A transgenic female killing system for the genetic control of Drosophila suzukii

Marc F Schetelig,Ying Yan,Jonas Schwirz

doi:10.1038/s41598-021-91938-1

Abstract

The spotted wing Drosophila (Drosophila suzukii) is an invasive pest of soft-skinned fruit crops. It is rapidly transmitted in Europe and North America, causing widespread agricultural losses. Genetic control strategies such as the sterile insect technique (SIT) have been proposed as environment-friendly and species-restricted approaches for this pest. However, females are inefficient agents in SIT programs. Here we report a conditional female-killing (FK) strategy based on the tetracycline-off system. We assembled sixteen genetic constructs for testing in vitro and in vivo. Twenty-four independent transgenic strains of D. suzukii were generated and tested for female-specific lethality. The strongest FK effect in the absence of tetracycline was achieved by the construct containing D. suzukii nullo promoter for early gene expression, D. suzukii pro-apoptotic gene hidAla4 for lethality, and the transformer gene intron from the Mediterranean fruit fly Ceratitis capitata for female-specific splicing. One strain carrying this construct eliminated 100% of the female offspring during embryogenesis and produced only males. However, homozygous females from these FK strains were not viable on a tetracycline-supplemented diet, possibly due to the basal expression of hidAla4. Potential improvements to the gene constructs and the use of such FK strains in an SIT program are discussed.

Highlights

IntroductionWe have previously reported the isolation and analysis of D. suzukii cellularization gene promoters, pro-apoptotic genes, and the nuclear localization signal (NLS) of the tra gene from the standard USA laboratory s train[27,28,29]
Cassette) to ensure that only females are killed[18,19,22,23]
We generated a range of AIO lethal constructs for in vitro cell culture assays and in vivo analysis, each containing the AmCyan marker gene controlled by the constitutive D. melanogaster polyubiquitin (PUb) promoter and an attP recombination site (Fig. 1a,b)

Summary

Introduction

We have previously reported the isolation and analysis of D. suzukii cellularization gene promoters, pro-apoptotic genes, and the nuclear localization signal (NLS) of the tra gene from the standard USA laboratory s train[27,28,29] In these studies, we confirmed the embryonic expression of the cellularization genes nullo, serendipity-α (sry-α), bottleneck (bnk), and slow-as-molasses (slam) and the functionality of the pro-apoptotic genes head involution defective (hid), grim, and reaper (rpr). We generated 16 all-in-one (AIO) piggyBac plasmids containing the driver and effector cassettes in a single genetic construct Some of these constructs were tested in vitro for their effect on cell survival, and the most promising ones were used to generate transgenic D. suzukii strains. We measured tTA expression levels and determined the developmental stage of lethality for females in the absence of tetracycline in several FK strains

Methods

Results

Conclusion