野牦牛和家牦牛粪便菌群与短链脂肪酸关系的研究
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摘要
短链脂肪酸(SCFA)是反刍动物吸收饲草、饲料中营养物质的重要形式。肠道菌群能够降解食物生成SCFA并影响其比例。本文通过16S r DNA测序和气相色谱质谱联用仪,分别测定了野牦牛(Bos mutus)和家牦牛(Bos grunniens)粪便菌群组成及SCFA含量,通过比较分析两种牦牛肠道菌群与SCFA的关系,筛选出野牦牛肠道中与SCFA高浓度有正相关关系的菌群。结果显示,野牦牛粪便菌群主要有厚壁菌门(Firmicutes)(66. 47%)、拟杆菌门(Bacteroidetes)(26. 00%)和变形菌门(Proteobacteria)(3. 48%),主要的科有瘤胃球菌科(Ruminococcaceae)(55. 18%)、拟杆菌科(Bacteroidaceae)(8. 75%)和毛螺菌科(Lachnospiraceae)(7. 57%),家牦牛的菌群结构和组成与野牦牛相似。野牦牛粪便中SCFA以乙酸和丙酸为主,乙酸、丙酸、异丁酸、正丁酸和正戊酸的含量均显著高于家牦牛(P <0. 01)。Spearman相关分析显示,野牦牛粪便菌群中紫单胞菌科(Porphyromonadaceae)、拟杆菌科(Bacteroidaceae)、普雷沃氏菌科(Paraprevotellaceae)、理研菌科(Rikenellaceae)和韦荣球菌科(Erysipelotrichaceae)与SCFA具有较强相关性(r> 0. 4),而家牦牛仅有弱相关性(r <0. 3)。说明牦牛后肠道具有丰富的能够促进SCFA生成的益生菌群,进而提高食物的转化效率。
Short chain fatty acids( SCFA),are a crucial form of nutrition for digestion and assimilation of grass for ruminants including the yaks. Gut microbiota degrades chyme,produces SCFA and influences its proportion. Here,we investigated the fecal microbiota and SCFA concentration in wild yak( Bos mutus) and domestic yak( Bos grunniens). We hypothesize that yaks' gut microbiota influence concentration of SCFA and in that way increase the assimilation of nutrients from the grass. Microbes and SCFA concentration of yaks were processed and measured by 16 S r DNA and Gas Chromatography-Mass Spectrometry respectively. The results show that the yak fecal microbes are mainly composed of Firmicutes( 66. 47%,68. 38%)( wild yak and domestic yak respectively,the same below),Bacteroidetes( 26. 00%,26. 16%)and Proteobacteria( 3. 48%,1. 52%) at the phylum level. Furthermore,Ruminococcaceae( 55. 18%,58. 48%),Bacteroidaceae( 8. 75%,9. 59%) and Lachnospiraceae( 7. 57%,6. 14%) at the family level; the domestic yak has the similar mainly composition with wild yak. SCFA concentration( including acetic acid,propionic acid,i-butyric acid,n-butyric acid and n-valeric acid) in wild yak's feces are significantly higher than in domestic yak( P < 0. 01). Spearman cor-relation relationship analysis shows that there is strong positive correlation between Porphyromonadaceae,Bacteroidaceae,Paraprevotellaceae,Rikenellaceae,Erysipelotrichaceae and SCFA concentration in wild yaks( r > 0. 4,P < 0. 05). However,there only has weak positive correlation in domestic yaks microbes. Our result suggests that the digestion of grass in wild yaks is more efficient than in domestic yaks.
