A pigment-deficient mosquito line with fluorescent salivary glands enables in vivo imaging of individual Plasmodium sporozoites

Plasmodium parasites are the causative agent of malaria, a major health burden in sub- Saharan Africa. A key step in the transmission process of Plasmodium is the colonization of the salivary glands of the female Anopheles mosquito by the parasite sporozoite stage. How sporozoites recognize and invade the salivary glands is still poorly understood. Here we generated salivary gland reporter lines in the African malaria mosquito Anopheles coluzzii using salivary gland-specific promoters of the genes encoding anopheline antiplatelet protein (AAPP), the triple functional domain protein (TRIO) and saglin (SAG). The observed expression pattern of the DsRed and roGFP2 fluorescent reporters revealed lobe-specific activity of these promoters within the salivary glands, restricted either to the distal lobes or the middle lobe. We characterized four mosquito lines (AAPP-DsRed, AAPP-roGFP2, TRIO- DsRed and SAG-EGFP) in terms of localization, expression strength and onset of expression, as well as potential influences of the expressed fluorescent reporters on the infection with Plasmodium berghei and salivary gland morphology. Furthermore, using crosses with a pigmentation deficient yellow(-) mosquito line, we demonstrate that our salivary gland reporter lines represent a valuable tool to study the process of salivary gland colonization by Plasmodium parasites in live mosquitoes. ### Competing Interest Statement The authors have declared no competing interest.