Background Anarchic and poorly controlled urbanization led to an increased risk of mosquito-borne diseases (MBD) in many African cities. Here, we evaluate the spatial heterogeneity of human exposure to malaria and arboviral disease vectors in an urban area of northern Senegal, using antibody-based biomarkers of exposure to Anopheles and Aedes mosquito bites. Methods A cross-sectional study was undertaken during the rainy season of 2014 in 4 neighborhoods of Saint-Louis, a city in northern Senegal. Among children aged 6–59 months in each neighborhood, the dried blood spot technique was used to evaluate immunoglobulin G (IgG) responses to both gSG6-P1 (Anopheles) and Nterm–34-kDa (Aedes) salivary peptides as validated biomarkers of respective mosquito bite exposure. Results IgG response levels to gSG6-P1 and Nterm–34-kDa salivary peptides varied significantly between the 4 neighborhoods (P < .0001). The level of exposure to Aedes bites also varied according to household access to sanitation services (P = .027), whereas that of exposure to Anopheles bites varied according to insecticide-treated bed net use (P = .006). In addition, spatial clusters of high contact between humans and mosquitoes were identified inside 3 neighborhoods. Conclusions Antibody-based biomarkers of exposure to Anopheles and Aedes mosquito bites could be helpful tools for evaluating the heterogeneity of exposure to malaria and arboviral disease vectors by national control programs.