Effects of Substituting Refined Forage with Beer Lees on Digestion and Microbial Diversity of Hu Sheep Rumen
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摘要:
目的 研究啤酒糟替代饲粮中不同比例精料对湖羊瘤胃发酵及微生物多样性的影响，促进啤酒糟在湖羊饲养中的合理利用。 方法 选择体况良好、相近体重（15.90±0.36 kg）的湖羊24只，随机分为４组，分别为饲喂基础饲粮（玉米+豆粕为精料，占比36%，Ⅰ组，CK）及啤酒糟替代1/3（Ⅱ组）、2/3（Ⅲ组）和全部（Ⅳ组）基础饲粮中精料。试验结束后，每组随机选取3只羊，测定羊瘤胃发酵参数和瘤胃微生物多样性。 结果 1）啤酒糟替代饲粮中不同比例精料对湖羊瘤胃的pH值、氨态氮、乙酸等挥发性脂肪酸的含量无显著影响（P＞0.05），试验Ⅱ组的异丁酸含量显著高于其他试验组（P＜0.05）。2）在门水平上，啤酒糟替代饲粮中不同比例精料对湖羊瘤胃微生物厚壁菌门（Firmicutes）、拟杆菌门（Bacteroidetes）、变形菌门（Proteobacteria）、互养菌门（Synergistetes）、纤维杆菌门（Fibrobacteres）、放线菌门（Actinobacteria）、变形杆菌门（Epsilonbacteraeota）、螺旋体门（Spirochaetes）和无壁菌门（Tenericutes）的丰富度均无显著影响（P＞0.05）。试验Ⅰ组（CK）中疣微菌门（Verrucomicrobia）的丰富度显著低于其他组（P＜0.05）。在属水平上，啤酒糟替代饲粮中不同比例精料对湖羊瘤胃微生物普雷沃氏菌属（Prevotella）、琥珀酸菌属（Succiniclasticum）、肠单胞菌属（Intestinimonas）、酪氨酸菌属（Tyzzerella）、瘤胃球菌属（Ruminococcus）、粪便菌属（Faecalicatena）、假丁酸弧菌属（Pseudobutyrivibrio）和特氏梭菌属（Hungateiclostridium）的丰富度无显著影响（P＞0.05）。 结论 啤酒糟替代精料提高了疣微菌门的相对丰富度，未对湖羊其他瘤胃群落结构和瘤胃微生物群落丰富度产生显著影响。 Abstract:Objective Effects of substituting refined meal in forage with beer brewing residues on the digestion and microbial diversity of rumen of Hu sheep were studied to explore the feasibility of utilization of the waste material. Methods Twenty-four healthy Hu sheep with similar body weight of (15.90±0.36) kg were randomly divided into 4 groups. The animals were fed with a basic forage that consisted of corn and soybean meal at 36% as control (Group I) or one that used beer lees to replace 1/3 (Group II), 2/3 (Group III), or total (Group IV) of the meal. At end of the feeding period, 3 sheep were randomly selected from each group to determine the digestion indicators and microbial diversity in the rumen fluid. Results (1) None of the substitutions showed significant differences on volatile fatty acids, pH, ammonia nitrogen, or acetic acid in the rumen (P＞0.05), but significantly higher on isobutyric acid in Group II than others (P＜0.05). (2) At phylum level, no significant differences on richness of diversity were introduced by the substitutions on Firmicutes, Bacteroidetes, Proteobacteria, Synergistetes, Fibrobacteres, Actinobacteria, Epsilonbacteraeota, Spirochaetes, and Tenericutes (P＞0.05). However, the diversity of Verrucomicrobia in Group I was significantly lower (P＜0.05). At genus level, the beer lees replacements in the forage did not significantly alter the diversity on Prevotella, Succiniclasticum, Intestinimonas, Tyzzerella, Ruminococcus, Faecalicatena, Pseudobutyrivibrio, and Hungateiclostridiumhad (P＞0.05). Conclusion The replacement of the refined meal in forage with beer lees reduced the relative richness of Verrucobacteria but did not significantly changed the microbial community structure and diversity of the rumen fluid. -
