Rapid diagnosis of bacteremia by universal amplification of 23S ribosomal DNA followed by hybridization to an oligonucleotide array

The rapid identification of bacteria in blood cultures and other clinical s pecimens is important for patient management and antimicrobial therapy. We describe a rapid (<4 h) detection and identification system that uses unive rsal PCR primers to amplify a variable region of bacterial 23S ribosomal DN A, followed by reverse hybridization of the products to a panel of oligonuc leotides. This procedure was successful in discriminating a range of bacter ia in pure cultures. When this procedure was applied directly to 158 unsele cted positive blood culture broths on the day when growth was detected, 125 (79.7%) were correctly identified, including 4 with mixed cultures. Nine ( 7.2%) yielded bacteria for which no oligonucleotide targets were present in the oligonucleotide panel, and 16 culture positive broths (10.3%) produced no PCR product. In seven of the remaining eight broths, streptococci were identified but not subsequently grown, and one isolate of Staphylococcus au reus was misidentified as a coagulase negative staphylococcus. The accuracy , range, and discriminatory power of the assay can be continually extended by adding further oligonucleotides to the panel without significantly incre asing complexity or cost.