关键词: 意大利蜜蜂, 吡虫啉, 学习行为, 转录组, 差异表达基因

Abstract: 【Aim】 This study aims to analyze the effect of imidacloprid treatment on the olfactory learning behavior and the gene transcription in the brain of Apis mellifera ligustica so as to provide evidence for the negative effects of neonicotinoid insecticides on honeybees. 【Methods】 Under laboratory conditions, A. m. ligustica adult workers were fed with 50% sucrose solution containing 4 ng imidacloprid at one time, with those fed with 50% sucrose solution without imidacloprid as the control, and its effect on the olfactory learning behavior of A. m. ligustica adult workers was measured via proboscis extension response (PER) behavior test. Total RNA was extracted from the brain of A. m. ligustica workers tested above for RNA-Seq sequencing and bioinformatics analysis. To verify the RNA-Seq sequencing results, the expression levels of six selected differentially expressed genes (DEGs) in the brain of A. m. ligustica adult workers were detected by real-time fluorescent quantitative PCR. 【Results】 The olfactory learning ability of A. m. ligustica adult workers fed with 50% sucrose solution containing 4 ng imidacloprid was significantly decreased as compared to the control group (fed with 50% sucrose solution). RNA-Seq sequencing results showed that there were 123 DEGs [adjusted P-value (padj)<0.05］ between the treatment group and the control group, including 82 down-regulated DEGs and 41 up-regulated DEGs. GO enrichment analysis revealed that the down-regulated DEGs were mainly enriched in S-adenosylmethionine-dependent methyltransferase activity, acid phosphatase activity, oxidoreductase activity, and protein heterodimerization activity. The up-regulated DEGs were mainly enriched in functional items such as transmembrane receptor activity, molecular transducer activity, and neurological system processes. KEGG enrichment analysis showed that the down-regulated DEGs were mainly enriched in such organelles as ribosome and lysosome, metabolism pathways like carbon metabolism and tryptophan metabolism, and Toll and IMD signaling pathways, while the up-regulated DEGs were not enriched in KEGG pathways. Real-time fluorescent quantitative PCR results showed that the relative expression levels of the six DEGs tested showed the same trend with the RNA-Seq sequencing results of FPKM (fragments per kilobase million) value, verifying the reliability of the sequencing results. 【Conclusion】 Exposure of sublethal dose of imidacloprid significantly reduces the olfactory learning ability of A. m. ligustica adult workers, and also affects the expression of immune and detoxification related genes, enzyme activity, redox and other biological metabolic processes in the brain of A. m. ligustica. Short-term stress of sublethal dose of imidacloprid can stimulate the olfactory sensory process and nerve signal transduction process of A. m. ligustica.

Key words: Apis mellifera ligustica, imidacloprid, learning behavior, transcriptome, differentially expressed genes