饲粮NDF水平对山羊营养物质表观消化率及瘤胃古菌结构与组成的影响研究
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摘要
【目的】旨在采用代谢试验和高通量测序技术,研究饲粮中性洗涤纤维(NDF)水平对山羊营养物质表观消化率及瘤胃古菌结构与组成的影响。【方法】选用6只8月龄健康雄性山羊进行3×3拉丁方试验,依据饲粮NDF水平设低(LN组)、中(MN组)和高(HN组)NDF水平共3个处理组进行代谢试验。代谢试验共分3期进行,每期试验结束后采集瘤胃内容物,提取瘤胃微生物总DNA后,用古菌通用引物进行PCR扩增,扩增产物用Illumina平台进行高通量测序后进行生物信息学分析。【结果】结果表明:(1)处理组间山羊日粮DM、OM、CP表观消化率差异均不显著(P>0.05);LN组EE的表观消化率显著高于MN组和HN组(P<0.05),HN组NDF的表观消化率显著高于LN组和MN组(P<0.05)。(2)LN组古菌Chao1、Shannon和Observed_species指数均显著高于其他两组(P<0.05),其他两组间的差异不显著(P>0.05)。(3)在门水平上,3个处理组间所有古菌的相对丰度差异均不显著(P>0.05),优势门均为广古菌门(Euryarchaeota)。(4)在属水平上,Methanobrevibacter,Candidatus_Methanomethylophilus和Candidatus_Methanoplasma均为3个处理组的优势属。LN组Methanobrevibacter的相对丰度极显著低于其他两组(P<0.01),但Candidatus_Methanomethylophilus和Candidatus_Methanoplasma的相对丰度显著高于其他两组(P<0.05),其余各菌属的相对丰度在3个处理组间差异均不显著(P>0.05)。【结论】研究结论:饲粮NDF水平在35%~45%之间变化时,显著影响山羊对日粮中EE、NDF的表观消化率,显著影响瘤胃Methanobrevibacter、Candidatus_Methanomethylophilus和Candidatus_Methanoplasma等古菌属的相对丰度。
【Objective】This study was conducted to examine the effects of dietary NDF levels on apparent digestibility of nutrients and the composition of rumen archaea in goats by using metabolic trial and high throughput sequencing techniques.【Method】Six eight-month-old healthy male goats were used in a 3×3 Latin square experiment. Three experimental treatments were set for the metabolic trial as follows:low dietary NDF level group(LN),middle dietary NDF level group(MN) and high dietary NDF level group(HN). The whole experiment contained three phases,and the rumen contents of each goat were collected after each phase. The total DNA of rumen microorganism in the rumen contents was extracted. Primers specific for archaea were used to amplified the V3-V4 high variable regions of 16 S r RNA gene. Bioinformatics analysis of PCR products were performed by high-throughput sequencing using Illumina platform.【Result】(1)There were no significant differences in the apparent digestibility of DM,OM and CP between the three groups(P>0.05). The apparent digestibility of EE in LN group was significantly higher than that in MN and HN group(P<0.05). The apparent digestibility of NDF in HN group was significantly higher than that in LN and MN group(P<0.05).(2)The Chao1,Shannon and Observed species indices of LN group were significantly higher than those of the other two groups(P<0.05),no significant difference was found between the other two groups(P >0.05).(3)At phylum level,the relative abundance of all taxonomy in the three treatment groups had no significant different(P>0.05),and the dominant phylum group was Euryarchaeota.(4)At genus level,Methanobrevibacter,Candidatus_Methanomethylophilus and Candidatus_Methanoplasma were the dominant genera groups in all of the three treatment groups. The relative abundance of Methanobrevibacter in LN group was remarkablely lower than that in the other two groups(P<0.01),but the relative abundances of Candidatus_Methanomethylophilus and Candidatus_Methanoplasma were significantly higher than those of the other two groups(P<0.05),and these archaea had no significant difference between the other two groups. The other archaeal genera had no significant difference between the three treatment groups(P >0.05).【Conclusion】The change of the NDF level(from 35% to 45%)significantly affected the apparent digestibility of EE and NDF,as well as the relative abundances of major ruminal archaea such as Methanobrevibacter in goats.
