Phylogeography Of TypicalRana Species In Northeast And North China

Biodiversity pattern and its formation mechanism is one of the key subjects of the biogeographic studies. It is the basis to understand the evolution of biodiversity, reveal the ecological and evolutionary process of speciation, and thus is critical important for setting biodiversity conservation and management strategies. Amphibians have been identified as the most threatened vertebrate group due to the highest percentage of the threated species among all species groups. Meanwhile, amphibian is the ideal species for phylogeographical and evolution of biodiveristy studies due to its low individual mobility, sensitive to habitat quality and its high loyalty to home range. Six Rana species are recorded in Northeast and North China, e.g., Rana amurensis Booulenger,1886, Rana kunyuensis Lu et Li,2002, Rana dybowskii Gunther,1876, a cryptic species of R. chensinensis(henceforth referred to as R. cf. chensinensis), Rana huanrenensis Liu, Zhang et Liu,1993, Rana culaiensis Li, Lu and Li,2008. Among the six known species to the region,Rana amurensis Booulenger,1886, Rana dybowskii Gunther,1876, a cryptic species of R. chensinensis(henceforth referred to as R. cf. chensinensis) are the widely distributed species, whereas Rana kunyuensis Lu et Li,2002is endemic to KunyuMountains in Shandong province. The other two species are extremely rare, and they were not found in the field survey, and thus, the study focus on the four Rana species (Ranidae) to address the following questions:(i) Whether the mtDNA mitochondrial genome structure and novel feature of the four Rana species is different from the known Rana species?(ii) What are the genetic structure and its demography of three widely distributed Rana species;(iii) What are the causes of phylogeographical patterns of three Rana species,? and whether the Climate barrier hypothesis, Riverrine barrier hypothesis and Ridge barrier hypothesis can explain such pattern?(iv) How to develop conservation strategies using the findings of this study?In order to address these questions, this study surveyed northest and northen China for the collection of experimental samples; applied mitochondrial genes and nuclear genes mark to obtain full gene sequences of the four species; reconstructed the phylogenetic relationship and population history using molecular clock, as well as the geological evidence, and obtained the following results:(i) Feature of complete mitochondrial genome structure of four Rana speciesThe mitogenome length of R. amurensis and R. kunyuensis were20,564bp and22,255bp, respectively, including13protein-coding genes,22transfer RNA genes,2ribosomal RNA genes, and2control region (D-loop).We first found a novel gene order arrangement with the translocation of tRNALeu(CUN) and ND5, and duplicated D-loop genes from this two species. The length of R. dybowskiiand R. cf. chensinensiswere18,864bp and18,808bp,respectively, including13protein-coding genes,22transfer RNA genes,2ribosomal RNA genes, and1control region (D-loop). The genes structure of above two species displayed a similar pattern to that of known Rana species. According to analysis of the8protein-coding genes and D-loop gene, we found that ND1, ND2, ND4, ND5and cytb contain more phylogenetic information than COI, COⅡ and COⅢ.(ii) The genetic structure and population demography of three common Rana speciesThe distribution pattern of genetic diversity showed phylogenetic discontinuities and spatial separation amongRana amurensis, R. dybowskii, and R. cf. chensinensis,. For R. amurensis, Clade A was distributed in the high-altitude and high-latitude of GreatHinganMountain, Clade B was distributed in NenjiangRiver and Songnen Plain. For R. dybowskii, Clade A was restricted to south slope of LessHinganMountain, Clade B was distributed in east slope of Great Hingan Mountain, north and south of Less Hingan Mountain, and Changbai Mountain. Four Clades were identified in R. cf. chensinensis. Clade A was distributed in Weihe River, Clade B was distributed in Mu us derset, Clade C was distributed in the left bank of Yellow River and Hai River Basin, and Clade D was distributed to north of Yan Mountain.There are some difference in the population demography among three Rana species. Population expansion of R. amurensis, R. dybowskii and clade A, B and D of R. cf. chensinensis were occurred since late-Pleistocene, mid-Pleistocene and late-Pleistocene, respectively. Clade C of R. cf. chensinensis have been indicated relative stable population demography.(iii) Factors that contributed to the phylogeographical patterns of three Rana species Climatic events, large river and mountain ridge are the major factors that influence the phylogeographic parttern of the three Rana species. The major divergence of Rana amurensis was trigged by climate oscillation of Xixiabangma Glaciation and the isolation of Great Hingan Mountains. The phylogeographical pattern of R. dybowskii was influenced by climate oscillation of Guxiang Glaciation and the isolation of Great Hingan Mountains, Less Hingan Mountains and Changbai Mountain. The phylogeographical pattern of R.cf.chensinensis indicated that the divergence time of clades were consistent withclimate oscillation of Xixiabangma Glaciation and the penultimate glaciation. Meanwhile, both Yellow River and Yan Mountain played an important role in shaping the phylogeograpohical pattern of R. cf.chensinensis.(iv) Conservation strategies for the three Rana speciesThe glacial refugia should be protected as wildlife habitat areas. The refugia of Rana amurensis are located in Hailar Basin and middle reach of Songhua River. The refugia and regions of unique haplotype for R. dybowskii are situated in middle Changbai Mountain, southern slopes of Less Hingan Mountains, northern slopes of Less Hingan Mountains, and eastern slopes of Great Hingan Mountains. Four refugia of R. cf. chensinensis are as follows: Weihe river, Mu us desert, Taihang Mountain and low-latitude area of Northeast China.In summary, this study determined the mitogenome of four Rana species, described phylogeographical pattern for the three widely distributed species, and analyzed causes for the spatial genetic distribution of the three studies species based on Mitochondrial DNA genes and nuclear genes. Clear clades were identified in three species distribution range, and climate oscillation, river and mountain ridge were found important in the phylogeographical pattern formulation. However, due to limited time and resources, a number of key questions have not yet addressed in this study, which include diversification history of R. chensinensis and R. cf. chensinensis, the phylogenetic study of Rana species, the spatial patterns of genetic diversity through ecological method, the speciation mechanism of endemic species in China, comparative biogeographical study in North region scale, and modeling of the species distribution patterns in the future environmental scenarios.