Key words:
- Beer lees /
- Hu sheep /
- volatile fatty acids /
- rumen microorganism
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表 1 基础饲粮成分及营养水平（干物质基础）
Table 1. Nutritional composition of basal diet (DM basis)
项目
Items试验Ⅰ组
Group Ⅰ试验Ⅱ组
Group Ⅱ试验Ⅲ组
Group Ⅲ试验Ⅳ组
Group Ⅳ原料 Ingredients 啤酒糟 Brewers dried grain/% 0.00 12.00 24.00 36.00 玉米 Corn/% 24.00 16.00 8.00 0.00 豆粕 Soybean meal/% 12.00 8.00 4.00 0.00 谷壳粉 Chaff bran/% 9.00 9.00 9.00 9.00 杂交狼尾草 Hybrid penisetum/% 51.50 51.50 51.50 51.50 磷酸氢钙 CaHPO4/% 0.50 0.50 0.50 0.50 石粉 Limestone/% 1.50 1.50 1.50 1.50 食盐 NaCl/% 0.50 0.50 0.50 0.50 预混料 Premix1)/% 1.00 1.00 1.00 1.00 合计 Total/% 100.00 100.00 100.00 100.00 营养水平 Nutrient levels2) 有机物 OM/% 90.18 90.12 90.11 89.75 代谢能 ME/（MJ·kg−1） 9.52 9.19 8.56 8.14 总能 GE/（MJ·kg−1） 17.00 16.80 16.68 16.56 中性洗涤纤维 NDF/% 46.98 49.38 54.07 57.13 酸性洗涤纤维 ADF/% 26.54 28.91 30.20 30.71 粗蛋白质 CP/% 12.20 12.60 13.27 13.71 钙 Ca/% 0.77 0.78 0.81 0.83 总磷 TP/% 0.23 0.24 0.24 0.25 1）每千克饲粮预混料中含有Cu 20.0 mg，Fe 80.0 mg，Mn 30.0 mg，Zn 80.0 mg，I 1 mg，Se 0.30 mg，VA 20 000 IU，VD 5 000 IU，VE 50.0 mg。2）代谢能为计算值，其他均为实测值。
1) The premix provid the following per kg of diets: Cu 20.0 mg, Fe 80.0 mg, Mn 30.0 mg, Zn 80.0 mg, I 1 mg, Se 0.30 mg, VA 20 000 IU, VD 5 000 IU, VE 50.0 mg. 2）ME is calculated, others are measured values.
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表 2 啤酒糟替代饲粮中不同比例精料对湖羊瘤胃发酵参数的影响
Table 2. Effects of different forages on Hu sheep rumen digestion indicators
项目
Items试验Ⅰ组
Group Ⅰ试验Ⅱ组
Group Ⅱ试验Ⅲ组
Group Ⅲ试验Ⅳ组
Group ⅣpH值 pH value 6.73±0.005 6.99±0.06 6.79±0.07 6.85±0.009 氨态氮 NH3-N/（mg·dL−1） 9.40±0.45 9.83±0.54 10.50±0.50 10.97±1.02 乙酸 Acetate/（mmoL·L−1） 47.35±3.52 49.31±3.68 51.79±3.86 50.97±3.89 丙酸 Propionate/（mmoL·L−1） 20.53±1.69 21.71±1.72 22.51±2.35 20.57±3.20 异丁酸 Isobutyrate/（mmoL·L−1） 4.12±0.85 b 6.74±0.71 a 2.47±0.80 b 3.60±0.95 b 丁酸 Butyrate/（mmoL·L−1） 15.84±2.05 16.34±3.56 18.36±1.20 17.59±1.07 异戊酸 Isovalerate/（mmoL·L−1） 3.60±1.01 6.48±2.56 5.22±2.20 3.78±2.60 戊酸 Valerate/（mmoL·L−1） 1.03±0.35 1.56±0.23 1.66±0.39 1.30±0.42 乙酸/丙酸 Acetate/Propionate 2.33±0.20 2.27±0.23 2.31±0.25 2.57±0.24 同行数据后不同小写字母表示差异显著（P＜0.05）。下表同。
In the same row, data with different lowercase letters indicate significant differences(P＞0.05). The same as below.
