Population Genetics And Phylogeny Of Lepus In Xinjiang Uygur Autonomous Region

Hares (Lepus) distribute throughout China and occupy a wide variety of habitats, including deciduous, boreal and temperate rain forest, prairie and shrub-steppe. Study about hare species is very worthful to understand the species evolution and assess the relationships between species and environment. Of the nine Chinese hare species,three distribute in Xinjiang: Lepus yarkandensis Gunther 1875, Lepus timidus Linnaeus 1758 and Lepus capensis Linnaeus, 1758. To date, studies about molecular evolution of these three species are rather limited.Yarkand hare and mountain hare were listed in the Second Category of State Key Protected Wildlife List in 1988. The Yarkand hare is endemic to China and restricted to the scattered oases around the Taklamakan Desert in Tarim Basin. Unfortunately, in recent years, habitat fragmentation of Yarkand hare is becoming serious. In this study, two mitochondrial DNA (mtDNA) markers, control region (D-loop) and cytochrome b (Cytb), were used to examine population genetic diversity and assess the impact of geological isolation on phylogeographical structure of Yarkand hare. Also, Quaternary climatic oscillations may have affected the ecosystems and, consequently, the distributions and genetic structuring of the Yarkand hare. In this study, we obtained 553bp D-loop and 1140bp Cytb full sequences from 224 Yarkand hares. Nucleotide diversity value of Lepus yarkandensis is relatively low (D-loop 3.3%, Cytb 0.8%) compared with that of other reported mammals. Mantel tests and molecular variance analysis (AMOVA) suggested that population divergence was significant because a large number of private haplotypes were identified in different populations of Yarkand hare. This differentiation indicated gene flow among populations was limited, which might result from geographical isolation (rivers, oases, deserts) and habitat fragmentation. On genealogical tree, haplotypes of Yarkand hare were divided into 5 deeply divergent clades with high bootstrap support. When the samples were combined into four geographical groups, phylogeographic differentiation between the southwest group and the northeast group was observed and this structure was supported by pairwise FST values and Median-joining network (MJN). Populations from the northern and eastern Tarim Basin shared a similar history, as did those from the western and southern regions. Moreover, Demographical analysis and genetic diversity estimation indicated that the western and southern regions might have served as glacial refugia for the Yarkand hare during Quaternary climatic oscillations. The distribution of the Yarkand hare, especially in the northern and eastern parts, probably represented 3 postglacial colonization events, dated to 0.21, 0.090 and 0.054MYA, which corresponded to known interglacial periods. Given the relatively complete geographic isolation between the eastern and southern populations, the Yarkand hare likely dispersed during postglacial periods from the southwest to the north, and then onward to the east. Significantly, availability of water, rather than displacement by glaciers, was the dominant driving force for the retreat of the Yarkand hare to refugia. This differed from the dynamic mechanism for refugia occupation in Europe, North America, and the Qinghai-Tibetan Plateau in China. The demographical and historical patterns have important implications for conservation. In addition, significant sequence divergence (0.7%-2.3%) among five clades might suggest that subspecies differentiation based on mtDNA existed in Yarkand hare. However, whether the division of subspecies valid or not will require additional evidence from morphology and behavior.To date, the genetic structure and genetic diversity of Lepus capensis in Xinjiang has not been systematically studied at the molecular level, and its subspecies taxonomic status has been under debate for years. According to traditional morphology, there are three subspecies of Lepus capensis distributed in Xinjiang: L.c. centrasiaticus, L.c. lehmanni and L.c. pamirensis. In this study, we determined 553bp D-loop and 1140bp Cytb sequences of 74 cape hares from Xinjiang Province. Nucleotide diversity of Cape hare in Xinjiang is 0.021±0.010 and haplotype diversity is 0.974±0.007. Mantel test and AMOVA analysis revealed a high level of differentiation among populations except central and eastern populations, suggesting that mountains and deserts may make an effective barrier against gene flow. Phylogenetic tree based on mtDNA grouped 74 Cape hare sequences into three clades with high bootstrap support. Each clade included samples from the same area or neighboring areas, which indicate a significant phylogeographic division pattern in Cape hare. Results from median-joining network analysis are in accordance with the phylogenetic analysis. Neutrality tests was insignificant and mismatch distrbution analysis showed a multimodal distribution which was caused by significant differentiation among populations. Our data supported the subspecies status of L. c. lehmanni. The fact that haplotypes of L. c. centrasiaticus were grouped into two distinct clades suggests that this traditional subspecies should be considered as two subspecies. In addition, L. c. pamirensis shows a significantly higher sequence divergence compared to other subspecies, and the difference even reached the level of species.The phylogenetic relationships among hare species in Xinjiang remain unclear. Here, we sequenced two mtDNA markers (D-loop and Cytb) in 353 samples and one nuclear fragment (MGF) in 304 samples to explore the evolutionary relationships of three hare species in Xinjiang. Gene tree based on mtDNA data clustered all haplotypes into three major clades with strong bootstrap support. One L. capensis clade included the majority of L. capensis haplotypes and one L. yarkandensis clade included the majority of L. yarkandensis haplotypes. However, the third clade contained haplotypes both from L. capensis and L. yarkandensis. This is a new mtDNA lineage. The only one L. timidus haplotype was placed into L. capensis clade. Of the three clades, the L. capensis clade branched first; The L. yarkandensis clade and the new lineage formed sister relationship. On nuclear gene tree, haplotypes of L. capensis and L. yarkandensis mixed together and grouped into two major groups. Haplotype of the only one L. timidus specimen formed a separate clade and branched first. The discrepancy between mtDNA and nuclear DNA analysis suggested that introgressive hybridization might occur among L. capensis, L. yarkandensis and L. timidus in Xinjiang. But evidence from morphology, especially skull characters, is essential to confirm this hypothesis.