小麦条锈菌对三唑酮敏感性生测体系优化及其抗药性分子标记开发

Optimization of bioassay system for sensitivity of Puccinia striiformis f.sp.tritici to triadimefon and development of molecular markers for fungicide-resistance

条锈病是小麦上重要的真菌病害,严重威胁小麦的生产安全,施用杀菌剂是防治小麦条锈病的重要措施。三唑酮是防治条锈病的主要药剂,但长期单一、大规模使用三唑酮势必导致病原菌抗药性出现,由于小麦条锈菌是严格的专性寄生菌,目前测定条锈菌对杀菌剂的敏感性实验主要在活体小麦上进行,实验结果易受外界环境影响。本研究通过培养皿内离体小麦叶段培养,采取定量接种和软件读数的方法,对2021年西北核心越夏区60个小麦条锈菌菌株进行三唑酮敏感性测定,生测结果表明供试菌株平均EC50值为0.543μg·mL^(-1),抗性菌株7株,敏感菌株53株,11.67%的供试菌株对三唑酮具有低至中等水平抗性,分子标记结果与生测结果高度吻合。本研究通过优化三唑酮对条锈病防治效果的室内生测体系,使得结果更加稳定、重复性好,同时基于敏感/抗药菌株靶标基因序列分析开发了高通量的抗药性竞争性等位基因特异性PCR-单核苷酸多态性分子标记(KASP-SNP),旨在为防治小麦条锈病筛选高效杀菌剂,同时开展田间小麦条锈菌抗药性监测,了解抗药性分布及抗性水平,为小麦条锈菌抗药性综合治理提供科学依据。

Stripe rust,caused by Puccinia striiformis f.sp.tritici,is an important fungal disease on wheat,which seriously threatens the production safety of wheat.Fungicide application is an important measure to control this disease where triadimefon is the main fungicide used for the disease control.However,with the long-term single and large-scale use of the triadimefon,it is inevitably to the emergence of resistant isolates.Since P.striiformis f.sp.tritici is a strict obligate parasite,the current experiments to determine the sensitivity of the pathogen to fungicides are mainly carried out in vivo,the results are easily affected by the external environment.In this study,based on the cultures of detached wheat leaves in petri dishes,quantitative inoculation and software reading methods,the sensitivities of 60 P.striiformis f.sp.tritici isolates from the core over-summering area of northwestern China in 2021 to triadimefon were evaluated.The bioassay test showed that the average EC50 of the tested isolates was 0.543μg·mL^(-1),where 7 isolates were resistant with 11.67%of the tested isolates had low to moderate resistance to triadimefon.The results of the molecular marker were consistent with those of the bioassay.Through optimizing the indoor bioassay system of the control effect of triadimefon on stripe rust,the results were more stable and reproducible.A high-throughput fungicide-resistant kompetitive allele-specific PCR-single nucleotide polymorphism(KASP-SNP)molecular marker based on the target gene sequence analysis of sensitive/resistant strains was developed.The results of this study provide a scientific basis to screen efficient fungicides for the control of wheat stripe rust,and to understand the distribution and resistance level of resistance based on monitoring the resistance of the pathogen in the field which helps for the comprehensive management of fungicide resistance.