Molecular Phylogenetic Study Of The Small Green Leafhopper Family In China

Empoascini belongs to Hemiptera,Auchenorryhncha,Cicadellidae,Typhlocybinae,it is the second largest tribe of Typhlocybinae,currently comprising about 1373 valid species in 83 extant genera.Widely distributed all over the world.At present,216 species of 42 genera of Empoascini have been recorded in China.In this study,specimens from 13 provinces of Guizhou,Guangxi,Guangdong,Hainan,Qinghai,Xinjiang,Sichuan,Yunnan,Gansu,Jiangsu,Henan,Inner Mongolia,and Zhejiang were examined,and 18 genera and 66 species were identified.The 16 S rDNA,CO?,CO?,Cytb and 28 S rDNA sequences were used to analyze the phylogeny of Empoascini.In this study,a total of 254 gene sequences were obtained,including 12 16 S rDNA sequences,64 CO? sequences,63 28 S rDNA sequences,64 CO? sequences and51 Cytb sequences.Through MEGA-X,Geneious ver.10.2.5,the sequences obtained from each gene were analyzed for base sequence composition,substitution frequency,genetic distance,substitution saturation:In terms of base composition,the content of A+T was significantly higher than that of C+G in 16 SrDNA,CO?,CO? and Cytb.However,in the nuclear gene 28 S rDNA,the content of A+T was lower than that of C+G,which proved that the base composition characteristics of insects mitochondrial genes were different from those of nuclear genes.In terms of the statistics of nucleotide of each gene,the Conserved sites of 28 S rDNA gene accounted for the highest proportion among all sites of gene,with 475 Conserved sites,accounting for 73%,indicating that 28 S rDNA evolved slowly among different species of Empoascini,with little difference among species.Among the five gene sequences,the highest proportion of Variable sites is CO?,which accounted for 81%.The results of genetic distance analysis based on each gene showed that the average genetic distance was 0.22-0.25,and there was no significant difference between the genetic distance calculated based on CO?,CO? and Cytb.The genetic distance calculated based on CO? gene was slightly larger than that of the other two gene fragments.In the CO? gene,the maximum genetic distance between inner groups was 0.407 and the minimum was 0.000.The maximum genetic distance between the inner and outer groups was 0.428 and the minimum was 0.271.The results of base substitution saturation analysis showed that the Transitions values(Ts)and Transversions(Tv)in seven data sets were linear with the increase of genetic distance,indicating that none of the data sets reached substitution saturation,which can be used for phylogenetic analysis.Phylogenetic trees were constructed based on the maximum likelihood method(ML)and Bayesian method(BI)by using MEGA-X and Mrbayaes ver.3.2.6.Based on the information of 14 phylogenetic trees,the phylogenetic analysis of the relationship among the species of Empoascini is as follows:Amrasca(Amrasca)biguttula,Amrasca(Quartasca)czerwcowa,Amrasca(Amrasca)bombaxia,Amrasca(Amrasca)apicoserrata,Amrasca(Amrasca)bella,Amrasca(Amrasca)sp.2 constitute a monophyletic group.Distantasca riora,Distantasca rokasa and Distantasca faciata form a monophyletic group.Jacobiasca formosana and Jacobiasca boninensis form a monophyletic group.Kybos rufescens and Kybos sp.6 form a sister group,which are far away from Kybos sp.7.Asymmetrasca dahaituoensis and Asymmetrasca lutowa are not together,show a great distance apart.Ghauriana sinensis is closely related to Membranacea stenoprocessa.The genus Dayus show a closer relationship with the genus Usharia.The genus Kyboasca is closely related to the genus Austroasca.Two main genera of Empoascini: genus Empoasca and genus Alebroides,are not monophyletic.