Multi-locus phylogeny and evolution of a Sky Island species complex reveal unexpected incongruence between morphological and molecular data

Sky islands archipelago is largely known to promote remarkable speciation rates and consequently holding high numbers of endemic species. Ololygon machadoi is a sky-island dweller reported to occur at the Espinhaço Mountain Range and at Serra da Canastra. However, these two mountains are separated by approximately 250km of lowlands, thus, being very unlikely that populations of both mountains correspond to the same species. Additionally, during field expeditions to sites along the Espinhaço, we have collected adult specimens morphologically similar to O. machadoi, but with tadpoles presenting conspicuous differences in color pattern. Through multi-locus phylogenetic analyses with samples from all known populations and many others, we recovered O. machadoi as paraphyletic. Within the large clade formed by samples from the Espinhaço, we have found that the differences of tadpole’s color pattern correspond to a remarkable intraspecific variation since one of the distinct morphotypes is nested within the other. We then performed three species delimitation methods (GMYC, BPP and STACEY) to investigate populational structure along the Espinhaço and a dated phylogenetic analysis. The most conservative method recovered 15 populations. Most of these populations are clearly in accordance with the sky island configuration of the Espinhaço Range, suggesting that geographic vicariant events may have played a major role in the current population structure. This possible explanation was also supported by the results of the dated species tree. The results suggest that the majority of the populational diversification occurred during the Pliocene, when average temperatures become 2–3°C warmer than they are today.