Abstract: Chinese chestnut (Castanea mollissima) dieback and canker diseases are common in plantations, affecting tree health and nut yield. Based on the nationwide disease survey of chestnut cultivation areas, fungal strains associated with diseases were obtained by tissue isolation and single-spore isolation methods from samples collected in Wangmo County, Qiannan Prefecture, Guizhou Province, China. Isolates were identified by combined morphological characteristics and molecular phylogeny. As a result, isolates CFCC 71068 and N317B represent a new species of the genus Cytospora, named Cytospora. minispora. The results enrich the category of chestnut pathogens and provide foundational data for subsequent taxonomic studies and disease control.