Study On The Difference Of Rhinolophus Sinicus From China Based On Morphology And Molecular Sequences Data

Rhinolophus sinicus is distributed six geographical subregions in the Oriental Region and a geographical subregions in the Palaearctic Region, within 12 provinces included. R. sinicus in China generally considered has two subspecies: R. s. sinius and R. s. septentrionalis. So far, there is no extensive comprehensive comparison and research on this species based on morphology and molecular sequences data.Whether the differentiation having reached the levels for subspecies classification requires further study. Therefore, the classification of the R. sinicus is very necessary. In order to explore the differences and classification of R. sinicus in China, we conducted a series of bat surveys in the provinces of South China from 2013 to 2016. We investigated 103 bat habitats, of which 17 habitats were living by R. sinicus.Then we collected samples of the R. sinicus and recorded the relevant data. We used morphological measurements for multivariate morphometric analysis and compared DNA sequence divergence. And discussed the differences between Chinese rhizobia and subspecies classification.99 R. sinicus specimens were captured .From the Min-Guang coastal subregion in South China,,Eastern hilly subregion in Central China, the Hainan Island subregion, , West mountain and plateau subregion in Central China, Southwest mountainous subregion in Southwest China and the South Yunnan subregion; Measure 16 external variables for all samples. 29 of the 99 bats were brought back to the laboratory for skull samples, 21 skull variables were measured, and the rest of the 100 bats were released immediately after the samples were collected and measured. Statistical analysis SPSS 20.0 statistical method, using 16 external and 21 brain variables, from six geographical areas of one-way analysis of variance and discriminant analysis.The one-way analysis showed no significant difference between male and female in R. sinicus.However, different sub-regions of the R. sinicus were significantly different. Discriminant analysis of external and skull measurements showed significant differences between the six geographic sub-regions.All individuals are divided into three groups in the scatter plot. The correct recognition rate is high, the cross validation is similar to the original accuracy, indicating that the authentication function is stable and effective.Therefore, there were significant differences in morphology between the six subregions.However, different sub-regions of the R. sinicus were significantly different. Discriminant analysis of external and skull measurements showed significant differences between the six geographic sub-regions.All individuals are divided into three groups in the scatter plot. The correct recognition rate is high, the cross validation is similar to the original accuracy, indicating that the authentication function is stable and effective.The Cytb and COI genes have compared and analyzed in R. sinicus between different regions. The genetic distances of Cytb and COI gene between the subregion of SY and other sub-regions were 4.7%-6.9% and 4.3%-5.3%, respectively. The differences indicated that the species in this region having reached the levels for subspecies classification. The genetic differences of Cytb and COI gene between the three subregions of SM, CE and SH were 0% -2.3% and 0%-1.4%, respectively. And the genetic differences between the three subregions and the other subregions were 2.5% -3.5 % and 2.5%-4.8%, respectively. The differences indicated that the species in this region having reached the levels for subspecies classification.The genetic differences of Cytb and COI gene between the WS and the WS subregion was 0.7% -1.1% and 0.5%-1.7%. And the genetic differences of Cytb and COI gene between the two subregions and the other subregions was 2.5% -6.9% and 2.5%-5.3%. The differences indicated that the species in this region having also reached the levels for subspecies classification. Based on the phylogenetic analysis of COI and Cytb genes, all the R. sinicus were clustered into three different branches, which indicated that R. sinicus in these six subregions should belong to three subspecies.Combined with the results of morphological and molecular sequence and phylogenetic tree, we can see that there should be three subspecies in China of the Oriental Region. The specimens from SM, CE and SH subregions should be R. s. sinius. The specimens from CW and WS subregions should be R. s.septentrionalis. So, R. sinius from SY may be evolving into a unique subspecies.Due to R. sinicus samples in the Tibetan area of the Palaearctic Region have not been collected, further study are still needed to study R. sinicus samples in this subregions.