Plant Virus Expression of Interfering Sequences Gives RNA Interference (RNAi) Effects in Hemipteran Phloem Feeders

RNA interference (RNAi) is a natural gene regulation and antiviral defense system of eukaryotes. Recently, RNAi technology has been proposed as a potentially useful tool for plant-associated insect control. Most insect RNAi studies reported to date have used in vitro-transcribed dsRNAs as effectors to induce RNAi activity in recipient insects. Previously, we demonstrated RNAi effects in the glassy winged sharpshooter (Homalodisca vitripennis）and the potato psyllid (Bactericera. cockerelli) by both injection and oral feeding of dsRNAs. Here, we investigated in planta expression of interfering sequences and resulting RNAi effects on the hemipteran phloem feeders, B. cockerelli and Planococcus citri by using recombinant plant viruses containing insect gene fragments. Our hypothesis is that the virus-infected plants will generate siRNAs in defense towards the infecting virus plus the recombinant cDNA sequence it carries. The recombinant virus causes a systemic infection in the plant, interacting with many cell types including those in the phloem, and the phloem is an effective conduit for transport of small and even large RNAs, and thus will provide a continuous source of silencing RNAs for the phloem-feeding insects used here. We show that recombinant Tobacco mosaic virus (TMV) containing insect sequence inserts can efficiently infect and generate interfering small RNAs in a variety of plant species including tomato (Solanum lycopersicum), tomatillo (Physalis philadelphica), tobacco (Nicotiana tabacum) and Nicotiana benthemiana plants, and more importantly delivery of interfering sequences via recombinant TMV induces gene-specific RNAi effects in the insects feeding on these plants, even yielding decreased progeny number as compared to plants infected by recombinant TMV containing GFP. These results showed that RNAi effects can be achieved in plants against hemipteran phloem feeders, and the TMV-plant system can provide a faster and more convenient method for screening of suitable RNAi target sequences in planta.