A new species of Aspidoras is described from the upper rio Itiquira and the upper rio Taquari, rio Paraguai basin, and represents the first record of the genus from this river system. The new species differs from its congeners by the following unique features: infraorbitals and preopercle covered by skin; nuchal plate reduced and covered by skin; and lateral line absent from body plates posterior to the two small lateral line ossicles. The new species exhibits the greatest body size observed for an Aspidoras species. The occurence of Aspidoras in tributaries of the rio Paraguai basin extends the range of the genus further southwest than previously known, encompassing most of the river basins that drain the Brazilian Shield.