Identification of Aspergillus fumigatus and related species by nested PCR targeting ribosomal DNA internal transcribed spacer regions

Aspergillus fumigatus is the most common species that causes invasive asper gillosis. In order to identify A. fumigatus, partial ribosomal DNA (rDNA) f rom two to six strains of five different Aspergillus species was sequenced. By comparing sequence data from GenBank, we designed specific primer pairs targeting rDNA internal transcribed spacer (ITS) regions of A. fumigatus. A nested PCR method for identification of other A. fumigatus-related specie s was established by using the primers. To evaluate the specificities and s ensitivities of those primers, 24 isolates of A. fumigatus and variants, 8 isolates of Aspergillus nidulans, 7 isolates of Aspergillus flavus and vari ants, 8 isolates of Aspergillus terreus, 9 isolates of Aspergillus niger, 1 isolate each of Aspergillus parasiticus, Aspergillus penicilloides, Asperg illus versicolor, Aspergillus wangduanlii, Aspergillus qizutongii, Aspergil lus beijingensis, and Exophiala dermatitidis, 4 isolates of Candida, 4 isol ates of bacteria, and human DNA were used. The nested PCR method specifical ly identified the A. fumigatus isolates and closely related species and sho wed a high degree of sensitivity. Additionally, four A. fumigatus strains t hat were recently isolated from our clinic were correctly identified by thi s method. Our results demonstrate that these primers are useful for the ide ntification of A. fumigatus and closely related species in culture and sugg est further studies for the identification of Aspergillus fumigatus species in clinical specimens.