Anopheles azevedoi (Ribeiro, 1969) in Angola: new geographical records, molecular characterization, and insecticide susceptibility profile of a commonly misidentified species

Abstract Background Anopheles azevedoi is a poorly studied member of the Paramyzomyia series, originally described from Angola in 1969. Since then, little information has been available regarding its distribution, ecology, vectorial importance, and susceptibility to insecticides. This study updates the distribution of An. azevedoi in Angola, provides the first molecular characterization of the species, and assesses its blood-feeding behavior, Plasmodium infection status, and insecticide susceptibility profile. Methods Entomological surveillance was conducted in Luanda, Benguela, Cuanza Sul, and Namibe provinces between 2015 and 2022. Adult mosquitoes were collected to characterize species composition, while larvae were reared to obtain live specimens for insecticide susceptibility bioassays. Anopheles mosquitoes were initially identified morphologically and subsequently confirmed using molecular markers (ITS2 and COI sequencing). A subset of wild-caught adults was screened for Plasmodium infection and blood-meal sources. Insecticide susceptibility was evaluated using WHO tube and bottle bioassays. Results An. azevedoi was confirmed in multiple habitats along the Atlantic coast of Angola, including new records from Luanda and Benguela. Populations exhibited resistance to pyrethroids and DDT, while two populations from Luanda remained susceptible to chlorfenapyr. No Plasmodium infections were detected among the screened adult mosquitoes. The species was frequently collected in urban and peri-urban settings, and human blood meals were identified in three field-collected specimens, providing evidence that An. azevedoi feeds on humans in the study area. Conclusions This study provides the first publicly available genetic markers (COI and ITS2) for the molecular identification of An. azevedoi and expands its known distribution in Angola. Evidence of human blood feeding and resistance to pyrethroids and DDT highlights the importance of including this species in routine entomological surveillance and future studies of its potential public health relevance.