Mitochondrial and Nuclear DNA-based Identification of Some Forensically Important Calliphoridae(Diptera)in Luoyang of China

Introduction:Calliphoridae plays a key role in forensic entomology research,which is one of the first insects to decompose animal carcasses.The mitochondrial cytochrome c oxidase subunit I and the ribosomal internal transcribed spacer 2(ITS2)are among the most widely used molecular markers for insect taxonomic characterization.Aim:The aim of the study was to test the suitability of two genetic markers based on conducting the molecular identification of six necrophagous Calliphorid flies.Materials and Methods:Fourteen Calliphoridae flies were collected and classified with traditional morphological characteristics.The DNA of flies was extracted and the fragments of COI and ITS2 were amplified and sequenced.All the sequences were aligned and analyzed by MEGA 7 software for NCBI BLAST,nucleotide composition,intra-and inter-specific divergence calculation,and phylogenetic tree inference successively.Results:The results indicated that COI and ITS2 genes were robust in the identification of Calliphoridae at the species level and ITS2 gene sequence possessed a strong resolution power as it showed higher variation values between Lucilia sericata and Lucilia cuprina,Calliphora vomitoria and Triceratopyga calliphoroides,C.vomitoria andAldrichina grahami,but inferior to COI fbrT.calliphoroides and A.grahami.Conclusions:Our results showed that combination of COI+ITS2 genes yields more accurate identification and diagnoses and better agreement with morphological data than the mitochondrial barcodes alone.As a supplementary method for morphological identification,we advocated for the combination of nuclear and mitochondrial gene approaches to address the taxonomy and phylogeny of forensic relevant flies,especially of closely related species and populations.