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Title: Double strand RNA-mediated RNA interference through feeding in larval gypsy moth, Lymantria dispar (Lepidoptera: Erebidae)

item Ghosh, Saikat Kumar
item Gundersen-Rindal, Dawn

Submitted to: European Journal of Entomology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/10/2017
Publication Date: 4/5/2017
Citation: Ghosh, S.B., Gundersen, D.E. 2017. Double strand RNA-mediated RNA interference through feeding in larval gypsy moth, Lymantria dispar (Lepidoptera: Erebidae). European Journal of Entomology. 114:170-178.

Interpretive Summary: The gypsy moth is a serious insect pest of the North American forests where larvae feed on over 300 tree and shrub species, especially hardwood and shade trees. Measures to control the pest and reduce its spread have been implemented in the United States, including augmentive release of natural enemies, use of chemical pesticides, mating disruption using a chemical pheromone, and use of microbial biopesticides or Bacillus thuringiensis. In order to slow the spread of this pest, we have successfully tested a novel biomolecular technique known as RNA interference. This technique uses extracellular RNA to target the in vivo RNA for degradation using the cells internal machinery. Degradation of RNA in turn blocks protein translation eventually adversely affecting the pest. This information will be of benefit to scientists and biopesticide companies that are interested in developing new biomolecule-based strategies towards control of gypsy moth.

Technical Abstract: RNA interference (RNAi) has gained popularity in several fields of research, silencing targeted genes by degradation of RNA. The objective of this study was to develop RNAi for use as a molecular tool in the control of the invasive pest Lymantria dispar (Lepidoptera: Erebidae), gypsy moth, which has been a serious forest pest, primarily of hardwood trees across North Eastern United States and Canada where it has caused substantial economic damage. Lepidopteran insects have been notoriously refractory to RNAi, thus the challenge to this novel technique relied on the method of delivery of dsRNA. Genes were selected based on specificity for RNAi from the transcriptome profiles of gypsy moth larval midgut (Sparks et al., 2011; 2013). An efficient method using a plasmid designed to express double stranded RNA (dsRNA) for these genes was induced in a specific bacterial strain. These bacterially expressed dsRNAs when fed to gypsy moth larvae were found to significantly deplete the expression of the targeted genes. Consistent with the depletion in expression we also observed approximately 60% drop in body mass especially in gene targets only known as locus 365 and locus 28365, which may be a result of decreased dietary intake. In testing the feasibility of per os delivered RNAi for imapct on gypsy moth larvae in vivo, we observed that in vitro synthesized dsRNA when ingested over the course of time by larvae, either individually or in combination, depleted the target gene(s) in a sequence specific manner. Interestingly, females that developed to adults after ingestion as larvae of locus 365-specific RNAi displayed a two-fold decrease in their egg mass compared to controls. This work has 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 to be used in the control of gypsy moth and other insect pests, especially when other approaches may prove to be nonviable.