Much remains unknown about the natural history of respiratory tract infection by Burkholderia cepacia complex (Bcc) in persons with cystic fibrosis (CF). Specifically, it is not clear whether infected CF patients typically harbor a single Bcc strain or multiple strains that may be phenotypically indistinguishable. We genotyped 912 Bcc isolates recovered from CF sputum culture from in excess of 100 patients to demonstrate that chronic infection generally involves a single strain. Transient coinfection with more than 1 strain occurs infrequently and may be more common early in the course of Bcc infection.