Stabilized Double-Stranded RNA Strategy Improves Cotton Resistance to CBW (Anthonomus grandis)

Abstract

Cotton is the most important crop for fiber production worldwide. However, the cotton boll weevil (CBW) is an insect pest that causes significant economic losses in infested areas. Current control methods are costly, inefficient, and environmentally hazardous. Herein, we generated transgenic cotton lines expressing double-stranded RNA (dsRNA) molecules to trigger RNA interference-mediated gene silencing in CBW. Thus, we targeted three essential genes coding for chitin synthase 2, vitellogenin, and ecdysis-triggering hormone receptor. The stability of expressed dsRNAs was improved by designing a structured RNA based on a viroid genome architecture. We transformed cotton embryos by inserting a promoter-driven expression cassette that overexpressed the dsRNA into flower buds. The transgenic cotton plants were characterized, and positive PCR transformed events were detected with an average heritability of 80%. Expression of dsRNAs was confirmed in floral buds by RT-qPCR, and the T1 cotton plant generation was challenged with fertilized CBW females. After 30 days, data showed high mortality (around 70%) in oviposited yolks. In adult insects fed on transgenic lines, chitin synthase II and vitellogenin showed reduced expression in larvae and adults, respectively. Developmental delays and abnormalities were also observed in these individuals. Our data remark on the potential of transgenic cotton based on a viroid-structured dsRNA to control CBW.

Agriculture evolution, sustainability and trends, focusing on Brazilian agribusiness: a review

“…In view of these possibilities, Embrapa Genetic Resources and Biotechnology is developing several scientific research projects aimed at developing transgenic-based technologies capable of 10.3389/fsufs.2023.1296337 Frontiers in Sustainable Food Systems 05 frontiersin.org controlling the CBW. In brief, these new technologies for cotton to control CBW use the expression of Cry toxins or the combination of Cry toxins with RNAi, both of which have already shown promising results in transgenic cotton lines evaluated under greenhouse conditions (Ribeiro et al, 2017(Ribeiro et al, , 2019(Ribeiro et al, , 2022Vasquez et al, 2023).…”

“…New versions of previously consolidated Cry or VIP3A proteins, as well as new molecules, have recently been released to ensure greater control efficacy and reduce the rate of resistance development of these technologies (Supplementary File S1; ISAAA, 2023). Also, the search for active molecules that have an effect against other insect pest orders is also ongoing, such as molecules for transgenic control of CBW in cotton (Ribeiro et al, 2017(Ribeiro et al, , 2019(Ribeiro et al, , 2022Vasquez et al, 2023). The main crops 10.3389/fsufs.2023.1296337…”

“…Together, these technologies collectively have contributed to the emergence of a new generation of seed treatments. In addition to this universe of new-generation seeds, the use of nanotechnologies combined with RNAi technologies, through topical application spraying bioprotected RNAi molecules that are active only in the target organism, is revolutionizing a new area of agriculture worldwide (Ribeiro et al, 2022;Távora et al, 2022;Vasquez et al, 2023). However, regulatory issues for the commercial use of topical RNAi technologies still need to be addressed on a global level, especially in Brazil (Dietz-Pfeilstetter et al, 2021).…”

A Review on Digestive System of Rhynchophorus ferrugineus as Potential Target to Develop Control Strategies

“…The inhibition of chitin synthase in cotton boll weevils ( Anthonomus grandis ) using RNAi recombinant transgenic cotton resulted in malformed first and third instar larvae. The report suggested that the knockdown of chitin synthase led to the obstruction of nutrient uptake in the gut [ 87 ].…”

State-of-the-Art Molecular Plant Sciences in Brazil

“…Various methodologies can be employed to identify and understand plant cellular mechanisms in response to biotic stress, as well as to develop distinct molecular strategies to reduce negative effects. The studies by Pinheiro et al [4] and Ribeiro et al [5] showed different methods to evaluate plant response to pathogens and limit their deleterious effects. The sterility and resulting susceptibility of bananas (Musa spp.)…”