Generation and rearing of axenic Aedes aegypti mosquitoes

Abstract This protocol describes the generation of axenic Aedes aegypti, which can be raised from larvae to adults in the absence of a microflora. The total protocol including the time taken to grow E. coli (used in one of the diets) takes two days to set up. The mosquitoes will develop normally with a slight developmental lag compared to conventionally raised mosquitoes (~1 week). Two larval diets are described, a liver:yeast (LY) extract diet and an LY diet supplemented with heat-inactivated E. coli. Axenic mosquitoes raised on food supplemented with heat-inactivated E. coli develop slightly faster than axenic mosquitoes raised without killed bacteria. This axenic system allows the manipulation of the mosquito microbiome and transitions the microbiome into a variable that can be controlled and manipulated in a mechanistic manner.