二斑叶螨抗甲氰菊酯种群解毒酶基因表达分析

doi:10.11686/cyxb2014321

草业学报 ›› 2015, Vol. 24 ›› Issue (8): 150-158.DOI: 10.11686/cyxb2014321

二斑叶螨抗甲氰菊酯种群解毒酶基因表达分析

杨顺义^{1**, *}, 岳秀利^{1**, *}, 王进军², 刘长仲¹, 沈慧敏^{1*, *}, 沈一凡¹, 郭金梅¹

1.甘肃农业大学草业学院,草业生态系统省部共建教育重点实验室,中-美草地畜牧业可持续发展中心,甘肃兰州 730070;
2.西南大学植物保护学院,昆虫学及害虫控制工程重点实验室,重庆400716

出版日期:2015-08-20 发布日期:2015-08-20
通讯作者: E-mail: ndshm@gsau.edu.cn
基金资助:
公益性行业(农业)科研专项 (201103020)和国家自然科学基金项目(31260442)资助; 作者简介:杨顺义(1972-),男,甘肃礼县人,副教授,硕士; E-mail:yangshy@gsau; edu; cn; 岳秀利(1988-),男,山东滕州人,硕士; E-mail:yuexiuli.01@163; com

Gene expression of a detoxification enzyme in Tetranychus urticae resistant to fenpropathrin

YANG Shun-Yi^{1, **}, YUE Xiu-Li^{1, **}, WANG Jin-Jun², LIU Chang-Zhong¹, SHEN Hui-Min^{1, *}, SHEN Yi-Fan¹, GUO Jin-Mei¹

1.College of Prataculture, Gansu Agricultural University, Key Laboratory of Grassland Ecosystem Education Ministry, The Sino-U.S. Centers for Grazingland Ecosystem Sustainability, Lanzhou 730070, China;
2.College of Plant Protection, Southwest University, China Key Laboratory of Entomology and Pest Control Engineering, Chongqing 400716, China

Online:2015-08-20 Published:2015-08-20

摘要/Abstract

摘要： 基因mRNA水平相对表达量的显著变化是二斑叶螨对拟除虫菊酯类药剂产生抗性的重要机制。为了揭示二斑叶螨对甲氰菊酯抗性产生的解毒酶分子机理,本研究采用实时荧光定量PCR(quantitative real time PCR, qRT-PCR)方法分析二斑叶螨实验室敏感(SS)和田间种群(LZ-R、GN-R、WW-R、TS-R和LX-R)主要解毒酶谷胱甘肽转移酶(glutathione s-transferases, GSTs)、细胞色素P450单加氧酶(cytocheome P450 monooxygenases, P450s) 及羧酸酯酶(carboxyl/cholinesterases, CCEs)基因mRNA水平相对表达量的差异。结果表明,二斑叶螨不同种群不同解毒酶基因的相对表达量不同。WW-R和TS-R种群中TuGSTd05以及LX-R种群中TuGSTd01和 TuGSTd06基因表达量均显著上调,为SS种群的1.42~2.34倍,而GN-R种群中TuGSTd04,LZ-R种群中TuGSTd05 和GSTd09表达量显著下调,为SS种群的0.41~0.70倍;P450s基因CYP406A1 和CYP4CL1表达量在LZ-R、GN-R以及WW-R种群中均显著上调,分别为SS种群的1.80~4.88倍,此外,CYP387A1在LZ-R种群中显著上调2.19倍,而在LX-R种群中显著下调0.42倍;CCEs基因TuCCE-35表达量在WW-R和TS-R种群中显著上调,分别为SS种群的2.82和3.09倍,而TuCCE-36基因在所有种群中的表达量均不显著。二斑叶螨不同种群中解毒酶基因GSTs、P450s 和CCEs的显著上调或下调可能与甲氰菊酯的抗性形成有关。

Abstract: The variation of mRNA gene expression is an important pyrethroid insecticide resistance mechanism in Tetranychus urticae. To identify the molecular mechanism for producing detoxification enzymes in T. urticaein response to fenpropathrin, the mRNA expression levels of glutathione s-transferases (GSTs), cytochrome (P450s) and carboxyl/cholinesterases (CCEs) genes in laboratory fenpropathrin susceptible (SS), fenpropathrin resistant (Sp-R) and field strains (LZ-R, GN-R, WW-R, TS-R and LX-R) of T. urticae were measured using quantitative real time PCR (qRT-PCR). The results showed that relative expression levels of different detoxification enzyme genes varied with different strains of T. urticae. Compared with the SS strain, the expression levels of TuGSTd05 in the WW-R and TS-R strains, TuGSTd01 and TuGSTd06 in the LX-R strain were significantly up-regulated,1.42-2.34 times greater than the SS strain, while TuGSTd04 in the GN-Rstrain, TuGSTd05 and TuGSTd09 in the LX-R strain were significantly down-regulated, 0.41-0.70 that of the SS strain. The expression levels of P450s genes CYP406A1 and CYP4CL1 in the LZ-R, GN-R and WW-R strains were significantly up-regulated, 1.80-4.88 times that of the SS strain. The expression level of P450s genes CYP387A1 in the LZ-R strain was also significantly higher (2.19 times) than the SS strain whereas that in the LX-R strain was significantly lower (0.42 times) than the SS strain. Similarly, the expression levels of CCEs gene TuCCE-35 in the WW-R and TS-R strains was significantly higher than the SS strain. However, the gene expression levels of TuCCE-36 did not change in all strains. The significant up-regulation or down-regulation of detoxification enzyme genes (GSTs, p450s and CCEs) among different strains of T. urticae may be associated with the formation of resistance to fenpropathrin.

