Spray-induced gene silencing (SIGS) can inhibit plant diseases by topical application of double- (dsRNA) or single-stranded (sRNA) RNA molecules onto plants to silence virulence-related pathogen genes. However, the on-field application of SIGS is limited by the instability of naked RNA and low RNA uptake by pathogens. Nanoparticles have been used as RNA carriers to enhance RNA silencing. Rice sheath blight caused by Rhizoctonia solani (R. solani) is one of the most devastating fungal diseases in rice (Oryza sativa L.). In this study, we aimed to explore the protective effects of nanoparticle-delivered dsRNA against rice sheath blight. The key pathogenic genes, RsAGO1 and RsAGO2, of R. solani were screened as targets for dsRNA. Chitosan (CS), polyethyleneimine (PEI), protamine, carbon quantum dot (CQD), polyamidoamine (PAMAM), and chitosan/SPc complex (CSC) were selected as dsRNA carriers. All the evaluated nanoparticles could assemble with dsRNA to form nanoparticle-dsRNA complexes, and CQD and CSC showed improved dsRNA load capacity. Particularly, CSC could enhance the stability of dsRNA and cause a 7% reduction in fluorescence intensity after nuclease treatment. CSC and CS effectively enhanced the efficiency of dsRNA uptake by pathogens. Furthermore, CSC could reduce pathogen infection and prolong the protection time of dsRNA by up to 20 days. Overall, this study provides a novel and efficacious SIGS-based strategy for producing RNA-based fungicides.

Rhizoctonia solani is one of the important pathogenic fungi causing several serious crop diseases, such as maize and rice sheath blight. Current methods used to control the disease mainly depend on spraying fungicides because there is no immunity or high resistance available in crops. Spraying double-strand RNA (dsRNA) for induced-gene silencing (SIGS) is a new potentially sustainable and environmentally friendly tool to control plant diseases. Here, we found that fluorescein-labelled EGFP-dsRNA could be absorbed by R. solani in co-incubation. Furthermore, three dsRNAs, each targeting one of pathogenicity-related genes, RsPG1, RsCATA, and RsCRZ1, significantly downregulated the transcript levels of the target genes after co-incubation, leading to a significant reduction in the pathogenicity of the fungus. Only the spray of RsCRZ1 dsRNA, but not RsPG1 or RsCATA dsRNA, affected fungal sclerotium formation. dsRNA stability on leaf surfaces and its efficiency in entering leaf cells were significantly improved when dsRNAs were loaded on layered double hydroxide (LDH) nanosheets. Notably, the RsCRZ1-dsRNA-LDH approach showed stronger and more lasting effects than using RsCRZ1-dsRNA alone in controlling pathogen development. Together, this study provides a new potential method to control crop diseases caused by R. solani.