Consequences of resistance evolution in a Cas9-based sex conversion-suppression gene drive for insect pest management

Resistance evolution caused by CRISPR/Cas9 gene-drive systems has a major impact on both the future scientific design of such gene-drive systems and on the politics of regulating experimentation and use of such systems. In our study, we show that in-frame drive-resistant alleles can be produced readily and inherently in a suppression gene-drive system. The rate at which such alleles emerge will determine the maximum size of the population that could be targeted for collapse and elimination. Here, we provide a potential target site and the modeling framework for implementation and optimization of a suppression gene-drive strategy to control Mediterranean fruit fly populations.The use of a site-specific homing-based gene drive for insect pest control has long been discussed, but the easy design of such systems has become possible only with the recent establishment of CRISPR/Cas9 technology. In this respect, novel targets for insect pest management are provided by new discoveries regarding sex determination. Here, we present a model for a suppression gene drive designed to cause an all-male population collapse in an agricultural pest insect. To evaluate the molecular details of such a sex conversion-based suppression gene drive experimentally, we implemented this strategy in Drosophila melanogaster to serve as a safe model organism. We generated a Cas9-based homing gene-drive element targeting the transformer gene and showed its high efficiency for sex conversion from females to males. However, nonhomologous end joining increased the rate of mutagenesis at the target site, which resulted in the emergence of drive-resistant alleles and therefore curbed the gene drive. This confirms previous studies that simple homing CRISPR/Cas9 gene-drive designs will be ineffective. Nevertheless, by performing population dynamics simulations using the parameters we obtained in D. melanogaster and by adjusting the model for the agricultural pest Ceratitis capitata, we were able to identify adequate modifications that could be successfully applied for the management of wild Mediterranean fruit fly populations using our proposed sex conversion-based suppression gene-drive strategy.