Short chain fatty acids( SCFA),are a crucial form of nutrition for digestion and assimilation of grass for ruminants including the yaks. Gut microbiota degrades chyme,produces SCFA and influences its proportion. Here,we investigated the fecal microbiota and SCFA concentration in wild yak( Bos mutus) and domestic yak( Bos grunniens). We hypothesize that yaks' gut microbiota influence concentration of SCFA and in that way increase the assimilation of nutrients from the grass. Microbes and SCFA concentration of yaks were processed and measured by 16 S r DNA and Gas Chromatography-Mass Spectrometry respectively. The results show that the yak fecal microbes are mainly composed of Firmicutes( 66. 47%,68. 38%)( wild yak and domestic yak respectively,the same below),Bacteroidetes( 26. 00%,26. 16%)and Proteobacteria( 3. 48%,1. 52%) at the phylum level. Furthermore,Ruminococcaceae( 55. 18%,58. 48%),Bacteroidaceae( 8. 75%,9. 59%) and Lachnospiraceae( 7. 57%,6. 14%) at the family level; the domestic yak has the similar mainly composition with wild yak. SCFA concentration( including acetic acid,propionic acid,i-butyric acid,n-butyric acid and n-valeric acid) in wild yak's feces are significantly higher than in domestic yak( P < 0. 01). Spearman cor-relation relationship analysis shows that there is strong positive correlation between Porphyromonadaceae,Bacteroidaceae,Paraprevotellaceae,Rikenellaceae,Erysipelotrichaceae and SCFA concentration in wild yaks( r > 0. 4,P < 0. 05). However,there only has weak positive correlation in domestic yaks microbes. Our result suggests that the digestion of grass in wild yaks is more efficient than in domestic yaks.
引文
Aziz Q,Dore J,Emmanuel A,Guarner F,Quigley E M.2013.Gut microbiota and gastrointestinal health:current concepts and future directions.Neurogastroenterol Motil,25:4-15.
Blekhman R,Goodrich J K,Huang K,Sun Q,Bukowski R,Bell J T,Spector T D,Keinan A,Ley R E,Gevers D,Clark A G.2015.Host genetic variation impacts microbiome composition across human body sites.Genome Biol,16:191.
Caporaso J G,Kuczynski J,Stombaugh J,Bittinger K,Bushman F D,Costello E K,Fierer N,Pena A G,Goodrich J K,Gordon J I,Huttley G A,Kelley S T,Knights D,Koenig J E,Ley R E,Lozupone C A,Mc Donald D,Muegge B D,Pirrung M,Reeder J,Sevinsky JR,Turnbaugh P J,Walters W A,Widmann J,Yatsunenko T,Zaneveld J,Knight R.2010.QIIME allows analysis of high-throughput community sequencing data.Nat Methods,7:335-336.
De Oliveira M N,Jewell K A,Freitas F S,Benjamin L A,Totola M R,Borges A C,Moraes C A,Suen G.2013.Characterizing the microbiota across the gastrointestinal tract of a Brazilian Nelore steer.Vet Microbiol,164:307-314.
Demigne C,Remesy C,Morand C.1999.Short Chain Fatty Acids.Springer Netherlands.
De Santis T Z,Hugenholtz P,Larsen N,Rojas M,Brodie E L,Keller K,Huber T,Dalevi D,Hu P,Andersen G L.2006.Greengenes,a chimera-checked 16S rRNA gene database and workbench compatible with ARB.Applied and Environmental Microbiology,72:5069-5072.
Edgar R C.2013.UPARSE:highly accurate OTU sequences from microbial amplicon reads.Nat Methods,10:996-998.
Edgar R C,Haas B J,Clemente J C,Quince C,Knight R.2011.UCH-IME improves sensitivity and speed of chimera detection.Bioinformatics,27:2194-2200.
Gomez A,Espinoza J L,Harkins D M,Leong P,Saffery R,Bockmann M,Torralba M,Kuelbs C,Kodukula R,Inman J,Hughes T,Craig J M,Highlander S K,Jones M B,Dupont C L,Nelson K E.2017.Host genetic control of the oral microbiome in health and disease.Cell Host Microbe,22:269-783.
Goodrich J K,Waters J L,Poole A C,Sutter J L,Koren O,Blekhman R,Beaumont M,Van Treuren W,Knight R,Bell J T,Spector TD,Clark A G,Ley R E.2014.Human genetics shape the gut microbiome.Cell,159:789-799.
Guan L L,Nkrumah J D,Basarab J A,Moore S S.2008.Linkage of microbial ecology to phenotype:correlation of rumen microbial ecology to cattle’feed efficiency.FEMS Microbiol Lett,288(1):85-91.
