Scientia Agricultura Sinica ›› 2022, Vol. 55 ›› Issue (8): 1685-1694.doi: 10.3864/j.issn.0578-1752.2022.08.018


Effects of Sublethal Doses of Imidacloprid on the Expression of Neurometabolic Genes in Apis cerana cerana

QIU YiLei(),WU Fan,ZHANG Li,LI HongLiang()   

  1. College of Life Sciences, China Jiliang University/Zhejiang Provincial Key Laboratory of Biometrology and Inspection & Quarantine, Hangzhou 310018
  • Received:2021-11-02 Accepted:2021-11-15 Online:2022-04-16 Published:2022-05-11
  • Contact: HongLiang LI;


【Background】The target of neonicotinoid insecticides is the acetylcholine receptor in the nervous system of insects. Due to its good systemicity and low toxicity to humans and animals, it has been widely used in agricultural production. Thus there is still a low residue in plants, and this sublethal dose residue can still cause adverse effects on the behavior and nervous system of flower-visiting insects like bees.【Objective】The objective of this study is to clarify the effect of sublethal dose imidacloprid (a kind of neonicotinoid insecticides) on the nervous physiological and metabolism system of Apis cerana cerana.【Method】In this study, the worker bees were treated with two sublethal concentration gradient doses of 5 and 10 μg·L-1 imidacloprid for 10 days (three biological replicates). After total RNA was extracted, the resulting library was analyzed by RNA-seq. Throughput sequencing, and the bioinformatics technology was used to assemble and annotate the sequence de novo, and the differentially expressed genes (DEGs) after sublethal doses imidacloprid treatment were analyzed by clustering and enrichment. Finally, real-time fluorescence quantitative PCR (RT-qPCR) technology was used to verify the DEGs related to the nervous and metabolic systems.【Result】A total of 9 sequencing libraries were obtained, the ratio of effective sequencing data exceeded 94.45%. From the obtained 37 364 unigenes, 571 DEGs were identified. The enrichment analysis of GO and KEGG found that the DEGs were mainly related to multiple pathways such as protein translation, redox, oxidative phosphorylation, and ribosome, indicating that sublethal doses of imidacloprid had an impact on multiple physiological processes and metabolic pathways of A. c. cerana. Nine DEGs (e.g. neuropeptide F (NPF), neuropeptide SIFamide receptor (SIFaR), phosphoinositide 3-dependent kinases (PDK1), A-kinase anchoring protein 1 (AKAP1), carbonic anhydrase 3 (CA3), superoxide dismutase 2 (SOD2), NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 10 (ND42), cuticle protein 12 (CP12), and odorant-binding protein (OBP17)) related to nerve signal transmission and metabolic function were selected for RT-qPCR verification. Their expression patterns were completely consistent with the transcriptome results.【Conclusion】The sublethal dose of imidacloprid can affect lots of aspects of A. c. cerana such as nerve signal transduction, cellular respiration, immune response, maintenance of homeostasis, and olfactory perception.

Key words: Apis cerana cerana, sublethal dose, imidacloprid, RNA-seq, RT-qPCR

Table 1

Real-time fluorescence quantitative PCR (RT-qPCR) primers"

Coding protein
Primer name
Primer sequence (5′→3′)
NPF 神经肽F
Neuropeptide F
SIFaR 神经肽SIFamide受体
Neuropeptide SIFamide receptor
PDK1 3-磷酸肌醇依赖性蛋白激酶
Phosphoinositide 3-dependent kinases
AKAP1 激酶(PRKA)锚蛋白1
A-kinase anchoring protein 1
CA3 碳酸酐酶
Carbonic anhydrase 3
SOD2 超氧化物歧化酶
Superoxide dismutase 2
ND42 NADH脱氢酶亚基
NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 10
CP12 昆虫表皮蛋白
Cuticle protein 12
OBP17 气味结合蛋白17
Odorant-binding protein 17
β-actin 肌动蛋白(内参)
Actin (reference)

Fig. 1

Clustering heat map of DEGs The abscissa represents different samples, and the ordinate represents different DEGs. The darker the color, the greater the difference in the gene among different samples"

Fig. 2

Histogram of DEGs GO enrichment"

Fig. 3

Scatter plot of DEGs KEGG enrichment Rich factor represents the number of differential genes located in the KEGG/the total number of genes located in the KEGG. The greater the Rich factor value, the greater the degree of KEGG enrichment. The closer the color of the dot is to red, the greater the degree of renaturation, and the size of the dot represents the number of genes enriched in the pathway"

Fig. 4

Transcriptome sequencing and RT-qPCR verification results of 9 DEGs The column height is the mean±SEM. The LSD method was used for one-way analysis of variance. * indicates a significant difference, P<0.05"

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