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The fall armyworm, Spodoptera frugiperda, a lepidopteran pest from the family Noctuidae, has become a major invasive pest since 2016. Using RNAi methods to control S. frugiperda is currently under investigation. This study is the first to target the V-ATPaseD gene of S. frugiperda using RNAi. Injection of dsRNA-V-ATPaseD into the hemolymph of 4th-instar larvae significantly suppressed gene expression at 24 and 48 h post-injection. Treated larvae showed delayed development and reduced pupation after 7 days. Subsequently, V-ATPaseD silencing was achieved through topical or oral administration of chitosan/dsRNA-V-ATPaseD nanoparticles. Larvae fed these nanoparticles exhibited significant reductions in V-ATPaseD mRNA at 72 h, persisting until 96 h before normalizing. Additionally, the treated larvae displayed disrupted molting and impaired pupation. Furthermore, larvae fed chitosan/dsRNA-V-ATPaseD were more susceptible to emamectin benzoate–lufenuron at LC30 concentrations, resulting in 68% mortality—27% higher than the pesticide alone—72 h post-exposure. Combining chitosan/dsRNA-V-ATPaseD nanoparticles with emamectin benzoate–lufenuron significantly enhanced pest control efficacy, providing new insights into pesticide reduction and sustainable pest control methods for this invasive species.

The responsiveness of insects to oral delivery of insecticidal dsRNA has been shown to be dependent on dsRNA length and sequence match. Previous work with the western corn rootworm (WCR, Diabrotica virgifera virgifera ; Coleoptera: Chrysomelidae) demonstrated that at least one ≥21 nt match must be present in the DvSnf7 dsRNA of approximately ≥60 base-pairs (bp) for activity. Further data is needed on the activity of <21 nt matches along with characterization of relationship between activity and the number of ≥21 nt matches. To characterize the sequence–activity relationship for insecticidal dsRNA further, the activity of orthologous Snf7 dsRNAs with 19, 20, and 21 nt contiguous matches against WCR was compared. Neither 19 nor 20 nt sequence matches were active, supporting that a ≥21 nt sequence match is required for activity. The relationship between the number of 21 nt matches with activity of Snf7 dsRNA orthologs from several Chrysomelid species was characterized using WCR and Colorado potato beetle (CPB, Leptinotarsa decemlineata; Coleoptera Chrysomelidae). For WCR, there was a strong relationship between an increasing number of 21 nt matches and increased activity ( i.e. , lower LC50 values). A similar relationship was observed for CPB with an exception for a single ortholog, which may be related to the exceptionally high rate of polymorphisms in CPB. Overall, these results demonstrate a general relationship between the number of 21 nt matches and activity, and this relationship could be used to inform a testing and assessment plan for an ecological risk assessment for an insecticidal dsRNA.

RNA interference insecticides have received increasing attention in recent years due to their classification as a reduced-risk biopesticide and their proposed faster path to registration compared with conventional synthetic insecticides. The goal of this study was to synthesize and compare efficacy of 62 double-stranded RNAs (dsRNAs) from 31 target genes against the pest termite species, Reticulitermes flavipes (Kollar) (Isoptera: Rhinotermitidae). Fifty-seven dsRNAs of ~125 base pairs each were successfully synthesized. First-tier screens using a combination immersion/feeding assay revealed 10 top candidates and also that dsRNAs coming from synthesis reactions with 80–90× yields were the most effective. Follow-up studies using uptake enhancers in combination with top candidate dsRNAs were unsuccessful. Subsequent concentration range feeding assays on the top candidates revealed two lead termiticidal dsRNAs (3 Hexamerin-2 and 3 Glycosyl Hydrolase Family [GHF] 9-2 cellulase) and another that enhanced feeding (5 GHF9-2 cellulase).Testing a matrix of combinations of these three dsRNAs revealed ultimately that the most consistently effective dsRNA combination was the 3 Hexamerin-2 + 3 GHF9-2 cellulase dsRNA combination. These results provide new information on candidate termiticidal dsRNAs and some apparent factors that have a bearing on their efficacy. Despite these successes, further research and development will be necessary to move dsRNA termiticides from pest management theory to real-world application.