Conclusion

Our study aimed to investigate the support for the two main speciation hypotheses (ecological and allopatric) used to explain the evolution ofM. festivus within the Amazon basin. We showed strong evidence that the divergent physicochemical characteristics between black and white water have a weak structuring power on M. festivuspopulations in the Amazonian watershed. Furthermore, our results challenge the recently suggested ecological speciation hypothesis explaining fish diversification in Amazonia. Unlike previous studies focusing on a single confluence between black and white water, our extensive sampling design comprising 12 populations of M. festivus detected a genetic structure congruent with isolation by unidirectional downstream water current, past geologic events, and waterways connectivity shift. While the Brazilian Amazon supports one of the richest fish faunas on Earth, our comprehension of the evolutionary processes which shaped its biodiversity is still lacking. Understanding the origin of such richness would help us protect its diversity. Our study not only constitutes a step forward in understanding these important processes but also provides a conceptual framework that should benefit the sampling designs of future investigations on this matter.