益生芽孢杆菌固体发酵工艺及其微生物肥料应用研究
发布时间：2018-10-30 12:42
【摘要】：目前我国传统农业生产中长期大量施用N、P、K元素肥料,虽然一定程度上缓解了我国粮食安全问题,但也由此产生了一系列环境问题,如土壤酸化、板结、盐渍化等。适当施用生物菌肥可以提高肥效、防治病害和修复土壤,是实现农业可持续发展的必要途径之一。生产低成本、高益生菌含量的微生物肥料具有广阔的市场前景,也非常符合我国绿色农业的发展策略。传统益生菌液体发酵工艺因其工艺复杂、成本高,在一定程度上限制了其在农业领域的应用,因此开发低成本、适用于农业生产的益生菌剂及其衍生产品,是推动微生物肥料领域技术进步的关键环节。本文首先对4株益生芽孢杆菌进行了形态学和分子生物学的鉴定及特性分析。随后采用固体发酵手段研究开发低成本、高密度益生芽孢杆菌生产工艺。通过单因素和正交工艺优化,最终确定Bacillus subtilis L7固体发酵工艺条件为：糠醛渣、废糖液和麦麸添加量分别为85%、10%和5%,初始水分60%,温度30℃,初始pH 7.0,接种量5×106CFU/g,翻堆频率为1次/d,L7发酵水平达到(活菌数和芽孢数分别为)2.48×1010CFU/g和2.40×1010CFU/g,芽孢率达到96.7%。随后对L7固体发酵工艺进行了100kg规模的中试放大,发酵后活菌数达到3.26×1010CFU/g,菌活水平比小试提高了31.1%,且发酵时间由6天缩短为3天。同时对其他三种益生芽孢杆菌进行固体发酵小试实验。结果表明Bacillus licheniformis D6、Bacillus megaterium J3和Bacillus mucilaginosus G14发酵后总活菌数分别为2.48×1010CFU/g、1.02×1010CFU/g和8.08×109CFU/g,发酵水平高于相应的液体工艺,且本方法成本低廉,具有明显的竞争优势。随后我们将固体发酵菌剂与复合肥(NPK 16：16：8)复配制得益生菌含量为1.25×108 CFU/g的微生物肥料；以苏州青油菜(Brassica campestris L. Sp)为实验样本开展盆栽实验,研究微生物肥料对其作用效果。结果表明,所制备的复合微生物肥料B3可以有效的提高实验作物根系和茎叶干重,与对照相比分别提高46.1%和34.1%,以上结果说明该复合微生物肥料能够显著促进植株生长。最后我们采用DGGE方法对盆栽作物根际土壤菌群进行了分析。结果表明,添加30天后,微生物肥料对细菌和真菌种类和种群结构都有明显的影响。其中复合微生物肥料B3组细菌丰富度和多样性指数与对照组相比分别降低206%和47.7%,真菌丰富度和多样性指数分别降低233%和47.8%。添加60天后,微生物肥料对土壤中真菌结构仍有显著影响。其中复合微生物肥料B3与对照组相比,真菌丰富度和多样性指数分别降低98.2%和26.7%。上述结果说明经固体发酵工艺所开发的复合微生物肥料能够明显抑制土壤真菌,在防治作物各种土传真菌病害方面具有广阔的开发前景。
[Abstract]:At present, the traditional agricultural production in our country has applied a large amount of fertilizer with NbPK element for a long time. Although it has alleviated the problem of food security in China to some extent, it has also resulted in a series of environmental problems, such as soil acidification, cementation, salinization and so on. Proper application of biofilm fertilizer can improve fertilizer efficiency, control diseases and repair soil, which is one of the necessary ways to realize the sustainable development of agriculture. The production of low cost and high probiotic microbial fertilizer has a broad market prospect and is also in line with the development strategy of green agriculture in China. The traditional liquid fermentation technology of probiotics, because of its complexity and high cost, limits its application in the field of agriculture to a certain extent, so it can develop low cost probiotics and its derivative products, which are suitable for agricultural production. It is the key link to promote the technical progress in the field of microbial fertilizer. In this paper, four strains of Bacillus probiotics were identified and characterized by morphological and molecular biology. Then the production process of low cost and high density probiotic bacillus was studied by solid-state fermentation. Through the optimization of single factor and orthogonal process, the technological conditions of Bacillus subtilis L7 solid fermentation were determined as follows: furfural residue, waste sugar solution and wheat bran were added to 85% and 5% respectively, the initial moisture content was 60%, the temperature was 30 鈩，
本文编号：2300042
[Abstract]:At present, the traditional agricultural production in our country has applied a large amount of fertilizer with NbPK element for a long time. Although it has alleviated the problem of food security in China to some extent, it has also resulted in a series of environmental problems, such as soil acidification, cementation, salinization and so on. Proper application of biofilm fertilizer can improve fertilizer efficiency, control diseases and repair soil, which is one of the necessary ways to realize the sustainable development of agriculture. The production of low cost and high probiotic microbial fertilizer has a broad market prospect and is also in line with the development strategy of green agriculture in China. The traditional liquid fermentation technology of probiotics, because of its complexity and high cost, limits its application in the field of agriculture to a certain extent, so it can develop low cost probiotics and its derivative products, which are suitable for agricultural production. It is the key link to promote the technical progress in the field of microbial fertilizer. In this paper, four strains of Bacillus probiotics were identified and characterized by morphological and molecular biology. Then the production process of low cost and high density probiotic bacillus was studied by solid-state fermentation. Through the optimization of single factor and orthogonal process, the technological conditions of Bacillus subtilis L7 solid fermentation were determined as follows: furfural residue, waste sugar solution and wheat bran were added to 85% and 5% respectively, the initial moisture content was 60%, the temperature was 30 鈩，
本文编号：2300042
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