Misidentification of Burkholderia cepacia in US cystic fibrosis treatment centers: an analysis of 1,051 recent sputum isolates.

BACKGROUND: Burkholderia cepacia remains a significant pathogen in persons with cystic fibrosis (CF). The medical and psychosocial consequences of pulmonary colonization with this bacterium are enormous. However, B cepacia may be frequently misidentified from CF sputum culture. STUDY OBJECTIVES: To determine the rate of misidentification of B cepacia recently recovered from CF sputum culture of persons receiving care in US treatment centers. DESIGN: Bacterial isolates cultured from CF sputum and putatively identified as B cepacia or other related nonlactose-fermenting Gram-negative species were referred from participating treatment centers. Isolates underwent polyphasic analyses employing phenotypic (selective media and biochemical testing) and genotypic (polymerase chain reaction) assays to determine species identification. Taxonomic evaluations were performed by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis of whole-cell proteins and amplified-fragment length polymorphism analysis. MEASUREMENTS AND RESULTS: A total of 1,051 isolates recovered from 608 patients were received from 115 treatment centers in 91 US cities. Among the isolates identified as B cepacia by referring laboratories, 11% could not be confirmed as B cepacia by polyphasic analyses. In addition, 36% of isolates not specifically identified by the referring laboratory or identified as a species other than B cepacia were, in fact, found to be members of the B cepacia complex. CONCLUSIONS: Rates of misidentification of B cepacia remain unacceptably high among US treatment centers. These data suggest the need for increased awareness of this problem among CF centers and their affiliated laboratories, better adherence to recommended protocols for evaluation of CF sputum, and greater use of reference laboratories equipped to provide advanced analyses.

Development of rRNA-based PCR assays for identification of Burkholderia cepacia complex isolates recovered from cystic fibrosis patients.

PCR assays targeting rRNA genes were developed to identify species (genomovars) within the Burkholderia cepacia complex. Each assay was tested with 177 bacterial isolates that also underwent taxonomic analysis by whole-cell protein profile. These isolates were from clinical and environmental sources and included 107 B. cepacia complex strains, 23 Burkholderia gladioli strains, 20 Ralstonia pickettii strains, 10 Pseudomonas aeruginosa strains, 8 Stenotrophomonas maltophilia strains, and 9 isolates belonging to nine other species. The sensitivity and specificity of the 16S rRNA-based assay for Burkholderia multivorans (genomovar II) were 100 and 99%, respectively; for Burkholderia vietnamiensis (genomovar V), sensitivity and specificity were 87 and 92%, respectively. An assay based on 16S and 23S rRNA gene analysis of B. cepacia ATCC 25416 (genomovar I) was useful in identifying genomovars I, III, and IV as a group (sensitivity, 100%, and specificity, 99%). Another assay, designed to be specific at the genus level, identified all but one of the Burkholderia and Ralstonia isolates tested (sensitivity, 99%, and specificity, 96%). The combined use of these assays offers a significant improvement over previously published PCR assays for B. cepacia.