A drug‑spiked blood meal provides substantial female elimination as a pretreatment step for mosquito sex separation in the sterile insect technique

The sterile insect technique (SIT) is a sustainable, species-specific mosquito control strategy but limited by inefficient sex-separation, particularly for Anopheles species. Using female obligate blood-feeding, we aimed to develop a pharmacological method to selectively eliminate females, preserving males for SIT. We conducted a large-scale screening of candidate compounds in Aedes albopictus, followed by systematic optimization of drug formulation and blood-feeding parameters, including feeding interval, temperature, frequency, duration, and mosquito age. The performance of surviving males was rigorously evaluated based on flight ability, longevity, mating capacity, and mating competitiveness. The optimized protocol was subsequently validated in Anopheles stephensi. An optimal formulation (deltamethrin 1,000 ppm, PBO 6,000 ppm, ATP 10 ppm) under optimized conditions (36 °C, two 1 h feedings at 5 to 6 d post-emergence) achieved ∼82.0% female elimination in Ae. albopictus while keeping male mortality below 4.0%. Surviving males showed no significant impairment in flight ability, longevity, insemination capacity, or mating competitiveness, indicating the method's suitability as a pretreatment to reduce female numbers prior to precision sorting. Efficacy was even higher in An. stephensi (∼90.0% female elimination; 45:1 female-to-male mortality ratio) without compromising male flight ability. We developed a simple, cross-species pretreatment using drug-spiked blood meals to support Aedes and Anopheles SIT programs. Although not meeting the