灰葡萄孢侵染垫缺失突变体的筛选及其相关生物学特性的研究

Screening and biological characterization of infection cushion-deficient mutants in Botrytis cinerea

【目的】从农杆菌介导获得的灰葡萄孢Rose BC-3的突变体库中筛选侵染垫缺失突变体菌株,并明确其相关生物学特性。【方法】将菌株接种于洋葱表皮,利用棉兰染色观察侵染垫形成情况,筛选得到一个侵染垫缺失突变体(AT19)。采用形态学方法、离体叶片接种法、钌红染色法、小麦种子幼芽生长抑制法分别对该菌株的菌落培养性状、侵染垫产生情况、致病力、产果胶酶能力以及产植物毒性代谢产物能力进行测定。【结果】筛选灰葡萄孢突变体168株,根据侵染垫形成可分为三类:快速形成侵染垫型(158株)、缓慢形成侵染垫型(9株)和侵染垫形成缺陷型(1株,AT19)。AT19在接种洋葱120 h后依然无法形成成熟侵染垫。该菌株生长较为缓慢,菌落扩展均匀,可以产生分生孢子,对烟草、草莓、蚕豆和豌豆叶片均不能致病,可以产生果胶酶和植物代谢毒性物质。【结论】突变体菌株AT19可以产生果胶酶和植物代谢毒性物质,其致病力缺失与侵染垫产生缺陷相关。研究结果为了解灰葡萄孢侵染垫形成分子机制提供基础材料。

[Objective] To screen infection cushion-deficient mutants in Botrytis cinerea strain Rose BC-3 transformed with Agrobacterium tumefaciens. [Methods] Mycelial plugs of B. cinerea were inoculated on onion epidermal strips, which were then incubated for 12 to 120 h, stained with Medan dye and finally examined with a light microscope. Colony morphology, formation of infection cushions, pathogenicity, and production of pectinase and toxic metabolites by the selected mutants were determined using routine methods. [Results] One hundred and sixty-eight transformed mutants of B. cinerea were divided into three types： the rapid formation type（158 mutants）, the slow formation type（9 mutants）, and the defective formation type（only one isolate, namely AT19）. On potato dextrose agar, isolate AT19 grew slowly, produced conidia and formed colonies with the normal appearance. However, it could hardly infect leaves of tobacco, strawberry, broadbean and pea, although it was detected to be able to produce pectinase and some toxic metabolites. [Conclusion] Formation of infection cushions appears important for B. cinerea to infect plant tissues.