【Objective】This study was conducted to examine the effects of dietary NDF levels on apparent digestibility of nutrients and the composition of rumen archaea in goats by using metabolic trial and high throughput sequencing techniques.【Method】Six eight-month-old healthy male goats were used in a 3×3 Latin square experiment. Three experimental treatments were set for the metabolic trial as follows:low dietary NDF level group(LN),middle dietary NDF level group(MN) and high dietary NDF level group(HN). The whole experiment contained three phases,and the rumen contents of each goat were collected after each phase. The total DNA of rumen microorganism in the rumen contents was extracted. Primers specific for archaea were used to amplified the V3-V4 high variable regions of 16 S r RNA gene. Bioinformatics analysis of PCR products were performed by high-throughput sequencing using Illumina platform.【Result】(1)There were no significant differences in the apparent digestibility of DM,OM and CP between the three groups(P>0.05). The apparent digestibility of EE in LN group was significantly higher than that in MN and HN group(P<0.05). The apparent digestibility of NDF in HN group was significantly higher than that in LN and MN group(P<0.05).(2)The Chao1,Shannon and Observed species indices of LN group were significantly higher than those of the other two groups(P<0.05),no significant difference was found between the other two groups(P >0.05).(3)At phylum level,the relative abundance of all taxonomy in the three treatment groups had no significant different(P>0.05),and the dominant phylum group was Euryarchaeota.(4)At genus level,Methanobrevibacter,Candidatus_Methanomethylophilus and Candidatus_Methanoplasma were the dominant genera groups in all of the three treatment groups. The relative abundance of Methanobrevibacter in LN group was remarkablely lower than that in the other two groups(P<0.01),but the relative abundances of Candidatus_Methanomethylophilus and Candidatus_Methanoplasma were significantly higher than those of the other two groups(P<0.05),and these archaea had no significant difference between the other two groups. The other archaeal genera had no significant difference between the three treatment groups(P >0.05).【Conclusion】The change of the NDF level(from 35% to 45%)significantly affected the apparent digestibility of EE and NDF,as well as the relative abundances of major ruminal archaea such as Methanobrevibacter in goats.
引文
[1]FLINT H J.The rumen microbial ecosystem---some recent development[sJ].Trends in Microbiology,1997,5(12):483-488.
[2]KAMRA D N.Rumen microbial ecosystem[J].Current Science,2005,89(1):124-135.
[3]王立志,徐谊英,蔡永华.高通量测序分析人工养殖成年林麝粪便古菌菌群多样性[J].动物营养学报,2016,28(2):477-484.
[4]HUNGATE R E.The rumen microbial ecosystem[J].Annu Rev Ecol Syst,1975,6:39-66.
[5]刘艳红,钟志军,艾生权,等.亚成体大熊猫肠道纤维素降解真菌的分离与鉴定[J].中国兽医科学,2015,45(1):43-49.
[6]SARNKLONG C,CONE J W,PELLIKAAN W,et al.Utilization of rice straw and different treatments to improve its feed value for ruminants:a review[J].Asian Australasian Journal of Animal Sciences,2010,23(5):680-692.
[7]DEHORITY B A.Effects of microbial synergism on fibre digestion in the rumen[J].Proceedings of the Nutrition Society,1991,50(2):149-59.
[8]RUSSELL J B,MUCK R E,WEIMER P J.Quantitative analysis of cellulose degradation and growth of cellulolytic bacteria in the rumen[J].Fems Microbiology Ecology,2009,67(2):183-197.
[9]王梦芝,徐爱秋,李世霞,等.瘤胃微生物对纤维素类物质降解的研究[J].饲料工业,2007,28(14):53-57.
[10]RUSSELL J B,MUCK R E,WEIMER P J.Quantitative analysis of cellulose degradation and growth of cellulolytic bacteria in the rumen[J].Fems Microbiology Ecology,2009,67(2):183-197.
[11]PATRA A K,YU Z.Essential oils affect populations of some rumen bacteria in vitro as revealed by microarray(RumenBactArray)analysi[sJ].Frontiers in Microbiology,2015,6:297.
[12]Fondevila M,Dehority B A.Interactions between Fibrobacter succinogenes,Prevotella ruminicola,and Ruminococcus flavefaciens in the digestion of cellulose from forages[J].Journal of Animal Science,1996,74(3):678-684
[13]WANG L Z,WANG Z S,XUE B,et al.Comparison of rumen archaeal diversity in adult and elderly yaks(Bos grunniens)using16S rRNA gene high-throughput sequencing[J].Journal of Integrative Agriculture,2017,16(5):1130-1137.
