阿南原等跳肠道细菌的分离鉴定及降解纤维素细菌的筛选

摘要

[目的]本研究旨在确定阿南原等跳Proisotoma ananevae成虫肠道细菌的组成, 并筛选降解纤维素细菌.[方法]运用传统培养与16S rDNA测序相结合方法, 分离鉴定阿南原等跳成虫肠道内可培养细菌;通过羧甲基纤维素钠筛选培养基 (CMC) 筛选能够降解纤维素的细菌, 并采用3, 5-二硝基水杨酸 (DNS) 法测定不同pH (5.0～9.0) 下的纤维素酶活力.[结果]从阿南原等跳成虫肠道共分离到20种不同的菌株, 隶属于厚壁菌门 (Firmicutes) 、变形菌门 (Proteobacteria) 和放线菌门 (Acinobacteria) 3门的10属, 即葡萄球菌属Staphylococcus, 芽孢杆菌属Bacillus, Terribacillus, Advenella, 赖氨酸芽孢杆菌属Lysinibacillus, 节杆菌属Arthrobacter, 肠杆菌属Enterobacter, Glutamicibacter, 无色杆菌属Leucobacter和不动杆菌属Acinetobacte;另有1株未鉴别细菌.10株纤维素降解细菌分别隶属于厚壁菌门 (Firmicutes) 和放线菌门 (Acinobacteria) 2门的6属, 即无色杆菌属Leucobacter, 芽孢杆菌属Bacillus, Terribacillus, 赖氨酸芽孢杆菌属Lysinibacillus, 节杆菌属Arthrobacter和Glutamicibacter.酶活力测定结果显示所有纤维降解素菌株在pH 7.0～9.0之间纤维素酶活性均相对较高, 且pH 8.0时酶活力最高.[结论]结果说明, 阿南原等跳成虫肠道内存在复杂的细菌结构, 在偏碱性条件下降解纤维素的细菌酶活力要高于酸性条件下的酶活力;跳虫作为生态系统中的分解者, 其肠道内大量降解纤维素细菌的存在不仅有助于跳虫利用环境中的大分子有机物满足自身的营养等需要, 同时对于饲料及工业生产也具有一定的应用价值.%[Aim] This study aims to determine the composition of intestinal bacteria in Proisotoma ananevae and to screen the cellulolytic bacteria. [Methods]The intestinal bacteria in P. ananevae adults were isolated and identified with traditional culturing method in combination with 16 S rRNA sequencing.The cellulolytic bacteria were screened with carboxymethyl cellulose (CMC) plate media, and the cellulase activity was measured by 3, 5-dinitrosalicylic acid (DNS) method under different pH conditions (pH 5. 0-9. 0). [Results]We isolated twenty bacterial strains from the intestinal tract of P. ananevae adults, which belong to 10 genera of three major phyla Firmicutes, Proteobacteria and Acinobacteria, i. e., Staphylococcus, Bacillus, Terribacillus, Advenella, Lysinibacillus, Arthrobacter, Enterobacter, Glutamicibacter, Leucobacter and Acinetobacter, and one unidentified bacterium. Among them, 10 strains were cellulolytic bacteria belonging to 6 genera of Firmicutes and Acinobacteria, i. e., Leucobacter, Bacillus, Terribacillus, Lysinibacillus, Arthrobacter and Glutamicibacter. The cellulase activities in all the cellulolytic strains were relatively higher under pH 7. 0-9. 0 and reached the maximum under pH 8. 0.[Conclusion]The results suggest that the bacteria in the intestinal tract of P. ananevae adults have complex structure. The presence of a large number of cellulolytic strains in P. ananevae, the disintegrator in ecosystem, not only helps springtails use macromolecules to meet their own nutrition requirements, but also has important application value especially in feed production and industrial application.