一株缩合单宁降解菌的筛选、鉴定及降解效果分析

Screening and Identification of a Condensed Tannin Degrading Bacteria and its Degradation Effect

为了筛选分离到能够降解缩合单宁的细菌并探究其降解能力,以牛羊粪土为菌源,用梯度驯化法最终分离筛选得到1株能够以缩合单宁为单一碳源生长的降解菌,经形态特征观察、Biolog生理生化测试及16S rRNA基因序列比对分析鉴定,菌B1为1株革兰氏阴性菌,属于肠杆菌属(Enterobacter sp.),在构建的系统发育树上与Enterobacter xiangfangensis 10-17T聚为一支。综合运用紫外分光光度法和高效液相色谱法检测,测得菌B1培养24 h最大OD600值达2.5左右,以计算出的原花青素、儿茶素变化量为指标分析菌B1对缩合单宁的降解能力,即菌B1对落叶松单宁降解率在72 h达90%以上且有儿茶素生成;进一步探究外加碳氮源对菌B1降解情况的影响,实验结果表明,外加氮源可以略微提升菌B1的降解能力,而外加碳源则会使降解率降低。研究结果对于丰富缩合单宁整体研究、提升缩合单宁的应用价值及进一步探究缩合单宁的微生物降解机制都有一定意义。

The purpose of this study was to screen and isolate a condensed-tannin degrading bacteria,then identify the strain,and explore its degrading ability. In this study,from cattle and sheep feces,a strain of degrading bacteria which could utilize condensed tannin as a sole carbon source was obtained by gradient acclimation method. Through morphological characteristics observation,Biolog physiological,biochemical tests and 16 S rRNA gene sequence analysis,strain B1 was identified as a gramnegative bacterium,which is belonging to the genus Enterobacter sp. and clustered with Enterobacter xiangfangensis 10-17 Tin a phylogenetic tree constructed. Comprehensive determination was detected by ultraviolet spectrophotometry and high performance liquid chromatography,the maximum OD_（600） value of B1 cultured for 24 hours was reached to 2.5. The degradation rate of larvae tannin by bacteria B1 was analyzed by the calculated changes in procyanidins and catechins,results showed that the degradation rate reached more than 90% at 72 hours and there was catechin generated. In addition,when explored the effects of additional nitrogen and carbon sources,the results showed that the addition of nitrogen source could slightly increase the degradation ability of bacterium B1,but the addition of carbon source would reduce the degradation rate. The results enriched the overall study of condensed tannins,enhanced the application value of condensed tannins,and further explored the mechanism of microbial degradation of condensed tannins.