Hostname: page-component-669899f699-7tmb6 Total loading time: 0 Render date: 2025-04-25T05:16:41.566Z Has data issue: false hasContentIssue false
  • English
  • Français

Diversity, abundance and novel 16S rRNA gene sequences of methanogens in rumen liquid, solid and epithelium fractions of Jinnan cattle

Published online by Cambridge University Press:  24 August 2009

Cai-Xia Pei ,
Sheng-Yong Mao ,
Yan-Fen Cheng  and
Wei-Yun Zhu
Cai-Xia Pei
Affiliation:
Laboratory of Gastrointestinal Microbiology, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, Jiangsu, 210095, China
Sheng-Yong Mao
Affiliation:
Laboratory of Gastrointestinal Microbiology, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, Jiangsu, 210095, China
Yan-Fen Cheng
Affiliation:
Laboratory of Gastrointestinal Microbiology, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, Jiangsu, 210095, China
Wei-Yun Zhu*
Affiliation:
Laboratory of Gastrointestinal Microbiology, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, Jiangsu, 210095, China
*
†E-mail: [email protected]
Get access

Abstract

Three methanogen 16S rRNA gene clone libraries were constructed from liquid (LM), solid (SM) and epithelium (EM) fractions taken from the rumen of Jinnan cattle in China. After the amplification by PCR using methanogen-specific primers Met86F and Met1340R, equal quantities of PCR products from the same fractions from each of the four cattle were mixed together and used to construct the three libraries. Sequence analysis showed that the 268 LM clones were divided into 35 phylotypes with 18 sequences of phylotypes affiliated with the genus Methanobrevibacter (84.3% of clones). The 135 SM clones were divided into 19 phylotypes with 11 phylotypes affiliated with the genus Methanobrevibacter (77.8%). The 267 EM clones were divided into 33 phylotypes with 15 phylotypes affiliated with the genus Methanobrevibacter (77.2%). Clones closely related to Methanomicrobium mobile and Methanobrevibacter wolinii were only found in the LM library, and those to Methanobrevibacter ruminantium and Methanobrevibacter gottschalkii only in the SM library. LM library comprised 12.4% unidentified euryarchaeal clones, SM library 23.7% and EM library 25.5%, respectively. Five phylotypes (accession number: EF055528 and EF055531–EF055534) did not belong to the Euryarchaeota sequences we had known. One possible new genus (represented by phylotype E17, accession number EF055528) belonging to Methanobacteriaceae was identified from EM library. Quantitative real-time PCR for the first time revealed that epithelium fraction had significantly higher density of methanogens, with methanogenic mcrA gene copies (9.95 log 10 (copies per gram of wet weight)) than solid (9.26, P < 0.01) and the liquid (8.44, P < 0.001). The three clone libraries also appeared different in Shannon index (EM library 2.12, LM library 2.05 and SM library 1.73). Our results showed that there were apparent differences in the methanogenic diversity and abundance in the three different fractions within the rumen of Jinnan cattle, with Methanobrevibacter species predominant in all the three libraries and with epithelium fraction having more unknown species and higher density of methanogens.

Keywords

rumen methanogenmolecular diversitydifferent fractions
Type
Full Paper
Information
animal , Volume 4 , Issue 1 , January 2010 , pp. 20 - 29
Copyright
Copyright © The Animal Consortium 2009

