Use of real-time PCR and the LightCycler system for the rapid detection ofPneumocystis carinii in respiratory specimens

Pneumocystis carinii pneumonia (PCP) remains a major cause of morbidity and mortality in immunocompromised patients, including those infected with hum an immunodeficiency virus (HIV). The advent of real-time PCR technology off ers the potential for rapid PCR results for the detection of P. carinii. In this report we describe the modification and evaluation of an existing PCR -based method for the detection of P. carinii DNA, into a real-time PCR ass ay suitable for use with the LightCycler system. Twenty eight induced sputu m and bronchial washing specimens from 28 patients were tested by both at c onventional PCR assay and a real-time PCR assay. Twelve specimens (42.9%) w ere positive in both the conventional and real-time PCR assays and sixteen (57.1%) were negative in both assays. The melting points of the amplified P . carinii DNA product obtained by melting curve analysis by the LightCycler of all P. carinii positive specimens ranged from 81.5 degreesC to 83.9 deg reesC. There were no discordant results between the two assays for any of t he specimens tested and results were available within 2 h for the real-time PCR assay compared to up to 11 h for the conventional PCR assay. (C) 2001 Elsevier Science Inc. All rights reserved.