Double-stranded RNA targeting white mold Sclerotinia sclerotiorum argonaute 2 for disease control via spray-induced gene silencing

Abstract Sclerotinia sclerotiorum , the causal agent of white mold infection, is a cosmopolitan fungal pathogen that causes major yield losses in many economically important crops. Spray induced gene silencing has recently been shown to be a promising alternative method for controlling plant diseases. Based on our prior research, we focus on developing spray-induced gene silencing approach to control white mold by silencing S. sclerotiorum argonaute 2 (SsAgo2), a crucial part of the fungal small RNA pathway. We compared the lesion size as a result of targeting each ~500-bp segments of SsAgo2 from 5’ to 3’ and found that targeting the PIWI/ RNaseH domain of SsAgo2 is most effective. External application of double-stranded RNA (dsRNA) suppressed white mold infection using either in vitro or in vivo transcripts was determined at the rate of 800 ng/0.2cm 2 area with a downregulation of SsAgo2 from infected leaf tissue confirmed by RT-qPCR. Furthermore, magnesium/iron-layered double hydroxides (MgFe-LDH) nanosheets loaded with in vitro and in vivo transcribed dsRNA segments significantly reduced the rate of S. sclerotiorum lesion expansion. In vivo produced dsRNA targeting the PIWI/RNaseH domain of the SsAgo2 transcript showed increased efficacy in reducing the white mold symptoms of S. sclerotiorum when combined with LDH nanosheets. This approach is promising to produce a large scale of dsRNA that can be deployed as an environmentally friendly fungicide to manage white mold infections in the field.