褐飞虱对氟啶虫胺腈早期抗性进化机理研究

The brown planthopper is one of the most serious pests of rice and causes severe losses in its production. Sulfoxaflor is a novel insecticide with new chemical structure, unique mechanism of action and has sound control effect against brown planthopper. Previous study demonstrated that the brown planthopper strain with medium level resistance to sulfoxaflor has higher detoxification enzyme activity than the susceptible strain, but the resistance mechanism is unclear. Study of the molecular mechanisms of sulfoxaflor resistance will provide an important basis for the management of resistance in the brown planthopper population. In this study, the theory and methods of molecular biology and biochemistry including high-throughput sequencing, qRT-PCR and RNA interference will be used to 1) study the resistance risk assessment of brown planthopper to sulfoxaflor, 2) understand the relationship between genes of detoxification enzymes and sulfoxaflor resistance, 3) explore the functional role of nAChRs subunit gene mutation in sulfoxaflor resistance. The subject of this research is to elucidate the molecular mechanism of early resistance evolution to sulfoxaflor in brown planthopper. This research would be useful for the in-depth analysis of the molecular mechanism of pest resistance to neonicotinoid insecticides, and have an important theoretical and practical significance for the scientifically rational application of sulfoxaflor and the effective management of brown planthopper resistance.

褐飞虱是目前水稻生产上最为严重的害虫，氟啶虫胺腈是具有新颖化学结构和独特作用机理的全新杀虫剂，对褐飞虱具有良好的防治效果，前期研究发现室内抗性选育的氟啶虫胺腈中等抗性水平褐飞虱品系解毒酶活力增强，但其具体抗性机理尚不明确。研究褐飞虱对氟啶虫胺腈抗性机理是进行抗性治理的重要前提。因此，本项目拟应用分子生物学与生物化学的理论与方法，结合高通量测序、实时荧光定量PCR及RNA干扰等技术，1）研究褐飞虱对氟啶虫胺腈的抗性风险；2）研究解毒酶基因与褐飞虱对氟啶虫胺腈抗性的关系；3）探索nAChRs亚基基因突变在褐飞虱对氟啶虫胺腈抗性中的作用。通过以上研究，明确褐飞虱对氟啶虫胺腈抗性产生的规律及抗性风险，揭示褐飞虱对氟啶虫胺腈早期抗性进化的分子机理。研究结果有助于深入解析害虫对新烟碱类杀虫剂抗性分子机制、氟啶虫胺腈的科学合理使用及褐飞虱抗性有效治理具有重要的理论和现实意义。

褐飞虱为当前水稻上为害最严重的害虫，氟啶虫胺腈是具有新颖化学结构和独特作用机理的全新杀虫剂，对褐飞虱具有良好的防治效果。抗性监测结果表明，当前我国褐飞虱田间种群对氟啶虫胺腈产生了低水平抗性，且抗性水平呈现明显上升趋势。通过室内连续39代筛选，获得了抗氟啶虫胺腈褐飞虱高水平抗性品系（RR=183.63），交互抗性测定结果表明氟啶虫胺腈与烯啶虫胺、吡虫啉、噻虫嗪、噻虫胺、呋虫胺和环氧虫啶之间存在明显交互抗性，与三氟苯嘧啶、异丙威、毒死蜱、醚菊酯和噻嗪酮之间不存在交互抗性。抗性遗传方式研究结果表明褐飞虱对氟啶虫胺腈抗性为多基因控制的常染色体不完全隐性遗传。抗性品系与初筛品系相比存在0.75的抗性适合度代价，抗性品系的1龄若虫历期、5龄若虫历期、成虫前期及总产卵前期（TPOP）均显著延长；3龄若虫历期、雌成虫寿命、产卵历期均显著缩短；同时，产卵量和孵化率显著降低。增效剂试验结果表明，在抗性品系中PBO和TPP对氟啶虫胺腈分别表现出2.69倍和1.48倍的相对增效倍数，表明褐飞虱对氟啶虫胺腈抗性可能与其体内P450和EST活性上升有关。解毒酶活力测定结果表明，抗性品系褐飞虱体内P450和EST活力分别是初筛品系的3.50倍和1.85倍。通过qRT-PCR检测，发现8条P450基因CYP15G1、CYP6CS1、CYP6CW1、CYP6ER1、CYP4C62、CYP4DE1、CYP417A1、CYP419A1在抗性品系SFX-SEL（G39）中的表达水平与初筛品系（UNSEL）相比上调2倍以上，其中CYP6ER1上调最明显，上调倍数达36.87倍。使用RNAi成功干扰CYP6ER1在抗性褐飞虱体内表达，被干扰后的抗性褐飞虱对氟啶虫胺腈的敏感性得到恢复，表明CYP6ER1上调表达是褐飞虱对氟啶虫胺腈产生抗性的关键因素。本研究明确了褐飞虱对氟啶虫胺腈抗性产生的规律、抗性风险、抗性遗传方式及适合度代价，揭示了褐飞虱对氟啶虫胺腈早期抗性进化的分子机理。研究结果对深入解析害虫对新烟碱类杀虫剂抗性分子机制、氟啶虫胺腈的科学合理使用及褐飞虱抗性有效治理具有重要的理论和现实意义。