Mosquito rearing

Anopheles coluzzii (SUA 2La/2La) originating from Suakoko, Liberia, previously known as Anopheles gambiae s.s. M form [9], was maintained in the Vanderbilt Insectary and mosquito rearing primarily followed a lab protocol developed in a previous study [32]. In brief, larvae were reared under standardized conditions (> 2 cm² water surface per larva with 1 liter of dH2O per larval pan) by adding food ad libitum in environmental chamber (27°C, 80% relative humidity, light:dark = 12:12 h) to ensure consistent larval development and large size adults. Pupae were collected and eclosed in a cage, and females and males were allowed to mate with constant access to 10% sucrose solution. For blood feeding, 6 to 7 days old females were provided with human blood (BioChemed, Winchester, VA) by using Hemotek membrane feeding system (Hemotek, Lancaster, UK), and 4.5% CO₂ was utilized to promote blood feeding.