Gene drives have enormous potential for the control of insect populations of medical and agricultural relevance. By preferentially biasing their own inheritance, gene drives can rapidly introduce genetic traits even if these confer a negative fitness effect on the population. We have recently developed gene drives based on CRISPR nuclease constructs that are designed to disrupt key genes essential for female fertility in the malaria mosquito. The construct copies itself and the associated genetic disruption from one homologous chromosome to another during gamete formation, a process called homing that ensures the majority of offspring inherit the drive. Such drives have the potential to cause long-lasting, sustainable population suppression, though they are also expected to impose a large selection pressure for resistance in the mosquito. One of these population suppression gene drives showed rapid invasion of a caged population over 4 generations, establishing proof of principle for this technology. In order to assess the potential for the emergence of resistance to the gene drive in this population we allowed it to run for 25 generations and monitored the frequency of the gene drive over time. Following the initial increase of the gene drive we observed a gradual decrease in its frequency that was accompanied by the spread of small, nuclease-induced mutations at the target gene that are resistant to further cleavage and restore its functionality. Such mutations showed rates of increase consistent with positive selection in the face of the gene drive. Our findings represent the first documented example of selection for resistance to a synthetic gene drive and lead to important design recommendations and considerations in order to mitigate for resistance in future gene drive applications.

Background The persistence of malaria in large parts of sub-Saharan Africa has motivated the development of novel tools to complement existing control programmes, including gene-drive technologies to modify mosquito vector populations. Here, we use a stochastic simulation model to explore the potential of using a driving-Y chromosome to suppress vector populations in a 10 6 km 2 area of West Africa including all of Burkina Faso. Results The consequence of driving-Y introductions is predicted to vary across the landscape, causing elimination of the target species in some regions and suppression in others. We explore how this variation is determined by environmental conditions, mosquito behaviour, and the properties of the gene-drive. Seasonality is particularly important, and we find population elimination is more likely in regions with mild dry seasons whereas suppression is more likely in regions with strong seasonality. Conclusions Despite the spatial heterogeneity, we suggest that repeated introductions of modified mosquitoes over a few years into a small fraction of human settlements may be sufficient to substantially reduce the overall number of mosquitoes across the entire geographic area. Electronic supplementary material The online version of this article (10.1186/s12915-019-0645-5) contains supplementary material, which is available to authorized users.

SummaryHoming endonuclease genes (HEGs) exist naturally in many single-celled organisms and can show extremely strong genetic drive allowing them to spread through populations into which they are introduced. They are being investigated as tools to manipulate the populations of important vectors of human disease, in particular the mosquitoes that transmit malaria. Before HEGs can be deployed, it is important to study their spatial spread in order to design efficient release strategies.A spatially explicit model is developed to study the spread of a HEG through a landscape whose structure is defined by the distribution of mosquito breeding and feeding sites. The model is motivated by the biology of the major vectors of malaria in Africa. The conditions for spread, fixation and loss of two major types of HEG are explored in different landscapes.In landscapes where mosquito resources are abundant, the conditions for spread are well approximated by a mean-field model. Where a HEG imposes a genetic load, it can cause population extinction, though spatial models more often predict population suppression.In certain types of landscapes where mosquito resources are rare, an introduced HEG may be prevented from moving between local mosquito populations and so a simple release strategy is unlikely to be effective, yet if the HEG succeeds in spreading population extinction is a feasible outcome. Increasing the number of release sites at the expense of releasing fewer mosquitoes per site reduces the probability that a HEG will fail.Synthesis and applications. The model presented asks for the first time how the spatial structure of mosquito populations will influence the effectiveness of a technology that is being rapidly developed for vector control. If homing endonuclease genes (HEGs) are to be used in this way, we have qualified the importance of accounting for landscape characteristics in both the execution and the expectation of their application. The next stage is to use the model to study the spread of HEGs through real landscapes where releases may take place, something that will be facilitated by the results of the present study.

Gene drives have enormous potential for the control of insect populations of medical and agricultural relevance. By preferentially biasing their own inheritance gene drives can rapidly introduce genetic traits even if these confer a negative fitness on the population.We have recently developed gene drives based on a CRISPR nuclease construct that is designed to disrupt key genes essential for female fertility in the malaria mosquito. The construct copies itself and the associated genetic disruption from one homologous chromosome to another during gamete formation, in a process called homing that ensures the majority of offspring inherit the drive. Such drives have the potential to cause long-lasting, sustainable population suppression though they are also expected to impose a large selection pressure for resistance in the mosquito. One of these population suppression gene drives showed rapid invasion of a caged population over 4 generations, establishing proof of principle for this technology. In order to assess the potential for the emergence of resistance to the gene drive in this population we allowed it to run for 25 generations and monitored the frequency of the gene drive over time. Following the initial increase of the gene drive we noticed a gradual decrease in its frequency that was accompanied emergence of small, nuclease-induced mutations at the target gene that are resistant to further cleavage and restore its functionality. Such mutations show rates of increase consistent with positive selection in the face of the gene drive. Our findings represent the first documented example of selection for resistance to a synthetic gene drive and lead to important design recommendations and considerations in order to mitigate for resistance for future gene drive applications.

Background: Gene drives based on CRISPR-Cas9 technology are increasingly being considered as tools for reducing the capacity of mosquito populations to transmit malaria, and one of the most promising options is driving endonuclease genes that reduce the fertility of female mosquitoes. In particular, there is much interest in constructs that target the conserved mosquito doublesex (dsx) gene such that the emergence of functional driveresistant alleles is unlikely. Proof of principle that these constructs can lead to substantial population suppression has been obtained in population cages, and they are being evaluated for use in sub-Saharan Africa. Here, we use simulation modelling to understand the factors affecting the spread of this type of gene drive over a one millionsquare kilometre area of West Africa containing substantial environmental and social heterogeneity. Results: We found that a driving endonuclease gene targeting female fertility could lead to substantial reductions in malaria vector populations on a regional scale. The exact level of suppression is influenced by additional fitness costs of the transgene such as the somatic expression of Cas9, and its deposition in sperm or eggs leading to damage to the zygote. In the absence of these costs, or of emergent drive-resistant alleles that restore female fertility, population suppression across the study area is predicted to stabilise at~95% 4 years after releases commence. Small additional fitness costs do not greatly affect levels of suppression, though if the fertility of females whose offspring transmit the construct drops by more than~40%, then population suppression is much less efficient. We show the suppression potential of a drive allele with high fitness costs can be enhanced by engineering it also to express male bias in the progeny of transgenic males. Irrespective of the strength of the drive allele, the spatial model predicts somewhat less suppression than equivalent non-spatial models, in particular in highly seasonal regions where dry season stochasticity reduces drive efficiency. We explored the robustness of these results to uncertainties in mosquito ecology, in particular their method of surviving the dry season and their dispersal rates. Conclusions: The modelling presented here indicates that considerable suppression of vector populations can be achieved within a few years of using a female sterility gene drive, though the impact is likely to be heterogeneous in space and time.