Identification of a novel resistance gene which provides insight into Vip3Aa mode of action in Helicoverpa armigera

Abstract

The global reliance on Bacillus thuringiensis (Bt) proteins for controlling lepidopteran pests in cotton, corn, and soybean crops underscores the critical need to understand resistance mechanisms. Vip3Aa, one of the most widely deployed and currently effective Bt proteins in genetically modified crops, plays a pivotal role in pest management. This study identifies the molecular basis of Vip3Aa resistance in Australian Helicoverpa armigera through genetic crosses, and integrated genomic and transcriptomic analyses. We identified a previously uncharacterized gene, LOC110373801 (designated HaVipR1), as a crucial determinant of Vip3Aa resistance in two field-derived resistant lines. Functional validation using CRISPR-Cas9 knockout in susceptible lines confirmed the genes role in conferring resistance. Despite extensive laboratory selection of Vip3Aa-resistant colonies in Lepidoptera, the biochemical mechanisms underlying resistance have remained elusive. Our research demonstrates that HaVipR1-mediated resistance operates independently of known resistance genes, including midgut-specific chitin synthase and the transcription factor SfMyb. The identification of HaVipR1 offers further insights into the Vip3Aa mechanism of action. This discovery is vital for devising strategies to counteract resistance and sustain the efficacy of Bt crops. Future research should focus on elucidating the biochemical pathways involving HaVipR1 and investigating its interactions with other resistance mechanisms. Our findings underscore the utility of analysing field-derived resistant lines in providing biologically relevant insights and stress the necessity for comprehensive management strategies to maintain agricultural productivity. Significance StatementThis is the first identification of a specific gene in Helicoverpa armigera which mediates resistance to the Bacillus thuringiensis (Bt) protein Vip3Aa. We identify that this gene is disrupted in two different ways in separate field-derived resistant lines, one being a large transposable element insertion in the first intron of the HaVipR1 gene, confirmed using long-read sequencing. Disruption of this gene using CRISPR-Cas9 knockout in a susceptible H. armigera line confers Vip3A resistance. The identification of a specific gene is important for molecular monitoring and management of H. armigera as well as other global pests of concern like Spodoptera frugiperda. These findings also offer insights for researchers aiming to understand how Vip3Aa functions, as the action pathway remains unclear.