Sublethal imidacloprid exposure disrupts olfactory-mediated host location in Hippodamia variegata through transcriptional and functional impairment of olfactory genes

Hippodamia variegata (Coleoptera: Coccinellidae) is an important predator of the cotton aphid, Aphis gossypii, and it plays an important role in its biological control. However, widespread use of neonicotinoid insecticides such as imidacloprid may exert sublethal effects on non-target natural enemies. In this study, we evaluated the impact of sublethal imidacloprid exposure on the olfactory behavior of H. variegata and explored the underlying molecular mechanisms. Adult beetles were fed cotton aphids treated with imidacloprid at 3.93 mg a.i./L (LC₂₀) and 13.62 mg a.i./L (LC₅₀), and their olfactory responses to aphid-infested versus uninfested cotton leaves were assessed using a Y-tube olfactometer. In parallel, transcriptomic profiling was conducted on the antennae and brain tissues of adult male and female beetles from both control and treatment groups, followed by GO/KEGG enrichment analyses and qRT-PCR validation of candidate genes. Finally, RNA interference (RNAi) was used to functionally verify the roles of selected odorant-binding protein (OBP) genes in the sublethal effects observed. Y-tube assays showed that control adults exhibited significant attraction to aphid-infested cotton plants, whereas beetles of both sexes exposed to LC₂₀ and LC₅₀ imidacloprid lost this olfactory preference. Transcriptomic analyses revealed extensive differential gene expression in the antennae and brains of beetles treated with LC₂₀ and LC₅₀ compared with the control group, with differentially expressed genes significantly enriched in pathways associated with metabolic processes, ribosomes, RNA transport, and neural signaling. Notably, several olfaction and neural function-related genes, including Obp2, Obp7, Obp8, Obp10, Ten-m, and Nlg4, were significantly downregulated after imidacloprid exposure, and these patterns were confirmed by qRT-PCR. RNAi-mediated silencing of Obp2, Obp7, Obp8, and Obp10, which significantly reduced transcript levels and eliminated the olfactory preference for aphid-infested cotton plants in behavioral assays, supporting their essential roles in prey-related odor perception. Overall, our findings demonstrate that sublethal imidacloprid exposure impaired olfactory function in H. variegata and was associated with transcriptional disruptions in olfactory and neural signaling pathways, which may compromise its efficiency as a biological control agent. These results highlight the need to consider sublethal ecological risks of insecticide use and to optimize pesticide application strategies to better align with integrated pest management programs.