Orally Delivered dsRNA-Derived siRNAs Reach the Central Nervous System in Leptinotarsa decemlineata

RNA interference (RNAi) has emerged as an eco-friendly approach to pest management and relies on the processing of exogenous double-stranded RNA (dsRNA). RNAi-based pest management is highly effective in the Colorado potato beetle (Leptinotarsa decemlineata); however, the tissue-specific distribution and processing of exogenous dsRNA following oral uptake remain incompletely understood. In this study, we investigated whether ingested dsRNA reaches the central nervous system (CNS) and is processed into active small interfering RNAs (siRNAs). Adult beetles were fed dsmGFP-coated leaf disks, and RISC-bound small RNAs were isolated from midgut, CNS, and remaining body tissues using a RISC-enrichment approach. Small RNA sequencing revealed abundant 21-nucleotide antisense guide-strand siRNAs in all analysed tissues, with relative proportions following the order midgut > CNS > remaining tissues. Notably, antisense siRNAs of consistent size were detected in CNS samples, indicating that exogenous dsRNA or its processed products can access neural tissue and enter the RNAi silencing machinery. These findings provide strong biochemical evidence that orally taken-up dsRNA is processed into AGO-loaded siRNAs in the L. decemlineata CNS. Together, our results offer a tissue-resolved view of functional RNAi activity in this species and contribute to a mechanistic understanding of systemic dsRNA transport in coleopteran pests. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=99 SRC="FIGDIR/small/711085v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1796858org.highwire.dtl.DTLVardef@1b183ceorg.highwire.dtl.DTLVardef@1447e91org.highwire.dtl.DTLVardef@1d17def_HPS_FORMAT_FIGEXP M_FIG C_FIG