Development of a DNA array for the simple identification of major filamentous fungi in the beverage manufacturing environment

Filamentous fungi were isolated from the indoor environment of a soft drink manufacturing plant and ordinary residences. The isolated strains were identified based on morphological observation and the nucleotide sequences of the region near the D2 region of the 26S rDNA. Three genera (Aspergillus, Penicillium, and Cladosporium) accounted for 48.1% of the fungal strains detected in the manufacturing plant and 75.3% in residences. A DNA array for identification of 15 genera and 26 species of filamentous fungi that were most frequently isolated from the manufacturing plant was developed. Genus- and species-specific probes with 13- to 20-mer were designed on the basis of the nucleotide sequences in the D2 region. The probes were affixed to a microscope slide after modifying an amino group at the 5′ or 3′ end. To prevent erroneous identification, 2 or 3 probes were designed for each of the target genera and species. The developed DNA array method correctly identified 9 genera (Alternaria, Aureobasidium, Cladosporium, Curvularia, Exophiala, Fusarium, Penicillium, Phoma, and Trichoderma) and 26 species belonging to 6 genera (Aspergillus, Neosartorya, Byssochlamys, Talaromyces, Paecilomyces, and Purpureocillium) in the strains isolated from the indoor environment. Identification results obtained by this DNA array method of fungi isolated from the manufacturing plant were consistent with those by the conventional method.