三株植物促生木霉的固体发酵工艺优化

Optimization of solid fermentation process of three Trichoderma strains with plant growth-promoting effects

摘要:

【背景】木霉是广泛分布于自然界中的一类真菌，能产生多种酶类和次生代谢产物，具有促进植物生长、提高土壤肥力、拮抗多种土传病原菌等作用。【目的】优化3株植物根际促生真菌(长枝木霉MD30、桔绿木霉JS84及贵州木霉NJAU4742)的固体发酵条件，探究不同发酵条件对木霉产孢量的影响，为木霉菌的生产提供参考。【方法】采用单因素试验和响应面法，对3种木霉在不同发酵条件下的产孢量进行测定并优化，分析了氮源添加、初始pH、物料厚度、接种量、温度等因子对固体发酵的影响。【结果】单因素试验表明，长枝木霉MD30、桔绿木霉JS84与贵州木霉NJAU4742固体发酵时，最佳发酵温度均为28℃、最优木霉菌液接种量均为10%、物料发酵厚度均为3.0cm，但最佳的初始物料pH与氨基酸水解液添加量有所不同，其中，长枝木霉MD30与贵州木霉NJAU4742发酵最佳的初始pH值为5.0，而桔绿木霉JS84为3.0；长枝木霉MD30与贵州木霉NJAU4742发酵最佳的氨基酸水解液添加量为10%，而桔绿木霉JS84为5%。通过试验分析，确定初始pH、物料厚度、温度为影响产孢量的3个重要因素。响应面分析得到最佳发酵条件：长枝木霉MD30固体发酵最佳条件为初始pH值为5.0，物料厚度3.0cm，温度27.4℃，在此优化条件下每克干重的固体产物中孢子数量高达6.3×10⁹CFU；桔绿木霉JS84固体发酵最佳条件为初始pH值为3.0，物料厚度3.0cm，温度28.8℃，在此优化条件下每克干重的固体产物中孢子数量高达6.9×10⁹CFU；贵州木霉NJAU4742固体发酵最佳条件为初始pH值为5.0，物料厚度3.3cm，温度26.7℃，在此优化条件下每克干重的固体产物中孢子数量高达5.1×10⁹CFU。【结论】响应面优化试验能提高试验效率，增大木霉产孢量，降低生产成本，有利于田间应用和菌剂的商业化生产。

Abstract:

[Background] Trichoderma is a kind of fungus widely distributed in nature, which produces a variety of enzymes and secondary metabolites, showing many benefits including promoting plant growth, improving soil fertility, and antagonizing various soil-borne pathogens. [Objective] To optimize the solid fermentation conditions of three plant growth-promoting fungi (Trichodermalongibrachiatum MD30, T. citrinoviride JS84, and T. guizhouense NJAU4742), the effects of different fermentation conditions on the spore production of Trichoderma spp. were explored to provide references for Trichoderma spp. production. [Methods] In this study, the spore production of three Trichoderma spp. under different fermentation conditions was measured and optimized by the single factor experiment and the response surface methodology. The effects of the addition of nitrogen source, initial pH, material thickness, inoculation amount, and temperature on solid fermentation were analyzed. [Results] The single factor experiment showed that the optimal fermentation temperature of T. longibrachiatum MD30, T. citrinoviride JS84, and T. guizhouense NJAU4742 was 28 ℃, and the optimal amount of Trichoderma liquid inoculum was 10% with 3.0 cm material fermentation thickness. However, the optimal initial pH of the material and the addition amount of amino acid hydrolyzate were different among these three strains. In detail, the optimal initial pH value of T. longbranchus MD30 and T. guizhouense NJAU4742 for solid fermentation was 5.0, while that of T. citrinoviride JS84 was 3.0. The optimum amino acid hydrolyzate addition amount for solid fermentation of T. longbranchus MD30 and T. guizhouense NJAU4742 was 10%, while that of T. citrinoviride JS84 was 5%. Based on the experimental analysis, the initial pH, material thickness, and temperature were selected as three top important factors in the determination of the spore production. Therefore, the optimal fermentation conditions for solid fermentation of these three strains were obtained based on the response surface analysis. The optimal conditions for solid fermentation of T. longbranchus MD30 were the initial pH value of 5.0, the material thickness of 3.0 cm, and thetemperature of 27.4 ℃. Under this optimized condition, the number of spores per gram dry weight of the solid fermentation product was as high as 6.3×10⁹ CFU. The optimal conditions for solid fermentation of T. citrinoviride JS84 were the initial pH value of 3.0, the material thickness of 3.0 cm, and the temperature of 28.8 ℃. Under this optimized condition, the number of spores per gram dry weight of the solid fermentation product was as high as 6.9×10⁹ CFU. The optimal conditions for solid fermentation of T. guizhouense NJAU4742 were the initial pH value of 5.0, the material thickness of 3.3 cm, and the fermentation temperature of 26.7 ℃. Under this optimized condition, the number of spores per gram dry weight of the solid fermentation product was as high as 5.1×10⁹ CFU. [Conclusion] Response surface optimization experiment improves the experimental efficiency, increases the spore production of Trichoderma spp., reduces the production cost, and is conducive to field application and commercial production of bacterial agents.