应用培养组学分离鉴定牛源芽孢杆菌及其系统发育性分析

The Analysis of Bovine-derived Spores and their System Developmental Properties Identified by the Separation of Culture Histology

在对动物肠道微生物领域的研究中,应用培养组学进行研究逐渐被人们所熟知。同时,应用其对肠道内益生菌的研究应用也越发广泛。实验基于培养组学对荷斯坦奶牛直肠粪便样品分进行分离纯化,得到芽孢杆菌菌株。实验使用6种培养方法分离培养荷斯坦奶牛肠道内的芽孢杆菌,并在纯化扩繁后对其进行形态特征、培养特性、扩增16S rRNA测序鉴定细菌菌属。结果表明:利用BLAST软件将14株(L1-L14)菌株测序后得到的16S r RNA序列与NCBI上Gen Bank中发布的已知芽孢杆菌参考株进行序列同源性分析,初步鉴定为14株芽孢杆菌。14个分离株与参考菌株的同源性为99.93%~100%。经MEGA-X软件操作系统发育树分析,得到的分离菌株分别为:2株蜡样芽孢杆菌,2株沙福芽孢杆菌,5株地衣芽孢杆菌,2株短小芽孢杆菌,1株海洋沉积物芽孢杆菌,1株高山芽孢杆菌,1株同温层芽孢杆菌;其中地衣芽孢杆菌和短小芽孢杆菌属于可添加饲喂功能益生菌。实验为未来对奶牛肠道内益生菌相关研究,奶牛肠道健康防控,开发有关益生菌类的微生态制剂等提供有效的实验支持。

In the research on the field of animal gut microbes,the application of culturomics for research had gradually become wellknown.At the same time,the application of it to the research and application of probiotics in the intestinal tract had become more extensive.The rectal feces samples of Holstein cows were separated and purified based on culture omics to obtain Bacillus strains.Six culture methods were used to isolate and cultivate Bacillus in the intestine of Holstein dairy cows,and after purification and propagation,the morphological characteristics,culture characteristics,and 16S rRNA sequencing of amplified bacteria were used to identify bacterial genera.The results showed that the 16S rRNA sequence obtained after sequencing 14 strains(L1-L14) using BLAST software was analyzed by sequence homology with the known Bacillus reference strains published in Gen Bank on NCBI,and14 strains of Bacillus were preliminarily identified.The homology between the 14 isolates and the reference strain was 99.93% to100%.Through MEGA-X software operating system development tree analysis,the isolated strains obtained were:2 Bacillus cereus,2Bacillus safensis,5 Bacillus licheniformis,2 Bacillus pumilus,1 Bacillus oceanisediminis,1 Bacillus altitudinis and 1 Bacillus stratosphericus;Bacillus licheniformis and Bacillus pumilus were probiotics that could add feeding function.The experiment provided effective experimental support for future research on probiotics in the intestinal tract of dairy cows,prevention and control of dairy cow intestinal health,and the development of probiotics-related microecological preparations.