杨顺义, 岳秀利, 王进军, 刘长仲, 沈慧敏, 沈一凡, 郭金梅. 二斑叶螨抗甲氰菊酯种群解毒酶基因表达分析[J]. 草业学报, 2015, 24(8): 150-158.

YANG Shun-Yi, YUE Xiu-Li, WANG Jin-Jun, LIU Chang-Zhong, SHEN Hui-Min, SHEN Yi-Fan, GUO Jin-Mei. Gene expression of a detoxification enzyme in Tetranychus urticae resistant to fenpropathrin[J]. Acta Prataculturae Sinica, 2015, 24(8): 150-158.

参考文献

[1] Grbic M, Van Leeuwen T, Clark R M, et al . The genome of Tetranychus urticae reveals herbivorous pest adaptations. Nature, 2011, 479(7374): 487-492.
[2] Zhang T W, Shen H M, Qian X J, et al . Effects of Tetranychus urticae feeding on the chlorophyll content and two kinds of protective enzyme of white clover. Chinese Journal of Applied Entomology, 2013, 50(2): 395-400.
[3] He D H, Zhao X P, Jin Q H, et al . Dispersion of two spotted spider mite, Tetranychus urticae Koch, and its selection of host plants on farmland in Ningxia. Chinese Journal of Applied and Environmental Biology, 2001, 7(5): 447-451.
[4] Van Leeuwen T, Vontas J, Tsagkarakou A, et al . Acaricide resistance mechanisms in the two-spotted spider mite Tetranychus urticae and other important Acari: A review. Insect Biochemistry and Molecular Biology, 2010, 40(8): 563-572.
[5] Van Leeuwen T, Tirry L. Esterase-mediated bifenthrin resistance in a multiresistant strain of the two-spotted spider mite, Tetranychus urticae . Pest Management Science, 2007, 63(2): 150-156.
[6] Van Leeuwen T, Van Pottelberge S, Tirry L. Comparative acaricide susceptibility and detoxifying enzyme activities in field-collected resistant and susceptible strains of Tetranychus urticae . Pest Management Science, 2005, 61(5): 499-507.
[7] Yorulmaz S, Ay R. Multiple resistance, detoxifying enzyme activity, and inheritance of abamectin resistance in Tetranychus urticae Koch (Acarina: Tetranychidae). Turkish Journal of Agriculture and Forestry, 2009, 33(4): 393-402.
[8] Van Pottelberge S, Van Leeuwen T, Khajehali J, et al . Genetic and biochemical analysis of a laboratory-selected spirodiclofen-resistant strain of Tetranychus urticae Koch (Acari: Tetranychidae). Pest Management Science, 2009, 65(4): 358-366.
[9] Kwon D H, Clark J M, Lee S H. Cloning of a sodium channel gene and identification of mutations putatively associated with fenpropathrin resistance in Tetranychus urticae . Pesticide Biochemistry and Physiology, 2010, 97(2): 93-100.
[10] Tsagkarakou A, Van Leeuwen T, Khajehali J, et al . Identification of pyrethroid resistance associated mutations in the para sodium channel of the two-spotted spider mite Tetranychus urticae (Acari: Tetranychidae). Insect Molecular Biology, 2009, 18(5): 583-593.
[11] Demaeght P, Dermauw W, Tsakireli D, et al . Molecular analysis of resistance to acaricidal spirocyclic tetronic acids in Tetranychus urticae : CYP 392 E 10 metabolizes spirodiclofen, but not its corresponding enol. Insect Biochemistry and Molecular Biology, 2013, 43(6): 544-554.
[12] Gao X J, Shen H M. Resistance selection of fenpropathrin and the change of detoxification enzyme activities in Tetranychus urticae Koch (Acari: Tetranychidae). Acta Entomologica Sinica, 2011, 54(1): 64-69.
[13] Duan X L, Zhang Z G, Gao X J, et al . Selection of Tetranychus urticae resistant to fenpropathrin and spirodiclofen and the synergistic action of the synergists to the resistant population. Plant Protection, 2011, 37(5): 106- 109.
[14] Dittrich V, Cranham J E, Jepson L R, et al . Revised method for spider mites and their eggs (eg Tetranychus spp. and Panonychus ulmi Koch). FAO Method No. 10a. FAO Plant Production and Protection Paper, 1980, 21: 49-53.
