Polymorphic internal transcribed spacer region 1 DNA sequences identify medically important yeasts

Species-specific polymorphisms in the noncoding internal transcribed spacer 2 (ITS2) region of the rRNA operon provide accurate identification of clin ically significant yeasts. In this study, we tested the hypothesis that ITS 1 noncoding regions contain diagnostically useful alleles. The length of IT S1 region PCR products amplified from 40 species (106 clinical strains, 5 r eference strains, and 30 type strains) was rapidly determined with single-b ase precision by automated capillary electrophoresis. Polymorphisms in the PCR product length permitted 19 species to be distinguished by ITS1 alone, compared with 16 species distinguished by using only ITS2. However, combina tion of both ITS alleles permitted identification of 30 species (98% of cli nical isolates). The remaining 10 species with PCR products of similar size s contained unique ITS alleles distinguishable by restriction enzyme analys is. DNA sequence analysis of amplified ITS1 region DNA from 79 isolates rev ealed species-specific ITS1 alleles for each of the 40 pathogenic species e xamined. This provided identification of unusual clinical isolates, and 53 diagnostic ITS1 sequences were deposited in GenBank. Phylogenetic analyses based on ITS sequences showed a similar overall topology to 26S rRNA gene-b ased trees. However, different species with identical 26S sequences contain ed distinct ITS alleles that provided species identification with strong st atistical support. Together, these data indicate that the analysis of ITS p olymorphisms can reliably identify 40 species of clinically significant yea sts and that the capacity for identifying potentially new pathogenic specie s by using this database holds significant promise.