Role of RNA interference (RNAi) as molecular biopesticide for control of the invasive pest gypsy moth (Lymantria dispar)

RNAi has gained popularity in several fields of research in silencing targeted genes by degradation of RNA into short RNAs.  Long double-stranded RNA (dsRNA) is diced into stretches of 21-nucleotide siRNA that in turn activate ribonucleases targeting homologous mRNA resulting in translational blockage.  The objective of this study was to use RNAi as a molecular tool in the control of the invasive pest Lymantria dispar, gypsy moth that has proved to be a serious pest to forests, primarily hardwood trees across much of North America causing substantial economic damage.  Lepidopteran insects have been notoriously refractory to RNAi, thus the challenge to this novel technique relies on the method of delivery of dsRNA.

Genes were selected based on specificity for RNAi from the transcriptome profiles of gypsy moth larval midgut and cloned into L4440 vector between two T7 promoters in inverted orientation.  dsRNA was synthesized by transforming the vector into a HT115(DE3) bacterial strain harboring IPTG-inducible expression of T7 polymerase but deficient for RNaseIII.  These bacterially expressed dsRNAs when fed to gypsy moth 3^rd instar larvae were found to significantly deplete the expression of selected genes.  Consistent with the depletion in expression we also observed approximately 60% drop in body mass especially in gene targets identified here as Genes B and C, which may be a result of decreased dietary intake.  We also observed that dsRNA synthesized in vitro when continuously ingested by the larvae depleted the target gene in a sequence specific manner.  Interestingly, a synergistic effect of reduced target gene expression was observed when dsRNA for Genes B and C were ingested as a combination by the larvae.  Here we have successfully demonstrated that ingestion of dsRNA can significantly deplete specific gene targets in gypsy moth larvae.  Consequently, RNAi may be considered as a molecular tool for development as molecular biopesticides in the control of insect pests, especially when other approaches may prove to be nonviable.