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表 3 啤酒糟替代饲粮中不同比例精料对湖羊瘤胃真菌Alpha多样性指数的影响
Table 3. Effects of different forages on fungal alpha diversity index of Hu sheep rumen
项目
Items试验Ⅰ组
Group Ⅰ试验Ⅱ组
GroupⅡ试验Ⅲ组
Group Ⅲ试验Ⅳ组
Group ⅣShannon指数 Shannon index 7.985±0.062 b 7.932±0.060 b 8.012±0.073 b 8.489±0.059 a Simpson指数 Simpson index 0.9887±0.002 0.9886±0.003 0.9851±0.002 0.9917±0.002 Chao1指数 Chao1 index 851.228±51.106 b 805.601±50.205 b 1056.283±61.488 a 1191.569±58. 476 a Ace指数 Ace index 929.949±57.366 bc 791.286±48.859 c 1092.112±48.547 ab 1146.678±46.839 a
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表 4 啤酒糟替代饲粮中不同比例精料处理湖羊瘤胃微生物门水平上菌群丰富度
Table 4. Effects of different forages on microbiota richness at phylum level of Hu sheep rumen （单位： %）
项目
Items试验Ⅰ组
Group Ⅰ试验Ⅱ组
Group Ⅱ试验Ⅲ组
Group Ⅲ试验Ⅳ组
Group Ⅳ厚壁菌门 Firmicutes 29.86±6.23 37.44±5.61 51.27±8.62 40.58±6.51 拟杆菌门 Bacteroidetes 18.10±3.23 15.76±4.10 11.89±4.28 14.56±3.87 变形菌门 Proteobacteria 1.43±0.23 0.59±0.39 0.86±0.51 0.43±0.21 互养菌门 Synergistetes 1.27±0.20 0.37±0.02 0.18±0.06 0.73±0.25 纤维杆菌门 Fibrobacteres 0.06±0.01 0.78±0.21 0.08±0.02 0.06±0.01 放线菌门 Actinobacteria 0.03±0.01 0.27±0.22 0.15±0.10 0.46±0.20 变形杆菌门 Epsilonbacteraeota 0.087±0.012 0.009±0.001 0.388±0.021 0.001±0.001 螺旋体门 Spirochaetes 0.195±0.052 0.044±0.068 0.080±0.058 0.124±0.060 无壁菌门 Tenericutes 0.008±0.034 0.052±0.038 0.019±0.032 0.026±0.045 疣微菌门 Verrucomicrobia 0.002±0.002 c 0.026±0.003 a 0.011±0.002 bc 0.017±0.005 ab
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表 5 啤酒糟替代饲粮中不同比例精料处理湖羊瘤胃微生物属水平上菌群丰富度
Table 5. Effects of different forages on microbiota richness at genus level of Hu sheep rumen （单位： %）
项目
Items试验Ⅰ组
Group Ⅰ试验Ⅱ组
GroupⅡ试验Ⅲ组
Group Ⅲ试验Ⅳ组
Group Ⅳ普雷沃氏菌属 Prevotella 17.94±3.95 15.68±3.94 10.61±3.95 14.46±3.95 琥珀酸菌属 Succiniclasticum 4.24±1.87 2.37±1.64 6.68±1.34 9.21±1.52 肠单胞菌属 Intestinimonas 3.34±1.82 4.43±1.42 4.67±2.10 6.62±2.87 酪氨酸菌属 Tyzzerella 4.07±1.56 4.05±1.30 6.05±0.89 3.32±1.20 Uncultured_bacterium_f_Ruminococcaceae 1.20±1.95 4.16±2.01 7.09±3.56 3.71±1.89 瘤胃球菌属 Ruminococcus 2.63±1.56 2.90±0.89 3.59±0.85 3.58±1.02 Uncultured_bacterium_o_Clostridiales 4.09±1.56 1.67±1.02 2.19±0.89 2.23±1.33 粪便菌属 Faecalicatena 0.03±0.00 1.58±1.14 3.63±0.87 0.52±0.23 假丁酸弧菌属 Pseudobutyrivibrio 0.59±0.23 1.36±0.65 1.21±0.56 2.30±0.75 特氏梭菌属 Hungateiclostridium 0.62±0.05 2.00±0.25 1.85±0.52 0.38±0.03
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表 6 啤酒糟替代饲粮中不同比例精料处理湖羊瘤胃微生物功能预测的基因占比
Table 6. Effects of different forages on genes proportion of predicted functions of microbial community in Hu sheep rumen （单位：%）
项目
Items试验Ⅰ组
Group Ⅰ试验Ⅱ组
GroupⅡ试验Ⅲ组
Group Ⅲ试验Ⅳ组
Group Ⅳ代谢途径 Metabolic pathways 17.43±0.89 17.25±0.89 17.19±0.89 17.34±0.89 次生代谢产物的生物合成 Biosynthesis of secondary metabolites 8.00±0.02 7.96±0.02 8.05±0.02 8.05±0.02 抗生素的生物合成 Biosynthesis of antibiotics 5.84±0.02 5.83±0.02 5.87±0.02 5.87±0.02 氨基酸的生物合成 Biosynthesis of amino acids 4.10±0.07 4.22±0.07 4.35±0.07 4.26±0.07 不同环境中的微生物代谢 Microbial metabolism in diverse environments 3.78±0.02 3.76±0.02 3.76±0.02 3.75±0.02 碳代谢 Carbon metabolism 2.62±0.02 2.60±0.02 2.56±0.02 2.58±0.02 ABC转运 ABC transporters 2.20±0.14 2.41±0.14 2.62±0.14 2.39±0.14 嘌呤代谢 Purine metabolism 2.11±0.01 2.12±0.01 2.10±0.01 2.10±0.01 嘧啶代谢 Pyrimidine metabolism 1.82±0.02 1.83±0.02 1.79±0.02 1.83±0.02
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