昆虫学报 ›› 2021, Vol. 64 ›› Issue (8): 943-955.doi: 10.16380/j.kcxb.2021.08.006

• 研究论文 • 上一篇    下一篇

西花蓟马GABA受体基因鉴定及FoRDL 在多杀霉素抗性中的作用

王京1, 何秉青2, 华登科1, 张坤1, 袁江江1, 郑晓斌1, 徐宝云1, 张友军1, 吴青君1,*    

  1.  (1. 中国农业科学院蔬菜花卉研究所, 北京 100081; 2. 北京市昌平区农业技术推广站, 北京 102200)
  • 出版日期:2021-08-20 发布日期:2021-08-12

Identification of GABA receptor genes and the role of FoRDL in spinosad resistance in Frankliniella occidentalis (Thysanoptera: Thripidae)

WANG Jing1, HE Bing-Qing2, HUA Deng-Ke1, ZHANG Kun1, YUAN Jiang-Jiang1, ZHENG Xiao-Bin1, XU Bao-Yun1, ZHANG You-Jun1, WU Qing-Jun1,*   

  1.  (1. Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing 100081, China; 2. Changping District Agricultural Technology Extension Station, Beijing 102200, China)
  • Online:2021-08-20 Published:2021-08-12

摘要:

【目的】γ-氨基丁酸(γ-aminobutyric acid, GABA)是动物神经系统中一种重要的神经递质。本研究旨在鉴定西花蓟马Frankliniella occidentalis γ-氨基丁酸受体(GABA receptors, GABAR)家族基因,明确离子型受体(GABAAR)在西花蓟马对多杀霉素抗性形成中的作用。【方法】根据西花蓟马基因组和转录组数据,鉴定并克隆GABAR基因,进行生物信息学分析;采用qPCR检测GABAAR亚基基因FoRDL, FoLCCH3和FoGRD在西花蓟马多杀霉素敏感品系不同发育阶段(1-2龄若虫、蛹和成虫)的表达模式及在敏感和抗性品系成虫期的相对表达量差异;对西花蓟马敏感品系3日龄成虫的FoRDL进行RNAi 24 h后,在0.250和0.400 mg/L多杀霉素处理下生物测定计算西花蓟马成虫死亡率。【结果】注释并克隆获得8个GABAR基因:FoRDL, FoLCCH3, FoGRD, FoGRDlike1, FoGRD-like2, FoB1, FoB2和FoB-like(GenBank登录号: MH148151-MH148158),其ORF长度介于1 080~3 720 bp;系统进化分析表明,西花蓟马的GABAR基因与其他昆虫物种相应基因聚类,具有很高的保守性。GABAAR亚基FoRDL, FoLCCH3和FoGRD具有典型的N端胞外区环结构(loop A-F)和4个跨膜区(TM 1-4);FoRDL外显子3存在互斥剪切。FoRDL, FoLCCH3和FoGRD的表达量随多杀霉素敏感品系西花蓟马发育阶段逐渐升高,成虫期达最高;FoRDL在西花蓟马抗性品系成虫期的表达量显著低于敏感品系成虫期的。对西花蓟马敏感品系的FoRDL进行RNAi后,在0.250和0.400 mg/L多杀霉素处理剂量下,西花蓟马成虫死亡率均显著下降,分别比对照下降了55.80%和43.00%。【结论】在西花蓟马体内鉴定到5个离子型和3个代谢型GABAR基因,其中FoRDL可能在西花蓟马对多杀霉素产生抗性中发挥一定作用。

关键词: 西花蓟马, γ-氨基丁酸受体, 多杀霉素, 抗药性, RNA干扰

Abstract:

【Aim】 Gammaaminobutyric acid (GABA) is an important neurotransmitter in animal nervous system. This study aims to identify the GABA receptor (GABAR) familygenesinthewestern flower thrips, Frankliniella occidentalis, and to clarify the role of ionotropic receptor (GABAAR) in the resistance evolution to spinosad in F. occidentalis. 【Methods】 Based on the genomeand transcriptome data of F. occidentalis, the GABAR genes were identified, cloned and analyzed with bioinformatics tools. The expression patterns of the GABAAR subunit genes, FoRDL, FoLCCH3, and FoGRD in the spinosad susceptible strain of F. occidentalis at different developmental stages (1st-2nd instar nymphal, pupal, and adult stages), and their expression differences between the spinosad susceptible and resistant strains of F. occidentalis at the adult stage were detected by qPCR. After treatment with 0.250 and 0.400 mg/L spinosad at 24 h post RNAi of FoRDL in the 3-day-old adults of the spinosad susceptible strain of F. occidentalis, the mortality rates of F. occidentalis adults were determined by bioassay. 【Results】 Eight GABAR genes including FoRDL, FoLCCH3, FoGRD, FoGRD-like1, FoGRD-like2, FoB1, FoB2, and FoB-like (GenBank accession numbers: MH148151-MH148158) were annotated and cloned, and their ORF lengths vary from 1 080 to 3 720 bp. Phylogenetic analysis showed that GABAR genes of F. occidentalis were clustered with the corresponding genes of other insect species, indicating high conservativeness. GABAAR subunits FoRDL, FoLCCH3 and FoGRD all have a typical nitrogen-terminal extracellular region loop structure (loop A-F) and four transmembrane regions (TM 1-4), and exon 3 of FoRDL has mutually exclusive splicing. The expression levels of FoRDL, FoLCCH3 and FoGRD in the spinosad susceptible strain of F. occidentalis increased with the developmental stage of F. occidentalis, and the expression peak occurred at the adult stage. The expression level of FoRDL in the spinosad resistant strain of F. occidentalis at the adult stage was significantly lower than that in the susceptible strain at the adult stage. After treatment with 0.250 and 0.400 mg/L spinosad following RNAi of FoRDL in the susceptible strain of F. occidentalis, the mortality rates of F. occidentalis adults decreased significantly by 55.80% and 43.00%, respectively, as compared to those of the control. 【Conclusion】 Five ionic and three metabotropic GABAR genes have been identified in F. occidentalis. FoRDL may play a role in the resistance evolution to spinosad in F. occidentalis.

Key words: Frankliniella occidentalis, γ-aminobutyric acid receptor, spinosad, insecticide resistance, RNA interference