Aim To investigate biogeographical patterns of the obligate freshwater fish order Characiformes.
Location South America.
Methods Parsimony analysis of endemicity, likelihood analysis of congruent geographical distribution, and partition Bremer support were used.
Results Areas of endemism are deduced from parsimony analysis of endemicity, and putative dispersal routes from a separate analysis of discordant patterns of distribution.
Main conclusions Our results demonstrate the occurrence of 11 major areas of endemism and support a preferential eastern–western differentiation of the characiforms in the Amazonian region, contrasting with the southern–northern differentiation of terrestrial organisms. The areas of endemism identified seem to be deeply influenced by the distribution of the emerged land during the 100-m marine highstand that occurred during the late Miocene and allow us to hypothesize the existence of eight aquatic freshwater refuges at that time. The raw distribution of non-endemic species supports nine patterns of species distribution across the 11 areas of endemism, two of which support a southern–northern differentiation in the eastern part of the Amazon. This result shows that the main channel of the Amazon limited dispersal between tributaries from each bank of the river. The levels of endemism further demonstrate that the aquatic freshwater refuges promoted allopatric speciation and later allowed the colonization of the lowlands. By contrast, the biogeographical pattern found in the western part of the Amazon is identified as a result of the Miocene Andean foreland dynamic and the uplift of the palaeoarches that promoted allopatric divergence across several sedimentary basins by the establishment of disconnected floodplains. The assessment of conflicting species distributions also shows the presence of seven putative dispersal routes between the Amazon, Orinoco and Paraná rivers. Our findings suggest that, rather than there being a single predominant process, the establishment of the modern South American freshwater fish biotas is the result of an interaction between marine incursions, uplift of the palaeoarches, and historical connections allowing cross-drainage dispersal.