烟草化感自毒物质降解复合菌剂的优化及应用效果评价

Optimization and evaluation of a compound bacterial agent degrading autotoxins of tobacco

摘要:

烟草(Nicotiana tabacum)长期连作导致土壤中烟草自身分泌的化感自毒物质的积累，加剧了烟草青枯病(tobacco bacterial wilt, TBW)的发生，严重威胁烟草的生产并造成了巨大的经济损失。【目的】获得对长期连作烟田青枯病具有显著防治效果的复合菌剂。【方法】利用筛选到的烟草化感自毒物质降解菌株构建复合菌剂，通过正交试验和单因素试验对菌种配比和助剂进行优化，对其在温室盆栽和连作烟田生产的应用效果进行评价。【结果】正交试验结果表明，复合菌剂中芽孢杆菌(Bacillus sp.) NO1、布鲁氏菌(Brucella sp.) NO8、芽孢杆菌(Bacillus sp.) NO9和NO10的最佳菌种配比为1:3:4:2；单因素试验结果表明最佳的载体为白炭黑；最佳润湿剂和分散剂分别为六偏磷酸钠和丁基萘磺酸钠，其最佳比例为1:1，最佳用量各为2%；稳定剂甘油的最佳浓度为1.0%。盆栽试验结果显示，复合菌剂对6种化感自毒物质的降解率和对青枯病的抑制率均达到78%以上。大田试验结果显示，100倍稀释的复合菌剂对连作15年烟田中的6种化感自毒物质具有显著的降解效果，并且能明显增加烟株的株高、茎围、腰叶长和腰叶宽，有效促进了烟草的生长发育；同时还调节了烟草根围微生物的丰度，显著抑制了狭义梭菌属1 (Clostridium_sensu_stricto_1)、劳尔氏菌属(Ralstonia)和纤维单胞属(Cellulomonas)等的相对丰度，大幅提高了戴沃斯菌属(Devosia)、黄杆菌属(Flavobacterium)和鞘氨醇单胞属(Sphingomonas)等的相对丰度。青枯病的发病率和病情指数也从处理前的92.22%和48.19%分别降低到18.15%和9.52%，防效超过了80.00%。【结论】本研究优化的复合菌剂显著降低了长期连作烟田青枯病的病情指数，可为长期连作烟田青枯病的防控提供解决方案。

Abstract:

[Background] Long-term continuous cropping of tobacco (Nicotiana tabacum) leads to the accumulation of autotoxins, which aggravates the incidence of tobacco bacterial wilt (TBW) caused by Ralstonia solanacearum and causing great economic losses of tobacco production. [Objective] To develop a compound bacterial agent capable of controlling TBW in the field with long-term continuous cropping of tobacco. [Methods] We formulated compound bacterial agents with screened strains capable of degrading autotoxins. Orthogonal design and single factor experiments were employed to optimize the strain ratio and the dosages of additives. The inhibition performance of the compound bacterial agent on TBW was evaluated in a greenhouse and in a field with continuous cropping of tobacco for 15 years. [Results] Orthogonal experiments showed that the optimal ratio of Bacillus sp. NO1, Brucella sp. NO8, Bacillus sp. NO9, and Bacillus sp. NO10 in the compound bacterial agent was 1:3:4:2. Single factor experiments showed that the best vector was silica. The best wetting agent and dispersant were sodium hexametaphosphate (SHMP) and sodium butylnaphthalene sulfonate (SBNS), respectively, which were optimized to be added at the dosages of 2%. The optimal concentration of the stabilizer glycerin was 1.0%. Pot experiments showed that both the degradation rate of six autotoxins and the inhibition rate of TBW by the compound bacterial agent reached over 78%. The results of the field experiment showed that the compound bacterial agent diluted by 100 folds had significant degradation effects on six autotoxins in the tobacco field with continuous cropping for 15 years. Moreover, the agent significantly increased the height, stem circumference, and waist leaf length and width of tobacco plants, thus promoting the growth and development of tobacco. In addition, the agent regulated the rhizosphere microbiota of tobacco, as manifested by the decreased relative abundance of Clostridium_sensu_stricto_1, Ralstonia, and Cellulomonas and the increased relative abundance of Devosia, Flavobacterium, and Sphingomonas. The agent decreased the incidence rate and disease index of TBW from 92.22% and 48.19% to 18.15% and 9.52%, respectively, with a control effect of 80%. [Conclusion] The optimized compound bacterial agent significantly reduces the incidence rate and disease index of TBW in the field with long-term continuous cropping of tobacco, which provides a solution for the prevention and control of TBW.