The ancient radiation of imperilled harlequin toads (Bufonidae: Atelopus) sheds light on historical biogeography of northern South America

Harlequin toads (Atelopus) are the most species-rich Neotropical bufonid genus. Their wide distribution across the tropical Andes, adjacent mountain systems, Chocoan and Amazonian lowlands makes them a promising model to study the historical assembly of northern South American biota, yet their phylogenetic relationships and their biogeographic history have remained poorly resolved. Here, we reconstruct a mitogenomic time-calibrated phylogeny of Atelopus covering 73 of the ~130 described and undescribed species. Our results support an ancient origin of the genus in northwestern South America during the late Eocene, followed by early divergences involving lineages associated with the Sierra Nevada de Santa Marta and the northeastern Andes. Extant diversity reflects a mixture of old, species-poor lineages and more extensive radiations from the late Miocene onwards. Biogeographic analyses indicate that the diversification of Andean lineages was shaped by progressive mountain building, regional isolation and repeated secondary range expansion, whereas the Amazonian clade likely originated in western Amazonia during Miocene wetland dynamics and drainage reorganization. The data further suggest that humid montane forests represented the ancestral environment for much of the genus, with repeated and more recent colonization of high-elevation habitats such as páramo and puna. Founder-event dispersal appears to have contributed to recent radiations into eastern Amazonia but also in the southern Andes. Together, these results identify Atelopus as an ancient radiation that preserves biogeographic signals of early diversification in northern South America and provide an evolutionary framework for conservation of one of the most threatened amphibian groups worldwide.