奶牛瘤胃Methanomassiliicoccales的微生态学及其调控宿主甲基化合物代谢的机制

Methanomassiliicoccales (Mmc) is a newly established methanogenic order. The members of this order are not only involved in the metabolism of hydrogen, but also involved in the metabolism of methyl-compounds. However, the ecology of the rumen Mmc community is unclear. The mechanism of Mmc regulating the metabolism of methyl-compounds in host is also unclear as well as the relationship between Mmc and host health. On the basis of previous studies on the diversity and isolation of Mmc, the following studies are carried out by combining the in vivo and in vitro studies as well as the metagenomics and metatranscriptomics technology. (1) The composition and function characteristics of the microbes involved in the metabolism of methyl-compounds (MIMM) in cow rumen will be analyzed, and the relationship between the diet and the composition of the MIMM will be also clarified; (2) The mechanism of the choline degradation by Mmc strains and other MIMM will be explored by the pure culture and co-culture technology of anaerobic microbes, and the mechanism of the interaction of Mmc strains and other MIMM will be elucidated; (3) The effect of the intermediate products of methyl-compounds, such as trimethylamine, on the host health will be studied by animal experiment, and the biological mechanism of the interaction of Mmc and host affecting the heath of the body will be revealed. The results will produce original innovation in theoretical understanding of rumen microorganisms, and have important guiding significance for promoting the utilization of feed nutrients and reducing methane emission in ruminants.

甲烷菌第七目菌群（Mmc）是近年来新发现的一类甲烷菌，其功能不仅涉及瘤胃氢代谢，还参与瘤胃内甲基化合物的代谢，但Mmc的生态组成及其调控宿主甲基化合物代谢的机制尚不清楚，其与宿主健康之间联系也尚未解析。本项目拟在申请人前期研究基础上，利用动物在体、微生物培养和组学技术：（1）解析奶牛瘤胃中甲基化合物代谢菌群的组成及其功能特点，明确日粮结构与甲基化合物代谢菌群组成间的内在联系；（2）采用厌氧微生物纯培养和共培养技术，探讨Mmc菌株和其它典型菌株对瘤胃代表性甲基化合物胆碱的降解机制，解析Mmc与甲基化合物代谢链中其它典型微生物的互作机制；（3）通过动物实验，解析甲基化合物代谢中间产物（如三甲胺）对宿主健康的影响，揭示Mmc与宿主互作影响机体健康的生物学机制。研究结果可在瘤胃微生物理论认识方面产生原始性创新，并对促进饲料营养素的高效利用和反刍动物甲烷减排具有重要指导意义。

瘤胃氢依赖甲基营养型甲烷菌（Mmc）是新建立的一个甲烷菌目，其菌群组成及调控宿主甲基化合物代谢的机制尚不清楚，与宿主健康之间联系也不清楚。本项目通过利用宏基因组测序技术，构建了包括奶牛在内的七种反刍动物全消化道微生物基因集和基因组集，在此基础上，利用关键酶基因筛选构建了胆碱降解菌和Mmc基因组集，重建了Mmc的甲基化合物代谢通路；发现奶牛瘤胃菌群降解胆碱生成三甲胺的速度比利用三甲胺的速度快；抑制甲烷菌活性后，三甲胺不能被瘤胃其他微生物利用，在发酵液中大量积累；日粮胆碱水平升高，提高了胆碱降解菌和Mmc丰度，同时提高了瘤胃液三甲胺浓度以及血液和乳中氧化三甲胺浓度；高精料日粮饲喂奶牛，降低了Mmc的相对丰度，降低了瘤胃菌群清除三甲胺的能力，导致血浆中氧化三甲胺浓度升高；分别从奶牛瘤胃和皱胃中分离获得一株胆碱降解菌DRC1和一株三甲胺利用菌DZ1（Mmc）。将两株菌共培养，DRC1为DZ1提供底物，DZ1为DRC1移除三甲胺，降低氢分压，保障DRC1代谢正常进行。此外，利用小鼠模型研究发现高水平的血液氧化三甲胺可能通过影响肝脏抗氧化能力和促炎因子水平影响宿主健康，而灌胃DZ1活菌降低了小鼠血液氧化三甲胺水平，同时提高了小鼠肝脏抗氧化能力，降低了血清促炎因子水平。综上，虽然Mmc不能将瘤胃中的三甲胺完全利用，但减少了三甲胺进入血液循环的量，对宿主健康具有保护作用。研究结果为反刍动物日粮胆碱高效利用的深入研究提供了基础；还为瘤胃甲烷减排策略制定提出警示，抑制瘤胃甲烷菌活性，可能导致大量有害物质进入血液循环，危害动物健康。项目执行过程中共发表7篇标注的论文，其中SCI论文4篇，中文核心期刊论文3篇，申请发明专利1项。