| Red squirrel (Sciurus vulgaris) is the most widespread of all tree squirrels. The species ranges throughout Palearctic forests from Iberia and Great Britain, east to Russia’s Kamchatka Peninsula and Sakhalin Island, and to Hokkaido in Japan, south to the Mediterranean and Black Seas, and across northern Mongolia, and in isolated forests in northwestern and northeast China. The number of S. vulgaris populations in China declined over recent decades due to deforestation and subsequent population fragmentation. For example, S. v. chiliensis, now considered synonymous with S. v. mantchuricus, has nearly disappeared in northern Hebei province of China. The genetic diversity of China’s S. vulgaris needs further attention for species conservation. This classification is based on morphological dissimilarity, and does not necessarily reflect real subspecies differentiation. Therefore, the further study of molecular biology, specifically its genetic structure, will provide a scientific basis for subspecies differentiation. China’s S. vulgaris is an important part of understanding the problem of Eurasian molecular phylogeography. Therefore, phylogeographical patterns of the China population should reflect ancient geographical and historical events, and lend themselves to investigation of the hypothesis of a postglacial population expansion in this region. In this study, our objectives were (1) quantify genetic diversity levels within S. vulgaris populations in China (2) determine genetic structure and phylogeographical patterns of S. vulgaris in China;(3) understand the species’postglacial expansion history in this region.Given the above problems, the three molecular markers were applied in this study: fragments of mtDNA control region sequences (252bp), fragments of mtDNA Cyt b gene sequences (359bp) and8microsatellite loci (Scv1、Scv3、Scv31、Scv32、RSμ1、 RSμ4、 RSμ5、RSμ6). The research was carried out for the following three aspects:genetic diversity of microsatellite markers for S. vulgaris in China, genetic diversity of mtDNA markers for S. vulgaris in China, and the study on the molecular phylogeography of S. vulgaris. The main results include:1. Based on microsatellite markers, S. vulgaris in China as a whole have the mean of effective number of alleles14.3750, the mean of observed heterozygosity0.7265, the mean of polymorphic information content0.8370, and the mean of Shannon’s Information index2.2333. The higher genetic diversity in overall S. vulgaris in China is found in this study. Genetic diversity from large to low order:Da Hinggan Ling population, Changbaishan population, Wandashan population, Xiao Hinggan Ling population, Altai population, and Tianshan population. In3subspecies, the S. v. mantchuricus has the highest genetic diversity, and the genetic diversity of S. v. altaicus is higher than S. v. exalbiduss. 2. Based on microsatellite markers, there are microsatellite loci of different situations do not meet the Hardy-Weinberg equilibrium in each population. Hardy-Weinberg disequilibrium are likely to be affected due to inbreeding. In each population, there are no obvious bottleneck effect in recent.3. Based on microsatellite markers, Analysis of molecular variances (AMOVA) show: only6.44%(P<0.05) genetic variation among subspecies,1.76%(P<0.05) genetic variation among populations within subspecies, and the remaining91.80%(P<0.05) genetic variation within populations. Genetic differentiation within populations was significantly higher than the genetic variation among populations. The three subspecies have the genetic structure of the distribution. The four populations (S. v. mantchuricus) from northeast China also show the genetic structure, which does not match the geographic distribution relationship.4. Based on mtDNA markers, S. vulgaris in China as a whole have haplotype diversity0.953±0.011and nucleotide diversity0.0103±0.0006.44haplotypes were obtained from mtDNA control region fragment of104samples analyzed.8haplotypes were obtained from mtDNA Cyt b gene fragment of100samples analyzed, in China as a whole have the higher genetic diversity. Having the higher genetic diversity is a common feature of Eurasia S. vulgaris. Genetic diversity of S. v. mantchuricus was significantly higher than the S. v. altaicus and S. v. exalbiduss.5. Based on mtDNA markers, Three phylogenetic analyses (maximum parsimony, MP; maximum likelihood, ML; Bayesian inference, BI) based on two data sets produced nearly the same topology trees. Haplotypes of Calabria in southern Italy produce significant differentiation branch. S. vulgaris of other Eurasian countries are clustering on a large branch, which show the lack of genetic structure in Eurasian.6. Based on mtDNA markers, Mismatch distributions show a nearly unimodal curve from S. vulgaris in China as a whole. Fu’s test for neutrality indicated population expansion for both gene regions (D-loop:FS=-19.62, P=0; cytochrome b:FS=-3.75, P=0.016<0.02) and for the combined data set (FS=-24.50, P=0). The result is consistent with the hypothesis of rapid population expansion. S. v. mantchuricus show having a significant population expansion history.7. Based on mtDNA markers, we found that several cytochrome b and also D-loop haplotypes were shared between two isolated and distant locations, one in Europe and one in China. Moreover, these haplotypes, generally displayed higher frequencies than the remainders not only in Chinese populations but also in European populations. Haplotypes of Calabria in southern Italy produce significant differentiation branch. S. vulgaris of other Eurasian countries are clustering on a large branch. These results lead us to suggest that China’s S. vulgaris population resulted from expansion from a glacial refugium that share characteristics with the refugium at Calabria.8. The result of molecular phylogeography based on microsatellite markers is consistent with the subspecies differentiation and geographic distribution of subspecies. This study propose evolutionary route of S. vulgaris:the squirrels migrated from Calabria via Central and Eastern Europe to Novosibirsk region (S. v. exalbiduss introduction land), then eastward into the Altai region, and then into the northeastern region of China through the forest of north Russia. |
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