Assessing the applicability of the COI for phylogenetic analysis of forensically important Calliphoridae (Diptera) flies from Argentina

Necrophagous blow flies (Calliphoridae) are among the most important insect groups in forensic investigations, making accurate species identification essential. The mitochondrial cytochrome c oxidase subunit I (COI) gene is widely used for molecular identification, although its diagnostic value and phylogenetic performance may vary among regional entomofaunas. To evaluate the COI marker's utility for Argentine forensic blow flies, sequences obtained from specimens of 10 local species (Calliphora vicina, Lucilia sericata, L. cuprina, L. ochricornis, L. purpurascens, Chrysomya megacephala, C. albiceps, Cochliomyia hominivorax, Co. macellaria, and Sarconesia chlorogaster) were analyzed together with 158 publicly available sequences representing 36 species of Calliphoridae from worldwide distributions, including three outgroup species belonging to closely related families. Phylogenetic analyses of aligned sequences were performed under Maximum Likelihood in IQ-TREE using ModelFinder for model selection, with group support estimated via 10,000 ultrafast bootstrap replicates (UFBoot) and Shimodaira–Hasegawa-like approximate Likelihood Ratio Test (SH-aLRT). The subfamilies Chrysomyinae, Toxotarsinae, and Calliphorinae were recovered as highly supported monophyletic clades (UFBoot ≥ 90%, SH-aLRT ≥ 95%). However, Luciliinae was not recovered because most of the 13 Lucilia species analyzed remained unresolved. Furthermore, Calliphoridae was not recovered as monophyletic, with very low bootstrap support, because species of Sarcophagidae and Tachinidae nested within it. These findings provide the first phylogenetic evaluation of the COI marker for Argentine forensic blow flies. Our results confirm that COI is reliable for delimiting subfamilies, while verifying that it is not suitable for resolving the species complexity within Lucilia. Overall, this work establishes a baseline for molecular identification within Calliphoridae and emphasizes the need for integrative approaches for complex genera.