Comparative Analysis Of The Determination Of The Full Mitochondrial Genome Of Four Species Of Hymenoptera

By means of Long-PCR and Sub-PCR methods,this research had sequenced four Neuroptera insects complete mitochondrial genomes of Bullanga florida(Navas,1913),Mantispa japonica(MacLachlan,1875),Eumantispa harmandi(Navas,1909)and Eudimacia badia(Okamoto,1911).And aslo finished annotation and analytic comparison to results.Meanwhile,combining the already published species complete mitochondrial genome sequences of 21 Neuropterida insects from GenBank.The research took Megaloptera and Raphidioptera as out-groups,reestablished the phylogenetic tree through the method of Maximum likelihood and Bayesian Inference,and conducted basic analysis of phylogenetic relationships of Neuroptera species.Below are main results for the study.1.The lengths of complete mitochondrial genome of four Neuroptera insects were 15937 bp(B.florida),16106 bp(M.japonica),15741 bp(E.harmmandi)and 15899 bp(E.badia),respectively,they shared consistent genes and transcription direction.They contained 13 protein-coding genes(cox 1-3,atp6,atp8,nadl-6,nad4l,cytb),22 transfer RNA genes,2 ribosomal RNA(rrnL,rrnS)and an A+T-rich region.While,the arrangement of mtDNA genes of these four species occurred WC indexable phenomenon with trnC gene in front of trnW gene.This arrangement exists in the other Neuroptera species excluding Osmylidae insects.The basic compositions of mtDNA sequences had obvious AT-skew,and A+T contents were 80.6%for E.harmandi,73.3%for B.florida,80.3%for E.badia and 79.3%for M.japonica.2.The A+T content of protein-coding genes were 79.0%for E.harmandi,71.2%for B.florida,78.1%for E.badia and 78.1%for M.japonica,among whose the highest A+T located at third locus of protein codon.The relative synonymous codon usage(RSCU)of protein could also reflect the deviation of A+T content to certain extent,the codon of starting or ending with A and T were highter than the frequency of other synonymous codon.The starting codons of M.japonica,E.badiaand and E.Harmandi were AGG,while ATG was starting codon of B.florida.The ending codons of coxl,cox2,nad5 and nad4 were incomplete terminal codon T,and all rest codons were complete terminaal codon TAA.3.Some non-coding gaps and overlaps existed in mtDNA of these four Neuroptera insects.The shortest non-coding gap between the gene cluster trnl and trnQ of B.florida was 57 bp,the longest gap for E.Badia was 91 bp;the non-coding gap between trnS(UCN)and nad1 of M.japonica was longer than other three species.The non-coding gaps within trnS(UCN)-nadl of M.japonica was larger than others.A relatively stable overlap regions,atp8-atp6 and nad4-nad4L with a length of 7 bp existed in four species,and the same overlap regions also happened in other insects.This phenomenon contributed to particular function of genes.4.All tRNAs genes of E.harmandi had typical cloverleaf structures,and B.florida,E.Badia and M.Japonica were lack the trnS(AGN)on DHU arm.All these four species had some mismatches in tRNA genes secondary structure,and B.florida topped the rank.The most mismatches were G-U,then U-U and A-C.The G-U mismatch mostly happened at the amino acid acceptor arm in this four species.5.The rrnL and rrnS genes were located on N-chain of four species.The lengths of rrnL and rrnSwere 1325 bp/776 bp(B.florzida),1307 bp/783 bp(M.japonica),1297 bp/786 bp(E.harmandi),1305 bp/791 bp(E.badia).A+T-rich region located between rrnS gene and gene cluster trnl-trnQ-trnM and the sizes were 1066 bp(B.florida),1281 bp(M.japonica),936 bp(E.harmandi),1092 bp(E.badia).The A+T content in A+T-rich region of B.florida was minimum.There was a stem and loop structure in four A+T-rich regions of mitochondrial genome.6.Based on PCGs and PCGs&rRNAs datasets,the study was reconstructed the phylogenetic relationships in Neuroptera using ML and Bayesian Inference methods.The topology structures of two phylogenetic trees were slightly different.Neuroptera were mainly divided into three clades and Osmylidae formed the basic of tree.The clades of Hemerobiformia and Myrmeleontiformia were monophyly,respectively.The relationships were complex in all families of Hemerobiformia.M.japonica,E.badia and E.harmandi were together formed sister group and Mantispidae was monophyly.The phylogenetic relationship in Myrmeleontiformia was as follows:(Nymphidae +(Myrmeleontidae + Ascalaphidae)).B.florida,Myrmeleon immanis and Epacanthaclisis banksi of Myrmeleontidae species did not get together,so we did not support the monophyly of Myrmeleontidae.