Population diversification in the frog Mantidactylus bellyi on an isolated massif in northern Madagascar based on genetic, morphological, bioacoustic and ecological evidence

In the processes that give rise to new species, changes first occur at the population level. But with the continuous nature of the divergence process, change in biological properties delimiting the shift from "individuals of divergent populations" towards "individuals of distinct species", as well as abiotic factors driving the change, remain largely ambivalent. Here we study diversification processes at the population level in a semi-aquatic frog, Mantidactylus (Brygoomantis) bellyi, across the diverse vegetation types of Montagne d'Ambre National Park (MANP), Madagascar. Genetic diversity was assessed with seven newly developed microsatellite markers as well as mitochondrial DNA sequences and concordance with patterns of ecological, morphological, and bioacoustic divergence evaluated. We found M. bellyi lacking mitochondrial differentiation within MANP, while microsatellite datasets partitioned them into three highly differentiated, geographically separated subpopulations (with indications for up to five subpopulations). The molecular grouping-primarily clustering individuals by geographic proximity-was coincident with differences in mean depth and width of waters, suggesting a possible role of fluvial characteristics in genetic exchange in this stream-breeding species. Genetic clustering not consistent with differences in call properties, except for dominant call frequencies under the two-subpopulations model. Morphological divergence was mostly consistent with the genetic clustering; subpopulations strongly differed by their snout-vent length, with individuals from high-elevation subpopulations smaller than those from populations below 1000 m above sea level. These results exemplify how mountains and environmental conditions might primarily shape genetic and morphological divergence in frog populations, without strongly affecting their calls.