大豆平头炭疽菌和胶孢炭疽菌对啶氧菌酯的抗性风险及抗性分子机制研究

doi:10.1016/j.jia.2023.07.037

摘要/Abstract

摘要：

炭疽病是大豆最重要的病害之一，平头炭疽菌和胶孢炭疽菌是其最主要的病原菌。啶氧菌酯属于QoI类杀菌剂，常用于作物炭疽病的防治。然而，我国大豆平头炭疽菌和胶孢炭疽菌对啶氧菌酯的敏感性、抗性风险和抗性分子机制尚不清楚。本研究建立了大豆平头炭疽菌和胶孢炭疽菌对啶氧菌酯的敏感基线，采用室内药剂驯化的方法诱导抗性突变体，并测定抗性突变体的适合度和交互抗性，同时比较分析抗性突变体和敏感菌株的cyt b基因cDNA序列，进一步利用分子对接明确敏感菌株和突变体Cyt b蛋白与啶氧菌酯的结合力。结果表明：供试的128株平头炭疽菌和121株胶孢炭疽菌对啶氧菌酯的EC₅₀值频率分布呈连续单峰曲线，平均EC₅₀值分别为0.7740和1.1561 μg mL^-1，该平均EC₅₀值可作为相对敏感基线监测未来田间大豆平头炭疽菌和胶孢炭疽菌对啶氧菌酯的敏感性变化。分别筛选获得11株平头炭疽菌和6株胶孢炭疽菌的抗性突变体，其EC₅₀值分别为5.40–152.96 µg mL^-1、13.53–28.30 μg mL^-1。抗性突变体在分生孢子的产生、萌发和致病性方面表现出与亲本菌株相似或更高的生存适合度。啶氧菌酯与吡唑醚菌酯之间存在交互抗性，而与氟啶胺、苯醚甲环唑及丙环唑之间无交互抗性。平头炭疽菌7株高抗突变体（RF > 100）cyt b基因427位碱基由G突变为A，所编码的Cyt b蛋白143位由甘氨酸（G）突变为丝氨酸（S），4株中抗突变体（RF < 50）cyt b基因409位碱基由G突变为A，所编码的Cyt b蛋白137位由甘氨酸（G）突变为精氨酸（R）。分子对接验证发现G143S和G137R突变是平头炭疽菌对啶氧菌酯产生抗药性的主要机制。胶孢炭疽菌抗性突变体cyt b基因未发生突变。温室试验结果表明，啶氧菌酯不能有效地防治高抗突变体，对中抗突变体的防效也显著下降。总之，G143S和G137R是大豆平头炭疽菌对啶氧菌酯产生抗药性的主要机制，大豆平头炭疽菌和胶孢炭疽菌对啶氧菌酯的抗性风险为中至高等，生产中应对QoI类杀菌剂实施抗性监测，并采取有效的抗性管理策略。

Abstract:

Anthracnose, caused by Colletotrichum truncatum and C. gloeosporioides, is amongst the most serious diseases of soybean in China. Picoxystrobin, a quinone outside inhibitor fungicide, is commonly used for the control of anthracnose. Its resistance risk and mechanism in C. truncatum and C. gloeosporioides are unclear. In this study, the sensitivities of 128 C. truncatum and 121 C. gloeosporioides isolates to picoxystrobin were investigated, and unimodal distributions were observed with average EC₅₀ values of 0.7740 and 1.1561 μg mL^–1, respectively. Eleven picoxystrobin-resistant mutants of C. truncatum and six mutants of C. gloeosporioides were acquired, with EC₅₀ values varying from 5.40–152.96 and 13.53–28.30 μg mL^–1, respectively. Compared to the parental isolates, mutants showed similar or higher relative fitness in conidial production and germination, and pathogenicity. Collectively, the resistance risk of C. truncatum and C. gloeosporioides to picoxystrobin is moderate to high. There was positive cross-resistance between picoxystrobin and pyraclostrobin, but not between picoxystrobin and fluazinam, difenoconazole, or propiconazole. The G143S mutation in Cyt b protein was detected in seven high-resistant mutants of C. truncatum (RF>100), and G137R occurred in four moderate-resistant mutants (RF<50). Contrastingly, there were no point mutations in Cyt b of any C. gloeosporioides mutants. Molecular docking confirmed that two mutations conferred different resistance levels to picoxystrobin. Under greenhouse trials, picoxystrobin did not control mutants with the G143S mutation, those bearing G137R or no point mutation were somewhat controlled, but at a lower level compared to wild-type isolates. These results showed that integrated management strategies should be implemented to preserve fungicide effectiveness.

Key words: Colletotrichum truncatum , Colletotrichum gloeosporioides , picoxystrobin , point mutation , Cyt b , molecular docking

SHI Niu-niu, LIAN Jin-pan, QIU De-zhu, CHEN Fu-ru, DU Yi-xin. 大豆平头炭疽菌和胶孢炭疽菌对啶氧菌酯的抗性风险及抗性分子机制研究[J]. Journal of Integrative Agriculture, 2023, 22(12): 3681-3693.

SHI Niu-niu, LIAN Jin-pan, QIU De-zhu, CHEN Fu-ru, DU Yi-xin. Resistance risk and molecular mechanism associated with resistance to picoxystrobin in Colletotrichum truncatum and Colletotrichum gloeosporioides[J]. Journal of Integrative Agriculture, 2023, 22(12): 3681-3693.

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