Molecular identification and genetic variation of hematophagous flies, (Diptera: Muscidae: Stomoxyinae) in Thailand based on cox1 barcodes

• First report on using DNA barcoding for identification of Stomoxyinae flies in Thailand
• DNA barcoding was successfully identified eight Stomoxyinae species.
• All species of Stomoxyinae flies formed a monophyletic group.
• Uncertain species Stomoxys pullus and Stomoxys uruma were discriminated by cox1 barcoding.
• Sub-species, Haematobia irritans irritans and Haematobia irritans exigua, were discriminated by cox1 barcoding.

Stomoxyinae flies (Diptera: Muscidae) transmit many pathogens to animals and humans. However, morphological identification of Stomoxyinae flies is difficult, and there is risk of misidentification. Therefore, this study analyzed cytochrome oxidase 1 (cox1) barcodes to test their ability to discriminate between taxonomic groups. cox1 barcodes (657 bp) from 66 specimens were classified into 8 species. cox1 sequences have typical insect mitochondrial characteristics including a high AT content (mean = 69%), especially at the third codon position. cox1 barcodes were able to discriminate between species with high interspecific divergence, especially those that are morphologically difficult to distinguish, such as certain Stomoxys species (S. indicus, S. pullus, and S. uruma) and Haematobia sub-species (H. irritans irritans and H. irritans exigua). Phylogenetic analysis and the genealogical sorting index revealed that all Stomoxyinae species formed a monophyletic group and were clearly separated into distinct clusters based on species. Species delimitation by the Poisson tree processes model indicated previously unrecognized diversity within Stomoxys and Haematobosca. Thus, DNA barcoding was able to discriminate between morphologically uncertain or misidentified Stomoxyinae flies and confirm the correct species, which are important steps for elucidating diversity.

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