Polymerase chain reaction amplification of Asp f I and alkaline protease genes from fungus balls: clinical application in pulmonary aspergillosis.

Asp fI(18 kDa) and alkaline protease (33 kDa) are the 2 major antigens which are derived from Aspergillus (A.) fumigatus and have been implicated as possible virulence factors in the pathogenesis of Aspergillus-induced diseases. We attempted to detect fragments of genes encoding both proteins from fungus balls obtained at surgery or autopsy by polymerase chain reaction (PCR) amplification and then used PCR to test clinical samples. Frozen-stored fungus ball samples from a patient with acute myeloid leukemia complicated by Aspergillus pneumonia and from a patient with pulmonary aspergilloma were studied. We successfully amplified a 315 bp PCR product, the target sequence for Asp f I, and a 747 bp PCR product as a target sequence for alkaline protease (ALP) in both cases. In addition, 13 clinical samples including sputum specimens from patients with pulmonary aspergillosis were also examined. PCR analysis for the Asp f I (ALP) gene in clinical samples showed positive results in 5/10 (6/10) patients with pulmonary aspergilloma and in 3/3 (1/ 3) patients with invasive pulmonary aspergillosis. Culture data on A. fumigatus revealed positive results in 3/9 patients with pulmonary aspergilloma and in 2/3 patients with invasive pulmonary aspergillosis. This method can be used to recognize the involvement of A. fumigatus in various clinical settings where conventional culture results are not readily available.