Aim We investigate the spatiotemporal context of the diversification ofAllobates, a widespread genus of Amazonian frogs with high species diversity particularly in western Amazonia. We tested if that diversity originated in situ or through repeated dispersals from other Amazonian areas and if this diversification took place during or after the Pebas system, a vast lacustrine system occupying most western Amazonia between 23 and 10 million years ago (Mya). Location Amazonia. Taxon Allobates(Anura: Aromobatidae). Methods We gathered a spatially and taxonomically extensive sampling of mitochondrial DNA sequences from 932Allobatesspecimens to delimit Operational Taxonomic Units (OTUs). Complete mitogenomes of these OTUs were assembled to reconstruct a time-calibrated phylogeny used to infer the historical and spatial origin of the AmazonianAllobateslineages. Results Phylogenetic analyses and ancestral area reconstruction suggested that most of the western Amazonian lineages resulted from in situ diversification and that these events occurred between the inferred origin of the genus (25 Mya) and 10 Mya, with a possible peak between 14 and 10 Mya. Dispersal among areas mostly occurred from western Amazonia towards the Brazilian and the Guiana Shields. Closely relatedAllobatesOTUs display an allopatric pattern of distribution, matching interfluves delimited by modern Amazonian rivers. Main Conclusions In western Amazonia, diversification ofAllobatesappears to have been simultaneous with the last stages of the Pebas system (14-10 Mya). Subsequently (within the last 10 Mya), modern Amazonian river courses shaped the distribution pattern ofAllobatesspecies and possibly promoted allopatric speciation.
