Freshwater fungi are a poly-phylogenetic group of taxonomically diverse organisms. In this study, we isolated diverse fungal strains from various environmental samples obtained from freshwaters in Korea. These strains were identified by performing molecular phylogenetic analyses of rDNA and/or other sequences (beta-tubulin, RNA polymerase II, and translation elongation factor 1). In addition, we examined their morphological characteristics microscopically and cultural characteristics using different media. We identified eleven unrecorded Pezizomycotina species: Cladosporium angulosum, Pseudorobillarda phragmitis, Paraconiothyrium estuarinum, Pseudopithomyces palmicola, Pyrenochaetopsis paucisetosa, Thelebolus globosus, Plagiostoma mejianum, Trichoderma cremeum, Fusarium tanahbumbuense, Coniochaeta endophytica, and Chaetomium tenue. Environmental samples obtained from different freshwater ecosystems in Korea could thus be a good source for isolating and investigating novel fungal species.