沙打旺黄萎病菌的分离鉴定及其生物学特性和遗传多样性

Isolation,identification and characterization of causal pathogen of Verticillium wilt of standing milkvetch Astragalus adsurgens

为明确沙打旺Astragalus adsurgens黄萎病病原菌的种类及其生物学特性和遗传多样性,结合形态学特征和分子生物学方法对分离自疑似感染黄萎病沙打旺植株的病原菌进行鉴定,并依照柯赫氏法则进行致病性验证,同时在室内条件下对病原菌的生物学特性进行测定,分别利用大丽轮枝菌Verticillium dahliae生理小种和交配型特异性引物对病原菌的生理小种和交配型进行PCR分子鉴定。结果表明,共分离到13株病原菌菌株,其形态学特征和培养性状基本相似,经鉴定引起沙打旺黄萎病的病原菌为大丽轮枝菌。该菌具有较宽的温度和酸碱度适应范围,最适生长和产孢的温度为25℃,最适生长和产孢的pH范围分别为pH 7~9和pH 6~7;该病原菌生长和产孢的最适碳源为可溶性淀粉和D-甘露醇,最适生长氮源为牛肉膏和胰蛋白胨,最适产孢氮源为甘氨酸。分离获得的13株大丽轮枝菌菌株均属于2号生理小种和MAT1-2-1交配型,表明无性繁殖可能是大丽轮枝菌的主要繁殖形式。

In order to clarify the species,biological characteristic and genetic diversity of the pathogen caused Verticillium wilt on standing milkvetch Astragalus adsurgens,the fungus isolated from the diseased samples was identified morphologically and molecularly and confirmed following Koch’s postulates.Meanwhile,the biological characteristics of the pathogen was studied under indoor conditions,the race and mating types of the pathogen were also identified using specific primers.The results showed that all 13 isolates obtained from diseased samples had basically similar morphological and culture characteristics.The pathogen caused Verticillium wilt on A.adsurgens was identified as Verticillium dahliae.The optimum temperature for its mycelia growth and sporulation was 25℃.The suitable pH values for its mycelia growth and sporulation were 7-9 and 6-7,respectively.The pathogen had a wide range of temperature and pH adaptation.It grew well on the medium with soluble starch and D-mannitol as the carbon source;beef extract and tryptone were the best nitrogen sources for its mycelia growth whiles the best nitrogen source for its sporulation was glycine.All the 13 isolates obtained in this study were characterized as race 2 and MAT1-2-1.This indicated that clonal propagation might be the main reproductive mode for these V.dahliae isolates.