New insights into transmission pathways and possible off-target effects of insecticidal dsRNA released by treated...

RNAi has shown great potential in controlling pests and pathogens, and dsRNA-based pesticides have been used in different ways. Due to off-target effects, the transmission pathways and possible impacts of dsRNA on non-target organisms after release should be researched. Here, we tested pathways of dsRNA transmission through the rice-hopper-spider food chain and their efficiency for triggering RNAi. The results revealed five new pathways by which plants transfer dsRNA into the environment through the food chain. We found that ingestion of the tissues or guttation droplets of treated plant could trigger both targeted and off-target RNAi both in consumers and predators. Ingestion of consumer hoppers could also result in localized RNAi in the midguts of the predator spiders. Trace amounts of dsRNA were detected in plant root excretions and in hopper honeydew. Cutting the root tips dramatically increased the levels of dsRNA in root excretions. Host shifting experiments proved that hoppers could transfer a trace amount of dsRNA via vomit. With specially designed dsRNAs, we showed that dsRNA sharing matching sequences of 29 bp or 32 bp in length with non-target genes could trigger off-target RNAi, but that dsRNA sharing 13 bp matching sequences could not. We conclude that field-released pesticidal dsRNA could be transmitted via the hydrophilic transport system in plants, and that this may pose a safety risk to non-target animal consumers that are closely related to target pests. Rational use of pesticidal dsRNAs should involve careful consideration of dsRNA design to manage the biosafety risk.