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      Comparative Analysis of the Methanogen Diversity in Horse and Pony by Using mcrA Gene and Archaeal 16S rRNA Gene Clone Libraries

      research-article
      Author(s): 1 , 2 , 1 , *
      Publication date (Electronic):
      Journal: Archaea
      Publisher: Hindawi Publishing Corporation

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          Abstract

          Comparative analysis of methanogen compositions in the feces of horse and pony was carried out by constructing the α -subunit of methyl coenzyme-M reductase ( mcrA) gene and 16S ribosomal RNA gene (16S rRNA) clone libraries. The mcrA clone library analysis indicated that Methanomicrobiales was predominant in both horse and pony. Furthermore, most of the clones of the 16S rRNA gene library showed that Methanomicrobiales was also predominant in horse and pony, but the LIBSHUFF analysis showed that the horse and pony libraries were significantly different ( P < 0.05). Most of operational taxonomic units (OTUs) showed low similarity to the identified methanogens in both the mcrA and the 16S rRNA clone libraries. The results suggest that horse and pony harbor unidentified and novel methanogens in their hindgut. The methanogen population was higher in horse than in pony; however, the anaerobic fungal population was similar in horse and pony. The methanogen diversity was different between two breeds of Equus caballus.

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          The Ribosomal Database Project (RDP-II): previewing a new autoaligner that allows regular updates and the new prokaryotic taxonomy.

          J. R. Cole, B. Chai, T Marsh … (2003)
          The Ribosomal Database Project-II (RDP-II) pro-vides data, tools and services related to ribosomal RNA sequences to the research community. Through its website (http://rdp.cme.msu.edu), RDP-II offers aligned and annotated rRNA sequence data, analysis services, and phylogenetic inferences (trees) derived from these data. RDP-II release 8.1 contains 16 277 prokaryotic, 5201 eukaryotic, and 1503 mitochondrial small subunit rRNA sequences in aligned and annotated format. The current public beta release of 9.0 debuts a new regularly updated alignment of over 50 000 annotated (eu)bacterial sequences. New analysis services include a sequence search and selection tool (Hierarchy Browser) and a phylogenetic tree building and visualization tool (Phylip Interface). A new interactive tutorial guides users through the basics of rRNA sequence analysis. Other services include probe checking, phylogenetic placement of user sequences, screening of users' sequences for chimeric rRNA sequences, automated alignment, production of similarity matrices, and services to plan and analyze terminal restriction fragment polymorphism (T-RFLP) experiments. The RDP-II email address for questions or comments is rdpstaff@msu.edu.
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            The mcrA gene as an alternative to 16S rRNA in the phylogenetic analysis of methanogen populations in landfill.

            Philip Luton, Jonathan Wayne, Richard J Sharp … (2002)
            Inferred amino acid sequences of the methyl coenzyme-M reductase (mcrA) gene from five different methanogen species were aligned and two regions with a high degree of homology flanking a more variable region were identified. Analysis of the DNA sequences from the conserved regions yielded two degenerate sequences from which a forward primer, a 32-mer, and a reverse primer, a 23-mer, could be derived for use in the specific PCR-based detection of methanogens. The primers were successfully evaluated against 23 species of methanogen representing all five recognized orders of this group of Archaea, generating a PCR product between 464 and 491 bp. Comparisons between the mcrA and 16S small subunit rRNA gene sequences using PHYLIP demonstrated that the tree topologies were strikingly similar. Methods were developed to enable the analysis of methanogen populations in landfill using the mcrA gene as the target. Two landfill sites were examined and 63 clones from a site in Mucking, Essex, and 102 from a site in Odcombe, Somerset, were analysed. Analysis revealed a far greater diversity in the methanogen population within landfill material than has been seen previously.
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              Quantitation and diversity analysis of ruminal methanogenic populations in response to the antimethanogenic compound bromochloromethane.

              Stuart E Denman, Nigel W Tomkins, Christopher S McSweeney (2007)
              Methyl coenzyme-M reductase A (mcrA) clone libraries were generated from microbial DNA extracted from the rumen of cattle fed a roughage diet with and without supplementation of the antimethanogenic compound bromochloromethane. Bromochloromethane reduced total methane emissions by c. 30%, with a resultant increase in propionate and branched chain fatty acids. The mcrA clone libraries revealed that Methanobrevibacter spp. were the dominant species identified. A decrease in the incidence of Methanobrevibacter spp. from the clone library generated from bromochloromethane treatment was observed. In addition, a more diverse methanogenic population with representatives from Methanococcales, Methanomicrobiales and Methanosacinales orders was observed for the bromochloromethane library. Sequence data generated from these libraries aided in the design of an mcrA-targeted quantitative PCR (qPCR) assay. The reduction in methane production by bromochloromethane was associated with an average decrease of 34% in the number of methanogenic Archaea when monitored with this qPCR assay. Dissociation curve analysis of mcrA amplicons showed a clear difference in melting temperatures for Methanobrevibacter spp. (80-82 degrees C) and all other methanongens (84-86 degrees C). A decrease in the intensity of the Methanobrevibacter spp. specific peak and an increase for the other peak in the bromochloromethane-treated animals corresponded with the changes within the clone libraries.
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                Author and article information

                Journal
                Journal ID (nlm-ta): Archaea
                Journal ID (iso-abbrev): Archaea
                Journal ID (publisher-id): ARCHAEA
                Title: Archaea
                Publisher: Hindawi Publishing Corporation
                ISSN (Print): 1472-3646
                ISSN (Electronic): 1472-3654
                Publication date Collection: 2014
                Publication date (Electronic): 30 January 2014
                Volume: 2014
                Electronic Location Identifier: 483574
                Affiliations
                1Graduate School of Bioresources, Mie University, 1577 Kurimamachiya-Cho, Tsu, Mie 514-8507, Japan
                2Livestock Breeding and Veterinary Department, Yangon Diagnostic Laboratory, Yangon 11121, Myanmar
                Author notes

                Academic Editor: Stefan Spring

                Author information
                http://orcid.org/0000-0002-0122-2735
                Article
                DOI: 10.1155/2014/483574
                PMC ID: 3941164
                PubMed ID: 24678264
                SO-VID: 88dec00a-79fc-4fa5-bcd7-ca6eb0c4fc1b
                Copyright © Copyright © 2014 K.-O. Lwin and H. Matsui.
                License:

                This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                Date received : 24 May 2013
                Date revision received : 15 October 2013
                Date accepted : 21 October 2013
                Categories
                Subject: Research Article

                ScienceOpen disciplines: Animal science & Zoology
                Data availability:
                ScienceOpen disciplines: Animal science & Zoology

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