A Comparative Phylogenetic Study Of Qinling Tree Frog And Hezheng Ji Frog

The Tsinling-Bashan and Dabieshan Mountains are located in central China,stretching from west to east.These Mountains are one of centers for the origin and evolution of biodiversity in China.These areas contain rich animal and plant resources.The Oriental and Palaearctic species congregate here forming a specific biotic province.The past climatic events,such as the Quaternary glaciation,could leave vestiges in geographical distribution of genetic diversity of population.Like other regions in the northern hemisphere,the Tsinling-Bashan Mountains also experienced several glacial-interglacial cycles.Understanding the pattern of genetic structure and diversity condition of fauna distributing on Tsinling-Bashan and the effects of palaeoglaciation on geographical distribution pattern have great value.Hyla tsinlingensis and Microhyla mixtura is an endemic frog restricted to a patchy distribution in the Tsinling-Bashan Mountains and Dabieshan mountains in china,a distribution that is becoming increasingly small.However,their molecular phylogeography and population genetic structure remain poorly investigated.In this study,we explored the genetic structure and phylogeography of H.tsinlingensis and M.mixtura based on partial sequences of the mitochondrial DNA(mtDNA)cytochrome b and cytochrome c oxidase subunit I genes.At the same time,we test whether the topography of the mountains affected the population genetic structure of H.tsinlingensis and M.mixtura according to partial sequences of the mitochondrial DNA cytochrome b(mtDNA cyt b)and cytochrome c-oxidase subunit I(mtDNA COI)genes.We also determined whether climatic oscillations during glacial periods in the Quaternary affected the distribution of them.And to explore these two sympatric species whether have the same phylogeographical structure.The results of investigations are offered as follows:1.A total of 212 adults were used in this study.32 haplotypes and four clades were found.Population comparisons showed significant or very significant genetic differentiation(FSTbetween most local populations.The AMOVA analysis suggests that most of the variation was attributed to among groups.Mantel tests indicated the existence of isolation-by-distance pattern of genetic divergence.The genetic barrier prediction analysis using Monmonier' maximum difference algorithm identified two putative barriers to gene flow that separate lineages of H.tsinlingensis.SAMOVA analysis indicated the existence of phylogeographic structure in this species.The maximum parsimony network yielded two unconnected subnetworks,Mismatch distribution and neutrality tests showed sudden population expansion in all clades but the total population and Tsinling population.Phylogenetic analyses revealed two deeply divergent and reciprocally monophyletic mtDNA lineages approximately corresponding to two geographical regions(Tsinling and Dabieshan Mountains)separated by complicated topography and long distance.The distinct geographic distributions of both lineages and the estimated divergence time suggested spatial and temporal separation coinciding with climatic changes during the Pleistocene.Analysis of molecular variance indicated that most of the observed genetic variation occurred between both groups(Tsinling and Dabieshan groups),implying long-term interruption of gene flow and the possible separation of H.tsinlingensis into two management units for conservation.BSP revealed past population dynamics and sudden expansion between 0.50 Myr and 0.23 Myr for H.tsinlingensis.The expansion times of individual haplogroups estimated according to T = ?/(2ut)were 0.326 Myr to 0.135 Myr,which are partially consistent with the result of BSP analysis.The results of the BEAST estimation showed the estimated age of H.tsinlingensis to be 3.26 Myr in the middle of Pliocene,with a 95%interval of confidence between 1.24 and 7.61 Myr.The divergence time between haplogroups A+B+C(Tsinling population)and haplogroup D(Dabieshan population)was calculated to have occurred in the early Pleistocene(1.74 Myr;95%HPD:0.71 Myr to 3.95 Myr).In conclusion,the topography of the Tsinling and Dabieshan Mountains significantly affects the population genetic structure of H.tsinlingensis,and climatic oscillations during glacial periods in the Quaternary affected the distribution of the frog species.2.A total of 229 adults were used in this study.61 haplotypes and two clades were found.The AMOVA analysis suggests that most of the variation was attributed to among groups.SAMOVA analysis detected the existence of phylogeographic structure in this species.Mantel tests indicated the existence of isolation-by-distance pattern of genetic divergence.Population comparisons showed significant genetic differentiation(FSTbetween most localities.Nested clade analysis(NCA)detected the existence of the restricted gene flow with isolation by distance.Harpending's raggedness index(R)and the sum of squared deviations(SSD)showed sudden population expansion in the Tsinling-Bashan population and the Dabieshan population.Both mismatch distribution and the neutrality tests rejected a sudden population expansion in the total population.Sixty-one haplotypes and two haplogroups were found among 229 samples.Phylogenetic analyses revealed two deeply divergent and reciprocally monophyletic mtDNA lineages approximately correspond to two geographic regions(Tsinling-Bashan and Dabieshan Mountains)separated by complicated topography and long distances.The distinct geographic distributions of both lineages and the estimated divergence time suggest spatial and temporal separation coinciding with climatic changes during the Pleistocene.Analysis of molecular variance indicated that most of the observed genetic variation occurred between both groups(Tsinling-Bashan and Dabieshan grous),implying long-term interruption of gene flow and the possible separation of M.mixtura into two management units for conservation.BSP revealed past population dynamics.and sudden expansion time was 0.05-0.015 Myr for M.mixtura.And the expansion time of individual clade estimated according to T= ?/(2ut)were 0.04808-0.03831Myr,which were consistent with the analysis of the BSP.The results of the BEAST estimation showed the divergence time between Haplogroup A and Haplogroup B to be 4.45 Myr in the early Pliocene,with a 95%interval of confidence between 2.73 and 6.26 Myr.The topography and the isolation by distance played an important role in the phylogeographical structure and the speciation event in M.mixtura.Therefore,The complexity of topography and isolation by distance are contributed to the genetic diversity and the phylogeographical structure.In conclusion,The Tsinling-Bashan Mountains and the Dabieshan Mountains revealed significant impact on the population genetic structure of M.mixtura,and climatic oscillations during glacial periods in the Quaternary affected the distribution of the frog species.3.we consider that there are significant genetic differentiations between populations of M.mixture and H.tsinlingensis distributing on the Tsinling-Bashan and Dabieshan Mountains,and we successfully reveal phylogeographic structure with them.The topography and the isolation by distance and climatic oscillations during glacial periods in the Quaternary played an important role in the phylogeographical structure and the distribution of them.By comparison,we found these two sympatric species of amphibians have a similar phylogeographical structure.