Identification of medically significant fungal genera by polymerase chain reaction followed by restriction enzyme analysis

Rapid non-culture-dependent assays for identification of fungi quicken diag nosis and prompt treatment of invasive fungal disease. Fungal DNA extracts from pure cultures of the most frequently isolated fungal pathogens belongi ng to the Genera Aspergillus, Candida and CI Cryptococcus along with less c ommon pathogenic Genera were amplified with the general fungal primer pair internal transcribed spacer-1/4. Subsequently, the amplicon was digested wi th the restriction endonucleases MspI, HaeIII, HinfI and EcoRI in order to generate genus- or species-specific patterns for identification of the fung us. HinfI produced indistinguishable fingerprints for all Aspergillus speci es tested, MspI produced species-specific patterns for: Cryptococcus neofor mans, Cryptococcus non-neoformans, Candida albicans and Candida tropicalis. EcoRI succeeded in differentiating penicillia from aspergilli and cryptoco cci from Candida spp, It is concluded that this procedure can differentiate genera and occasionally species of medically important fungi and that foll owing the necessary validation experiments, it can be used directly on clin ical samples to assist prompt diagnosis of systemic fungal infections. (C) 1999 Federation of European Microbiological Societies. Published by Elsevie r Science B,V. All rights reserved.