Aim: Pelophylax frogs in East Asia provide an opportunity to explore the impact of glacial cycling on demographic and genetic dynamics, because it has been suggested that they experienced distribution shifts and subsequent mtDNA introgression from Pelophylax plancyi to Pelophylax nigromaculatus in association with climatic oscillations. However, their evolutionary history, including the pattern of introgression, is incompletely understood. We used phylogenetic analyses based on multiple markers to address their evolutionary history, and palaeodistribution modelling to test whether the predicted distribution can explain the pattern of introgression suggested by molecular phylogenetics. Location: East Asia, including far-eastern Russia, mainland China, the Japanese archipelago, the Korean Peninsula and Taiwan. Methods: Ninety-nine samples of the Pelophylax nigromaculatus species complex were collected from 75 localities throughout the distributional range of each species. Phylogenetic analyses were performed with the mitochondrial cytochrome b (cytb) gene and six nuclear genes using two European Pelophylax frogs as outgroups. Additionally, palaeodistributions of P. nigromaculatus and P. plancyi were predicted using bioclimatic variables. Results: Differences were observed between the mitochondrial and nuclear DNA trees in the phylogenetic position of P. plancyi. Similar phylogenetic discordance was also observed within P. nigromaculatus. Distribution modelling suggested that a considerable distributional shift of P. nigromaculatus occurred in association with glacial cycling and that the distributional range of P. plancyi was relatively narrow during glacial periods. Main conclusions: The analyses detected five historical mtDNA introgression events and determined their directions, none of which had been deduced in previous studies. In association with glacial cycling, the distributional ranges of P. nigromaculatus and P. plancyi appear to have shifted repeatedly, resulting in multiple complex introgressions. By combining phylogenetic analyses with palaeodistribution modelling, our results supported the hypothesis that the history of mtDNA introgression among Pelophylax frogs was shaped by glacial cycling.
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