Table_2_Effect of an anti-methanogenic supplement on enteric methane emission, fermentation, and whole rumen metagenome in sheep.DOCX

A study was conducted to investigate the impact of an anti-methanogenic product supplementation on enteric methane emissions, whole rumen metagenome and ruminal fermentation in sheep. Twelve adult male sheep were randomly divided into two groups of six animals each. Animals were fed ad libitum on a total mixed ration either without (CON) or with an anti-methanogenic supplement (Harit Dhara-HD). The anti-methanogenic supplement contained 22.1% tannic acid in a 3: 1 ratio of condensed and hydrolysable tannins. The supplementation of product revealed a significant reduction in daily enteric methane emission (21.9 vs. 17.2 g/d) and methane yield (23.2 vs. 18.2) without affecting the nutrient intake and digestibility. However, the propionate concentration in the HD treatment group was significantly higher than in the CON group. On the contrary, the ammonia nitrogen concentration was lower. The anti-methanogenic supplement significantly decreased the ruminal protozoa in the HD treatment group. Whole rumen metagenome analysis revealed that the core bacterial (Bacteroidetes and Firmicutes) and archaeal communities (Methanobrevibacter and Methanosarcina) were comparable between the CON and HD treatment groups. However, the supplementation of anti-methanogenic product led to a considerable reduction in the abundance of Proteobacteria, whereas the abundance of Lentisphaerae was greater. The supplementation significantly decreased the abundance of Methanocaldococcus, Methanococcoides, Methanocella, and Methanoregula methanogens. A total of 36 KO related to methanogenesis were identified in this study. The activities of formate dehydrogenase (EC 1.8.98.6) and tetrahydromethanopterin S-methyltransferase (EC 2.1.1.86) were significantly lowered by the anti-methanogenic product supplementation in sheep. In conclusion, the anti-methanogenic supplement has the potential to decrease enteric methane emission (~22%) at the recommended level (5% of DM) of supplementation. The contribution of minor methanogens vulnerable to supplementation to rumen methanogenesis is not known; hence, the culturing of these archaea should be taken on priority for determining the impact on overall rumen methanogenesis.

本研究旨在探究抗产甲烷产品补饲对绵羊肠道甲烷排放、全瘤胃宏基因组及瘤胃发酵的影响。试验选取12只成年公绵羊，随机分为两组，每组6只。试验羊自由采食全混合日粮（Total Mixed Ration, TMR），其中对照组（CON）不添加任何补剂，处理组（HD）添加抗产甲烷补剂Harit Dhara-HD。该抗产甲烷补剂中单宁酸占比为22.1%，缩合单宁与水解单宁的比例为3:1。结果显示，补饲该产品可显著降低绵羊每日肠道甲烷排放量（21.9 vs. 17.2 g/d）及甲烷产率（23.2 vs. 18.2），且对营养物质采食量与消化率无显著影响。但HD处理组的丙酸浓度显著高于CON组，反之氨氮浓度则显著低于CON组。此外，HD处理组的瘤胃原虫数量因补饲该抗产甲烷补剂而显著降低。全瘤胃宏基因组分析结果表明，CON组与HD组的核心细菌群落（拟杆菌门Bacteroidetes、厚壁菌门Firmicutes）以及古菌群落（甲烷短杆菌属Methanobrevibacter、甲烷八叠球菌属Methanosarcina）并无显著差异。但补饲抗产甲烷产品可显著降低变形菌门Proteobacteria的丰度，同时提升Lentisphaerae门的丰度。此外，补饲该产品可显著降低甲烷球菌属（Methanocaldococcus）、甲烷囊菌属（Methanococcoides）、甲烷胞菌属（Methanocella）与甲烷杆状菌属（Methanoregula）这四类产甲烷古菌的丰度。本研究共鉴定出36个与产甲烷作用相关的KEGG直系同源基因（KO）。绵羊补饲该抗产甲烷产品后，甲酸脱氢酶（EC 1.8.98.6）与四氢甲烷蝶呤S-甲基转移酶（EC 2.1.1.86）的活性均显著降低。综上，在推荐补饲水平（占日粮干物质（Dry Matter, DM）的5%）下，该抗产甲烷补剂可使绵羊肠道甲烷排放量降低约22%。但目前仍不清楚对补饲敏感的小型产甲烷古菌对瘤胃产甲烷过程的贡献，因此后续应优先开展这类古菌的分离培养研究，以明确其对瘤胃整体产甲烷过程的影响。