尖孢镰刀菌黏团专化型β-葡萄糖苷酶Foglu1的功能  

Functional study of aβ-glucosidase Foglu1 in Fusarium oxysporum f.sp.conglutinans

作　　者：王腾 王碧香 李诗瑶 尉婧 李二峰 WANG Teng;WANG Bixiang;LI Shiyao;WEI Jing;LI Erfeng(College of Horticulture and Landscape Architecture,Tianjin Agricultural University,Tianjin 300392,China)

机构地区：[1]天津农学院园艺园林学院,天津300392

出　　处：《浙江农业学报》2023年第2期373-382,共10页Acta Agriculturae Zhejiangensis

基　　金：国家自然科学基金(31801682);国家重点研发计划(2018YFD0201204)。

摘　　要：甘蓝枯萎病是由尖孢镰刀菌黏团专化型(Fusarium oxysporum f.sp.conglutinans,FOC)引起的。β-葡萄糖苷酶(β-glucosidase)是纤维素降解过程中的限速酶。本研究中,为明确FOC中一种β-葡萄糖苷酶在其生长发育和致病中的生物学功能,通过split-marker同源重组技术对该病原菌中一种β-葡萄糖苷酶基因(Foglu 1)进行敲除与回补,分析野生型菌株与ΔFoglu 1敲除突变体菌株、ΔFoglu 1-C回补菌株间的表型差异。结果表明,在细胞膜胁迫试验(0.05%SDS)中,ΔFoglu 1突变体生长5 d后的菌落直径与野生型相比降低19.6%,推测Foglu 1基因对病原菌细胞膜功能具有一定的作用;氧胁迫试验(0.1%H_(2)O_(2))中菌落直径比野生型减小14%,表明ΔFoglu 1突变体比野生型对氧胁迫更加敏感;ΔFoglu 1突变体产孢量较野生型降低约22.3%,说明Foglu 1基因影响FOC的产孢能力;在接种寄主甘蓝时,发现接种ΔFoglu 1菌株的甘蓝植株初期发病缓慢,接种后第8天的病情指数比接种野生菌株的低39.5%,接种后第11、15和18天,野生菌株和ΔFoglu 1突变体致病力无显著差异,说明Foglu 1基因影响病原菌与寄主间的早期互作,延缓发病但对其致病力无影响。上述结果显示,β-葡萄糖苷酶在FOC细胞膜完整性、氧化胁迫、产孢和致病过程中可能起重要作用。Fusarium oxysporum f.sp.conglutinans(FOC)is a causal agent of cabbage Fusarium wilt.Theβ-glucosidase is a rate-limiting enzyme for cellulose degradation.In this study,to determine biological roles of aβ-glucosidase(Foglu1)in fungal development and virulence of FOC,the Foglu 1 gene was knocked out by split-marker homologous recombination and complementation strains were generated by reintroducing Foglu 1 gene intoΔFoglu 1 mutant.Then,phenotypes of the wild type strain,deletion mutantΔFoglu 1 and complemented strainΔFoglu 1-C were analyzed.The result showed that the colony diameter ofΔFoglu 1 mutant was significantly reduced by 19.6%compared to that of the wild type in cell membrane stress experiment(0.05%SDS),indicating that Foglu 1 was required for cell integrity in FOC.Also,in oxygen stress experiment(0.1%H_(2)O_(2)),colony diameter ofΔFoglu 1 mutant significantly decreased by 14%compared with the wild type,indicating that deletion of Foglu 1 resulted in the increase of sensitivity to oxygen stress.Meanwhile,conidial production ofΔFoglu 1 mutant was reduced by 22.3%compared to the wild type,indicating that Foglu 1 gene was important for sporulation in FOC.Interestingly,at 8 days after inoculation,disease index ofΔFoglu 1 mutant was significantly reduced and 39.5%lower than that of the wild type.The disease indexes of seedlings inoculated with the wild type,ΔFoglu1 andΔFoglu 1-C were no differences at 11,15 and 18 days after inoculation.It showed that Foglu 1 played roles in the early interaction between the pathogen and the host.It could delay the disease development but had no effect on pathogenicity.In conclusion,these findings suggested thatβ-glucosidase(Foglu1)played an important role in FOC cell membrane integrity,oxidative stress,sporulation and pathogenesis.

关 键 词：甘蓝枯萎病 基因敲除 Β-葡萄糖苷酶 致病力 

分 类 号：S432.1[农业科学—植物病理学]