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• Integrative tools support species identification in a problematic group. • Previously considered endemic of Argentina, Opuntia bonaerensis has a broader distribution. • Pleistocene events have impacted the distribution of cacti in southern South America. • Update on distribution reveal new insights for the historical biogeography of the species. • Knowledge regarding species distribution will affect its assessment for conservation. Geographical range is one of the critical features for species conservation assessment. Nevertheless, species distribution is frequently unknown, undervalued or overlooked. During a broad taxonomic and floristic study of the southern South American prickly pear species (Opuntia spp.), new records of a species previously considered endemic to Argentina have been found in Uruguay and Brazil. Molecular phylogenetic inference was carried out to further evaluate the identity of the new records, and ecological niche models were implemented to test how the new records would fit in the previous known distribution of the species. Through molecular and morphological evidence, we confirmed the new records of Opuntia bonaerensis for Brazilian and Uruguayan floras and discussed its phylogenetic relationship and morphological similarities with closely related species. Our new records uncovered a distribution pattern congruent with the Neotropical Peripampasic Orogenic Arc, which must be further explored to better determine the biogeographic history of the species. Ecological niche models (ENM) revealed that O. bonaerensis likely had a putative ancient distribution across the grasslands and shrublands in the Pampean region largely congruent with the populations found in Brazil and Uruguay, suggesting relictual Pleistocene populations of the species and the role of glacial/interglacial cycles on the distribution of the species. In a prospective climate change scenario, ENM suggests that the species would in general be more restricted to the southernmost limits of the Pampa region and previous outlying records from Mendoza (Argentina) are a putative future refuge for O. bonaerensis. The importance of these new records for biodiversity and conservation assessment efforts that are ongoing at different scales in Brazil and neighboring areas is highlighted. [ABSTRACT FROM AUTHOR]