Hypostomus commersoni Valenciennes 1836, Hypostomus cordovae (Günther 1880) and Hypostomus laplatae (Eigenmann 1907) have been little studied since their original descriptions. This study shows a comprehensive review of these species from the Lower La Plata Basin, including their taxonomic history, distribution, color patterns, morphology, and ecological and molecular phylogenetic data. Morphological and phylogenetic analyses based on D-loop sequences suggested that H. commersoni can be separated into two subclades, or subgroups. Based on these results and on the non-overlapping distribution range of the two subclades, we conclude that they represent two distinct species, thereby revalidating H. spiniger. The results also suggest that H. paranensis should be considered as species inquirenda and H. cordovae as valid species. This integrated approach provides key information for assessing the conservation status and biogeographic aspects of the genus Hypostomus in the Lower La Plata Basin.
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