In vertebrates and plants, dsRNA plays crucial roles as PAMP and as a mediator of RNAi. How higher fungi respond to dsRNA is not known. We demonstrate that Magnaporthe oryzae (Mo), a globally significant crop pathogen, internalizes dsRNA across a broad size range of 21 to about 3000 bp. Incubation of fungal conidia with 10 ng/µL dsRNA, regardless of size or sequence, induced aberrant germ tube elongation, revealing a strong sequence-unspecific effect of dsRNA in this fungus. Accordingly, the synthetic dsRNA analogue poly(I:C) and dsRNA of various sizes and sequences elicited canonical fungal stress pathways, including nuclear accumulation of the stress marker mitogen-activated protein kinase Hog1p and production of ROS. Leaf application of dsRNA to the cereal model species Brachypodium distachyon suppressed the progression of leaf blast disease. Notably, the sequence-unspecific effect of dsRNA depends on higher doses, while pure sequence-specific effects were observed at low concentrations of dsRNA ( < 0.03 ng/µL). The protective effects of dsRNA were further enhanced by maintaining a gap of at least seven days between dsRNA application and inoculation, and by stabilising the dsRNA in alginate-chitosan nanoparticles. Overall, our study opens up additional possibilities for the development and use of dsRNA pesticides in agriculture.

The hemibiotrophic fungus Magnaporthe oryzae (Mo) is the causative agent of rice blast and can infect aerial and root tissues of a variety of Poaceae, including the model Brachypodium distachyon (Bd). To gain insight in gene regulation processes occurring at early disease stages, we comparatively analyzed fungal and plant mRNA and sRNA expression in leaves and roots. A total of 310 Mo genes were detected consistently and differentially expressed in both leaves and roots. Contrary to Mo, only minor overlaps were observed in plant differentially expressed genes (DEGs), with 233 Bd-DEGs in infected leaves at 2 days post inoculation (DPI), compared to 4978 at 4 DPI, and 138 in infected roots. sRNA sequencing revealed a broad spectrum of Mo-sRNAs that accumulated in infected tissues, including candidates predicted to target Bd mRNAs. Conversely, we identified a subset of potential Bd-sRNAs directed against fungal cell wall components, virulence genes and transcription factors. We also show a requirement of operable RNAi genes from the DICER-like (DCL) and ARGONAUTE (AGO) families for fungal virulence. Overall, our work elucidates the extensive reprogramming of transcriptomes and sRNAs in both plant host (Bd) and fungal pathogen (Mo), further corroborating the critical role played by sRNA species in the establishment of the interaction and its outcome.