typing by PFGE. The PCR test should facilitate a greater understanding of the distribution of the Midlands 1 strain in the UK and elsewhere.