[14]张丽英.饲料分析及饲料质量检测技术[M].北京:中国农业大学出版社,2003:52-70
[15]JAMI E,MIZRAHI I.Composition and similarity of bovine rumen microbiota across individual animals[J].Plos One,2012,7(3):e33306.
[16]LI M,ZHOU M,ADAMOWICZ E,et al.Characterization of bovine ruminal epithelial bacterial communities using 16S rRNAsequencing,PCR-DGGE,and qRT-PCR analysis[J].Veterinary Microbiology,2012,155(1):72-80.
[17]WRIGHT A D,WILLIAMS A J,WINDER B,et al.Molecular diversity of rumen methanogens from sheep in Western Australia[J].Applied&Environmental Microbiology,2004,70(3):1263-1270.
[18]CAPORASO J G,KUCZYNSKI J,STOMBAUGH J,et al.QIIMEallows analysis of high-throughput community sequencing data[J].Nature methods,2010,7(5):335-336.
[19]KRISTIANDALY,STEWART C S,FLINT H J,et al.Bacterial diversity within the equine large intestine as revealed by molecular analysis of cloned 16S rRNA genes[J].Fems Microbiology Ecology,2001,38(2/3):141-151.
[20]张立涛,李艳玲,王金文,等.不同中性洗涤纤维水平饲粮对肉羊生长性能和营养成分表观消化率的影响[J].动物营养学报,2013,25(2):433-440.
[21]周汉林,莫放,李琼,等.日粮中性洗涤纤维水平对中国荷斯坦公牛营养物质消化率的影响[J].海南大学学报(自然科学版),2006,24(3):275-283.
[22]TJARDES K E,BUSKIRK D D,ALLEN M S,et al.Neutral detergent fiber concentration in corn silage influences dry matter intake,diet digestibility,and performance of Angus and Holstein steer[sJ].Journal of Animal Science,2002,80(3):841-846.
[23]孔祥浩,郭金双,朱晓萍,等.不同NDF水平肉羊日粮养分表观消化率研究[J].动物营养学报,2010,22(1):70-74.
[24]LIN C,RASKIN L,STAHL D A.Microbial community structure in gastrointestinal tracts of domestic animals:comparative analyses using rRNA-targeted oligonucleotide probes[J].Fems Microbiology Ecology,1997,22(4):281-294.
[25]WRIGHT A D,TOOVEY A F,PIMM C L.Molecular identification of methanogenic archaea from sheep in Queensland,Australia reveal more uncultured novel archaea[J].Anaerobe,2006,12(3):134-139.
[26]RICHARDSHARP,ZIEMER C J,STERN M D,et al.Taxonspecific associations between protozoal and methanogen populations in the rumen and a model rumen system[J].Fems Microbiology Ecology,1998,26(1):71-78.
[27]JANSSEN P H,KIRS M.Structure of the Archaeal community of the rumen[J].Applied&Environmental Microbiology,2008,74(12):3619-3625.
[28]WRIGHT A D,MA X,OBISPO N E.Methanobrevibacter phylotypes are the dominant methanogens in sheep from Venezuela[J].Microbial Ecology,2008,56(2):390-394.
[29]ZHOU M,MCALLISTER T A,GUAN L L.Molecular identification of rumen methanogens:Technologies,advances and prospects[J].Animal Feed Science&Technology,2011,166-167(7):76-86.
[2]KAMRA D N.Rumen microbial ecosystem[J].Current Science,2005,89(1):124-135.
[3]王立志,徐谊英,蔡永华.高通量测序分析人工养殖成年林麝粪便古菌菌群多样性[J].动物营养学报,2016,28(2):477-484.
[4]HUNGATE R E.The rumen microbial ecosystem[J].Annu Rev Ecol Syst,1975,6:39-66.
[5]刘艳红,钟志军,艾生权,等.亚成体大熊猫肠道纤维素降解真菌的分离与鉴定[J].中国兽医科学,2015,45(1):43-49.
[6]SARNKLONG C,CONE J W,PELLIKAAN W,et al.Utilization of rice straw and different treatments to improve its feed value for ruminants:a review[J].Asian Australasian Journal of Animal Sciences,2010,23(5):680-692.
[7]DEHORITY B A.Effects of microbial synergism on fibre digestion in the rumen[J].Proceedings of the Nutrition Society,1991,50(2):149-59.
[8]RUSSELL J B,MUCK R E,WEIMER P J.Quantitative analysis of cellulose degradation and growth of cellulolytic bacteria in the rumen[J].Fems Microbiology Ecology,2009,67(2):183-197.
