Testing molecular date estimates using an ecological and vicariant case study in treefrogs: The Geological‐Ecological Molecular Calibration.

Aim: To date a vicariant event through the mutualistic relationship of a hylid frog and its bromeliad host found across two isolated mountain ranges as part of an island–continental split and to use this information to calibrate a molecular dating analysis of hylids. Location: South America. Time Period: Pliocene. Major Taxa Studied: Treefrogs, Arboranae. Methods: Pliocene tectonic movements sank a portion of a mountain range that connected present‐day Trinidad (island) and northern Venezuela, eventually isolating populations of the golden tree frog Phytotriades auratus and its host bromeliad Glomeropitcairnia erectiflora on both sides of a saltwater barrier. We estimated the submersion rate timing of the vicariant event, then employed ecological niche modelling to establish the distribution of the frog and its host to the Pliocene. We generated nucleotide sequence data for the hylid on each side of the marine barrier and proposed a biogeographic calibration for the split between populations. Using Bayesian phylogenetic analysis, we evaluated the impact of incorporating this biogeographic calibration on estimates of treefrog divergence times. Results: A relatively continuous, high‐elevation mountain range existed before the Pliocene and the opening of the Gulf of Paria (4.45 ± 0.85 Ma). Phytotriades auratus and G. erectiflora would have been distributed across the Paria and Northern Range mountains as a single population since the Pliocene. Divergence times among treefrogs (Arboranae) were older than those inferred using fossil calibrations alone, with mean age estimates for subfamilies and genera being ~2 and ~1.2 million years older, respectively. Main Conclusions: Our study presents a biogeographic calibration based on a vicariant model that connects the evolution of the populations of a hylid through geology and ecological data. The implementation of this date as a molecular clock calibration reduces the uncertainty in date estimates for the shallower nodes in the phylogeny of Arboranae. [ABSTRACT FROM AUTHOR]