Phylogeography Study Of The Species Debregeasia Orientalis Based On Chloroplast Genome And Ribosomal ITS Gene

The Southwest China is complicated in terrain and topography,abundant in mountains and river system,and is a hotspot for global biodiversity research.Although domestic and foreign scholars have conducted a large number of studies on the phylogeography of plants in this region,and significant progress has been made,a completely clear and universal phylogeographic pattern has not yet been resolved.Debregeasia orientalis belongs to the genus Debregeasia Gaud.in the family Urticaceae,and there are few reports of population studies based on large-scale population sampling in this genus and family.In this study,79natural populations of D.orientalis were sampled from a wide range of populations in Southwest China.The study of phylogeography using chloroplast genome and nuclear ITS and ecological niche modeling（ENM）was conducted.By analyzed the genetic diversity and genetic structure,lineages differentiation and geographic distribution patterns,population dynamic history and the refuges of glacial period,and further to analyze the spatial and temporal evolutionary history of D.orientalis and then provide clues for the formation of plant genetic diversity patterns in Southwest China.The main results are as follows:（1）Based on genome re-sequencing data,chloroplast genomes of 281 individuals and nuclear ITS sequences of 283 individuals of D.orientalis were obtained using Get Organelle for assembly.The results using Dna SP and PERMUT indicated that 138 haplotypes were detected based on chloroplast genome,with total haplotype diversity of 0.964,total nucleotide diversity of 0.00038,average number of nucleotide differences of 59.571,total genetic diversity at the species level of 0.969,average intra-population genetic diversity of 0.698;and48 haplotypes were detected based on ITS,with total haplotype diversity of 0.890,total nucleotide diversity of 0.00067,average number of nucleotide differences of 3.859,total genetic diversity at the species level of 0.903,average intra-population genetic diversity of0.599.The results indicate that D.orientalis has a high level of genetic diversity at both species and populations.（2）AMOVA analysis based on chloroplast genome and ITS indicated that genetic variation in D.orientalis occurred mainly among populations(75.32%and 54.85%of genetic variation among populations,F_STare 0.75 and 0.55,respectively);Mantel test analysis indicated that the genetic and geographical distance between D.orientalis populations were significantly correlated;phylogeographic structure analysis based on chloroplast genome and ITS indicated the genetic differentiation indexs N_STand G_STwere N_ST=0.759 and G_ST=0.279,N_ST>G_ST（P<0.01）;N_ST=0.560 and G_ST=0.337,N_ST>G_ST（P<0.05）,respectively.All of which support that the distribution of D.orientalis haplotypes has a significant phylogeographic structure.Phylogenetic analysis and network of chloroplast genome haplotypes indicated that D.orientalis haplotypes generally indicated a phylogeographic pattern with two major clades and nine sub-clades;geographical distribution analysis based on each sub-clade indicated that Nujiang River and Lancangjiang River,Jinshajiang River and its tributary Yalongjiang River,and the middle region of Yangtze River Basin to the distribution of D.orientalis have a spread effect,while the Himalayas and Yunling Mountain in Hengduan Mountainous Region have an isolating effect to the distribution of D.orientalis.The results also indicate that distribution pattern of D.orientalis is related to current river system patterns formed after the Paleo-Red River was diverted.（3）Divergence time of two clades and nine sub-clades of D.orientalis were estimated between 0.83 Ma and 0.07 Ma based on molecular clock model.The results indicate that first clade-divergence of D.orientalis was affected by Xixiabangma Glaciation,further divergence of two clades was affected by Naynayxungla Glaciation,and nine sub-clades were formed by divergence was affected mainly by Guxiang Glaciation and secondly by Baiyu Glaciation.The earliest divergence of sub-clade B3 and B4 were located in Northwest Yunnan-Central Yunnan region,which was presumed to be origin center of D.orientalis.（4）Results of neutrality test and mismatch distribution analysis indicated that D.orientalis had experienced recent expansion events at A2 sub-clade level;the A2 sub-clade pattern with star-shaped radial shape of network relationship map also supported its expansion,while other eight sub-clades did not be supported events of rapid population growth.Further combining the geographic distribution of chloroplast genome and ITS haplotypes with the results of ecological niche model by Maxent,it was indicated that all sub-clades of D.orientalis had experienced two different patterns of refuge in glacial period,that are A2 sub-clade populations were mainly concentrated in refuges where are southern part of its distribution-northern part of Guizhou,such as Ta-lou Mountains,and then a large area of spread to north after glacial period,which belonged to expansion-contraction pattern.Other sub-clades populations took refuge in multiple micro-refuges,and then moved back with a small area,which belonged to be in situ survival or in nearby survival pattern.Hengduan Mountains,Yunnan-Guizhou Plateau,and northern Taiwan are main glacier refuges for D.orientalis.