核桃黑斑病生防链霉菌YNF36的鉴定及其抑菌活性测定

【背景】由野油菜黄单胞菌(Xanthomonas campestris)和成团泛菌(Pantoea agglomerans)侵染引起的核桃黑斑病是一种严重的细菌性病害，给核桃产业带来了极大损失。【目的】从根际土壤中筛选出对核桃黑斑病病原菌野油菜黄单胞菌和成团泛菌均具有拮抗效果的放线菌菌株，可作为创制生防菌剂的出发菌株。【方法】采用稀释涂布法、平板对峙法和改良牛津杯法筛选拮抗菌株，通过形态学特征、生理生化特性和16S rRNA基因序列分析进行鉴定，测定无菌发酵液抗菌谱，离体叶片试验验证其对核桃黑斑病的防治效果。【结果】筛选到一株对2种病原菌均有较强拮抗作用的放线菌菌株YNF36。经形态学特征观察、生理生化特性试验及16S rRNA基因序列分析，将菌株YNF36鉴定为沙场链霉菌(Streptomyces arenae)。该菌株在SYP培养基上产量最高，抑菌活性最强，其无菌发酵液对金黄色葡萄球菌、大肠杆菌、黑曲霉、白色念珠菌、枯草芽孢杆菌、铜绿假单胞菌、蜡样芽孢杆菌这7种指示菌，以及链格孢菌、黑腐皮壳菌、胶孢炭疽菌、灰葡萄孢菌、黄褐孢霉菌、辣椒刺盘孢菌、腐皮镰孢菌这7种植物病原菌均有抑制作用，抗菌谱广。发酵液原液对离体叶片上的由野油菜黄单胞菌和成团泛菌造成的核桃黑斑病防效分别为75.69%和62.39%。【结论】沙场链霉菌YNF36补充了一种防治核桃黑斑病的生防材料，具有良好的开发价值和应用前景。

Identification and activity of Streptomyces YNF36 against walnut blight

Background] Walnut blight, caused by Xanthomonas campestris and Pantoea agglomerans, is a bacterial disease resulting in severe damage to walnut tree and substantial economic loss. Objective] To isolate and screen out the actinomycete strains with inhibitory activity on these two pathogens from the rhizosphere soil of walnut tree for the development of biocontrol agents. Methods] The actinomycetes were isolated by spread plate method and further screened by plate confrontation method and improved Oxford-cup test. The target strain was identified based morphological, physiological, and biochemical properties, as well as 16S rRNA gene sequence. The antibacterial spectrum and biocontrol effect in vitro of the cell-free fermentation broth were evaluated. Results] An actinomycete strain YNF36 with strong antagonistic effect on both pathogens was screened out and identified as Streptomyces arenae. The strain YNF36 showed antimicrobial activity to seven indicator bacterial species: Staphlococcus aureus, Escherichia coli, Aspergillus niger, Monilia albicans, Bacillus subtilis, Pseudomonas aeruginosa, and Bacillus cereus. Furthermore, it had inhibitory activity against seven plant pathogens: Alternaria alternata, Valsa mali, Colletotrichum gloeosporioides, Botrytis cinerea, Fulvia fulva, Colletotrichum capsica, and Fusarium solani. The strain YNF36 had the largest yield and the strongest antimicrobial activity on SYP medium. In addition, the biocontrol experiment in vitro showed that the cell-free fermentation broth of the strain YNF36 had the biocontrol effects of 75.69% and 62.39% against X. campestris and P. agglomerans, respectively. Conclusion] The strain YNF36 could be considered as the actinomycete resource with biocontrol potential against walnut blight and demonstrates a wonderful prospect for development and application.