Malaria Resistant Mosquitos by Design

Malaria Resistant Mosquitos by Design

Larry H. Bernstein, MD, FCAP, Curator



CRISPR-Powered Malaria Mosquito Gene Drive

Using the precision gene-editing tool, researchers demonstrate an ability to create large populations of malaria parasite–resistant mosquitoes.

By Tracy Vence | November 24, 2015

Using CRISPR, investigators at the Universities of California (UC) in San Diego and Irvine have engineered transgenic Anopheles stephensimosquitoes carrying an anti-malaria parasite effector gene “capable of introgressing the genes throughout wild vector populations,” they wrote in a PNAS paper published this week (November 23). The resulting gene-drive system could help wipe out the malaria pathogen (Plasmodium falciparum) within a targeted population of A. stephensi vectors, Anthony James of UC Irvine and his colleagues wrote.

“We know the gene works,” James said in a statement. “The mosquitoes we created are not the final brand, but we know this technology allows us to efficiently create large populations.”

As Nature noted, this study is not the first to report engineered Anopheles that stifle the malaria parasite but, until now, “researchers lacked a way to ensure that the resistance genes would spread rapidly through a wild population.” CRISPR/Cas9 gene-editing enabled this feat. “Males and females derived from transgenic females . . . produce progeny with a high frequency of mutations in the targeted genome sequence, resulting in near-Mendelian inheritance ratios of the transgene,” James and his colleagues wrote in their paper. (See “Reining in Gene Drives,” The Scientist, November 2015.)

“This work suggests that we’re a hop, skip, and jump away from actual gene-drive candidates for eventual release,” Kevin Esvelt of the Wyss Institute who was not involved in the work told Nature. “This is a major advance because it shows that gene drives will likely be effective in mosquitoes,” Esvelt told MIT Technology Review. “Technology is no longer the limitation.”

In the UC Irvine statement, study coauthor Ethan Bier of UC San Diego added that “the ability of this system to carry large genetic payloads should have broad applications to the future use of related CRISPR-based ‘active genetic’ systems.”

Reining in Gene Drives

Researchers have developed two methods to avoid the unchecked spread of engineered genes through wild populations.

By Karen Zusi | November 18, 2015

“Gene drive” is a phenomenon that causes a gene to be inherited at a rate faster than Mendelian principles would dictate. It relies on genes that can copy themselves onto a corresponding location in a paired chromosome, thereby overriding typical allele inheritance patterns. In conjunction with CRISPR/Cas9, gene drives can be created with almost any DNA sequence, raising questions about the risk of engineered genes spreading quickly through a population. But a team of researchers from Harvard University published a study this week (November 16) in Nature Biology that offers some safety constraints on the system.

A description of the first CRISPR/Cas9 gene drive system was published in March by a team at the University of California, San Diego, and showed rapid spreading of a normally recessive phenotype inDrosophila. Other labs are researching the system’s potential to wipe out insect-borne diseases such as malaria by spreading mutated genes throughout a mosquito population. But the strategy carries the risk of accidental contamination of wild populations.

“We have a responsibility to keep our experiments confined to the laboratory,” Kevin Esvelt, an evolutionary engineer and coauthor on the paper, told Nature. “The basic lesson is: if you don’t have to build a gene drive that can spread through a wild population, then don’t.”


‘Gene drive’ mosquitoes engineered to fight malaria

Mutant mozzies could rapidly spread through wild populations.

Heidi Ledford & Ewen Callaway!/image/1.18858%20.jpg_gen/derivatives/landscape_630/1.18858%20.jpg

The Anopheles stephensi mosquito can spread the malaria parasite to humans.

Mutant mosquitoes engineered to resist the parasite that causes malaria could wipe out the disease in some regions — for good.

Humans contract malaria from mosquitoes that are infected by parasites from the genusPlasmodium. Previous work had shown that mosquitoes could be engineered to rebuff the parasiteP. falciparum1, but researchers lacked a way to ensure that the resistance genes would spread rapidly through a wild population.

In work published on 23 November in the Proceedings of the National Academy of Sciences, researchers used a controversial method called ‘gene drive’ to ensure that an engineered mosquito would pass on its new resistance genes to nearly all of its offspring2 — not just half, as would normally be the case.

The result: a gene that could spread through a wild population like wildfire.

“This work suggests that we’re a hop, skip and jump away from actual gene-drive candidates for eventual release,” says Kevin Esvelt, an evolutionary engineer at Harvard University in Cambridge, Massachusetts, who studies gene drive in yeast and nematodes.

For Anthony James, a molecular biologist at the University of California, Irvine, and an author of the paper, such a release would spell the end of a 30-year quest to use mozzie genetics to squash malaria.

James and his laboratory have painstakingly built up the molecular tools to reach this goal. They have worked out techniques for creating transgenic mosquitoes — a notoriously challenging endeavour — and isolated genes that could confer resistance to P. falciparum. But James lacked a way to ensure that those genes would take hold in a wild population.

Fast forward

The concept of engineering a gene drive has been around for about a decade, and James’s laboratory had tried to produce them in the past. The process was agonizingly slow.

Then, in January, developmental biologists Ethan Bier and Valentino Gantz at the University of California, San Diego, contacted James with a stunning finding: they had engineered a gene drive in fruit flies, and wondered whether the same system might work in mosquitoes. James jumped at the opportunity to find out.

Bier and Gantz had used a gene-editing system called CRISPR–Cas9 to engineer a gene drive. They inserted genes encoding the components of the system that were designed to insert a specific mutation in their fruit flies. The CRISPR–Cas9 system then copied that mutation from one chromosome to the other3. James used that system in mosquitoes to introduce two genes that his past work showed would cause resistance to the malaria pathogen.

