Rapid identification of fungi by using the ITS2 genetic region and an automated fluorescent capillary electrophoresis system

Invasive fungal disease often plays an important role in the morbidity and mortality of immunocompromised patients. The poor sensitivity of current fu ngal blood culture and histological practices has led to the development of highly sensitive and specific molecular techniques, such as the PCR. Seque nce variability of the internal transcribed spacer 2 (ITS2) region of fungi is potentially useful in rapid and accurate diagnosis of clinical fungal i solates. PCR with fungus-specific primers targeted toward conserved sequenc es of the 5.8S and 28S ribosomal DNA (rDNA) results in amplification of the species-specific ITS2 regions, which are variable in amplicon length. We h ave made use of the ABI PRISM 310 genetic analyzer and the ABI PRISM 310 Ge neScan analysis software for the determination of variable size differences of the ITS2 region of clinically important fungi, including Candida and no n-Candida yeasts, Aspergillus species, and a variety of dermatophytes. No c ross-reaction occurred when samples were tested against human and bacterial genomic DNA. We have found that most clinically significant fungal isolate s can be differentiated by this method, and it therefore serves to be a pro mising tool for the rapid (<7 h) diagnosis of fungemia and other invasive f ungal infections.