Morphological differences in the otoliths of Macrodon atricauda from Rio Grande do Sul State (southern Brazil) and of M. ancylodon from Pará State (northern Brazil) were examined. Until recently, these were considered two populations of the same species. Although relatively similar in their general shape, those of large M. atricauda (>300 mm total length) were shorter and thinner and their outer surface was convex (those of the smallest specimens had a more lobullated antero-dorsal margin) whereas those of similarly sized M. ancylodon were larger, thicker and less lobullated. These features of the otoliths allow these morphologically similar species to be discriminated. The pattern of translucent and opaque bands in sectioned otoliths also differed. The otoliths of M. atricauda showed clear annual banding which allowed ageing, while those of M. ancylodon had no distinct alternate bands.