The Colorado potato beetle (Leptinotarsa decemlineata Say, CPB) is an important agricultural pest. It is well known for its ability to rapidly evolve resistance to insecticides and is predicted to expand to the north of Europe due to the prospects of climate change. In the last decade, double-stranded RNAs (dsRNAs) were shown to have the potential to become environmentally acceptable insecticidal agents. These molecules activate the RNA interference (RNAi) mechanism that mediates a sequence-specific suppression of gene expression, also more generally called gene silencing.

In CPB, RNAi-mediated gene silencing can be effectively triggered and spread systemically by ingestion of dsRNA with a sequence complementary to the target’s mRNA. If the targeted gene silencing can disrupt an essential life-sustaining process in the beetle, the dsRNAs can act as an insecticide. The advantages of RNAi insecticides include high specificity for target species, low probability of resistance development and short environmental degradation period into nontoxic compounds. However, crucial for efficiency as well as the environmental safety of dsRNA insecticides is the selection of target genes. To select appropriate target genes we will examine the results of RNAi screening experiments in model organisms (e.g. Drosophila melanogaster, Tribolium castaneum) as well as employ our previous data and literature knowledge on CPB’s gene expression and physiology.

CPB feeding trial (Photo: Marko Petek)

This project’s main objective is to identify and validate gene targets for CPB specific RNAi insecticides. To achieve that, the methodology for identification and validation of RNAi insecticide efficiency will be established at NIB. This will include a candidate gene selection procedure, establishment of dsRNA production methodology and the evaluation of dsRNAs’ insecticidal potential in insect feeding trials. Additionally, a Next-Generation Sequencing (NGS) RNA-Seq analysis will be performed to explore the global changes induced by the RNAi insecticide on the CPB gut transcriptome. To explore how an RNAi insecticide changes the bacterial community structure in CPB guts, larval gut contents will be examined by NGS to obtain bacterial metagenomic/metatranscriptomic profiles.

Available statistical tools will be used for evaluation of RNAi insecticide efficiency and already developed NGS pipelines for the analysis of RNA-Seq data will be adopted, whereas for metagenomics data, a dedicated analysis pipeline will be established at NIB.

The project aims at:

• exploring the potential of selected candidate genes as new targets for RNAi insecticides,
• validating computationally predicted gene functions in an agricultural pest,
• following the expression of selected target genes in different CPB tissues and life stages,
• studying the global transcriptional patterns by RNA-Seq in CPB guts after RNAi insecticidal treatment,
• investigating the effect of an RNAi insecticide on CPB’s gut metagenome.

The prospect of this omics-supported validation of RNAi insecticide target genes is to provide invaluable insights into beetle’s response to dsRNA and to contribute to the knowledge base on mechanisms of action of RNAi-based insecticides.

We are open for collaborations, if you’re interested please contact dr Marko Petek.