The Comparative Phylogeography Of Two Co-distributed Cyprinid Species From Southern China

In this paper, mtDNA (D-loop region and Cyt b gene) and SSR wereused to analyze phylogeographic patterns and population genetic structureof8Garra orientalis Populations and12Osteochilus salsburyi Populations.The results were shown as follows:1. Comparative phylogeography of Garra orientalis and Osteochilussalsburyi based on nucleotide sequences of the mtDNA sequences.Genetic variation and phylogeographic patterns of8G. orientalis and12O. salsburyi populations were investigated based on mtDNA sequences,with157G. orientalis individuals and236O. salsburyi individuals.(1)Genetic Diversity. As for G. orientalis, there were110variable sites (5.82%of the full sequences) among1887base pairs of the mtDNA sequences(750bp D-loop region and1137bp Cyt b gene sequences), including60(3.17%) parsimony informative sites and50(2.64%) singleton. Thesamples fall into88haplotypes, with0.981haplotype diversity and0.00744nucleotide diversity. As for O. salsburyi, there were59variable sites (3.12% of the full sequences) among1888bp of the mtDNA sequences (751bpD-loop region and1137bp Cyt b gene sequences), including39(2.06%)parsimony informative sites and20(1.05%) singleton. The samples fallinto44haplotypes, with0.833haplotype diversity and0.00233nucleotidediversity.(2) Phylogenetic Analysis. NJ tree of157individuals for G.orientalis revealed that the8populations can be divided into three majorlineages. NJ tree of236individuals for O. salsburyi revealed that the12populations can be divided into four major lineages. Both of them showedthat there were significantly genetic differentiation between population ofChanghua River and other populations.(3) Genetic differentiation andgenetic structure among Populations. The pairwise Fst value of G.orientalis and O. salsburyi were0.649and0.428, respectively. AMOVAand SAMOVA revealed that both of the two species have strongphylogeographic structure due to restricted gene flow among geographicpopulations. Qiongzhou Strait and Wuzhi Mountains was geographicbarriers for G. orientalis, and YunKai Mountains was geographic barriersfor O. salsburyi.(4) Demographic History. Neutral test and mismatchanalysis revealed that the two species had experienced a populationexpansion. Bayesian skyline plot of G. orientalis and O. salsburyi showedthe population expansion may happen18,000and11,000years ago,respectively. 2. Isolation and characterization of polymorphic microsatellitemarkers in the fish Garra orientalis and Osteochilus salsburyi andpopulation genetic structure analysisBiotinylated magnetic enrichment beads method was used to isolatethe microsatellite loci.14primer pairs of G. orientalis and13primer pairsof O. salsburyi successfully amplified DNA products and showed highlevels of polymorphism. Then, we used23individuals of G. orientalis and20individuals of O. salsburyi to quantify the levels of polymorphism. Asfor G. orientalis, the number of alleles (NA), polymorphism informationcontent (PIC), observed and expected heterozygosity (HOand HE) perlocus ranged from8to25,0.6788to0.9342,0.5652to1.0000and0.7179to0.9585, respectively. Four loci are deviated from Hardy-Weinbergequilibrium due to the heterozygote deficiency, no evidence of genotypiclinkage disequilibrium was detected among loci. As for O. salsburyi, NA,PIC, HOand HEper locus ranged from10to21,0.8020to0.9354,0.7500to1.0000and0.8436to0.9628, respectively. No locus was found tosignificantly deviate from Hardy–Weinberg equilibrium, no evidence ofgenotypic linkage disequilibrium was detected among loci.The microsatellite markers were used to study Genetic diversity andgenetic structure of the populations of G. orientalis and O. salsburyi. Theresults showed that:①Genetic Diversity. As for G. orientalis, the average number of alleles (A), the average allele richness (AR), HEand HOwere11.923,10.001,0.8581and0.8477, respectively. As for O. salsburyi, A, AR,HEand HOwere12.090,14.385,0.8720and0.8415.②Geneticdifferentiation and genetic structure among Populations. The differentiallevel between Populations of the two species is medium or lower. SoftwareStructure was used to rebuild the cluster structure of the populations, fourmain groups in G. orientalis and three main groups in O. salsburyi wereidentified.③Software Bottleneck was used to analysis the“Bottleneckeffect”, the results showed that some populations of the two species hadexperienced bottleneck effect.