[9]王梦芝,徐爱秋,李世霞,等.瘤胃微生物对纤维素类物质降解的研究[J].饲料工业,2007,28(14):53-57.
[10]RUSSELL J B,MUCK R E,WEIMER P J.Quantitative analysis of cellulose degradation and growth of cellulolytic bacteria in the rumen[J].Fems Microbiology Ecology,2009,67(2):183-197.
[11]PATRA A K,YU Z.Essential oils affect populations of some rumen bacteria in vitro as revealed by microarray(RumenBactArray)analysi[sJ].Frontiers in Microbiology,2015,6:297.
[12]Fondevila M,Dehority B A.Interactions between Fibrobacter succinogenes,Prevotella ruminicola,and Ruminococcus flavefaciens in the digestion of cellulose from forages[J].Journal of Animal Science,1996,74(3):678-684
[13]WANG L Z,WANG Z S,XUE B,et al.Comparison of rumen archaeal diversity in adult and elderly yaks(Bos grunniens)using16S rRNA gene high-throughput sequencing[J].Journal of Integrative Agriculture,2017,16(5):1130-1137.
[14]张丽英.饲料分析及饲料质量检测技术[M].北京:中国农业大学出版社,2003:52-70
[15]JAMI E,MIZRAHI I.Composition and similarity of bovine rumen microbiota across individual animals[J].Plos One,2012,7(3):e33306.
[16]LI M,ZHOU M,ADAMOWICZ E,et al.Characterization of bovine ruminal epithelial bacterial communities using 16S rRNAsequencing,PCR-DGGE,and qRT-PCR analysis[J].Veterinary Microbiology,2012,155(1):72-80.
[17]WRIGHT A D,WILLIAMS A J,WINDER B,et al.Molecular diversity of rumen methanogens from sheep in Western Australia[J].Applied&Environmental Microbiology,2004,70(3):1263-1270.
[18]CAPORASO J G,KUCZYNSKI J,STOMBAUGH J,et al.QIIMEallows analysis of high-throughput community sequencing data[J].Nature methods,2010,7(5):335-336.
[19]KRISTIANDALY,STEWART C S,FLINT H J,et al.Bacterial diversity within the equine large intestine as revealed by molecular analysis of cloned 16S rRNA genes[J].Fems Microbiology Ecology,2001,38(2/3):141-151.
[20]张立涛,李艳玲,王金文,等.不同中性洗涤纤维水平饲粮对肉羊生长性能和营养成分表观消化率的影响[J].动物营养学报,2013,25(2):433-440.
[21]周汉林,莫放,李琼,等.日粮中性洗涤纤维水平对中国荷斯坦公牛营养物质消化率的影响[J].海南大学学报(自然科学版),2006,24(3):275-283.
[22]TJARDES K E,BUSKIRK D D,ALLEN M S,et al.Neutral detergent fiber concentration in corn silage influences dry matter intake,diet digestibility,and performance of Angus and Holstein steer[sJ].Journal of Animal Science,2002,80(3):841-846.
[23]孔祥浩,郭金双,朱晓萍,等.不同NDF水平肉羊日粮养分表观消化率研究[J].动物营养学报,2010,22(1):70-74.
[24]LIN C,RASKIN L,STAHL D A.Microbial community structure in gastrointestinal tracts of domestic animals:comparative analyses using rRNA-targeted oligonucleotide probes[J].Fems Microbiology Ecology,1997,22(4):281-294.
[25]WRIGHT A D,TOOVEY A F,PIMM C L.Molecular identification of methanogenic archaea from sheep in Queensland,Australia reveal more uncultured novel archaea[J].Anaerobe,2006,12(3):134-139.
[26]RICHARDSHARP,ZIEMER C J,STERN M D,et al.Taxonspecific associations between protozoal and methanogen populations in the rumen and a model rumen system[J].Fems Microbiology Ecology,1998,26(1):71-78.
[27]JANSSEN P H,KIRS M.Structure of the Archaeal community of the rumen[J].Applied&Environmental Microbiology,2008,74(12):3619-3625.
[28]WRIGHT A D,MA X,OBISPO N E.Methanobrevibacter phylotypes are the dominant methanogens in sheep from Venezuela[J].Microbial Ecology,2008,56(2):390-394.
[29]ZHOU M,MCALLISTER T A,GUAN L L.Molecular identification of rumen methanogens:Technologies,advances and prospects[J].Animal Feed Science&Technology,2011,166-167(7):76-86.