The Taxonomic Of Quasipaa Spinosa Based On Morphology And Molecular Properties

Quasipaa spinosais,is a large frog with high economic and nutritional value,which is mainly distributed in the southern part of China and is distributed in a small amount in the northern part of Vietnam.Previous molecular data indicate that Q.spinosa can be divided into three branches according to the geographical population.In this study,multi-gene technique combined with morphological study was used to verify the differentiation level of different branches,which provided the basis for the classification of Q.spinosa.The main results are as following:(1)In terms of morphology,based on sixteen morphological characters of populations of Q.spinosa from Longsheng Guangxi,Jinhua Zhejiang,Wuyishan Fujian,Daiyunshan Dehua,Le'an and Yichun Jiangxi of China,multivariate morphometrics(Principal component analysis,Discriminant analysis and Hierarchical cluster analysis)were used to investigate their morphological variations among the six different geographical populations.Principal component analysis results showed that Longsheng geographical population was markedly differ in morphology from other different geographical populations.The first 4 principal components extracted from external measurements explained 56.6% of the total variation,and Longsheng geographical population was separated from other geographical populations in the scatter plots.Discriminant analysis of external measurements showed differences between the 6 geographical subregions.All individuals were separated into 3 groups in a scatter plot.and the correct discriminate rates of Longsheng Q.spinosa is 96.2%.The discriminant function can be used for accurate discrimination.Hierarchical cluster analysis results showed that the taxonomic status and genetic relationship of the geographical populations of Fujian,Jiangxi and Zhejiang were relatively close.There has been a certain degree of differences between Longsheng population and other geographical populations,and a certain degree of difference in morphology.(2)we measured 166 samples of mitochondrial 12 S rRNA,16 S rRNA genes and nuclear genes Rag-1 fragments in 6 populations.Sequences analysis combined with 12 S rRNA,16 S rRNA and Rag-1 gene.We selected haplotype based on the sequence data.Phylogenetic analyses were conducted using maximum likelihood and Bayesian inference,and calculated the genetic distance of interspecies by MEGA6.0.The molecular phylogenetic trees which were constructed by using three gene fragments showed that Longsheng thorny frog is a single tree.Genetic distance showed that the intraspecific differentiation between Longsheng geographical population and other 5 geographical populations has reached the level of interspecific differentiation of Quasipaa,and it can be used as a single lineage.(3)Based on the analysis of microsatellite data,the results showed that the six geographical populations were most likely to be 3 populations.There is a great genetic difference among different geographical groups of Q.spinosa,especially in Longsheng geographical population.It is highly differentiated between Longsheng and the other 5 geographical populations.And the migration rate between Longsheng population and other populations in Guangxi is far less than 1.According to the morphological and molecular data analysis results of this study,the morphological and molecular differences of Longsheng Q.spinosa have reached the level of interspecific differentiation.Especially,the foot length of Longsheng Q.spinosa is more than 3/5 of the body length,which is obviously different from other populations.Therefore,it is suggested that Longsheng Q.spinosa should be identified as a new species,and named Quasipaa longshengensis sp.nov..