Retrospective Analysis of Clinical Filamentous Basidiomycetes and Development of Multiplex PCR for Rapid Identification

Pathogenic basidiomycetes are increasingly recognized as causative agents of mycoses, particularly allergic bronchopulmonary mycosis (ABPM), in humans. To clarify their clinical significance, we analyzed the fungal species, their sources and associated diseases of 650 basidiomycetous strains preserved at the Medical Mycology Research Center, Chiba University, from 2010 to 2024. Schizophyllum commune was the most frequently isolated species (60%), followed by Trametes spp. (9.5%), Irpex spp. (5.2%), and Bjerkandera adusta (4.3%). Most isolates originated from respiratory specimens and were associated with ABPM, pneumomycosis, or sinusitis. These fungi often cannot be identified in culture because of their limited morphological characteristics. Therefore, we developed a multiplex polymerase chain reaction (PCR) assay targeting the internal transcribed spacer region for the rapid discrimination of S. commune, B. adusta, Irpex spp., and Trametes spp. Species- and genus-specific forward and reverse primers yielded distinct amplicon sizes for each target. The assay demonstrated high specificity and showed no cross-reactivity with non-target fungi; the results were concordant with those of rDNA sequencing. Multiplex PCR provides a rapid, cost-effective tool for identifying clinically important basidiomycetes, facilitating the timely diagnosis and management of fungal respiratory diseases.