水稻秸秆青贮饲料中可培养微生物多样性分析及优势乳酸菌的分离鉴定

doi:10.11733/j.issn.1007-0435.2014.03.022

草地学报 ›› 2014, Vol. 22 ›› Issue (3): 586-592.DOI: 10.11733/j.issn.1007-0435.2014.03.022

水稻秸秆青贮饲料中可培养微生物多样性分析及优势乳酸菌的分离鉴定

王彦苏¹, 张一凡¹, 严振亚¹, 丁成龙², 戴传超¹

1. 南京师范大学生命科学学院江苏省微生物与功能基因组学重点实验室江苏省微生物资源产业化工程技术研究中心, 江苏南京 210023;
2. 江苏省农业科学院畜牧研究所, 江苏南京 210014

收稿日期:2013-09-26 修回日期:2014-01-13 出版日期:2014-06-15 发布日期:2014-06-04
通讯作者: 戴传超
作者简介:王彦苏（1989- ），女，江苏徐州人，硕士研究生，主要从事微生物学研究，E-mail：wangyansu1990@163.com
基金资助:
江苏省农业自主创新项目“区域秸秆全量利用关键技术研究与应用推广”（CX（12）1002）资助

Isolation and Identification of Lactic Acid Bacteria, Analysis of Cultivable Microbial Diversity in Rice Straw Silage

WANG Yan-su¹, ZHANG Yi-fan¹, YAN Zhen-ya¹, DING Cheng-long², DAI Chuan-chao¹

1. Jiangsu Engineering and Technology Research Center for Industrialization of Microbial Resources; Jiangsu Key Laboratory for Microbes and Functional Genomics; College of Life Sciences, Nanjing Normal University, Nanjing, Jiangsu Province 210023, China;
2. Institute of Animal Science Jiangsu Academy of Agricultural Sciences, Nanjing, Jiangsu Province 210014, China

Received:2013-09-26 Revised:2014-01-13 Online:2014-06-15 Published:2014-06-04

摘要/Abstract

摘要：

为探究影响水稻（Oryza sativa L.）秸秆青贮质量的腐败菌种类并筛选适合其青贮的优质乳酸菌，研究了青贮前后水稻秸秆中可培养微生物的变化并分离鉴定2株乳酸菌。使用稀释平板涂布法将青贮前后可培养微生物计数并分离纯化，限制性酶切分析ARDRA技术分析分离纯化后的微生物多样性，结合ARDRA和RAPD-PCR技术对青贮中优势乳酸菌进行分离鉴定。结果表明：青贮前后好氧或兼性厌氧细菌菌落总数由7.6×10⁷cfu·g^-1下降到2.43×10⁶cfu·g^-1，真菌菌落数变化较为明显，从4.43×10⁵cfu·g^-1下降到86 cfu·g^-1，乳酸菌菌落数从4.16×10⁵cfu·g^-1上升到6.61×10⁶ cfu·g^-1。ARDRA聚类分析及16SrDNA测序显示青贮前存在腐败细菌芽孢杆菌（Bacillus）和致病菌阴沟肠杆菌（Enterobacter），青贮后腐败细菌主要是芽孢杆菌，腐败真菌主要为青霉菌属（Penicillium）。筛选出的2株乳酸菌分别属于乳杆菌属（Lactobacillus casei）和片球菌属（Pediococcus ethanolidurans），需进一步验证是否可作为青贮饲料添加剂。水稻秸秆青贮后依然有腐败菌的存在，需进一步提高其青贮发酵工艺，防治有氧恶化。

关键词: 水稻秸秆青贮, 腐败菌, 乳酸菌, 分离鉴定

Abstract:

The diversities of cultivable microorganisms in both pre-and post-ensiled straws were investigated and two strains of lactic acid bacteria from post-ensilage straw were isolated. According to morphological characteristics of bacterial colonies, bacterial strains subjected to amplified ribosomal DNA restriction analysis (ARDRA). Then lactic acid bacteria were separated and identified from rice straw silage by ARDRA and RAPD-PCR technology. The results show that the number of aerobic bacteria decreases from 7.6×10⁷cfu·g^-1to 2.43×10⁶cfu·g^-1, the populations of fungi significantly decrease from 4.43×10⁵ cfu·g^-1 to 86 cfu·g^-1, whereas the number of lactic acid bacteria increases from 4.16×10⁵cfu·g^-1 up to 6.61×10⁶cfu·g^-1after ensiled. There are not only spoilage organisms (Bacillus), but also pathogen pathogenic bacterium (Enterobacter) in pre-ensiled straw. After ensiled, the mainly putrefying bacterium is Bacillus and corruption fungi consist of Penicillium and Fusarium. Two strains of isolated lactic acid bacteria are Lactobacillus casei and Pediococcus ethanolidurans. There are still spoilage organisms in post-ensiled straw. Therefore, fermentation process should be improved to prevent aerobic deterioration.

Key words: Rice straw silage, Spoilage organisms, Lactic acid bacteria, Isolation and identification

中图分类号:

S816.53

王彦苏, 张一凡, 严振亚, 丁成龙, 戴传超. 水稻秸秆青贮饲料中可培养微生物多样性分析及优势乳酸菌的分离鉴定[J]. 草地学报, 2014, 22(3): 586-592.

WANG Yan-su, ZHANG Yi-fan, YAN Zhen-ya, DING Cheng-long, DAI Chuan-chao. Isolation and Identification of Lactic Acid Bacteria, Analysis of Cultivable Microbial Diversity in Rice Straw Silage[J]. Acta Agrestia Sinica, 2014, 22(3): 586-592.

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水稻秸秆青贮饲料中可培养微生物多样性分析及优势乳酸菌的分离鉴定

Isolation and Identification of Lactic Acid Bacteria, Analysis of Cultivable Microbial Diversity in Rice Straw Silage

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