The global fauna of Didymozoidae infecting fishes is very diverse and includes 270 species. Integrative taxonomic studies in this group are rare, and genetic data are lacking for the molecular identification of species. In this study, a new genus and species, Platodidymocystis yamagutii n. gen., n. sp., is described based on morphological and genetic analyses. The new genus is allocated to Didymozoinae but differs from all other genera of this subfamily, mainly by the morphology of the testes and ovary. The genetic analysis and molecular phylogeny using 28S rDNA and ITS2 markers showed P. yamagutii n. gen., n. sp. in a unique cluster in monophyly and most closely related to Platocystis vivipara (Yamaguti, 1970), followed by Didymocystis spp. Additionally, Didymosulcus philobranchiarca (Yamaguti, 1970) was also characterized by integrative taxonomy and Koellikerioides internogastricus Yamaguti, 1970, Didymocystis neothunni (Yamaguti, 1970), and P. vivipara were analyzed by molecular taxonomy. This is the first integrative taxonomic study of Didymozoidae from the Atlantic coast and the first survey to provide novel genetic data for five species of Didymozoidae trematodes, contributing to the increment of the knowledge and expansion of the geographical distribution of didymozoid species parasites of Scombridae in the Southern Atlantic Ocean.