Detection of leaf-associated fungi based on PCR and nucleotide sequence of the ribosomal internal transcribed spacer (ITS) in Miscanthus

PCR amplification of the internal transcribed spacer region (ITS) of the nuclear ribosomal DNA from the genomic DNA isolated from the leaf tissue of Miscanthus sinensis varieties and M. floridulus resulted in two amplified fragments. Via FASTA search, the shorter length of about 500 base pairs was identified as the ITS region of fungi symbiotic in the grass foliage. Two Cladosporium species, one Fusarium and one basidiomycete were added to the mycoflora of Miscanthus grassland. Using a Southern blotting and hybridization, fungal ITS was also detected from the genomic DNAs isolated from sterilized seedling, a suggestive seed transmission of endophytic fungi. The "contaminated" genomic DNA may be a hindrance to the phylogenetic analysis of Miscanthus itself, but may provide other insights into the biodiversity of grassland ecosystems. The endophytic foliage fungi were not detected in M. sinensis var. condensatus, probably a result of its apomictic reproduction.