Studies On Population Divergence And Molecular Phylogenetics Of Fejervarya Limnocharis And Bufo Gargarizans In Zhoushan Archipelago

The Zhoushan Archipelago has significant fragmentation of ecogeographical landscape,but few researches to date were conducted on phylogenetics and genetic conservation of amphibians in this area.Fejervarya limnocharis and Bufo gargarizansare most conspicuous amphibians distributing in Zhoushan archipelago.Due to the three times transgressive and regressive events happened in the late Pleistocene,ancestors of both amphibians might had experienced a longtime isolation,and thus formed a unique significant phylogeographical genetic structure.In this thesis,the F.limnocharis and B.gargarizanswas used to analyses on(1)the sexual dimorphism of the F.limnocharis and the relationship betweenthe body size and island features.(2)The phylogographical structure and population divergence of F.limnocharis and B.gargarizans.(3)the relationship between genetic diversity and island characteristics of both amphibians.A total of 339 F.limnocharis species collected from 30 sample sites were used to analysis morphological variance.We measured 8 morphological characteristics including snout-vent length(SVL),head length(HL),head width(HW),diameter of eye(ED),diameter of tympanum(TD),belly length(BL),foreleg length(FLL)and posterior leg length(PLL).Our results showed that the adults of F.limnocharis are sexual dimorphic in body size with female are larger than male on all morphological characteristics.The body size of F.limnocharis was significantly negatively associated with island area and perimeter,significantly positively associated with the nearest distance between island and mainland.The body size of small islands was larger than big islands.This result strongly support the hypothesis that the body size of rice frogs on island populations turned to be significant increases.In this work,a total of 2600 bp in length of mitochondiral DNA including two partial fragments of NDl-ND2 and Cytb-CR(control region)were used to study the phylogenetic evolution and conservative genetics of both amphibian species.Results of phylogenetics and haplotype analyses suggested that F.limnocharis clustered into two deeply diverged major clades respectively distributes in mainly Zhoushan Archipelago and coastal continent.The clade of mainland was furthur devided into two subclades,with clade A distributes in south coastal area of Qiantang river.TheArchipelago main clade of F.limnocharis was subdivided into two clades which respectively occupied Shensi islands and Qushan islands by clade D,as well as clade C occupied Daishan island and Zhoushan mainland.Analyses on phylogenetic relationship among species of genus Fejervarya showed that our samples from the Zhoushan Archipelago mainly divided into two branches,the mainland clade and the island clade.Specimen of F.limnocharis from Japan Hiroshima was nested in Zhoushan island clade with another individual of F.sakishimaensis from Japan formed the outgroup.Our results illustrated that populations of F.limnocharis from Zhoushan island has a closer phylogenetic relationship with the Japan F.limnocharis population than that of the mainland branches.Phylogenetic tree based on partial Cytb support that the island population of F.limnocharis from Zhoushan archpelago are conspecific with the Japan mainland F.limnocharis.Our phylogenetics based on combined ND1-ND2 and Cytb-CR fragments discovered five clades of B.gargarizans with three clades A.B and C diverged earlier distributing in Zhoushan Archipelago except one popultaibn locality(PIH).Two later diverged sister clades with clade D distributing in Zhoushan islands and clade E occupying both costal areas and the islands.The isolation and disperse of ancestors of these clades was probably affected by the formation and disappearance of isolated factors correlated with environmental changes in this area.The haplotype diversity(h)of F.limnocharis population on the island of Zhoushan,Qushan and Daishan was found to be very high(h of Cytb-CR are 0.789±0.055,0.923d±0.057 and 0.846±0.089,respectively;h of ND1-ND2 are 0.820±0.036,0.297±0.115 and 0.412±0.119,respectively).Nucleotide diversity(?)of the F.limnocharis population on Zhoushan was also higher(? of Cytb-CR is 0.05286±0.00941 and ? ofND1-ND2 is 0.03200±0.00746).In addition,the Qushan,Shengsi Dongji islands which are the major overlap areas of different clades based on Cytb-CR and ND1-ND2 also showed higher haplotype diversity and nucleotide diversity.The Cytb-CR haplotype diversities of B.gargarizans on all the islands are relatively higher(from 0.795±0.085 to 0.964Q0.077).Qushan,-Daishan and Zhoushan had the higer h and ? based on ND1-ND2 gene.The overlap area of clades of B.gargarizans based on phylogenetic analyses using Cytb-CR and ND1-ND2 included Huaniao,Lvhua from Shengsi islands,Xiaoyangshan,Qushan,Nanpu from Daishan island,Zhanmao and Yandun from Zhoushan island.All these overlapped areas showed higher genetic diversities.The clarification of significant evolutionary clades and population genetic diversities in this thesis are valuable for guiding the principles of genetic conservation of both species in this area.