Study On Microevolution Of Eothenomys Sp. Among Different Populations

Eothenomys, an extraordinary member of Cricetidae in Rodentia, which is a kind of small mammal inhabiting mountain forests and bushwood, is inherent taxa and the typical species in Hengduan Mountain areas. Since then establishing of Eothenomys, biologist and systematist have been being arguing for its taxonomic status. Especially, taxon under the genus Eothenomys is more confused. Animals living in environment, will possess physiological and genetic variation, which caused by adaptation to environment and geographical isolation.There were 3 parts in this paper:1 The taxonomic status of Eothenomys.2 Study of body mass, thermogenesis and relative fatness in Eothenomys sp. among different populations from Yunnan province3 Study of genetic divergence based on mtDNA COI of Eothenomys sp. from 4 areas, Yunnan Province4 Study of genetic divergence based on mtDNA D-loop of Eothenomys sp. from 4 areas, Yunnan ProvinceThe results were outlined as follows:1. The taxonomic status of EothenomysWe downloaded the COI and Cyt b sequences of different species from GeneBank, together with sequences that are amplificated by myself, and then constructed NJ and ME trees among Eothenomys、Clethrionomys、Alticola、Caryomys, respectively, based on kimura-2-paramater genetic distance. The trees topology results showed:(1) Cyt b:all species clustered into 2 big clad-genus Eothenomys divided into subgenus Eothenomys and subgenus Anteliomys; Clethrionomys、Alticola and Caryomys clustered together, meanwhile Eothenomys andersoni and Eothenomys smithii were contained in Clethrionomys. Our results supported that genus Eothenomys should be divided into subgenus Eothenomys and subgenus Anteliomys; Eothenomys andersoni and Eothenomys smithii should belong to genus Clethrionomys rather than become an independent genus Phaulomys. Otherwise, Caryomys also should be regard as an independent genus according to our consequence. Our specimens was closed to Eothenomys fidelis infer from NJ phylogenetic tree.(2) COI:the phylogenetic tree clustered into 2 clades, namely-subgenus Eothenomys and subgenus Anteliomys. Based on Cyt b analysis, our specimens clustered together with Eothenomys fidelis; on the other hand, it was closed to Eothenomys miletus in COI. Different Eothenomys sp. can separate each other preferablely-Eothenomys custos and Eothenomys chinensis clustered into a clade, another clad was composed by Eothenomys melanogaster、Eothenomys miletus、 Eothenomys proditor、Eothenomys olitor、Eothenomys eleusis and Eothenomys cachinus. (3) all haplotypes from specimens didn’t mix with Eothenomys melanogaster, which suggested that our specimens might be independent species, not subspecies of Eothenomys melanogaster implied from D-loop NJ phylogenetic tree.Different oriental voles can be separated each other in phylogenetic tree based on morphological data——E. chinensis and E. custos clustered into a big clade, another 6 oriental voles clustered into other big clade.Based on results above, we suggested that different morphological measurement system analysis of Eothenomys can be regarded as an available means used to identify. Phyletic relationship under subgenus Eothenomys was still unclear. Combining previous studies with mine, we suggested that different species of subgenus Eothenomys was still unclear, but our specimens could be seen as independent species that inferred from molecular systematics, which need integrated research, furtherly.(2) Study of body mass, thermogenesis and relative fatness in Eothenomys sp. from Yunnan provinceMammals will be able to form different adaptive mechanism based on the variable ambient pressure. In order to investigate the physiological adaptation in different environments of Eothenomys sp., which were captured from Lijiang (n=19), Dali (n=11) and Kunming (n=17) in November, respectively. Body mass, basal metabolic rate (BMR), nonshivering thermogenesis (NST) and relative fatness (K) were measured. The results showed:(1) body mass of Lijiang population and Kunming population were lower than Dali populationas well as showed significant; (2) BMR and NST of Lijiang and Dali all showed significant comparing with Kunming population; whereas (3) relative fatness, just Lijiang populationwas higher than Dali population and Kunming population, and it was no striking between Dali and Kunming. The variations among three populations, which indicated that specific physiological characteristics of Eothenomys sp. to adapt the corresponding ;nvironments.