[15] Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 1976, 72(1): 248-254.
[16] Clark A G, Dick G L, Smith J N. Kinetic studies on a glutathione S-transferase from the larvae of Costelytra zealandica . Biochemical Journal, 1984, 217(1): 51-58.
[17] Hansen L G, Hodgson E. Biochemical characteristics of insect microsomes: N-and O-demethylation. Biochemical Pharmacology, 1971, 20(7): 1569-1578.
[18] Van Asperen K. A study of housefly esterases by means of a sensitive colorimetric method. Journal of Insect Physiology, 1962, 8(4): 401-416.
[19] Pfaffl M W. A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Research, 2001, 29(9): 2002-2007.
[20] Gao X J. Resistance Molecular Mechanism of Tetranychus urticae (Koch) to Fenpropathrin[D]. Lanzhou: Gansu Agricultural University, 2012.
[21] Tirello P, Pozzebon A, Cassanelli S, et al . Resistance to acaricides in Italian strains of Tetranychus urticae : toxicological and enzymatic assays. Experimental and Applied Acarology, 2012, 57(1): 53-64.
[22] Niu J Z, Liu G Y, Dou W, et al . Susceptibility and activity of glutathione S-transferases in nine field populations of Panonychus citri (Acari: Tetranychidae) to pyridaben and azocyclotin. Florida Entomologist, 2011, 94(2): 321-329.
[23] He L, Xue C H, Wang J J, et al . Resistance selection and biochemical mechanism of resistance to two Acaricides in Tetranychus cinnabarinus (Boiduval). Pesticide Biochemistry and Physiology, 2009, 93(1): 47-52.
[24] Zhong D B, Chang X L, Zhou G F, et al . Relationship between knockdown resistance, metabolic detoxification and organismal resistance to pyrethroids in Anopheles sinensis . PLoS ONE, 2013, 8(2): 1-10.
[25] Riveron J M, Irving H, Ndula M, et al . Directionally selected cytochrome P450 alleles are driving the spread of pyrethroid resistance in the major malaria vector Anopheles funestus. Proceedings of the National Academy of Sciences of the United States of America, 2013, 110(1): 252-257.
[26] Yang S Y, Yue X L, Wang J J, et al . Testing two methods to detect voltage-gated sodium channels gene mutation in Tetranychus urticae . Acta Prataculturae Sinica, 2014, 23(5):153-160.
[27] Van Leeuwen T, Dermauw W, Grbic M, et al . Spider mite control and resistance management: does a genome help. Pest Management Science, 2013, 69(2): 156-159.
[28] Jia X P, Ye X Q, Liang L J, et al . Transcriptome characteristics of Paspalum vaginatum analyzed with Illumina seguencing technology. Acta Prataculturae Sinica, 2014, 23(6): 242-252.
[29] Zhang J, Song L L, Guo D L, et al . Genome-wide identification and investigation of the MADS-box gene family in Medicago truncatula . Acta Prataculturae Sinica, 2014, 23(6): 233-241.
[2] 张廷伟, 沈慧敏, 钱秀娟, 等. 二斑叶螨刺吸胁迫对白三叶叶绿素含量和两种保护酶的影响. 应用昆虫学报, 2013, 50(2): 395-400.
[3] 贺达汉, 赵晓萍, 靳巧红, 等. 宁夏地区二斑叶螨的寄主植物选择及其季节转移. 环境与应用生物学报, 2001, 7(5): 447-451.
[12] 高新菊, 沈慧敏. 二斑叶螨对甲氰菊酯的抗性选育及解毒酶活力变化. 昆虫学报, 2011, 54(1): 64-69.
[13] 段辛乐, 张志刚, 高新菊, 等. 二斑叶螨对甲氰菊酯和螺螨酯的抗性选育及增效剂的增效作用. 植物保护, 2011, 37(5): 106-109.
[20] 高新菊. 二斑叶螨对甲氰菊酯的抗性机理研究[D]. 兰州:甘肃农业大学, 2012.
[26] 杨顺义, 岳秀利, 王进军, 等. 两种方法在二斑叶螨电压门控钠离子通道基因突变检测中的应用. 草业学报, 2014, 23(5):153-160.
[28] 贾新平, 叶晓青, 梁丽建, 等. 基于高通量测序的海滨雀稗转录组学研究. 草业学报, 2014, 23(6):242-252.
[29] 张军, 宋丽莉, 郭东林, 等. MADS-box基因家族在蒺藜苜蓿的全基因组分析. 草业学报, 2014, 23(6):233-241.
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二斑叶螨抗甲氰菊酯种群解毒酶基因表达分析

Gene expression of a detoxification enzyme in Tetranychus urticae resistant to fenpropathrin

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