CRISPR/Cas9 gene drive (CGD) promises to be a highly adaptable approach for spreading genetically engineered alleles throughout a species, even if those alleles impair reproductive success. CGD has been shown to be effective in laboratory crosses of insects, yet it remains unclear to what extent potential resistance mechanisms will affect the dynamics of this process in large natural populations. Here we develop a comprehensive population genetic framework for modeling CGD dynamics, which incorporates potential resistance mechanisms as well as random genetic drift. Using this framework, we calculate the probability that resistance against CGD evolves from standing genetic variation, de novo mutation of wild-type alleles, or cleavage repair by nonhomologous end joining (NHEJ)—a likely by-product of CGD itself. We show that resistance to standard CGD approaches should evolve almost inevitably in most natural populations, unless repair of CGD-induced cleavage via NHEJ can be effectively suppressed, or resistance costs are on par with those of the driver. The key factor determining the probability that resistance evolves is the overall rate at which resistance alleles arise at the population level by mutation or NHEJ. By contrast, the conversion efficiency of the driver, its fitness cost, and its introduction frequency have only minor impact. Our results shed light on strategies that could facilitate the engineering of drivers with lower resistance potential, and motivate the possibility to embrace resistance as a possible mechanism for controlling a CGD approach. This study highlights the need for careful modeling of the population dynamics of CGD prior to the actual release of a driver construct into the wild.

Evolutionary rescue occurs when a population that is threatened with extinction by an environmental change adapts to the change sufficiently rapidly to survive. Here we extend the mathematical theory of evolutionary rescue. In particular, we model evolutionary rescue to a sudden environmental change when adaptation involves evolution at a single locus. We consider adaptation using either new mutations or alleles from the standing genetic variation that begin rare. We obtain several results: i) the total probability of evolutionary rescue from either new mutation or standing variation; ii) the conditions under which rescue is more likely to involve a new mutation versus an allele from the standing genetic variation; iii) a mathematical description of the U-shaped curve of total population size through time, conditional on rescue; and iv) the time until the average population size begins to rebound as well as the minimal expected population size experienced by a rescued population. Our analysis requires taking into account a subtle population-genetic effect (familiar from the theory of genetic hitchhiking) that involves “oversampling” of those lucky alleles that ultimately sweep to high frequency. Our results are relevant to conservation biology, experimental microbial evolution, and medicine (e.g., the dynamics of antibiotic resistance). Changes to an organism's environment may have such adverse effects on fitness that the population begins to decline in size. To survive, the population must adapt before it goes extinct. Such “evolutionary rescue” is characterized by a U-shaped curve: population size declines and then recovers as a beneficial allele increases in number. Here we describe this U-shaped curve mathematically when the rescuing allele starts out rare. We obtain several results. First, we calculate when evolutionary rescue is more likely to come from new mutation than from the standing genetic variation. Second, by describing the entire U-shaped curve mathematically, we derive the time until the average rescued population begins to rebound in size as well as the smallest average population size experienced before rescue. We also find that evolutionary rescue from new mutation takes longer and involves a smaller minimum population size than rescue from the standing genetic variation.

The use of recombinant genetic technologies for population manipulation has mostly remained an abstract idea due to the lack of a suitable means to drive novel gene constructs to high frequency in populations. Recently Gantz and Bier showed that the use of CRISPR/Cas9 technology could provide an artificial drive mechanism, the so-called mutagenic chain reaction (MCR), which could lead to rapid fixation of even a deleterious introduced allele. We establish the near equivalence of this system to other gene drive models and review the results of simple models showing that, when there is a fitness cost to the MCR allele, an internal equilibrium may exist that is usually unstable. In this case, introductions must be at a frequency above this critical point for the successful invasion of the MCR allele. We obtain estimates of fixation and invasion probabilities for the appropriate scenarios. Finally, we discuss how polymorphism in natural populations may introduce sources of natural resistance to MCR invasion. These modeling results have important implications for application of MCR in natural populations.

