Transmissible Pseudomonas aeruginosa clones potentially pose a serious threat to cystic fibrosis (CF) patients. The AES-1 clone has been found to infect up to 40 % of patients in five CF centres in eastern Australia. Studies were carried out on clonal and non-clonal (NC) isolates from chronically infected CF patients, and the reference strain PAO1, to gain insight into the properties of AES-1. The transcriptomes of AES-1 and NC isolates, and of PAO1, grown planktonically and as a 72 h biofilm were compared using PAO1 microarrays. Microarray data were validated using real-time PCR. Overall, most differentially expressed genes were downregulated. AES-1 differentially expressed bacteriophage genes, novel motility genes, and virulence and quorum-sensing-related genes, compared with both PAO1 and NC. AES-1 but not NC biofilms significantly downregulated aerobic respiration genes compared with planktonic growth, suggesting enhanced anaerobic/microaerophilic growth by AES-1. Biofilm measurement showed that AES-1 formed significantly larger and thicker biofilms than NC or PAO1 isolates. This may be related to expression of the gene PA0729, encoding a biofilm-enhancing bacteriophage, identified by PCR in all AES-1 but few NC isolates (n=42). Links with the Liverpool epidemic strain included the presence of PA0729 and the absence of the bacteriophage gene cluster PA0632-PA0639. No common markers were found with the Manchester strain. No particular differentially expressed gene in AES-1 could definitively be ascribed a role in its infectivity, thus increasing the likelihood that AES-1 infectivity is multi-factorial and possibly involves novel genes. This study extends our understanding of the transcriptomic and genetic differences between clonal and NC strains of P. aeruginosa from CF lung.