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Updated: Aug 25, 2025
Quantifying Fitness Costs in Transgenic Aedes aegypti Mosquitoes
Published on: September 15, 2023
Assessing single-locus CRISPR/Cas9-based gene drive variants in the mosquito Aedes aegypti via single-generation
William Reid1, Adeline E Williams2, Irma Sanchez-Vargas2
1Department of Veterinary Pathobiology, University of Missouri, Columbia, MO 65211, USA.
Abstract:
The yellow fever mosquito Aedes aegypti is a major vector of arthropod-borne viruses, including dengue, chikungunya, and Zika viruses. A novel approach to mitigate arboviral infections is to generate mosquitoes refractory to infection by overexpressing antiviral effector molecules. Such an approach requires a mechanism to spread these antiviral effectors through a population, for example, by using CRISPR/Cas9-based gene drive systems. Critical to the design of a single-locus autonomous gene drive is that the selected genomic locus is amenable to both gene drive and appropriate expression of the antiviral effector. In our study, we used reverse engineering to target 2 intergenic genomic loci, which had previously shown to be highly permissive for antiviral effector gene expression, and we further investigated the use of 3 promoters (nanos, β2-tubulin, or zpg) for Cas9 expression. We then quantified the accrual of insertions or deletions (indels) after single-generation crossings, measured maternal effects, and assessed fitness costs associated with various transgenic lines to model the rate of gene drive fixation. Overall, MGDrivE modeling suggested that when an autonomous gene drive is placed into an intergenic locus, the gene drive system will eventually be blocked by the accrual of gene drive blocking resistance alleles and ultimately be lost in the population. Moreover, while genomic locus and promoter selection were critically important for the initial establishment of the autonomous gene drive, it was the fitness of the gene drive line that most strongly influenced the persistence of the gene drive in the simulated population. As such, we propose that when autonomous CRISPR/Cas9-based gene drive systems are anchored in an intergenic locus, they temporarily result in a strong population replacement effect, but as gene drive-blocking indels accrue, the gene drive becomes exhausted due to the fixation of CRISPR resistance alleles.
Insights
CRISPR/Cas9 gene drive systems in mosquitoes show temporary population replacement but are ultimately lost due to resistance alleles and fitness costs. Careful locus and promoter selection are important, but line fitness is key for persistence.
Area of Science:
- Genetics
- Vector Control
- Molecular Biology
Background:
- Aedes aegypti mosquitoes transmit arboviruses like dengue and Zika.
- CRISPR/Cas9 gene drive systems offer a novel method to control mosquito populations by spreading antiviral effectors.
- Successful gene drive implementation requires careful genomic locus and promoter selection for effector expression and drive efficiency.
Purpose of the Study:
- To investigate the potential of intergenic genomic loci for CRISPR/Cas9 gene drive systems in Aedes aegypti.
- To evaluate the impact of different promoters on Cas9 expression and gene drive performance.
- To model the long-term persistence and potential failure of autonomous gene drives in mosquito populations.
Main Methods:
- Utilized reverse engineering to target two intergenic loci in Aedes aegypti.
- Tested three promoters (nanos, β2-tubulin, zpg) for Cas9 expression.
- Quantified indel accrual, measured maternal effects, and assessed fitness costs.
- Employed MGDrivE modeling to simulate gene drive fixation and population dynamics.
Main Results:
- Autonomous gene drives in intergenic loci were predicted to be blocked by resistance alleles and lost from populations.
- Genomic locus and promoter choice influenced initial gene drive establishment.
- Mosquito line fitness was the most critical factor determining gene drive persistence.
- CRISPR resistance alleles ultimately led to gene drive exhaustion.
Conclusions:
- CRISPR/Cas9 gene drives in intergenic loci provide temporary population replacement effects.
- Gene drive systems become exhausted due to the accumulation of resistance alleles.
- Fitness costs significantly impact the long-term viability of gene drive strategies in mosquito vectors.
