Identification of highly effective target genes for RNAi-mediated control of emerald ash borer, Agrilus planipennis

Abstract

Recent study has shown that RNA interference (RNAi) is efficient in emerald ash borer (EAB), Agrilus planipennis, and that ingestion of double-stranded RNA (dsRNA) targeting specific genes causes gene silencing and mortality in neonates. Here, we report on the identification of highly effective target genes for RNAi-mediated control of EAB. We screened 13 candidate genes in neonate larvae and selected the most effective target genes for further investigation, including their effect on EAB adults and on a non-target organism, Tribolium castaneum. The two most efficient target genes selected, hsp (heat shock 70-kDa protein cognate 3) and shi (shibire), caused up to 90% mortality of larvae and adults. In EAB eggs, larvae, and adults, the hsp is expressed at higher levels when compared to that of shi. Ingestion of dsHSP and dsSHI caused mortality in both neonate larvae and adults. Administration of a mixture of both dsRNAs worked better than either dsRNA by itself. In contrast, injection of EAB.dsHSP and EAB.dsSHI did not cause mortality in T. castaneum. Thus, the two genes identified cause high mortality in the EAB with no apparent phenotype effects in a non-target organism, the red flour beetle, and could be used in RNAi-mediated control of this invasive pest.

Prospects, challenges and current status of RNAi through insect feeding

“…In Agrilus planipennis larvae and adults, oral treatment of high concentrations of dsRNA (10 μg μL −1 ) targeting two vital genes, heat shock protein‐70 and shibire , showed ∼90% mortality. However, co‐treatment of both dsRNA at lower concentrations (1 μg μL −1 ) showed a synergistic effect with similar mortality rates (∼90%) . Similar effects were observed in T. castaneum fed with both Bip and Armet dsRNAs complexed with the BAPCs nanoparticle.…”

“…However, co-treatment of both dsRNA at lower concentrations (1 μg μL −1 ) showed a synergistic effect with similar mortality rates (∼90%). 114 Similar effects were observed in T. castaneum fed with both Bip and Armet dsRNAs complexed with the BAPCs nanoparticle. The individual treatment of both dsR-NAs complexed with nanoparticles showed 25% mortality, but combined treatment resulted in 60% mortality.…”

Bacteria-Mediated RNA Interference for Management of Plagiodera versicolora (Coleoptera: Chrysomelidae)

“…Interestingly, it was demonstrated that oral treatment of a combination of dsSHI and dsHSP70 at lower concentration by neonates and adults of Agrilus planipennis efficiently elicited mortality. Exposing adults to 1 µg/µL of dsSHI plus dsHSP70 caused 90% mortality after two weeks, whereas, 10 µg/µL of single dsRNA treatment exhibited lower mortality (up to 40%) [34]. Therefore, it may be worthwhile to test whether there is a combined effect by silencing both SHI and HSP70 in P. versicolora.…”

Oral Ingestion of Bacterially Expressed dsRNA Can Silence Genes and Cause Mortality in a Highly Invasive, Tree-Killing Pest, the Emerald Ash Borer

“…Newly hatched neonates were used in all bioassays. To determine the biological activity of the recombinant bacteria expressing dsRNA, neonate EAB larvae were fed using a modified droplet feeding bioassay [ 5 ], where 1 ml of bacterial culture was centrifuged at 3000 rpm for 15 min and the pellet was dissolved in 100 μL of 1% sucrose solution with green tracking dye (Kroger, Co., USA). For each assay, 3μL of bacterial suspension were fed to individual neonate larvae using the droplet assay.…”

“…Coleopteran insects are known to exhibit robust RNAi responses [ 2 , 4 , 5 ]. RNAi efficiency varies between insect species, insect life stages, target genes, and modes of dsRNA delivery [ 6 ]; dsRNA can be delivered in several ways, including by injection, orally, and through absorption [ 7 ].…”