Genetic diversity and phylogeographic structure of Anopheles kochi, Anopheles maculatus, and Anopheles vagus: ITS2-based analysis of highland transboundary populations in the Menoreh Hills, Java, Indonesia
1Department of Environmental Health, Faculty of Public Health, Universitas Diponegoro, Semarang 50275, Indonesia 2Public Health Genomics Lab, Faculty of Public Health, Universitas Diponegoro, Semarang 50275, Indonesia 3Department of Epidemiology and Tropical Diseases, Faculty of Public Health, Universitas Diponegoro, Semarang 50275, Indonesia 4Department of Health Entomology, Faculty of Public Health, Universitas Diponegoro, Semarang 50275, Indonesia 5Environmental Health Laboratory Center, Directorate General of Primary and Community Health, The Ministry of Health of Indonesia, Salatiga 50721, Indonesia 6Department of Medical Environmental Biology and Tropical Medicine, School of Medicine, Kangwon National University, Chuncheon 24341, Korea 7Department of Biomedical Laboratory Science, College of Software and Digital Healthcare Convergence, Yonsei University, Wonju 26493, Korea 8Vector-Borne and Zoonotic Research Group, Research Center for Public Health and Nutrition, National Research and Innovation Agency Indonesia, Salatiga 50721, Indonesia 9Division of Vector-Zoonotic, Health Office of Purworejo District, Central Java Province 54111, Indonesia
Baseline genetic characterization of malaria vector populations provides critical data for evidence-based surveillance in persistent transmission foci. This pilot study generated preliminary genetic baseline data for Anopheles populations in the Menoreh Hills border region between Central Java and Yogyakarta provinces, Indonesia, addressing a key geographic gap in regional vector research. Adult female mosquitoes were collected from 3 houses with reported malaria cases in Ngadirejo Village using standardized entomological methods, including human landing, animal landing, and resting collections. Specimens were morphologically identified and molecularly characterized via ITS2 gene sequencing. Phylogenetic analyses were assessed using maximum likelihood methods, and genetic diversity indices were calculated to examine population structure. A total of 62 specimens representing 3 species were collected exclusively through animal landing collections: Anopheles vagus (48 specimens, 77.4%), Anopheles maculatus (9 specimens, 14.5%), and Anopheles kochi (5 specimens, 8.1%). An. kochi exhibited high haplotype diversity (Hd=0.709) with low nucleotide diversity (π=0.004), while An. maculatus showed lower haplotype diversity (Hd=0.480) and higher nucleotide diversity (π=0.026). Phylogenetic analysis revealed Purworejo specimens clustered with regional populations: An. kochi grouped within Clade I with Indonesian isolates; An. maculatus distributed across multiple clades; An. vagus formed a cohesive unit with other Indonesian populations. The exclusive success of animal landing collections in the Menoreh Hills highlands provides key methodological insights. This study offers essential baseline reference data, validates cost-effective genetic surveillance approaches, and supports future large-scale population connectivity studies across the Menoreh Hills malaria transmission complex.
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