Studies On The Comparative Molecular Phylogeography Of Two Frogs In Zhoushan Archipelago

Zhoushan Archipelago with unique geographical environments has been consider to be an ideal platform for researches on animal evolution and biogeography.It had experienced marine transgression,regression and tectonic events since the Pleistocene,which could probably influence on the migration,isolation and evolution of various animals distributed on the islands.Amphibians are widely distributed in Zhoushan Archipelago and are very sensitive to climate change and are ideal materials for molecular phylogeographical researches.The study selected two frogs,Fejervarya limnocharis and Rana zhenhaiensis,to analyze the origin,genetic differentiation and historical dynamics of populations in Zhoushan Archipelago and China mainland.Through the dating estimation,the role of vicariance vs.dispersal in shaping genetic structures of populations was discussed,and the underlying biogeographic events affecting the tempo-spatial distribution of the ancestral populations were revealed.The main results are as follows:(1)Population molecular phylogeography of F.limnocharisA total of two mitochondrial DNA(mtDNA)fragments(ND1-ND2,Cytb-CR)and eight nuclear DNA markers(CXCR4,Tyr,Rhod,3RAG-1,5RAG-1,BDNF,NCX1,RAG-2)were sequenced from 270 individuals collected from 46 localities of China continent and Zhoushan Archipelago to study the isolation,divergence and historical dynamics of populations of a rice frog F.limnocharis.Two major topological groups including 12 subgroups were estimated by phylogenetic analyses,which originated in different areas in southeastern China.Affected by the Dongsha movement(10-5Ma),the ancestors of the F.limnocharis diverged earliest about 5.86 Ma in southern China.The Zhoushan Archipelago subgroup(D)and the Zhoushan Archipelago coastal continent subgroup(L),respectively,located in the top branch of each of both major groups in the tree,indicating populations of Zhoushan Archipelago and the costal continent had different original ancestors.The ancestors of one major group diffused along the ancient Chinese continent to northward and emerged as an independent subgroup(D)in Zhoushan Archipelago during the period of Himalayan tectonic events(2.2-1.8Ma).Moreover,the ancestor of Zhoushan populations and the costal populations had undergone rapid range expansions since 25,000 years ago approximated at the end of Pseudorotalia transgression.Both subgroups D and L were geographically isolated with subgroup L emerged most recently which could diffuse very lightly to a small number of islands closest to the continent.Influenced by the transgression of the Jiangzhen(62-560,000 years ago),the ancestor population of the Hainan Island began to diverge from the mainland population.Analyses of phylogenetics and haplotype network concluded that Zhoushan populations has a closer relationship with populations in Japan main island rather than that of China continent,which supports the consideration of Zhoushan populations as an "hidden" evolutionary significant unit or genetic protection unit.(2)Molecular phylogeography of R.zhenhaiensisA total of two mitochondrial DNA(mtDNA)fragments ND2 and Cytb were sequenced from 118 individuals collected from 24 localities of China continent and Zhoushan Archipelago to study the isolation,divergence and historical dynamics of populations of R.zhenhaiensis.The species was considered to be originated in southeast China with two major clades including 7 subclades in phylogenetic trees.Populations of Zhoushan Archipelago and its adjacent coastal areas are originated by dispersion from the west parts of China.Affected by the Xuefeng Uplift(1.8-0.73Ma)and Wuling Uplift(1.8-0.73Ma),the ancestors of the R.zhenhaiensis in southwest China diverged earliest about 0.95 Ma.Influenced by the transgression of the Jiangzhen(62-560,000 years ago),the ancestor population of the southwest China began to diverge from the north and south population.Moreover,the ancestor of Zhoushan populations(G)and the mainland populations(C)had undergone rapid range expansions since approximate 50,000 years ago approximated at the end of Zhousha transgression(65,000-53,500 years ago).Affected by the Spirillina transgression(30-200,000 years ago),the ancestor population of Zhoushan Archipelago and the adjacent coastal areas began to split from the central China.(3)Suitability evaluation of potential geographic distribution for F.limnocharis and R.zhenhaiensis based on MAXENTIn this work,a total of 116 distributing localities of F.limnocharis and 55 distributing localities of R.zhenhaiensis collected from field investigation or literatures were utilized to predict the potential historical distribution of four distinct evolutionary clades of the rice frog F.limnocharis and R.zhenhaiensis through application of the maximum ecological niche model by using 19 biological environments.The prediction results had higher credibilities(AUC values>0.95),and the Jackknife test was used to test the contribution of 19 environmental variables to estimation of potential habitats.Our results showed that the environmental variable factors contributed with some difference to the perspective distribution of different populations among different historical periods,but the precipitation associated variables contributed majority on the estimation.The perspective range of F.limnocharis four evolutionary clades expanded in a large scale in the Last Glacial Maximum,and further undergone obviously retreat after glaciation and finally reached relative stable since middle Holocene.But no range expansion evidence of R.zhenhaiensis after the Last Glacial Maximum.The prospective range predicted by the model were similar but with some difference to the presently known.