Aims: Burkholderia cepacia is a Gram-negative bacterium associated with increasing morbidity and mortality and is readily transmitted among infected cystic fibrosis (CF) patients. The B. cepacia complex consists of five distinct subgroups, termed genomovars. A collection of 17 presumptive B . cepacia isolates, obtained from three national CF referral centres located in different geographical regions in Ireland, was studied. The aim of this study was to investigate these isolates using molecular subtyping protocols for evidence of genetic relationships and for the presence of antibiotic resistance-encoding class 1 integron structures. Methods and Results: Genomovar classifications were assigned to each isolate based on Hae III enzyme profiles of their recA locus. Genetic relationships among this collection were also assessed after restriction fragment length polymorphism (RFLP)-mediated analysis of the 16S rDNA locus and DNA amplification fingerprinting (DAF). The surface expression of the cable pilus gene (cblA ) may facilitate an early step in the infection process. All isolates were tested by amplification strategies for this marker. Burkholderia cepacia is known to be resistant to several antimicrobial agents. Resistance typing showed that the majority were resistant to three or more common antimicrobial agents. Five of the 17 isolates were resistant to sulphonamide, a characteristic linked with the presence of class 1 integrons. Gene cassettes containing beta-lactamase (oxa ) and aminoglycoside acetyltransferase (aac (6')-1a) encoding genes were identified by polymerase chain reaction amplification. Conclusions: Most of the isolates in this study were classified as genomovar III and were indistinguishable based on their corresponding 16S rDNA-RFLP profiles, whilst DAF further subtyped the collection. The cblA marker was identified in 47% of the isolates, many of which clustered in the genomovar III group. Class 1 integrons with recombined gene cassettes containing bla- (OXA) and aac (6')-1a genes were identified. Significance and Impact of the Study: This study demonstrates the application of molecular methods to investigate B. cepacia , a well-recognized human pathogen, cultured from Irish CF patients. Genomovar III was the most common genomic type identified. DNA fingerprinting further subtyped the latter isolates, facilitating a more detailed description of the molecular epidemiology. Drug resistance in these organisms can be explained, at least in part, by the presence of class 1 integrons. Development of targeted infection control strategies could be facilitated using these applied methods.