Apolipoprotein D3 is required for long-chain cuticular hydrocarbon accumulation and water retention in Acyrthosiphon pisum.

BACKGROUND: The pea aphid, Acyrthosiphon pisum, is a damaging phloem-feeding pest of legume crops, and RNA interference (RNAi)-based targeting of stress-adaptation genes offers a promising route for target-specific pest management. Apolipoprotein D (ApoD) proteins are putative lipid carriers in insects, but the function of the chromosome-3 paralog ApApoD3 in aphid cuticular waterproofing remains unknown. We investigated whether ApApoD3 regulates cuticular hydrocarbon (CHC) accumulation and desiccation tolerance in A. pisum using RNAi, GC-MS-based CHC profiling, water-balance and survival assays, fecundity assessment, and RNA sequencing. RESULTS: Injection of dsApApoD3 significantly suppressed ApApoD3 expression, producing 47.47% knockdown at 48 h and maintaining significant silencing through 96 h. ApApoD3 depletion reduced total CHC abundance by 28.69% and significantly reduced long-chain n-alkanes, specifically n-C25-n-C30 and n-C32. This disruption of the cuticular lipid barrier was accompanied by a 19.44% reduction in body water content and an approximately 2.05-fold increase in water loss rate. Under severe desiccation, no ApApoD3-silenced aphids survived at 36 h, whereas 26.67% of controls remained alive. Separately, cumulative fecundity was significantly reduced at all observation times from 12 to 120 h after ApApoD3 silencing. Transcriptome analysis identified 187 differentially expressed genes and revealed enrichment of neuropeptide signaling, hormone activity, and G protein-coupled receptor binding, indicating a broad transcriptional response to ApApoD3 silencing. CONCLUSION: ApApoD3 is required for long-chain CHC homeostasis, water retention, desiccation tolerance, and reproductive fitness in A. pisum. These findings identify ApApoD3 as a mechanistically supported RNAi candidate for stress-sensitization-based aphid management. © 2026 Society of Chemical Industry.