黄瓜枯萎病生防菌HD-087产抗菌物质条件的 优化及抑菌作用初探

Fermentation conditions for the biocontrol strain HD-087 against cucumber Fusarium wilt and it’s antibacterial effect

摘要:

【目的】优化比基尼链霉菌HD-087摇瓶发酵条件, 提高菌株发酵液对黄瓜枯萎病菌HU-M的抑制率, 并通过拮抗试验初步评价发酵液的抑菌作用效果。【方法】采用单因素筛选及正交试验对HD-087的发酵培养基和发酵条件进行优化。经发酵液处理后, 光学显微镜观察HU-M菌丝形态和孢子萌发抑制率, 测定HU-M菌丝电导率。【结果】改进的发酵培养基配方为: 淀粉1.00%、黄豆粉0.80%、酵母粉0.12%、CaCO3 0.40%。对发酵条件的研究表明: pH为6.8, 180 r/min、28 °C条件下, 250 mL三角瓶装液量为40 mL, 接种1 mL种龄为2 d的种子, 发酵5 d为最佳培养条件。抑菌结果表明, HD-087产抗菌物质能造成病原菌HU-M菌丝细胞质渗漏, 菌丝畸形, 分生孢子萌发受抑制, 5倍发酵稀释液孢子抑制率达72.1%; 除此之外还能引起菌丝电解质渗漏, 造成菌丝细胞膜受损。【结论】优化后的摇瓶发酵条件能提高生防菌HD-087发酵液抑菌效果, 并且发酵液可破坏细胞膜明显抑制病原菌HU-M生长, 具有较大开发应用潜力。

Abstract:

[Objective] In order to improve the inhibitory rate of fermentation broth of Streptomyces bikiniensis strain HD-087 against Fusarium oxysporum strain HU-M, the shake-flask fermentation conditions of biocontrol agent HD-087 were optimized. Restraining effectiveness of broth towards HU-M has been tested at the same time. [Methods] The liquid fermentation medium and conditions of HD-087 were optimized by single factor experiments and orthogo-nal experiments. After inoculation with broth, HU-M hyphae morphology and germination of conidia were then examined under the light microscope, and the relative conductivity of HU-M hyphae was measured using a conductivity meter. [Results] The composition of the optimal medium was 1.00% (W/V) starch, 0.80% (W/V) soybean flour, 0.12% (W/V) yeast extract, 0.40% (W/V) CaCO3. The optimal fermentation conditions were the combination of temperature 28 °C, initial pH 6.8, 180 r/min, incubation time 5 d, 1 mL inoculum volume and medium volume 40 mL/250 mL. The result of antibacterial effect can be concluded as follows: abnor-mality of pathogen hyphae and leakage of cytoplasm after treated with broth; meanwhile the inhibitive rate to conidia germination reached 72.1% when the broth diluted by 5 times. In ad-dition to that, exosmosis of hyphae content, conductance rate increased. [Conclusion] All these changes indicate that optimal shake-flask fermentation conditions can improve the in-hibitory activity of HD-087 broth against F. oxysporum HU-M, and the broth has been proved efficient for suppression of HU-M by severely destroying cell membrane structure, it’s of great potential application value.