The resulting mosquitoes passed on the modified genes to more than 99% of their offspring. Although the researchers stopped short of confirming that all the insects were resistant to the parasite, they did show that the offspring expressed the genes.

“It’s a very significant development,” says Kenneth Oye, a political scientist who studies emerging technologies at the Massachusetts Institute of Technology in Cambridge. “Things are moving rapidly in this field.”

Other teams are developing gene drives that could control malaria. A team at Imperial College London has developed a CRISPR-based gene drive in Anopheles gambiae, the mosquito species that transmits malaria in sub-Saharan Africa. The group’s gene drive inactivates genes involved in egg production in female mosquitoes, which could be used to reduce mosquito populations, according to team member Austin Burt, an evolutionary geneticist. Their results will be published inNature Biotechnology next month, Burt says.

Oye notes that such technological advances are outpacing the regulatory and policy discussionsthat surround the use of gene drive to engineer wild populations. Gene drives are controversial because of the potential that they hold for altering entire ecosystems.

Before testing gene drive in the field, Oye hopes that researchers will study the long-term consequences of the changes, such as their stability and potential to spread to other species, as well as methods to control them. “I’m less worried about malevolence than getting something wrong,” he says.

Esvelt says that the US-based researchers made a wise decision in selecting a non-native mosquito species for their experiments. (The team worked with Anopheles stephensi, which is native to the Indian subcontinent.) “Even if they escaped the lab, there’d be no one to mate with and spread the drive,” Esvelt says.

James predicts that it will take his team less than a year to prepare mosquitoes that would be suitable for field tests, but he is in no rush to release them. “It’s not going to go anywhere until the social science advances to the point where we can handle it,” he says. “We’re not about to do anything foolish.”



  1. Isaacs, A. T. et al. PLoS Pathog. 7, e1002017 (2011).
  2. Gantz, V. M. et al. Proc. Natl Acad. Sci. USA




With This Genetic Engineering Technology, There’s No Turning Back

Designers of a “selfish” gene able to spread among mosquitoes say it could wipe out malaria, but the scientific community is at odds over whether or not we should do it.

The students in Anthony James’s basement insectary at the University of California, Irvine, knew they’d broken the laws of evolution when they looked at the mosquitoes’ eyes.

By rights, the bugs, born from fathers with fluorescent red eyes and mothers with normal ones, should have come out only about half red. Instead, as they counted them, first a few and then by the hundreds, they found 99 percent had glowing eyes.

More important than the eye color is that James’s mosquitoes also carry genes that stop the malaria parasite from growing. If these insects were ever released in the wild, their “selfish” genetic cargo would spread inexorably through mosquito populations, and potentially stop the transmission of malaria.

The technology, called a “gene drive,” was built using the gene-editing technology known as CRISPR and is being reported by James, a specialist in mosquito biology, and a half dozen colleagues today in the Proceedings of the National Academy of Sciences.

A functioning gene drive in mosquitoes has been anticipated for more than a decade by public health organizations as a revolutionary novel way to fight malaria. Now that it’s a reality, however, the work raises questions over whether the technology is safe enough to ever be released into the wild.

“This is a major advance because it shows that gene drives will likely be effective in mosquitoes,” says Kevin Esvelt, a gene drive researcher at Harvard University’s Wyss Institute. “Technology is no longer the limitation.”


Highly efficient Cas9-mediated gene drive for population modification of the malaria vector mosquito Anopheles stephensi


Malaria continues to impose enormous health and economic burdens on the developing world. Novel technologies proposed to reduce the impact of the disease include the introgression of parasite-resistance genes into mosquito populations, thereby modifying the ability of the vector to transmit the pathogens. Such genes have been developed for the human malaria parasite Plasmodium falciparum. Here we provide evidence for a highly efficient gene-drive system that can spread these antimalarial genes into a target vector population. This system exploits the nuclease activity and target-site specificity of the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) system, which, when restricted to the germ line, copies a genetic element from one chromosome to its homolog with ≥98% efficiency while maintaining the transcriptional activity of the genes being introgressed.

Genetic engineering technologies can be used both to create transgenic mosquitoes carrying antipathogen effector genes targeting human malaria parasites and to generate gene-drive systems capable of introgressing the genes throughout wild vector populations. We developed a highly effective autonomous Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated protein 9 (Cas9)-mediated gene-drive system in the Asian malaria vector Anopheles stephensi, adapted from the mutagenic chain reaction (MCR). This specific system results in progeny of males and females derived from transgenic males exhibiting a high frequency of germ-line gene conversion consistent with homology-directed repair (HDR). This system copies an ∼17-kb construct from its site of insertion to its homologous chromosome in a faithful, site-specific manner. Dual anti-Plasmodium falciparum effector genes, a marker gene, and the autonomous gene-drive components are introgressed into ∼99.5% of the progeny following outcrosses of transgenic lines to wild-type mosquitoes. The effector genes remain transcriptionally inducible upon blood feeding. In contrast to the efficient conversion in individuals expressing Cas9 only in the germ line, males and females derived from transgenic females, which are expected to have drive component molecules in the egg, produce progeny with a high frequency of mutations in the targeted genome sequence, resulting in near-Mendelian inheritance ratios of the transgene. Such mutant alleles result presumably from nonhomologous end-joining (NHEJ) events before the segregation of somatic and germ-line lineages early in development. These data support the design of this system to be active strictly within the germ line. Strains based on this technology could sustain control and elimination as part of the malaria eradication agenda.


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