不同尿素添加水平对育肥湖羊瘤胃发酵及微生物菌群结构的影响

Effects of incremental urea supplementation in diet on rumen fermentation and microbial communities in fattening Hu lambs

[目的]本试验旨在研究低蛋白日粮添加不同水平尿素对育肥湖羊瘤胃发酵及微生物菌群结构的影响。[方法]70只育肥公湖羊(23.6~26.0 kg)随机分为5组,分别饲喂以下日粮:正常豆粕(SBM,170 g·kg^(-1)),低豆粕(40 g·kg^(-1))基础上添加0、10、20和30 g·kg^(-1)尿素(U0、U10、U20、U30)。试验预饲期1周,正试期8周。采集瘤胃液测定发酵参数和功能菌群数量,同时扩增细菌16S rRNA基因V3—V4区进行Illumina-MiSeq测序。[结果]与U10组相比,U20和U30组瘤胃液pH值显著升高(P<0.05)。氨态氮(NH_(3)-N)浓度随尿素添加量的增加显著升高(P<0.05)。与U0和U30组相比,SBM、U10和U20组总菌、甲烷菌和原虫数量显著升高(P<0.05)。Illumina-MiSeq测序结果显示U30组Shannon指数显著高于U0和U10组(P<0.05),但Chao1指数没有差异(P>0.05)。U10组Bacteroidetes的丰度显著高于SBM、U20和U30组(P<0.05),但Firmicutes却显著降低(P<0.05)。各组的主要优势菌属为Prevotella 1、Muribaculaceae norank、Rikenellaceae RC9 gut group和Christensenellaceae R-7 group。U10组Prevotella 1的丰度显著高于U20和U30组(P<0.05),而Rikenellaceae RC9 gut group、Christensenellaceae R-7 group和Ruminococcaceae NK4A214 group显著低于U30组(P<0.05)。RDA结果显示pH值、NH_(3)-N和总挥发性脂肪酸(TVFA)对瘤胃不同菌属的影响程度存在差异,其中pH值的影响最显著(P<0.05)。[结论]在低蛋白日粮中添加适量尿素(10 g·kg^(-1))可增加育肥湖羊瘤胃功能菌群数量,促进相关代谢产物的产生与利用,但日粮氮不足或尿素添加水平过高则会降低瘤胃功能菌群数量和改变细菌群落组成。

[Objectives]This study was conducted to evaluate the effects of incremental urea supplementation on rumen fermentation and microbial communities in fattening Hu lambs.[Methods]Seventy fattening male Hu lambs were randomly divided into two blocks according to body weight(23.6-26.0 kg)and assigned to one of five dietary treatments:soybean meal(SBM)at 170 g·kg^(-1) or reduced SBM(40 g·kg^(-1))plus 0,10,20 and 30 g·kg^(-1) urea(U0,U10,U20,U30).The experiment consisted of 1 week of adaptation followed by 8 weeks of dietary treatments.The rumen fluid of each Hu lamb was collected to measure rumen fermentation parameters and functional microbial populations.Sequences were determined using Illumina-MiSeq through amplifying the bacterial 16S rRNA gene V3-V4 region.[Results]Compared with U10 group,the ruminal pH value was significantly higher for U20 and U30 groups(P<0.05).The concentration of ammonia(NH_(3)-N)increased with incremental urea supplementation.Compared with U0 and U30 groups,the populations of total bacteria,methanogens and protozoa greatly increased in SBM,U10 and U20 groups(P<0.05).High-throughput sequencing results showed that the Shannon index of U30 group was higher(P<0.05)than U0 and U10 groups,whereas no difference(P>0.05)was detected in the Chao1 index among U30,U0 and U10 groups.The relative abundance of Bacteroidetes was higher(P<0.05)for U10 group than for SBM,U20 and U30 groups,whereas that of Firmicutes was significantly lower for U10 group than for SBM,U20 and U30 groups(P<0.05).The dominant genus in each group were Prevotella 1,Muribaculaceae norank,Rikenellaceae RC9 gut group and Christensenellaceae R-7 group.The relative abundance of Prevotella 1 was greatly higher for U10 than for U20 and U30 groups(P<0.05),while Rikenellaceae RC9 gut group,Christensenellaceae R-7 group and Ruminococcaceae NK4A214 group were lower for U10 than for U30 group(P<0.05).The RDA results indicated that pH value,NH_(3)-N and total volatile fatty acids(TVFA)showed different impacts on rumen bacteria,but the most significant effect was found in pH value.[Conclusions]Appropriate urea addition level(10 g·kg^(-1))could increase the functional microbial populations and promote the production and utilization of related metabolites on the basis of low protein diet.However,insufficient nitrogen or high urea supplementation in diets reduced the functional microbial populations and changed the bacterial communities.