(3) Study of genetic divergence based on mtDNA COI of Eothenomys sp. from 4 reas, Yunnan Province1323bp COI sequences, the percent of A、T、G and C was 28.2%、27.4%、27.7% and 16.8%, respectively.120 variation sites were detected (accounted for 9.07% of all sites), of which including 96 parsimony-informative sites and 24 singleton sites. There were 26 transitons and 3 transversion, Ts/Tv 8.28. We found 28 haplotypes from 88 individuals, Hd 0.935, the average genetic distance among 28 haplotypes was 0.02124±0.00237. Pi was from 0.00249(Lijiang)-0.02419(Ailaoshan), average Pi was 0.02234. The discrepancy of nucleotide divergence (Dxy)、level of differentiation (Fst)、net genetic distance (Da)、coefficient of differentiation (Gst) between Hengduan Mountain populations and Kunming population was very remarkable, whereas the discrepancy was unconspicuous within Hengduan Mountain populations.Phylogenetic tree of different populations based on neighbor-joining method, UPGMA method and kimura-2-paramater genetic distance showed that it contained: main parts.The distribution curves of the mismatch analysis on different populations may be the multimodal curves that encovered populations without expansion. Furthermore, Tajima’s D、Fu and Li’s D and Fu’s Fs test for population showed that red-back voles didn’t go through the event of population expansion.(4) Study of genetic divergence based on mtDNA D-loop of Eothenomys sp. from 4 areas, Yunnan ProvinceIn this study, 1010bp valid sequence was obtained. The percent of A、T、G and C was 30.5%、29.2%、13.3% and 27.0%, respectively.158 variation sites were detected (accounted for 15.64% of all sites), of which including 116 parsimony-informative sites and 24 singleton sites. Transiton (Ts) 29 and Transversion (Tv) 8, Ts/Tv 3.83. We found 49 haplotypes from 88 individuals, had no common haplotypes among these populations, The average genetic distance among 49 haplotypes was 0.04256±0.00492.Among different populations, nucleotide diversity (Pi) was located between 0.006 (Lijiang population) and 0.034 (Ailaoshan population), average Pi was 0.0364. Analysis of Molecular Variance (AMOVA) encovered that differentiation index (Fst) was significant differences and Variance components among populations (Va) was higher than Variance components within populations (Vb). The gene flow (Nm) was less-than 1, except the relationship between Dali and Kunming, which indicated Eothenomys sp. was developing evident genetic differentiation.Infered from NJ and UPGMA phylogenetic tree, different haplotype clustered together according to its attribution, approximately. From large scale,4 clades was included — Clade Ⅰ:consistented of Dali population and Lijiang population, fundamentally; Clade II included Dali population/Ailaoshan population/Kunming population; Clade Ⅲ-Kunming population; Clade Ⅳ-Ailaoshan population.The distribution curves of the mismatch analysis on different populations may be the multimodal curves that encovered populations without expansion. Furthermore, Tajima’s D、Fu and Li’s D and Fu’s Fs test for population showed that oriental voles Eothenomys didn’t go through the event of population expansion, the population was still stable.Conclusion:Phylogenetic analysis supported the genus Eothenomys should be divided into 2 subgenera-subgenus Eothenomys and subgenus Anteliomys. But we failed to solve the taxon under genus just by molecular analysis. Phyletic relationship under subgenus Eothenomys was still unclear.After long-term adaptation and geographical isolation, Eothenomys sp. from different areas will possess new physiological and genetic variation, finally fixed in the different populations, finally. Among different populations, genetic differentiation index (Fst) showed significant, which possessed distinct. We infered from above, Eothenomys sp.may remain the phase of ecological speciation. Integrating fossil evidence with COI and D-loop, which illuminated red-backed voles without going through population expansion. The results supported Eothenomys was the inherent groups, Eothenomys can be viewed as typical representatives to research small mammals in Hengduan Mountain Region.