Construction And Applications Of DNA Barcode Library Of Leaf-mining Beetles In Agriculture And Forestry

Coleoptera leaf-mining beetles have a high number of species,a wide range of distribution,and a wide range of host plants.Some leaf-mining beetles are economic pests in agriculture and forestry.Most leaf-mining beetles have small body size,which may make them difficult to identify with morphological characteristics only.In this paper,both DNA barcode library and mitochondrial genome library for leaf-mining beetles were constructed,which could provide a useful background for scientific researchers in the fields of macro-biology and reduce the costs of both material resources and human resources.In order to provide technical support for the identification and control of leaf-mining pests,we analyzed the interspecific phylogenetic relationship among leafmining Dactylispa beetles using either COI barcoding sequences or the whole mitochondrial genomic sequences.Our main findings are as follows:(1)COI,CYTB and ITS2 genes were used to construct DNA barcode library of leaf-mining beetles.The lengths of COI gene amplified fragment of leaf-mining beetles were about 700 bp,the lengths of CYTB gene amplified fragment of leaf-mining beetles were about 500 bp,and the lengths of ITS2 gene amplified fragment of leaf-mining beetles were about 750 bp.So far,240 individuals of leaf-mining beetles have been sequenced in our lab,including 100 individuals with COI gene,101 individuals with ITS2 gene,and 39 individuals with CYTB gene.(2)The biological characteristics of mitochondrial genomes such as AT content and codon usage,were analyzed for Trachys auricollis,eight Dactylispa species,and Downesia tarsata.33 individuals of leaf-mining beetles with the complete mitochondrial genome have been sequenced in our lab,and the framework of mitogenome library has been basically constructed.(3)Based on the COI gene sequences of Dactylispa species,two phylogenetic trees of leafmining beetles were respectively constructed using the maximum likelihood method and the Bayesian method.Both phylogenetic trees of Dactylispa beetles seemed to support the two subgenera(Dactylispa s.str.and Platypriella)in Chen’s et al.(1986)system.However,the subgenus Triplispa seemed to contain transitional forms between Dactylispa s.str.and Platypriella,therefore the placement of different Triplispa species required further verifications.Adult body shape and host plant relationship might somewhat explain the interspecific relationships among Dactylispa species.COI could be the DNA barcode for leaf-mining beetles.(4)Compared with the phylogenetic tree constructed by single gene,the mitogenomic phylogenetic trees of eight Dactylispa species had more accurate results and higher branch supporting rates.The mitogenomic phylogenetic trees of Dactylispa beetles also implied that Triplispa might be transitional forms between Dactylispa s.str.and Platypriella,and the classification of the three subgenera could be related to body shape and host plant.To sum up,we have completed the construction of the DNA barcode library for some leafmining beetles.However,more data,including genetic sequences,morphological characteristics,geographical distribution,host plants and ecological behaviors,are still needed to realize the comprehensive and rapid identification of leaf-mining beetles.COI gene is one ideal candidate DNA barcodes for leaf-mining beetles,which can accurately reflect the phylogenetic relationship of leafmining beetles.The phylogenetic relationships of leaf-mining beetles based on the whole mitogenomes should be more accurate.