# Engineering resistance to Zika virus in Aedes aegypti for Cas9 driven population modification

> **NIH NIH R01** · COLORADO STATE UNIVERSITY · 2021 · $574,502

## Abstract

Project Summary. Zika virus (ZIKV; Flavivirus) is transmitted to humans by the mosquito vector Aedes
aegypti and poses significant health risks to the Americas (1, 2).This phase I project from Colorado State
University (CSU; Olson and Black), University of Missouri (MU; Franz) and University of Notre Dame (UND;
Fraser) will apply gene drive technology to generate Ae. aegypti populations refractory to ZIKV and other
arboviruses transmitted by this vector species. Our gene drive approach is based on Clustered Regularly
Interspaced Short Palindromic Repeats (CRISPR)-Cas9 for rapid, effective, genetics-based population
modification of Aedes aegypti. Our principal goals are: 1) apply two validated Anti-Viral Trans Gene (AVTG)
strategies that target ZIKV to minimize or ablate ZIKV infection of Ae. aegypti, 2) generate transgenic Ae.
aegypti that express the AVTGs in the context of a CRISPR/Cas9 (CC) gene drive system, 3) show that the
genetically modified mosquitoes have a stable, refractory phenotype for ZIKV infection, and 4) validate gene
drive (in indoor cage studies) by introgressing the AVTGs into an Ae. aegypti population to convert the
population from a competent to a refractory infection phenotype. Previously, Drs. Olson and Franz have
genetically modified Ae. aegypti (with no drive) that stably express DENV2-specific, inverted repeat (IR) RNAs
forming dsRNA to trigger the small interfering (si)RNA pathway of RNAi in the mosquito (3, 4). Dr. Fraser has
developed mosquito cell lines that express group I intron (GrpI) ribozymes targeted to a highly conserved
region of arboviral genomes. He has already developed GrpI ribozymes that target all four DENV serotypes
and chikungunya virus (CHIKV; Alphavirus) (6, 7). In this proposal, we will initially develop siRNA- and
ribozyme-based anti-viral genes to target ZIKV in the context of CC gene drive. We will compare the two AVTG
strategies with CC gene drive to see which is the most effective at suppressing mosquito infection. If the GrpI
AVTG genes successfully suppress vector competence and spread the ZIKV AVTG into caged populations of
Ae. aegypti, we will use other GrpI AVTGs with the CC drive system to additionally target DENVs and CHIKV.
CC gene drive has already been used to efficiently modify cage-populations of Anopheles stephensi with anti-
pathogen genes (8). We are confident we can adapt this approach to drive antiviral IR RNA and ribozyme
AVTG genes to greatly reduce or ablate Ae. aegypti competence for transmitting ZIKV. The CC-AVTG system
we propose will target two Ae. aegypti genome sites (TIMP P4 on chromosome aem 2q and 3'UTR
polyadenylate binding protein (PABP on arm 3q). We use the β2-tubulin promoter to express Cas9 to achieve
gene drive through the male germ-line. This will maximize homology-dependent repair (HDR) to maintain drive
over non-homologous end-joining (NHEJ) which impairs drive (9). This proposal represents a significant step
towards developing and optimizing CC drive w...

## Key facts

- **NIH application ID:** 10112159
- **Project number:** 5R01AI130085-04
- **Recipient organization:** COLORADO STATE UNIVERSITY
- **Principal Investigator:** KENNETH E OLSON
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $574,502
- **Award type:** 5
- **Project period:** 2018-04-15 → 2023-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10112159

## Citation

> US National Institutes of Health, RePORTER application 10112159, Engineering resistance to Zika virus in Aedes aegypti for Cas9 driven population modification (5R01AI130085-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10112159. Licensed CC0.

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