Wolbachia are the most widespread maternally-transmitted bacteria in the animal kingdom. Their global spread in arthropods and varied impacts on animal physiology, evolution, and vector control are in part due to parasitic drive systems that enhance the fitness of infected females, the transmitting sex of Wolbachia. Male killing is one common drive mechanism wherein the sons of infected females are selectively killed. Despite decades of research, the gene(s) underlying Wolbachia-induced male killing remain unknown. Here using comparative genomic, transgenic, and cytological approaches in fruit flies, we identify a candidate gene in the eukaryotic association module of Wolbachia prophage WO, termed WO-mediated killing (wmk), which transgenically causes male-specific lethality during early embryogenesis and cytological defects typical of the pathology of male killing. The discovery of wmk establishes new hypotheses for the potential role of phage genes in sex-specific lethality, including the control of arthropod pests and vectors. Male killing is an adaptive trait for bacteria that are maternally transmitted through host populations. Such bacteria are common in arthropods and resultantly have significant impacts on host population size, mating strategy, and evolution. Moreover, male-killing bacteria are under recent scrutiny as a symbiotic strategy for arthropod pest and vector control. Despite decades of research, the microbial genetic basis of Wolbachia-induced male killing remains elusive. Here we demonstrate that a single gene from the eukaryotic association module in prophage WO of Wolbachia is a candidate for male killing as it recapitulates many aspects of the phenotype when transgenically expressed in fruit flies. This discovery represents a step forward in understanding new roles of phage WO genes in shaping arthropod hosts and may inform the potential use of male killing in worldwide pest and vector control strategies.

Both malnutrition and undernutrition can lead to compromised immune defense in a diversity of animals, and “nutritional immunology” has been suggested as a means of understanding immunity and determining strategies for fighting infection. The genetic basis for the effects of diet on immunity, however, has been largely unknown. In the present study, we have conducted genome-wide association mapping in Drosophila melanogaster to identify the genetic basis for individual variation in resistance, and for variation in immunological sensitivity to diet (genotype-by-environment interaction, or GxE). D. melanogaster were reared for several generations on either high-glucose or low-glucose diets and then infected with Providencia rettgeri, a natural bacterial pathogen of D. melanogaster. Systemic pathogen load was measured at the peak of infection intensity, and several indicators of nutritional status were taken from uninfected flies reared on each diet. We find that dietary glucose level significantly alters the quality of immune defense, with elevated dietary glucose resulting in higher pathogen loads. The quality of immune defense is genetically variable within the sampled population, and we find genetic variation for immunological sensitivity to dietary glucose (genotype-by-diet interaction). Immune defense was genetically correlated with indicators of metabolic status in flies reared on the high-glucose diet, and we identified multiple genes that explain variation in immune defense, including several that have not been previously implicated in immune response but which are confirmed to alter pathogen load after RNAi knockdown. Our findings emphasize the importance of dietary composition to immune defense and reveal genes outside the conventional “immune system” that can be important in determining susceptibility to infection. Functional variation in these genes is segregating in a natural population, providing the substrate for evolutionary response to pathogen pressure in the context of nutritional environment. Previous studies have indicated that dietary nutrition influences immune defense in a variety of animals, but the mechanistic and genetic basis for that influence is largely unknown. We use the model insect Drosophila melanogaster to conduct an unbiased genome-wide mapping study to identify genes responsible for variation in resistance to bacterial infection after rearing on either high-glucose or low-glucose diets. We find the flies are universally more susceptible to infection when they are reared on the high-glucose diet than when they are reared on the low-glucose diet, and that metabolite levels genetically correlate with quality of immune defense after rearing on the high-glucose diet. We identify several genes that contribute to variation in defense quality on both diets, most of which are not traditionally thought of as part of the immune system. The genetic variation we observe can be important for evolved responses to pathogen pressure, although the effectiveness of natural selection will be partially determined by the host nutritional state.