Transplant of cryopreserved mosquito gut microbiota reveals strain-specific patterns of microbial acquisition and assembly

Mosquitoes are important vectors of human pathogens, and their gut microbiome influences traits such as survival, fecundity, pathogen resistance, insecticide resistance, and vector competence. Manipulating this microbiome offers a promising strategy for reducing the mosquito-borne disease. While 16S rRNA gene amplicon sequencing has revealed global patterns of gut microbial diversity, the factors driving these patterns remain poorly understood. Here, we describe a method to collect, cryopreserve, and transplant whole adult microbial communities into axenic (germ-free) mosquitoes. Using reciprocal transplants between Aedes aegypti Liverpool strains maintained for almost 50 years at the University of Wisconsin-Madison (UW) and the Liverpool School of Tropical Medicine (LSTM), we found striking strain-specific responses: UW mosquitoes exhibited altered bacterial composition and developmental defects, whereas LSTM mosquitoes did not. Transplant fidelity depended on donor community composition and recipient genotype, and we observed conserved assembly patterns across the mosquito life cycle. This approach enables mechanistic studies of microbiome assembly and supports the development of microbiome-based vector control strategies.