The tomato brown rugose fruit virus (ToBRFV) is an emerging pathogen that severely damages the global tomato industry. Topical application of double-stranded RNA (dsRNA) has shown promise as an effective tool to control many pathogens, including viruses. However, it was not yet demonstrated for the control of ToBRFV. Here, we utilized in vivo synthesis of dsRNA molecules for the protection of tomato plants against ToBRFV. ToBRFV-specific long dsRNA molecules were produced by incorporating parts of ToBRFV genome into the genome of bacteriophage phi6 enabling the amplification of the chimeric dsRNA in Pseudomonas syringae bacteria. Application of purified, high quality (hq)-dsRNA onto tomato plants resulted in efficient ToBRFV protection by reducing both viral RNA levels and disease symptoms. Functional analysis of the hq-dsRNA response against the virus revealed its independence of RNA-dependent RNA polymerase 6 (RDR6) and suppressor of gene silencing 3 (SGS3). In addition, non-infected plants showed a mild activation of innate immune responses upon hq-dsRNA treatment, including callose accumulation at plasmodesmata. Together, this work provides evidence for hq-dsRNA as an effective tool for controlling ToBRFV in tomato plants and the potential of in vivo produced dsRNA in the battle against emerging crop pathogens.
Keywords: RNA silencing; double-stranded RNA; plant immunity; plant protection; plasmodesmata; synthetic biology; tobamovirus; tomato.
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