Van Houtert M F J.1993.The production and metabolism of volatile fatty acids by ruminants fed roughages:a review.Animal Feed Science and Technology,43(3-4):189-225.
Huda-Faujan N,Abdulamir A S,Fatimah A B,Anas O M,Shuhaimi M,Yazid A M,Loong Y Y.2010.The impact of the level of the intestinal short chain fatty acids in inflammatory bowel disease patients versus healthy subjects.The Open Biochemistry Journal,4:53-58.
Koh A,De Vadder F,Kovatcheva-Datchary P,Backhed F.2016.From dietary fiber to host physiology:short-chain fatty acids as key bacterial metabolites.Cell,165:1332-1345.
Kurilshikov A,Wijmenga C,Fu J Y,Zhernakova A.2017.Host genetics and gut microbiome:challenges and perspectives.Trends in Immunology,38:633-647.
Leamy L J,Kelly S A,Nietfeldt J,Legge R M,Ma F,Hua K,Sinha R,Peterson D A,Walter J,Benson A K.2014.Host genetics and diet,but not immunoglobulin A expression,converge to shape compositional features of the gut microbiome in an advanced intercross population of mice.Genome Biol,15:552.
Louis P,Hold G L,Flint H J.2014.The gut microbiota,bacterial metabolites and colorectal cancer.Nat Rev Microbiol,12:661-672.
Lu D,Tiezzi F,Schillebeeckx C,Mc Nulty N P,Schwab C,Shull C,Maltecca C.2018.Host contributes to longitudinal diversity of fecal microbiota in swine selected for lean growth.Microbiome,6:4.
Nicholson J K,Holmes E,Kinross J,Burcelin R,Gibson G,Jia W,Pettersson S.2012.Host-gut microbiota metabolic interactions.Science,336:1262-1267.
Nie Y,Zhou Z,Guan J,Xia B,Luo X,Yang Y,Fu Y,Sun Q.2017.Dynamic changes of yak(Bos grunniens)gut microbiota during growth revealed by polymerase chain reaction-denaturing gradient gel electrophoresis and metagenomics.Asian-Australas J Anim Sci,30:957-966.
Parker A,Lawson M,Vaux L,Pin C.2017.Host-microbe interaction in the gastrointestinal tract.Environ Microbiol,Suppl 1.
Qiu Q,Wang L,Wang K,Yang Y,Ma T,Wang Z,Zhang X,Ni Z,Hou F,Long R,Abbott R,Lenstra J,Liu J.2015.Yak wholegenome resequencing reveals domestication signatures and prehistoric population expansions.Nat Commun,6:10283.
Qiu Q,Zhang G,Ma T,Qian W,Wang J,Ye Z,Cao C,Hu Q,Kim J,Larkin D M,Auvil L,Capitanu B,Ma J,Lewin H A,Qian X,Lang Y,Zhou R,Wang L,Wang K,Xia J,Liao S,Pan S,Lu X,Hou H,Wang Y,Zang X,Yin Y,Ma H,Zhang J,Wang Z,Zhang Y,Zhang D,Yonezawa T,Hasegawa M,Zhong Y,Liu W,Zhang Y,Huang Z,Zhang S,Long R,Yang H,Wang J,Lenstra J A,Cooper D N,Wu Y,Wang J,Shi P,Wang J,Liu J.2012.The yak genome and adaptation to life at high altitude.Nat Genet,44:946-949.
Racine J S.2012.Studio:a platform-independent IDE for R and Sweave.Journal of Applied Econometrics,27:167-172.
Rodriguez-Carrio J,Salazar N,Margolles A,Gonzalez S,Gueimonde M,de Los Reyes-Gavilan C G,Suarez A.2017.Free fatty acids profiles are related to gut microbiota signatures and short-chain fatty acids.Front Immunol,8:823.
R Core Team.2017.R:A Language and Environment for Statistical Computing.(ed by R Foundation for Statistical Computing V,Austria).https://www.R-project.org/.
