Phyllomys is the most diverse genus of the subfamily Echimyinae, and is represented by 12 described species endemic to the Atlantic forest of eastern South America. We name and describe a new species of Phyllomys based on specimens collected in southern Brazil. This species is closely related to P. dasythrix and has been referred to as “Phyllomys aff. dasythrix” in the literature. Chromosomal restructuring was probably involved in their speciation because genetic and morphological similarities of the 2 contrast with their karyotypic distinctiveness. The new species of Phyllomys is diagnosed by a unique combination of external, cranial, and karyotypic characters. We propose the conservation status of “Least Concern” for Phyllomys sp. nov., but “Vulnerable” for P. dasythrix. We present an identification key to the living species of Phyllomys.