Developing CRISPR-based self-limiting strategies for population suppression of the malaria vector Anopheles gambiae
File(s)
Author(s)
Strampelli, Anna
Type
Thesis
Abstract
Genetic vector control tools constitute potential complementary alternatives to current control measures for malaria and other vector-borne diseases. Such strategies could be leveraged to replace or suppress vector populations, either in a self-sustaining manner, whereby they propagate over time, or in a self-limiting one, whereby their spread is limited in space/time. Self-limiting strategies, in addition to exerting replacement or suppressive effects, can constitute intermediary products in the development pathway of self-sustaining counterparts. I generated and tested three self-limiting strategies for population suppression of the malaria vector An. gambiae: male-drive female-sterile (MDFS), split drive and split X-shredder. These were all engineered leveraging the CRISPR-Cas9 system to target and disrupt the female-specific exon of doublesex, a highly conserved gene involved in sex determination.
MDFS was tested in a multi-generational cage trial, which confirmed its potential as a self-limiting population suppression strategy. Specifically, two replicate caged populations of wildtype mosquitoes crashed following eight consecutive releases of MDFS males (at a 1:2 ratio of MDFS:wildtype males). Conversely, cages where only three releases were performed did not crash. A deterministic model simulating field releases suggests MDFS is more efficient than all its self-limiting comparators. The split drive was tested in a fertility assay, which revealed it is also a valid self-limiting system. Modelling suggests it is also superior to the other self-limiting comparators, though less so than MDFS. Conversely, the split X-shredder was not successful in its present configuration, but different approaches could be taken to improve it in the future. Finally, these experiments allowed to add to the existing body of knowledge on the promoters used to drive Cas9 expression in the different strategies, namely zpg and vasa2.
Overall, the results presented here contribute to the growing body of knowledge on genetic strategies for vector control and to their responsible deployment in the field.
MDFS was tested in a multi-generational cage trial, which confirmed its potential as a self-limiting population suppression strategy. Specifically, two replicate caged populations of wildtype mosquitoes crashed following eight consecutive releases of MDFS males (at a 1:2 ratio of MDFS:wildtype males). Conversely, cages where only three releases were performed did not crash. A deterministic model simulating field releases suggests MDFS is more efficient than all its self-limiting comparators. The split drive was tested in a fertility assay, which revealed it is also a valid self-limiting system. Modelling suggests it is also superior to the other self-limiting comparators, though less so than MDFS. Conversely, the split X-shredder was not successful in its present configuration, but different approaches could be taken to improve it in the future. Finally, these experiments allowed to add to the existing body of knowledge on the promoters used to drive Cas9 expression in the different strategies, namely zpg and vasa2.
Overall, the results presented here contribute to the growing body of knowledge on genetic strategies for vector control and to their responsible deployment in the field.
Version
Open Access
Date Issued
2023-08-07
Date Awarded
01/07/2024
License URL
Advisor
Crisanti, Andrea
Fasulo, Barbara
Sponsor
Imperial College London
Publisher Department
Life Sciences
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)