Persistence in peripheral refugia promotes phenotypic divergence and speciation in a rainforest frog

It is well established from the fossil record and phylogeographic analyses that late Quaternary climate fluctuations led to substantial changes in species' distribution, but whether and how these fluctuations resulted in phenotypic divergence and speciation is less clear. This question can be addressed through detailed analysis of traits relevant to ecology and mating within and among intraspecific lineages that persisted in separate refugia. In a biogeographic system (the Australian Wet Tropics [AWT]) with a well-established history of refugial isolation during Pleistocene glacial periods, we tested whether climate-mediated changes in distribution drove genetic and phenotypic divergence in the rainforest frog Cophixalus ornatus. We combined paleomodeling and multilocus genetics to demonstrate long-term persistence within, and isolation among, one central and two peripheral refugia. In contrast to other AWT vertebrates, the three major lineages differ in ecologically relevant morphology and in mating call, reflecting divergent selection and/or genetic drift in the peripheral isolates. Divergence in mating call and contact zone analyses suggest that the lineages now represent distinct species. The results show that climate shifts can promote genetic and phenotypic divergence and, potentially, speciation and direct attention toward incorporating adaptive traits into phylogeographic studies to better resolve the mechanisms of speciation.