Wing geometric morphometrics and DNA barcoding to distinguish three closely related species of Armigeres mosquitoes (Diptera: Culicidae) in Thailand

Armigeres subalbatus, a mosquito species widely found in Thailand and other Asian countries, serves as a vector for filarial parasites, affecting both humans and animals. However, the surveillance of this vector is complicated because of its morphological similarity to two other species, Armigeres dohami and Armigeres kesseli. To differentiate these morphologically similar species, our study employed both wing geometric morphometrics (GM) and DNA barcoding, offering a comprehensive approach to accurately identify these closely related Armigeres species in Thailand. Our GM analyses based on shape demonstrated significant accuracy in differentiating Armigeres species. Specifically, the outline-based GM method focusing on the 3rd posterior cell exhibited an accuracy rate of 82.61%, closely followed by the landmark-based GM method with 81.54%. Both these GM techniques effectively distinguished Ar. subalbatus from Ar. dohami and Ar. kesseli. Regarding DNA barcoding, our investigation of pairwise intra- and interspecific divergences revealed a "barcoding gap". Furthermore, the results of species confirmation using both species delimitation methods including the automatic barcode gap discovery method (ABGD) and the Multi-rate Poisson tree process (mPTP) were consistent with those of morphological identification, sequence comparisons with the GenBank and Barcode of Life Data System (BOLD) databases, and the neighbor-joining tree construction. These consistent results emphasize the efficacy of DNA barcoding in the precise identification of Armigeres species.