马盲肠纤维素分解菌的分离鉴定与酶活分析

Isolation and Identification of Cellulose Degrading Bacteria from Horse Cecum and Analysis of Enzyme Activity

利用羧甲基纤维素钠初筛培养基和LB发酵复筛培养基从马盲肠内容物中筛选具有高纤维素酶活性的菌株,通过16S rDNA测序分析对分离的纤维素降解菌进行分类鉴定并构建系统进化树,然后测定其酶活性、生长特征及理化特性,研究马盲肠纤维素分解菌对提高饲料中纤维素利用率的影响。结果表明,从马盲肠内容物中分离出66株高产纤维素酶的菌株,经过16S测序鉴定为10个物种,除了大肠杆菌(E.coli)之外,其余9种均为芽孢杆菌,分别是短小芽孢杆菌(Bacillus pumilus)、高地芽孢杆菌(Bacillus altitudinis)、解淀粉芽孢杆菌(Bacillus amyloliquefaciens)、枯草芽孢杆菌(Bacillus subtilis)、莫来芽孢杆菌(Bacillus mojavensis)、沙福芽孢杆菌(Bacillus safensis)、同温层芽孢杆菌(Bacillus stratosphericus)、维尔兹芽孢杆菌(Bacillus velezensis)、漳州芽孢杆菌(Bacillus zhangzhouensis)。其中酶活最高的达到14.81 U/mL,最低的为1.91 U/mL。本研究从马盲肠内容物中分离出了酶活性高的纤维素分解菌66株,为进一步研究马盲肠消化机制提供了思路,并为饲料添加菌奠定了重要的基础。

This study used sodium carboxymethyl cellulose primary screening medium and LB fermentation secondary screening medium to select strains with high cellulase activity from the caecal contents.Classify and identify the isolated cellulose-degrading bacteria through 16S rDNA sequencing analysis and construct a phylogenetic tree.The DNS method was used to determine their enzyme activity,and the top 5 cellulose-degrading bacteria with higher enzyme activity were selected for the determination of growth characteristics and physical and chemical characteristics.A total of 66 cellulase-producing strains were isolated from the caecal contents of horse.The molecular identification determined 10 strains,except for Escherichia coli,the rest are Bacillus sp.,including Bacillus pumilus,Bacillus altitudinis,Bacillus amyloliquefaciens,Bacillus subtilis,Bacillus mojavensis,Bacillus safensis,Bacillus stratosphericus,Bacillus velezensis,Bacillus zhangzhouensis.Among them,the highest enzyme activity reached 14.81 U/mL,and the lowest was 1.91 U/mL.In this study,66 cellulolytic bacteria with high enzyme activity were isolated from the contents of horse cecum for the first time,which provided ideas for further research on the digestive mechanism of horse cecum and laid an important foundation for adding bacteria to feed.