ASSESSMENT OF A PCR TECHNIQUE FOR THE DETECTION AND IDENTIFICATION OFCRYPTOCOCCUS-NEOFORMANS

The 18S ribosomal RNA gene of Cryptococcus neoformans was amplified by polymerase chain reaction (PCR). The primers CPL1 and CPR4 were teste d for their ability to amplify DNA from 30 strains of C. neoformans an d 27 specimens of cerebrospinal fluid (CSF) from patients with cryptoc occal meningitis. A 343 bp product was obtained and its specificity co nfirmed by Southern hybridization with an internal sequence (INSR4) pr obe. The sensitivity was 100 fg by Southern analysis and 1 pg using th e PCR. Neither human nor a variety of other fungal and bacterial strai ns (n = 78) gave an amplified product. This PCR method can detect as f ew as 5 cells ml(-1) of C. neoformans in spiked-CSF following a simple processing procedure. The developed system of PCR was more sensitive than the culture method and revealed a very high specificity. The PCR was easy to perform and needed only 4 h for all processes from receivi ng the CSF to detection of a specific DNA band after agarose gel elect rophoresis. This would provide another rapid laboratory method for the diagnosis of cryptococcal meningitis.