人肠道梭菌和甲烷马赛球菌协同代谢甜菜碱和胆碱产甲烷研究  

Clostridium and Methanomassiliicoccus isolated from humanintestine synergistically convert betaine and choline tomethane

作　　者：田栩萍 李凌燕[3,4] 李洁 高健[1,2] 邓锴 东秀珠 TIAN Xuping;LI Lingyan;LI Jie;GAO Jian;DENG Kai;DONG Xiuzhu(Biomedical Engineering College,Hubei University of Medicine,Shiyan 442000,Hubei,China;Reproductive Medicine Center,Renmin Hospital,Hubei University of Medicine,Shiyan 442000,Hubei,China;State Key Laboratory of Microbial Resources,Institute of Microbiology,Chinese Academy of Sciences,Beijing 100101,China;College of Life Sciences,University of Chinese Academy of Sciences,Beijing 100049,China)

机构地区：[1]湖北医药学院生物医学工程学院,湖北十堰442000 [2]湖北医药学院附属人民医院生殖医学中心,湖北十堰442000 [3]中国科学院微生物研究所微生物资源前期开发国家重点实验室,北京100101 [4]中国科学院大学生命科学学院,北京100049

出　　处：《微生物学报》2023年第8期3144-3156,共13页Acta Microbiologica Sinica

基　　金：国家自然科学基金(32070061)。

摘　　要：【目的】根据人肠道富含胆碱和甜菜碱,同时肠道微生物组中具有裂解胆碱和还原甜菜碱产三甲胺的细菌,以及利用三甲胺产甲烷的古菌,本研究探讨肠道细菌与古菌协同代谢甜菜碱和胆碱产甲烷的可能性。【方法】调查不同年龄段人群粪便中的16S rRNA基因多样性,分析肠道中古菌的菌群组成;利用定量PCR(quantitative PCR,qPCR)定量甲烷马赛球菌(Methanomassiliicoccus)特异的甲醇甲基转移酶基因mtaB和甲烷八叠球菌(Methanosarcina)及细菌的16S rRNA基因拷贝数,分析肠道中甲基营养型产甲烷古菌及总细菌的含量;宏基因组组装基因组(metagenome-assembled genomes,MAGs)分析携带甜菜碱还原酶基因grdH和胆碱裂解酶基因cutC的细菌组成。从粪便中分离代谢甜菜碱及胆碱产生三甲胺的细菌,并与分离自人肠道的甲烷马赛球菌构建共培养物,测定其协同转化甜菜碱和胆碱产甲烷的能力。【结果】年轻人粪便中含有甲烷杆菌科(Methanobacteriaceae,82.16%)的甲烷短杆菌属(Methanobrevibacter,49.18%)和甲烷杆菌属(Methanobacterium,33.34%)、甲基营养型的甲烷八叠球菌科(Methanosarcinaceae,5.67%)的甲烷八叠球菌属(Methanosarcina,5.70%),以及甲烷马赛球菌科(Methanomassiliicoccaceae,3.13%)的甲烷马赛球菌属(Methanomassiliicoccus,3.14%)。而中老年人粪便中的甲烷古菌多样性较低,也未检测到甲烷马赛球菌。qPCR定量分析显示年轻人比中老年人肠道的总古菌含量高3.11倍,其中甲烷马赛球菌高6.53倍、甲烷八叠球菌高5.52倍,总细菌含量高2.90倍。宏基因组分析组装了229个细菌基因组,其中42个携带基因grdH和cutC,这些细菌属于毛螺菌科(Lachnospiraceae)、肠杆菌科(Enterobacteriaceae)和梭菌科(Clostridiaceae)等。从粪便中分离到恶名梭菌(Clostridium malenominatum)B8,菌株B8与卢米尼甲烷马赛球菌(Methanomassiliicoccus luminyensis)B10共培养物可降解47.03%的甜菜碱和25.83%胆碱,并�[Objective]To explore the feasibility of using Clostridium and Methanomassiliicoccus from human intestine to synergistically convert betaine and choline to methane.[Methods]Illumina sequencing of the 16S rRNA gene was performed to survey the diversity of archaea in the feces from healthy people of 20–40 years old and over 40 years old.The Methanomassiliicoccus-specific mtaB gene and Methanomassiliicoccus-specific 16S rRNA gene were quantitated by quantitative PCR(qPCR)to quantify the trimethylamine-utilizing methanogens in human intestine.Metagenome-assembled genomes(MAGs)were reconstructed from metagenome data for the identification of the intestinal bacteria carrying the betaine reductase gene grdH and choline trimethylamine-lyase gene cutC.The bacteria that reduced betaine and choline were isolated from feces and used to construct the coculture with Methanomassiliicoccus.The potential of the coculture for producing methane from betaine and choline was then determined.[Results]The main methanogenic archaea in the intestine of the 20–40 years old included Methanobrevibacter(49.18%)and Methanobacterium(33.34%)affiliating to Methanobacteriaceae(82.16%),Methanosarcina(5.70%)of Methanosarcinaceae(5.67%),and Methanomassiliicoccus(3.14%)of Methanosassilicoccaceae(3.13%).The methanogen diversity was lower in the feces from the people over 40 years old,from whom Methanosassilicoccaceae was not detected.Quantitative PCR determined that the total abundance of archaea and bacteria in the people of 20–40 years old was 3.11 and 2.90 folds,respectively,higher than in those over 40 years old.Specifically,the abundance of Methanomassiliicoccus and Methanosarcina was 6.53 and 5.52 folds higher,respectively.A total of 229 bacterial MAGs were obtained from the fecal specimens,in which 42 MAGs carried genes grdH and cutC and were affiliated to Lachnospiraceae,Enterobacteriaceae,and Clostridiaceae.Clostridium malenominatum B8 was isolated from the fecal specimens.The co-culture of this strain with Methanomassiliicoccus luminye

关 键 词：人粪便 甜菜碱 胆碱 三甲胺 梭菌 甲烷马赛球菌 

分 类 号：R37[医药卫生—病原生物学]