| Summary: | The accurate identification and proper typing of basidiomycetes are required in medical, sanitary maintenance, agriculture, and biotechnology fields. A diagnostic method based on information from whole-cell proteins acquired by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was investigated to identify wood-rotting fungi, a group of filamentous fungi. In this study, mass spectra of intracellular peptides obtained from cultured mycelia of 50 strains of 10 wood-rotting fungal species were obtained multiple times and mass spectral patterns (MSPs) consisting of peaks that characterized the fungal species or strain was created to construct an in-house database. The species identification was conducted by comparing the newly obtained raw mass spectra with the MSPs in the database using the MALDI Biotyper. The results showed that the peak patterns of the mass spectra were reproducible and matched at the strain level. A cluster analysis based on the MSPs was also conducted to examine inter-and intraspecific diversity among the tested wood-rotting basidiomycetes. Most of the fungal strains examined in this study could be identified to a species level; however, the strains belonging to <i>Pleurotus</i> could only be identified to a genus level. This was due to an intraspecific variation, so the identification accuracy could be amendable with a more enhanced database.
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