Functional validation of pathogenicity genes in rice sheath blight pathogen Rhizoctonia solani by a novel host-induced gene silencing system

Rice sheath blight, caused by the soilborne fungus Rhizoctonia solani, causes severe yield losses worldwide. Elucidation of the pathogenic mechanism of R. solani is highly desired. However, the lack of a stable genetic transformation system has made it challenging to examine genes' functions in this fungus. Here, we present functional validation of pathogenicity genes in the rice sheath blight pathogen R. solani by a newly established tobacco rattle virus (TRV)–host-induced gene silencing (HIGS) system using the virulent R. solani AG-1 IA strain GD-118. RNA interference constructs of 33 candidate pathogenicity genes were infiltrated into Nicotiana benthamiana leaves with the TRV-HIGS system. Of these constructs, 29 resulted in a significant reduction in necrosis caused by GD-118 infection. For further validation of one of the positive genes, trehalose-6-phosphate phosphatase (Rstps2), stable rice transformants harbouring the double-stranded RNA (dsRNA) construct for Rstps2 were created. The transformants exhibited reduced gene expression of Rstps2, virulence, and trehalose accumulation in GD-118. We showed that the dsRNA for Rstps2 was taken up by GD-118 mycelia and sclerotial differentiation of GD-118 was inhibited. These findings offer gene identification opportunities for the rice sheath blight pathogen and a theoretical basis for controlling this disease by spray-induced gene silencing.