水牛瘤胃Methanomassiliicoccales富集培养条件研究

Research of Enrichmen Condition of Methanomassiliicoccales in Buffalo Rumen

试验旨在对来源于水牛瘤胃的Methanomassiliicoccales富集培养条件进行探索,在BRN培养基中分别添加甲醇、一甲胺、二甲胺和三甲胺连续传代培养,利用气相色谱、实时荧光定量PCR和16S rRNA高通量测序,对甲烷产量、Methanomassiliicoccales数量、总甲烷菌数量和甲烷菌多样性指数进行测定。结果表明,三甲胺组甲烷平均产气量显著高于其他组(P<0.05);与对照组相比富集第1代添加甲醇和三甲胺组Methanomassiliicoccales、总甲烷菌拷贝数显著升高(P<0.05);第7代添加三甲胺组Methanomassiliicoccales、总甲烷菌拷贝数显著高于对照、一甲胺组、二甲胺组(P<0.05);第23代仅有添加三甲胺组和二甲胺组检测到Methanomassiliicoccales,且三甲胺组总甲烷菌拷贝数显著高于其他组(P<0.05);Alpha多样性分析表明,与对照组相比第7代添加三甲胺组Ace、Chao1指数显著升高(P<0.05)。甲烷菌目水平分析表明,对照组、二甲胺组、三甲胺组Methanomassiliicoccales相对丰度均超过90%。由此可见Methanomassiliicoccales富集传代培养中添加三甲胺能提高甲烷产量以及Methanomassiliicoccale拷贝数和相对丰度。

The aim of this study was to investigate the enrichmen condition of Methanomassiliicoccales from buffalo rumen,methanol,monomethylamine,dimethylamine or trimethylamine were used as substrates in BRN culture medium respectively.Methane production,population and diversity indices of Methanomassiliicoccales and methanogen,wasmeasure by gas chromatography,Real-time quantitative PCR and high-throughput sequencing of the 16S rRNA gene.The results showed that:The methane production of trimethylamine group was higher than other group significantly(P<0.05).The copy of Methanomassiliicoccales and methanogen of methanol group and trimethylamine group were higher than control group significantly(P<0.05)in 1st generation.The copy of Methanomassiliicoccales and methanogen of trimethylamine group were higher than control group,monomethylamine group,dimethylamine group significantly(P<0.05)in 7th generation.The copy of Methanomassiliicoccales was detected in trimethylamine group and dimethylamine group only in 23rd generation,and the copy of methanogen of trimethylamine group was highest.The result of Alpha diversity indices showed that,the Ace and Chao1 index of trimethylamine group were higher than control group significantly(P<0.05).The relative abundance of methanogen at order level were more than 90%.The methane production,population and relative abundance of Methanomassiliicoccales were increased while trimethylamine was used as substratesduring transfers.