【作者】 刘翠娟；

【导师】 安德荣；

【作者基本信息】 西北农林科技大学 ， 植物病理学， 2004， 硕士

【摘要】 本试验首次从陕西渭北旱原的不同类型的土壤中采集了80个土样，并从中分离到了130株放线菌，通过皿内平板初筛、液体发酵复筛和单孢分离，得到了一株对多种植物病原真菌颉颃性较好且遗传稳定的新菌株S-159-05。 为了提高从土壤中分离放线菌的效率，解决放线菌分离培养中细菌和真菌的污染问题，研究了重铬酸钾、放线菌酮、制霉素和多菌灵等4种抑制剂对细菌、真菌及放线菌的作用，在此基础上提出了用重铬酸钾作为高效、方便、廉价的放线菌杂菌抑制剂的新观点。重铬酸钾不仅对真菌、细菌有明显抑制作用，显著提高土壤放线菌的分离效果，还有刺激某些放线菌生长的作用。应用正交试验对S-159-05菌株的发酵培养基进行优化，确定其最佳配方为：黄豆饼粉2%,葡萄糖1%,淀粉1.5%,碳酸钙0.25%,磷酸氢二钾0.05%。并对其发酵条件进行了研究，结果发现，S-159-05菌株在发酵时间为72h，发酵培养基pH8.0,培养温度30-34℃，摇床转速150r/min时更适于活性物质的产生。S-159-05活性产物粗提物有较广的抑菌谱，对十几种病原菌有较强的抑制作用，尤其对水稻稻瘟、番茄灰霉、小麦根腐、烟草赤星等病原真菌及十字花科软腐等细菌抑制率较高，1mg/l药剂浓度的抑制率都在45%以上，这为广谱性生物农药的研制提供了理论依据。 通过对S-159-05的活性产物进行分离纯化，得到一个新的单组分化合物，其理化特性及结构初步分析结果为：此化合物紫外最大吸收波长为316nm；易溶于水、乙醇、甲醇等低级醇，几乎不溶于丙酮、乙醚、乙酸乙脂、乙酸丁脂、苯、三氯甲烷；熔点241-244℃；分子量为314；比旋光度：|(|D=__-32(_，用正丁醇：冰乙酸：水=4：1：1的条件下进行硅胶薄层层析，其Rf=0.40；与双缩脲试剂、品红和三氯化铁试剂反应为阴性，邻苯二甲酸-苯胺、茚三酮反应呈阳性，推知此新化合物为氨基糖苷酸类化合物，结合波谱分析结果将该化合物命名为1-(3(呋喃基-4(-酰胺)-2-N-氨基脒基葡萄苷，为新的氨基糖苷类农用抗生素的研发打下了良好基础。对S-159-05的活性产物颉颃机理的初步研究表明： S-159-05活性产物能强烈抑制病原菌菌丝的生长和孢子的萌发，引起病原菌菌丝扭曲或膨大、分枝增多、分枝顶端细胞壁破裂、原生质外溢，产生溶菌作用；分生孢子数减少，孢子萌发率降低、芽管畸形；温室条件下，S-159-05活性产物不仅能阻止番茄灰霉病菌菌丝入侵，对番茄灰霉病有预防作用，而且对已侵入的菌丝也有很好的治疗效果。更多还原

【Abstract】 In this study, 130 strains of actinomycetes were isolated from 80 soil samples which collected from various areas in the south-wei dry plateau.One genetically stable actinomycetes strain S-159-05was selected after comparison on inhibition activity of pathogenic fungi by means of selected in solied culture and fermental filtrate. In order to improve the efficiency of isolating actinomycetes from soil and solve the problem of contamination by bacteria and fungus when actinomycetes isolated and cultivated, the effect of K2Cr2O7, actidinoe, nystatin and curbenduzim on bacteria, fungus and actinomycetes was studied in this article. The result showed that K2Cr2O7 had distinct inhibition effect on soil fungus and bacteria, which can stimulates the growth of some actinomycetes, is a high effect, convenient and cheap inhibitor.The optimum fermentation medium of S-159-05 was screened on the base of NMM, and it was composed of glucose 1%, starch 1.5%, peanut powder 2%,CaC03 0.25%，K2HPO4 0.05%.The optimum fermentation condition which can produce the most effective fermental filtrate was 72 hour for rotational culture time,pH 8.0 of medium,30-34℃,rotational velocity is 150r/min.The crude active-product of S-159-05 has a wide-range of inhibition. 1mg/l materials has an inhibition efficiency above 45% against Pyricularia oryzae、Bipolaris sorokiniana、Botrytis cirerea、Alternaria alternata and Erwinia carotovora et al.The avtive product was isolated and purified,it was a mono-component,named An-2.The physico-chemical property was studied:it was soluble in water ,methyl alcohol ,ethyl alcohol and insoluble in acetone,ether,ethyl acetate,benzene,chloroform,petroleum etc. UVmax is 316nm;melt-point is 241-244℃; According to the physical and chemical properties as well as the UV , IR, 1H-NMR, and Mass spectrums,It was classified to glucoside-containing antibiotic family.The inhibiting range and mechanism of the active product of S-159-05 was studied. The result showed that the active product of S-159-05 has a strongly inhibition of the mycelia growth and spore germination which lead to the mycelia twisted and inflated, the abnormal branching increased, the mycelia wall broke and the material of the cell let out, the spores was reduced and deformation. The rate of germination and length of the tube was decreased. The active product of S-159-05 not only can prevent the invasion of Botrytis cirerea mycelia,and has a good control efficacy, showed a good curing efficacy.更多还原