Characterization of PCR-ribotyping for Burkholderia (Pseudomonas) cepacia

Abstract

Ribotyping, a method of genotyping bacterial isolates for epidemiologic study, uses rRNA as a probe to detect chromosomal restriction fragment length polymorphisms. Although ribotyping is accurate, its utility is limited by the labor and time necessary for Southern blot analysis. PCR-ribotyping uses PCR to amplify the 16S-23S intergenic spacer region of the bacterial rRNA operon. Length heterogeneity in the spacer region has previously been found to be useful as an alternative to standard ribotyping in a study of Burkholderia (Pseudomonas) cepacia. To further analyze the accuracy of PCR-ribotyping, three groups of previously characterized isolates of B. cepacia were investigated. PCR-ribotyping grouped 90 isolates recovered from seven well- defined epidemics into the correct outbreak group with a mean concordance of 93%. Both standard ribotyping and PCR-ribotyping separated 15 unrelated isolates into 14 types. In an analysis of 83 B. cepacia isolates from chronically colonized cystic fibrosis patients, the concordance of PCR- ribotyping with standard ribotyping ranged from 83 to 100%, with a mean of 98%. One isolate from a chronically colonized patient had a different type by standard ribotyping but was identical to the other isolates from this patient by PCR-ribotyping. Thus, PCR-ribotyping is a rapid and accurate method for typing B. cepacia and is less labor intensive than standard ribotyping.