Sasson G,Kruger Ben-Shabat S,Seroussi E,Doron-Faigenboim A,Shterzer N,Yaacoby S,Berg Miller M E,White B A,Halperin E,Mizrahi I.2017.Heritable bovine rumen bacteria are phylogenetically related and correlated with the cow’s capacity to harvest energy from its feed.m Bio,8(4):703-717.
Seymour W M,Campbell D R,Johnson Z B.2005.Relationships between rumen volatile fatty acid concentrations and milk production in dairy cows:a literature study.Animal Feed Science and Technology,119(1-2):155-169.
Shabat S K,Sasson G,Doron-Faigenboim A,Durman T,Yaacoby S,Berg Miller M E,White B A,Shterze N,Mizrahi I.2016.Specific microbiome-dependent mechanisms underlie the energy harvest efficiency of ruminants.ISME J,10(12):2958-2972.
Tan J,Mc Kenzie C,Potamitis M,Thorburn A N,Mackay C R,Macia L.2014.The role of short-chain fatty acids in health and disease.Adv Immunol,121:91-119.
Topping D L,Clifton P M.2001.Short-chain fatty acids and human colonic function:roles of resistant starch and nonstarch polysaccharides.Physiological Reviews,81:1031-1064.
Wang Q,Garrity G M,Tiedje J M,Cole J R.2007.Naive Bayesian classifier for rapid assignment of rRNA sequences into the new bacterial taxonomy.Applied and Environmental Microbiology,73:5261-5267.
Zhang H,Shao M,Huang H,Wang S,Ma L,Wang H,Hu L,Wei K,Zhu R.2018.The dynamic distribution of small-tail han sheep microbiota across different intestinal segments.Front Microbiol,9:32.
Zhang Z,Xu D,Wang L,Hao J,Wang J,Zhou X,Wang W,Qiu Q,Huang X,Zhou J,Long R,Zhao F,Shi P.2016.Convergent evolution of rumen microbiomes in high-altitude mammals.Curr Biol,26:1873-1879.
Zheng X,Qiu Y,Zhong W,Baxter S,Su M,Li Q,Xie G,Ore B M,Qiao S,Spencer M D,Zeisel S H,Zhou Z,Zhao A,Jia W.2013.A targeted metabolomic protocol for short-chain fatty acids and branched-chain amino acids.Metabolomics,9:818-827.
Zhu B,Wang X,Li L.2010.Human gut microbiome:the second genome of human body.Protein&Cell,1:718-725.
蔡立.1992.中国牦牛.北京:中国农业出版社.
国家林业局.2009.中国重点陆生野生动物资源调查.北京:中国林业出版社.
刘敏雄.1991.反刍动物消化生理学.北京:北京农业大学出版社.
陆仲璘,李孔亮.2005.中国野牦牛的分布、类型及利用.中国草食动物科学,(增刊2):245-247.
Blekhman R,Goodrich J K,Huang K,Sun Q,Bukowski R,Bell J T,Spector T D,Keinan A,Ley R E,Gevers D,Clark A G.2015.Host genetic variation impacts microbiome composition across human body sites.Genome Biol,16:191.
Caporaso J G,Kuczynski J,Stombaugh J,Bittinger K,Bushman F D,Costello E K,Fierer N,Pena A G,Goodrich J K,Gordon J I,Huttley G A,Kelley S T,Knights D,Koenig J E,Ley R E,Lozupone C A,Mc Donald D,Muegge B D,Pirrung M,Reeder J,Sevinsky JR,Turnbaugh P J,Walters W A,Widmann J,Yatsunenko T,Zaneveld J,Knight R.2010.QIIME allows analysis of high-throughput community sequencing data.Nat Methods,7:335-336.
De Oliveira M N,Jewell K A,Freitas F S,Benjamin L A,Totola M R,Borges A C,Moraes C A,Suen G.2013.Characterizing the microbiota across the gastrointestinal tract of a Brazilian Nelore steer.Vet Microbiol,164:307-314.
Demigne C,Remesy C,Morand C.1999.Short Chain Fatty Acids.Springer Netherlands.
