Avoiding CRISPR-Mediated Gene-Drive–Evolved Resistance in Mosquitoes

Scientists at UC Berkeley and UC Riverside have demonstrated a way to edit the genome of disease-carrying mosquitoes that brings us closer to suppressing them on a continental scale. [NIH]

The discovery and development of improved gene-editing techniques in recent years have led to the revamping of a technique that promotes the inheritance of a particular gene or set of genes to increase the prevalence within a population. This so-called gene-drive technology has the potential to eliminate disease within a population of disease-carrying organisms, such as mosquitoes, but has been met with varying degrees of circumspection, as the technique also can potentially wipe out entire species.

Now, researchers at UC Berkeley and UC Riverside have demonstrated a way to edit the genome of disease-carrying mosquitoes that brings us closer to suppressing them on a continental scale. Findings from the new study were published recently in Scientific Reports, in an article entitled “Overcoming Evolved Resistance to Population-Suppressing Homing-Based Gene Drives.” The study utilized mathematical modeling and CRISPR/Cas9 gene-editing technology to insert and spread genes designed to suppress wild insects (the fruit fly Drosophila), while at the same time avoiding the resistance to these efforts that evolution would typically favor.

“We develop a mathematical model to estimate tolerable rates of homing-resistant allele generation to suppress a wild population of a given size,” the authors wrote. “Our results suggest that to achieve meaningful population suppression, tolerable rates of resistance allele generation are orders of magnitude smaller than those observed for current designs for CRISPR-Cas9-based homing systems.”

The authors continued stating that “to remedy this, we theoretically explore a homing system architecture in which guide RNAs (gRNAs) are multiplexed, increasing the effective homing rate and decreasing the effective resistant allele generation rate. Modeling results suggest that the size of the population that can be suppressed increases exponentially with the number of multiplexed gRNAs and that, with four multiplexed gRNAs, a mosquito species could potentially be suppressed on a continental scale.”

While the proof-of-concept study was demonstrated in fruit flies, the researchers believe this technology could be used in mosquitoes to help fight malaria and other mosquito-borne diseases in the next decade, pending public and regulatory approval.

“What we showed is that, if you disrupt a gene required for fertility in female mosquitoes at multiple sites all at once, it becomes much harder for the population to evolve around that disruption,” explained lead study investigator John Marshall, Ph.D., assistant professor of biostatistics and epidemiology at the UC Berkeley School of Public Health. “As a result, you can suppress a much larger population. It's much the same as combination drug therapy—but for CRISPR-based gene drive.”

An example of the use of gene drive in a mosquito population. [DOI:http://dx.doi.org/10.7554/eLife.03401.002]

Gene drives are used to bias genetic inheritance in favor of rapidly spreading, self-destructive genes and could be an environmentally friendly and cost-effective way to suppress populations of disease-spreading insects. The rise of CRISPR/Cas9 gene-editing technology has recently revolutionized gene-drive systems because it offers a rapid, efficient, and reliable way to make precise, targeted changes to the genome.

The new study based its calculations on past gene-drive findings that resulted in up to 99% of offspring inheriting the inserted gene. However, the few offspring that don't inherit the gene present a big problem for this technology. Since a fraction of these offspring is immune to the gene drive, any attempt to eliminate a mosquito species in this manner would result in a rapid rebound of those that are gene-drive immune. The impact of this resistance on the ability of gene drive to spread and suppress populations had previously been discussed—but had not been thoroughly evaluated.

The mathematical modeling that the investigators utilized found that the gene-drive–evolved resistance would have a major impact on attempts to eliminate a mosquito species on a continent-wide scale. To address this issue, the research team devised a technique that they determined could potentially suppress mosquito species continent-wide.

Employing a strategy called multiplexing, which involves using one of the components of the CRISPR system, a gRNA, to target multiple locations in a gene at once, the research team suggested that the size of the population that could be suppressed increases exponentially with the number of these gRNAs utilized. It also shows that with four or five multiplexed gRNAs, a mosquito species could potentially be suppressed on a continental scale.

“Knowing that we can potentially overcome the issues of resistance through careful engineering and multiplexing is huge,” noted senior study investigator Omar Akbari, Ph.D., assistant professor of entomology at UC Riverside.

The researchers demonstrated the technology was feasible using a fruit fly model. Now they are working to adapt this technology to the mosquito species that transmit malaria, dengue, and Zika.

“The potential of multiplexing is vast. With one gRNA, we could suppress a room of mosquitoes,” Dr. Marshall concluded. “With four, we could potentially suppress a continent and the diseases they transmit. But nature has a knack for finding a way around hurdles, so assessing that potential will require a lot more work.”