Comparative Mitogenomes And Phylogenetic Analysis Of Braconidae (Hymenoptera) Based On Mitochondrial Genomes

The mitochondrial genomes have their unique characteristics, such as the stable gene organization, small genome size, maternally inheritance and accelerated rate of nucleotide substitution. Therefore they are widely used in phylogenetic and evolutionary studies. The development of the sequencing technologies and phylogenetic analysis methods have resulted in extensive phylogenetic studies based on mitochondrial genomes. Braconidae is one of the most important groups of natural enemies because they parasitize other insects, usually the major pests in agriculture. The phylogenetic relationships within Braconidae, however, are confused. Only7braconid mitochondrial genomes have been sequenced compared with its huge number of species, therefore it has great values of sequencing and analyzing the mitochondrial genomes of more species in Braconidae, which will be useful in analyzing phylogenetic relationship, estimating divergence times, and deducing evolution of parasitic habits in Braconidae.In this study we sequenced, annotated and analyzed the mitochondrial genomes of28species representing25subfamilies of Braconidae, including7downloaded from GenBank. Then we explored phylogenetic relationship, evolution of parasitism and divergence time of Braconidae. Consequently we obtained six main points as follows:(1) We sequenced mitochondrial genomes of21species from18subfamilies of the family Braconidae and analysed traits of the genomes, including organization of the genomes, genome contents, overlap and intergenic regions, codon usage, and secondary structure of tRN A gene. Due to high AT content of the Braconidae, regions around A+T-rich region and nad2had not been sequenced. The length of the sequenced Braconidae range from9160bp to14216bp and AT%range from73.38%to87.63%。The highest and lowest AT content appear in Therophlius festivus and Elasmosoma sp. respectively. All start codons of protein-coding genes in the sequenced mitogenomes of Braconidae are ATN. The most common used start codon is ATT, the next is ATG and ATC is the least. All sequenced7coxl use ATC as start codon and only3other genes use ATC. The mainly used stop codon is TAA, and24genes use TAG as stop codon while incomplete stop codons T or TA appear in38genes. The overlap and intergenic regions in Braconidae mitogenomes are generally short but with exceptions. The longest intergenic region is629bp appearing in Eumacrocentrus sp. between cox3and trnG. The intergenic regions between trnG and nad3in Eumacrocentrus sp. and between nad4and nad4L in Mirax sp. are369bp and499bp respectively. Except for Cotesia vestalis, the gene rearrangement regions mainly are tRNA genes, and rearrangement hot-spots are tRNAAsp-tRNAHis-tRNALy tRNAAla-tRNAArg-tRNAAsn-tRNASerl-tRNAGlu-tRNAphe and tRNAThr-tRNAPro. Most used amino acids are Leu, Ile and Phe. Most used anti-codons are TTA (Leu), ATT (Ile) and TTT (Phe). Secondary structure of tRNA genes are clover.(2) In "Cyclostome" most gene rearrangement region is tRNAAsp-tRNAHis-tRNALvs while in "Noncyclostome" gene rearrangement regions are to tRNAAla-tRNAArg-tRNAAsn-tRNASerl-tRNAGla-tRNAphe and tRNAThr-tRNAPro. AT-skew and GC-skew reflect the base composition bias. In the sequenced mitogenomes of Braconidae, most of species have a negtive AT-skew and a positive GC-skew. However, Xiphozele sp., Proterops sp. and Eumacrocentrus sp. are exceptions. The values of nucleotide diversity (Pi), nucleotide substitution (S) and ratio of the rate of non-synonymons substitution to the rate of synonymons (Ka/Ks) reflect the evolution of the mitogenomes of the Braconidae. All Ka/Ks ratio are greater than1except Proterops sp. The fastest evolved species is T.festivus and gene is trnH.(3) The phylogeny of the Braconidae is reconstructed based on all sequenced mitochondrial genes. There are some differences of the topologies between DNA and amino acids results. All the results support Aphidiinae as a sister group to "cyclostome" and at the base of the Braconidae. Ichneutinae belongs to "microgastroid complex of subfamilies" and is a paraphyletic group. The relationship within "helconoid complex of subfamilies" is well supported based on amino acid analyses. The relationships within "cyclostome" and "microgastroid complex of subfamilies" are not well resolved.(4) We predicted the divergence time of the Braconidae based on the oldest known "cyclostome" fossil Protorhyssalus goldmani which is93mya. The divergence time of Braconidae is121.07mya."Cyclostome" and "noncyclostome" was splited at98.32mya. The divergence time of "Non-cyclostome" is92.07, a little later than "cyclostome". The earliest divergenced groups are "helconoid complex of subfamilies" and "microgastroid complex of subfamilies", and the youngest is "cyclostome". As for the parasitic habits, koinobiont endoparasitism appeared earlier than idiobiont ectoparasitism. I the early evolutionary history, most species of Braconidae displayed solitary parasitism and then followed by gregarious parasitism. We predicted the divergence of Braconidae is in step with their host based on the analyses of their divergence time.