Developing genetic tools in an endoparasitoid wasp

CRISPR/Cas gene editing is a common tool for genetic studies that has been increasingly applied in non-model organisms. However, CRISPR has not yet been applied in any endoparasitoids, despite their vast diversity and relevance to pest control. Trichogramma pretiosum is a minute endoparasitoid wasp less than 300 microns long and is important in applied lepidopteran biological control . Additionally, some populations of T. pretiosum are infected with parthenogenesis-inducing Wolbachia, making it a great system to study asexual reproduction. Currently, a major challenge for CRISPR-based gene editing lies in enzyme delivery. In insects, many CRISPR protocols rely on direct injection of developing embryos, or on yolk-pathway meditated uptake into developing oocytes. Unfortunately, Trichogramma embryos are simply too small to be injected with the tools available to most labs, and they do not produce yolk. We are developing alternative approaches for enzyme delivery in this species, ultimately allowing us to circumvent the massive physical trauma associated with direct injection and creating a simple and reliable method of embryonic Cas9-RNP delivery. Through this process we have successfully generated visually distinct mutants through an eye pigmentation gene knockout. Going forward, we will optimize the technique and begin using it to generate experimental knockouts for genes related to reproductive fitness, as part of our research into the evolutionary consequences of asexual reproduction.