红树林放线菌抗沃柑病原真菌的研究

Study on mangrove actinomycetes against pathogenetic fungi of Orah

目的分离鉴定沃柑叶斑病害中的病原真菌,并从广西红树林来源放线菌中筛选高效抑制病原真菌的菌株,为开展沃柑真菌病害的防治奠定基础。方法采用组织分离法对病原真菌进行分离和纯化培养,通过形态学、ITS基因序列分析及构建系统发育树相结合的方法对病原真菌进行鉴定,并按照柯赫氏法则对健康沃柑叶片进行回接验证。采用平板对峙法对24株红树林放线菌进行抑菌活性初筛,并检测活性菌株发酵产物的抗菌活性及防病害效果。通过antiSMASH对目标菌株的基因组序列进行分析,预测其合成次级代谢产物的能力。结果从沃柑叶斑病害中分离出4种植物病原真菌,其中大豆茎点霉(Boeremia exigua)和腐皮镰刀菌(Fusarium solani)可导致沃柑叶片出现叶斑病症;从24株放线菌中筛选到1株链霉菌M2020,其对4种病原真菌C1~C4表现出很好的抑菌活性,且其发酵产物在沃柑真菌病害(真菌C4侵染)防治试验中效果显著;菌株M2020基因组中含有38个负责聚酮类、非核糖体多肽类和萜烯类等次级代谢产物生物合成的基因簇。结论致使沃柑出现叶斑病症的病原真菌至少有大豆茎点霉和腐皮镰刀菌,链霉菌M2020具有防治沃柑叶斑病害的潜力。

Objective To provide the pathogenic fungi causing Orah leaf spot disease,and the mangroves actinomycete strains against pathogenic fungi to prevent and control Orah fungal disease.Methods The pathogens were isolated and purified from Orah leaves and further identified by morphological observation and ITS sequence analysis coupling with the phylogenetic tree.The pathogenicity was verified according to Koch’s rule with healthy Orah leaves.Twenty-four actinomycetes isolated from mangrove soil were used to screen inhibitory activity against plant pathogenetic fungi by plate confrontation method.The antifungal and disease prevention effects of the fermentation products from representative strains were tested.The genome sequence of the target strain was analyzed by antiSMASH to evaluate the biosynthetic potential of secondary metabolites.Results Pathogenicity was confirmed by inoculating these four kinds of fungi on healthy Orah leaves,as a result,the symptoms of leaf spot appeared by infection of Boeremia exigua and Fusarium solani.Streptomyces strain M2020 showed good antifungal activities against four different plant pathogenic fungi,and its fermentation products were effective in the control of Orah fungal diseases caused by fungal strain C4.A total of 38 secondary metabolite biosynthetic gene clusters for biosynthesis of Polyketides,Non-Ribosomal Polypeptides and Terpene,etc.were identified in the genome of strain M2020.Conclusion The pathogenic fungi causing Orah leaf spot disease are Boeremia exigua and Fusarium solani.The Streptomyces strain M2020 can be a good choice for a biological control of Orah leaf spot disease.