De Santis T Z,Hugenholtz P,Larsen N,Rojas M,Brodie E L,Keller K,Huber T,Dalevi D,Hu P,Andersen G L.2006.Greengenes,a chimera-checked 16S rRNA gene database and workbench compatible with ARB.Applied and Environmental Microbiology,72:5069-5072.
Edgar R C.2013.UPARSE:highly accurate OTU sequences from microbial amplicon reads.Nat Methods,10:996-998.
Edgar R C,Haas B J,Clemente J C,Quince C,Knight R.2011.UCH-IME improves sensitivity and speed of chimera detection.Bioinformatics,27:2194-2200.
Gomez A,Espinoza J L,Harkins D M,Leong P,Saffery R,Bockmann M,Torralba M,Kuelbs C,Kodukula R,Inman J,Hughes T,Craig J M,Highlander S K,Jones M B,Dupont C L,Nelson K E.2017.Host genetic control of the oral microbiome in health and disease.Cell Host Microbe,22:269-783.
Goodrich J K,Waters J L,Poole A C,Sutter J L,Koren O,Blekhman R,Beaumont M,Van Treuren W,Knight R,Bell J T,Spector TD,Clark A G,Ley R E.2014.Human genetics shape the gut microbiome.Cell,159:789-799.
Guan L L,Nkrumah J D,Basarab J A,Moore S S.2008.Linkage of microbial ecology to phenotype:correlation of rumen microbial ecology to cattle’feed efficiency.FEMS Microbiol Lett,288(1):85-91.
Van Houtert M F J.1993.The production and metabolism of volatile fatty acids by ruminants fed roughages:a review.Animal Feed Science and Technology,43(3-4):189-225.
Huda-Faujan N,Abdulamir A S,Fatimah A B,Anas O M,Shuhaimi M,Yazid A M,Loong Y Y.2010.The impact of the level of the intestinal short chain fatty acids in inflammatory bowel disease patients versus healthy subjects.The Open Biochemistry Journal,4:53-58.
Koh A,De Vadder F,Kovatcheva-Datchary P,Backhed F.2016.From dietary fiber to host physiology:short-chain fatty acids as key bacterial metabolites.Cell,165:1332-1345.
Kurilshikov A,Wijmenga C,Fu J Y,Zhernakova A.2017.Host genetics and gut microbiome:challenges and perspectives.Trends in Immunology,38:633-647.
Leamy L J,Kelly S A,Nietfeldt J,Legge R M,Ma F,Hua K,Sinha R,Peterson D A,Walter J,Benson A K.2014.Host genetics and diet,but not immunoglobulin A expression,converge to shape compositional features of the gut microbiome in an advanced intercross population of mice.Genome Biol,15:552.
Louis P,Hold G L,Flint H J.2014.The gut microbiota,bacterial metabolites and colorectal cancer.Nat Rev Microbiol,12:661-672.
Lu D,Tiezzi F,Schillebeeckx C,Mc Nulty N P,Schwab C,Shull C,Maltecca C.2018.Host contributes to longitudinal diversity of fecal microbiota in swine selected for lean growth.Microbiome,6:4.
Nicholson J K,Holmes E,Kinross J,Burcelin R,Gibson G,Jia W,Pettersson S.2012.Host-gut microbiota metabolic interactions.Science,336:1262-1267.
Nie Y,Zhou Z,Guan J,Xia B,Luo X,Yang Y,Fu Y,Sun Q.2017.Dynamic changes of yak(Bos grunniens)gut microbiota during growth revealed by polymerase chain reaction-denaturing gradient gel electrophoresis and metagenomics.Asian-Australas J Anim Sci,30:957-966.
Parker A,Lawson M,Vaux L,Pin C.2017.Host-microbe interaction in the gastrointestinal tract.Environ Microbiol,Suppl 1.
Qiu Q,Wang L,Wang K,Yang Y,Ma T,Wang Z,Zhang X,Ni Z,Hou F,Long R,Abbott R,Lenstra J,Liu J.2015.Yak wholegenome resequencing reveals domestication signatures and prehistoric population expansions.Nat Commun,6:10283.