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

Article purchase

Temporarily unavailable

References

Bonin, AS, Boone, DR 2006. The order Methanobacteriales. In The prokaryotes (ed. M Dworkin, S Falkow, E Rosenberg, KH Schleifer and E Stackenbrandt), 3rd edition pp. 231–243. Springer, New York.CrossRefGoogle Scholar
Chargan, I, Tokura, M, Jouany, JP, Ushida, K 1999. Detection of methanogenic archaea associated with rumen ciliate protozoa. The Journal of General and Applied Microbiology 45, 305–308.Google Scholar
Chargan, I, Ushida, K 2004. Detection of methanogens and proteobacteria from a single cell of rumen ciliate protozoa. The Journal of General and Applied Microbiology 50, 203–212.Google Scholar
Cheng, KJ, McAllister, TA 1997. Compartmentation in the rumen. In The rumen microbial ecosystem (ed. PN Hobson and CS Stewart), pp. 492–522. Blackie Academic and Professional Publishers, London.Google Scholar
Cheng, YF, Mao, SY, Pei, CX, Liu, JX, Zhu, WY 2006. Detection and diversity analysis of rumen methanogens in co-cultures with anaerobic fungi. Acta Microbiologica Sinica 46, 879–883.Google Scholar
Cheng, YF, Edwards, JE, Allison, GG, Zhu, WY, Theodorou, MK 2009. Diversity and activity of enriched ruminal cultures of anaerobic fungi and methanogens grown together in consecutive batch culture. Bioresource Technology 100, 4821–4828.Google Scholar
Dehority, BA, Grubb, JA 1981. Bacterial population adherent to the epithelium on the Roo of the dorsal rumen of sheep. Applied and Environmental Microbiology 41, 1424–1427.Google Scholar
Denman, SE, Tomkins, NW, McSweeney, CS 2007. Quantitation and diversity analysis of ruminal methanogenic populations in response to the antimethanogenic compound bromochloromethane. FEMS Microbiology Ecology 62, 313–322.Google Scholar
Janssen, PH, Kirs, M 2008. Structure of the archaeal community of the rumen. Applied and Environmental Microbiology 74, 3619–3625.Google Scholar
Johnson, KA, Johnson, DE 1995. Methane emissions from cattle. Journal of Animal Science 73, 2483–2492.Google Scholar
Lin, C, Raskin, L, Stahl, DA 1997. Microbial community structure in gastrointestinal tracts of domestic animals: comparative analyses using rRNA-targeted oligonucleotide probes. FEMS Microbiology Ecology 22, 281–294.Google Scholar
McAllister, TA, Bae, HD, Jones, GA, Cheng, KJ 1994. Microbial attachment and feed digestion in the rumen. Journal of Animal Science 72, 3004–3018.Google Scholar
McCowan, RP, Cheng, KJ, Bailey, CBM, Costerton, JW 1978. Adhesion of bacteria to epithelial cell surfaces within the reticulo-rumen of cattle. Applied and Environmental Microbiology 35, 149–155.CrossRefGoogle ScholarPubMed
Miller, TL, Wolin, MJ, Hongxue, Z, Bryant, MP 1986. Characteristics of methanogens isolated from bovine rumen. Applied and Environmental Microbiology 51, 201–202.Google Scholar
Nicholson, MJ, Evans, PN, Joblin, KN 2007. Analysis of methanogen diversity in the rumen using temporal temperature gradient gel electrophoresis: identification of uncultured methanogens. Microbial Ecology 54, 141–150.Google Scholar
Ouwerkerk, D, Turner, AF, Klieve, AV 2008. Diversity of methanogens in ruminants in Queensland. Australian Journal of Experimental Agriculture 48, 722–725.Google Scholar
Schloss, PD, Handelsman, J 2005. Introducing DOTUR, a computer program for defining operational taxonomic units and estimating species richness. Applied and Environmental Microbiology 71, 1501–1506.CrossRefGoogle ScholarPubMed
Sharp, R, Ziemer, CJ, Stern, MD, Stahl, DA 1998. Taxon-specific associations between protozoal and methanogen populations in the rumen and a model system. FEMS Microbiology Ecology 26, 71–78.CrossRefGoogle Scholar
Shin, EC, Choi, BR, Lim, WJ, Hong, SY, An, CL, Cho, KM, Kim, YK, An, JM, Kang, JM, Lee, SS, Kim, H, Yun, HD 2004. Phylogenetic analysis of archaea in three fractions of cow rumen based on the 16S rDNA sequence. Anaerobe 10, 313–319.CrossRefGoogle ScholarPubMed
Skillman, LC, Evans, PN, Naylor, GE, Morvan, B, Jarvis, GN, Joblin, KN 2004. 16S ribosomal DNA-directed PCR primers for ruminal methanogens and identification of methanogens colonising young lambs. Anaerobe 10, 277–285.Google Scholar
Skillman, LC, Evans, PN, Strömp, C, Joblin, KN 2006. 16S rDNA directed PCR primers and detection of methanogens in the bovine rumen. Letters in Applied Microbiology 42, 222–228.CrossRefGoogle ScholarPubMed
Tajima, K, Nagamine, T, Matsui, H, Nakamura, M, Rustam, I, Aminov, RI 2001. Phylogenetic analysis of archaeal 16S rRNA libraries from the rumen suggests the existence of a novel group of archaea not associated with known methanogens. FEMS Microbiology Letters 200, 67–72.Google Scholar
Theodorou, MK, Zhu, WY, Rickers, A, Nielsen, BB, Gull, K, Trinci, APJ 1996. Biochemistry and ecology of anaerobic fungi. In Mycota VI (ed. DH Howard and JD Miller), pp. 265–295. Springer-Verlag, Berlin.Google Scholar
Tokura, M, Chagan, I, Ushida, K, Kojima, Y 1999a. Phylogenetic study of methanogens associated with rumen ciliates. Current Microbiology 39, 123–128.Google Scholar
Tokura, M, Tajima, K, Ushida, K 1999b. Isolation of Methanobrevibacter sp. as a ciliate-associated ruminal methanogen. The Journal of General and Applied Microbiology 45, 43–47.Google Scholar
Whitford, MF, Teather, RM, Forster, RJ 2001. Phylogenetic analysis of methanogens from the bovine rumen. BMC Microbiology 1, 5. doi:10.1186/1471-2180-1-5.Google Scholar
Wright, ADG, Williams, AJ, Winder, B, Christophersen, CT, Rodgers, SL, Smith, KD 2004. Molecular diversity of rumen methanogens from sheep in Western Australia. Applied and Environmental Microbiology 70, 1263–1270.Google Scholar
Wright, ADG, Toovey, AF, Pimm, CL 2006. Molecular identification of methanogenic archaea from sheep in Queensland, Australia reveals more uncultured novel archaea. Anaerobe 12, 134–139.CrossRefGoogle ScholarPubMed
Wright, ADG, Auckland, CH, Lynn, DH 2007. Molecular diversity of methanogens in feedlot cattle from Ontario and Prince Edward Island, Canada. Applied and Environmental Microbiology 73, 4206–4210.Google Scholar
Wright, ADG, Ma, X, Obispo, NE 2008. Methanobrevibacter phylotypes are the dominant methanogens in sheep from Venezuela. Microbial Ecology 56, 390–394.Google Scholar
Yang, WZ, Beauchemin, KA, Rode, LM 2001. Effect of dietary factors on distribution and chemical composition of liquid- or solid-associated bacterial populations in the rumen of dairy cows. Journal of Animal Science 79, 2736–2746.Google Scholar
Zoetendal, EG, Akkermans, AD, De Vos, WM 1998. Temperature gradient gel electrophoresis analysis from human fecal samples reveals stable and host-specific communities of active bacteria. Applied and Environmental Microbiology 64, 3854–3859.Google Scholar