Daniele H Pinheiro1,2, Caitlin E Taylor1, Ke Wu1, Blair D Siegfried1. 1. Entomology and Nematology Department, University of Florida, Gainesville, FL, USA. 2. Embrapa Genetic Resources and Biotechnology, Brasília, DF, Brazil.
Abstract
BACKGROUND: RNA interference (RNAi) is widely used in entomological research for functional analysis of genes and is being considered as a new tool for insect pest management. Sri Lanka weevil (SLW) is a highly polyphagous pest of agronomically important plants, but currently only a few control methods are available for this insect. RESULTS: In the present study, we evaluated the stability of candidate reference genes β-ACT, α-TUB, EF1-α, RPL12 and GAPDH, and identified EF1-α as the most reliable for gene expression normalization. Four target genes involved in different cellular processes, including Prosα2, RPS13, Snf7 and V-ATPase A were selected to evaluate whether RNAi response in SLW adults can be triggered by microinjection and oral feeding of their double-stranded RNAs (dsRNAs). Three days after injection of the dsRNAs for the target genes, their transcript levels were significantly reduced (up to 91.4%) when compared to the control. Additionally, weevils fed with the target dsRNAs showed significant decreases in gene transcript levels and significant mortality was observed in insects treated with Prosα2 and Snf7 dsRNAs (78.6 to 92.7%). CONCLUSION: Our data demonstrate that microinjection and feeding of dsRNA produce a strong RNAi response in SLW, indicating that RNAi-based strategies could be explored to develop a selective and environmentally safe control method against SLW.
BACKGROUND: RNA interference (RNAi) is widely used in entomological research for functional analysis of genes and is being considered as a new tool for insect pest management. Sri Lanka weevil (SLW) is a highly polyphagous pest of agronomically important plants, but currently only a few control methods are available for this insect. RESULTS: In the present study, we evaluated the stability of candidate reference genes β-ACT, α-TUB, EF1-α, RPL12 and GAPDH, and identified EF1-α as the most reliable for gene expression normalization. Four target genes involved in different cellular processes, including Prosα2, RPS13, Snf7 and V-ATPase A were selected to evaluate whether RNAi response in SLW adults can be triggered by microinjection and oral feeding of their double-stranded RNAs (dsRNAs). Three days after injection of the dsRNAs for the target genes, their transcript levels were significantly reduced (up to 91.4%) when compared to the control. Additionally, weevils fed with the target dsRNAs showed significant decreases in gene transcript levels and significant mortality was observed in insects treated with Prosα2 and Snf7 dsRNAs (78.6 to 92.7%). CONCLUSION: Our data demonstrate that microinjection and feeding of dsRNA produce a strong RNAi response in SLW, indicating that RNAi-based strategies could be explored to develop a selective and environmentally safe control method against SLW.
Authors: Jonathan Willow; Liina Soonvald; Silva Sulg; Riina Kaasik; Ana Isabel Silva; Clauvis Nji Tizi Taning; Olivier Christiaens; Guy Smagghe; Eve Veromann Journal: Commun Biol Date: 2021-04-06