An investigation of freshwater fungi in Thailand resulted in the collection of one new monotypic genus, Neoxylomyces, and a novel species each in Camposporium, Brunneofusispora, Rattania, Neoxylomyces, and Phaeoacremonium. Camposporium dulciaquae resembles C. septatum in conidial morphology and number of septa but differs in conidial sizes. Brunneofusispora hyalina is similar to B. sinensis in conidiogenesis and conidial shape but differs in the sizes of conidiomata and conidiogenous cells. Rattania aquatica is the second species in Rattania, while Phaeoacremonium thailandense is the third species recorded from freshwater habitats. A new genus, Neoxylomyces, typified by N. multiseptatus, is similar to Xylomyces giganteus, but differs in the number of septa, chlamydospore measurements, and absence of a mucilaginous coating around the chlamydospores. These novel taxa form an independent lineage distinct from other species based on multi-loci phylogenetic analyses. Descriptions, illustrations, and notes are provided for each taxon. These new freshwater ascomycetes add to the increasing number of fungi known from Thailand and it is now evident that there are numerous novel taxa awaiting to be described as new freshwater habitats are explored. An update of newly discovered taxa in the widely studied freshwater habitats of Thailand over the last five years is also provided.