Clustered cases of nosocomial pulmonary infections were observed in a French intensive care unit. Biochemical tests showed the etiologic agents to be part of the Bcc (Bcc). recA polymerase chain reaction-restriction fragment length polymorphism analysis and molecular phylogeny positioned the isolates into Burkholderia cenocepacia. Their recA sequences were found identical to those of ET12 strains responsible of necrotic pneumonia in cystic fibrosis patients. Analyses of a multi locus sequence typing genes set confirmed this proximity and suggested a wide distribution among occidental countries but could not resolve their phylogenetic position unambiguously. A novel marker, ecfB, indicated a significant phylogenetic divergence from ET12 strains. Pulse field gel electrophoresis analysis of SpeI-restricted total genomic DNA of the strains showed a unique profile indicative of a clonal outbreak. Environmental hospital screenings indicated cross-contamination between staff and patients. Bcc strains from outdoor environments were not related to this clone but indicated the presence of Burkholderia multivorans and Burkholderia vietnamiensis.