Analysis of geometric morphometrics and molecular phylogeny for Anopheles species in the Republic of Korea.

Human malaria, transmitted by Anopheles mosquitoes, is the most predominant mosquito-borne disease that is responsible for hundreds of thousands of deaths worldwide each year. In the Republic of Korea (ROK), there are currently several hundred malaria cases annually, mostly near the demilitarized zone (DMZ). Eight species of Anopheles mosquitoes are currently known to be present in the ROK. Similar to other major malaria vectors in Africa and India, it is very challenging to morphologically differentiate Anopheles mosquitoes in the ROK due to their extremely similar morphology. In this study, wing geometric morphometrics (WGM) were used to differentiate the eight Anopheles species collected at six locations near the DMZ, Seoul and Pyeongtaek from April-October 2021. Phylogenetic analysis was also performed using cytochrome c oxidase subunit 1 (COI), internal transcribed spacer 2 (ITS2), and tyrosine hydroxylase (TH) genes for comparison with WGM analysis and to infer evolutionary relationships. The results of cross-validation (overall accuracy = 74.8%) demonstrated that species identification using WGM alone was not possible with a high accuracy for all eight species. While phylogenetic analyses based on the COI region could not clearly distinguish some species, the analysis based on ITS2 and TH was more useful for resolving the phylogenetic correlation of the eight species. Our results may improve Anopheles species identification strategies for effective identification and control of malaria vectors in the ROK.