Qiu Q,Zhang G,Ma T,Qian W,Wang J,Ye Z,Cao C,Hu Q,Kim J,Larkin D M,Auvil L,Capitanu B,Ma J,Lewin H A,Qian X,Lang Y,Zhou R,Wang L,Wang K,Xia J,Liao S,Pan S,Lu X,Hou H,Wang Y,Zang X,Yin Y,Ma H,Zhang J,Wang Z,Zhang Y,Zhang D,Yonezawa T,Hasegawa M,Zhong Y,Liu W,Zhang Y,Huang Z,Zhang S,Long R,Yang H,Wang J,Lenstra J A,Cooper D N,Wu Y,Wang J,Shi P,Wang J,Liu J.2012.The yak genome and adaptation to life at high altitude.Nat Genet,44:946-949.
Racine J S.2012.Studio:a platform-independent IDE for R and Sweave.Journal of Applied Econometrics,27:167-172.
Rodriguez-Carrio J,Salazar N,Margolles A,Gonzalez S,Gueimonde M,de Los Reyes-Gavilan C G,Suarez A.2017.Free fatty acids profiles are related to gut microbiota signatures and short-chain fatty acids.Front Immunol,8:823.
R Core Team.2017.R:A Language and Environment for Statistical Computing.(ed by R Foundation for Statistical Computing V,Austria).https://www.R-project.org/.
Sasson G,Kruger Ben-Shabat S,Seroussi E,Doron-Faigenboim A,Shterzer N,Yaacoby S,Berg Miller M E,White B A,Halperin E,Mizrahi I.2017.Heritable bovine rumen bacteria are phylogenetically related and correlated with the cow’s capacity to harvest energy from its feed.m Bio,8(4):703-717.
Seymour W M,Campbell D R,Johnson Z B.2005.Relationships between rumen volatile fatty acid concentrations and milk production in dairy cows:a literature study.Animal Feed Science and Technology,119(1-2):155-169.
Shabat S K,Sasson G,Doron-Faigenboim A,Durman T,Yaacoby S,Berg Miller M E,White B A,Shterze N,Mizrahi I.2016.Specific microbiome-dependent mechanisms underlie the energy harvest efficiency of ruminants.ISME J,10(12):2958-2972.
Tan J,Mc Kenzie C,Potamitis M,Thorburn A N,Mackay C R,Macia L.2014.The role of short-chain fatty acids in health and disease.Adv Immunol,121:91-119.
Topping D L,Clifton P M.2001.Short-chain fatty acids and human colonic function:roles of resistant starch and nonstarch polysaccharides.Physiological Reviews,81:1031-1064.
Wang Q,Garrity G M,Tiedje J M,Cole J R.2007.Naive Bayesian classifier for rapid assignment of rRNA sequences into the new bacterial taxonomy.Applied and Environmental Microbiology,73:5261-5267.
Zhang H,Shao M,Huang H,Wang S,Ma L,Wang H,Hu L,Wei K,Zhu R.2018.The dynamic distribution of small-tail han sheep microbiota across different intestinal segments.Front Microbiol,9:32.
Zhang Z,Xu D,Wang L,Hao J,Wang J,Zhou X,Wang W,Qiu Q,Huang X,Zhou J,Long R,Zhao F,Shi P.2016.Convergent evolution of rumen microbiomes in high-altitude mammals.Curr Biol,26:1873-1879.
Zheng X,Qiu Y,Zhong W,Baxter S,Su M,Li Q,Xie G,Ore B M,Qiao S,Spencer M D,Zeisel S H,Zhou Z,Zhao A,Jia W.2013.A targeted metabolomic protocol for short-chain fatty acids and branched-chain amino acids.Metabolomics,9:818-827.
Zhu B,Wang X,Li L.2010.Human gut microbiome:the second genome of human body.Protein&Cell,1:718-725.
蔡立.1992.中国牦牛.北京:中国农业出版社.
国家林业局.2009.中国重点陆生野生动物资源调查.北京:中国林业出版社.
刘敏雄.1991.反刍动物消化生理学.北京:北京农业大学出版社.
陆仲璘,李孔亮.2005.中国野牦牛的分布、类型及利用.中国草食动物科学,(增刊2):245-247.