青贮饲料微生态系统发酵过程中微生物群落演替机制研究及优良菌株筛选

Silage fermentation process is the process of microbial population’s growth phase change and microflora succession in silage microecosystem. The fermentation quality is effected by microbial population growth and microflora succession in silage microecosystem during fermentation. Understanding the microbial population growth, the relationship between populations, and the community succession in silage microecosystem during fermentation will provide important scientific basis for regulating microecology of silage and producing better silage. In this applied proposal, whole plant corn silage will be research material. The microbial population’s growth phase is defined by determining the count of microbial population (lactic acid bacteria, escherichia coli, yeast, bacteria and fungi) in silage microecosystem during fermentation. Adopting PacBio high throughout 16S/ITS rRNA sequence reveals the composition change law of microflora and the microbial diversity in silage microecosystem during fermentation. It verifies the correlation ship between microbial populations, and responding mechanism of microflora and matrix/fermentation products, and clears the succession mechanism of microflora using mathematical statistics and ecology multivariate statistics in silage microecosystem during fermentation. Meanwhile, to screen the yeast that has strong ability to multiply, less acid-resistant ability and short stationary phase, and the lactic acid bacteria that multiplies earlier and has strong ability to multiply and produce acid from silage regulates the growth phase of lactic acid bacteria, the time of aerobic fermentation and the microflora succession in silage microecosystem during fermentation. Those will lay the theoretical basis for studying regulating microecology of silage and the material basis for developing silage probiotics.

青贮饲料发酵过程是青贮微生态系统中微生物种群增长阶段变化和群落演替过程，而二者又影响着青贮的发酵品质；了解青贮饲料微生态系统发酵过程中微生物种群增长、种群间关系和群落演替机制可为青贮的微生态学调控及优质青贮调制提供重要的科学依据。本项目以全株玉米青贮饲料为研究对象，通过动态测定乳酸菌、大肠杆菌、酵母菌、细菌和真菌等种群数量，明确青贮微生态系统发酵过程中各微生物种群增长阶段；采用PacBio高通量16S/ITS rRNA全长测序（细菌/真菌种水平测定）技术，揭示微生物多样性和群落变化规律；利用数理统计和生态学多元统计分析技术，探明微生物种群间、群落与基质和发酵产物间的相互关系并明确微生物群落演替机制；同时，筛选增殖能力强、耐酸性弱且稳定期短的酵母菌和启动快、产酸与增殖能力强的乳酸菌，对乳酸菌种群增长阶段、有氧发酵和群落演替进行调控；为青贮生态学调控的研究及微生态制剂的研发奠定理论和物质基础。

青贮饲料发酵过程是青贮微生态系统中微生物种群增长阶段变化和群落演替过程，而二者又影响着青贮的发酵品质；了解青贮饲料微生态系统发酵过程中微生物种群增长、种群间关系和群落演替机制可为青贮的微生态学调控及优质青贮调制提供重要的科学依据。本项目以全株玉米青贮饲料为研究对象，通过动态测定乳酸菌、大肠杆菌、细菌和真菌等种群数量，明确青贮微生态系统发酵过程中各微生物种群增长阶段；采用高通量测序技术，揭示微生物多样性和群落变化规律；利用数理统计和生态学多元统计分析技术，探明微生物种群间、群落与基质和发酵产物间的相互关系并明确微生物群落演替机制；同时，筛选产酸与增殖能力强的乳酸菌，对青贮饲料发酵过程进行调控。项目组从发酵过程中的全株玉米青贮饲料中分离出乳酸菌10株，分别为植物乳杆菌（2株）、干酪乳杆菌（1株）、短乳杆菌（1株）、戊糖片球菌（1株）、乳酸片球菌（1株）、肠膜明串珠菌（2株）、蒙氏肠球菌（2株）。青贮饲料发酵的前3h，为所有微生物的加速期；3 h – 24 h，乳酸菌的指数期；24 h – 3 d，乳酸菌的稳定期；3 h – 3 d，细菌和酵母菌的指数期；3 d – 60 d，为所有微生物的衰弱期。全株玉米青贮饲料发酵过程的微生物演替分为四个阶段，分别为：乳酸菌为小众菌群（0-5 h），Weissella、Lactococcus和Leuconostoc起主导作用（5 h – 24 h），Lactobacillus起主导作用且丰富度升高（24 h – 10 d），Lactobacillus起主导作用且丰富度降低 (10 d – 60 d)。Weissella、Lactococcus和Leuconostoc等异型发酵乳酸菌的活动诱发了Lactobacillus的快速增值。Lactobacillus的种对全株玉米青贮饲料发酵品质的形成和保持起到决定性作用。第四阶段所有微生物活性均收到抑制。从全株玉米青贮饲料中筛选出优良乳酸菌2株，分别为Lactobacillus plantarum和Lactobacillus brevis，具有较强的产酸能力和耐酸能力。添加两株乳酸菌，可促进至青贮饲料发酵和加快微生物演替。为青贮饲料微生物演替调控技术研发奠定理论和物质基础。