Molecular Diversity of Rumen Methanogens from Sheep in Western Australia

Abstract

The molecular diversity of rumen methanogens in sheep in Australia was investigated by using individual 16S rRNA gene libraries prepared from the rumen contents obtained from six merino sheep grazing pasture (326 clones), six sheep fed an oaten hay-based diet (275 clones), and five sheep fed a lucerne hay-based diet (132 clones). A total of 733 clones were examined, and the analysis revealed 65 phylotypes whose sequences (1,260 bp) were similar to those of cultivated methanogens belonging to the order Methanobacteriales. Pasturegrazed sheep had more methanogen diversity than sheep fed either the oaten hay or lucerne hay diet. Methanobrevibacter strains SM9, M6, and NT7 accounted for over 90% of the total number of clones identified. M6 was more prevalent in grazing sheep, and SM9, despite being found in 16 of the 17 sheep, was more prevalent in sheep fed the lucerne-based diet. Five new species were identified. Two of these species exhibited very little sequence similarity to any cultivated methanogens and were found eight times in two of the six sheep that were grazing pasture. These unique sequences appear to represent a novel group of rumen archaea that are atypical for the rumen environment.The rumen is a unique environment and is home to billions of microbes, including bacteria, methanogenic archaea, protozoa, and fungi. These different microbes form a complex community of organisms that interact with one another and play an important role in the digestion of feed and the supply of energy to the host in the form of volatile fatty acids and microbial protein. In the past decade, there has been an increasing amount of interest in the rumen methanogenic archaea. This has primarily resulted from the role of these organisms in global warming due to the production of methane by domesticated livestock.In Australia, ruminant livestock are the single largest source of agricultural greenhouse gas emissions and alone account for at least 12% of Australian's total anthropomorphic national emissions (3). In New Zealand, enteric emissions are responsible for approximately 60% of that country's total greenhouse gas emissions (27). Approximately 95.5% of the methane emitted by ruminants is produced in the rumen (3), and the associated loss of energy for the ruminant has been estimated to be between 2 and 15% of the gross energy intake (18,25,40).Methane production is influenced by feed intake and the digestibility of the dry matter in the feed that is consumed. The effects of diet on changes in the diversity and numbers of a wide range of bacterial species in the rumen are known (20,30,36,37,45), but there is little information available concerning the composition of the methanogen population and their numbers of methanogens with regard to diet. Therefore, it is necessary to understand the diversity of methanogens in the rumen.In the past, methanogens from the digestive tracts of domesticated ruminants were identified by classical microbiological techniques (46). However, because of the fastidious growth requirements of ru...

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

“…The most predominant species of methanogens in the three groups were related to the genus Methanobrevibacter. This is in agreement with Pei, Mao, Cheng and Zhu previous studies indicating species belonging to the genus Methanobrevibacter as the major methanogens in the rumen of sheep (Wright et al, 2004 andSkillman et al, 2004;Nicholson et al, 2007) and cattle (Whitford et al, 2001;Skillman et al, 2006;Nicholson et al, 2007;Ouwerkerk et al, 2008), of which some were associated with the rumen protozoa (Tokura et al, 1999a;Sharp et al, 1998;Chargan and Ushida, 2004). The predominance of this genus has also been suggested before based on cultivation studies (e.g.…”

“…Lin et al (1997) targeting the small subunit ribosomal RNA through oligonucleotide probes demonstrated that animal species and diet affected the quantity and dominance of methanogens in the rumen. The molecular diversity of methanogens also confirmed this (Wright et al, 2004 and. Other factors such as the environment, fitness and age of animal could also have some effect on the microbial diversity (Wright et al, 2006).…”

“…The PCR primers used to amplify 16S rRNA genes were the methanogen-specific primers, Met86F (5 0 -GCT CAG TAA CAC GTG G-3 0 ) and Met1340R (5 0 -CGG TGT GTG CAA GGA G-3 0 ) (Wright et al, 2004). The amplification conditions were: initial denaturation at 948C for 3 min, then 40 cycles of 948C for 30 s, 588C for 30 s and 728C for 90 s, and last extension at 728C for 10 min.…”

“…In the rumen of merino wethers in Western Australia, more than 97% (713 of 733) of the clones were identified as Methanobrevibacter, while Mbr. ruminantium only accounted for 1.6% (Wright et al, 2004). However, the rumen methanogenic population of cattle of Chinese native breeds has received little attention.…”

Diversity of gut methanogens in herbivorous animals

“…Under the three diets that were investigated by Wright et al (2004), Methanobrevibacter-related archaea were found to be the dominant methanogens (95.7% to 100.0%) in the rumen of Merino sheep from Queensland, Australia. Overall, Wright et al (2004) found M. millerae to be the most prevalent, but its representation varied noticeably between diets (44.0% to 96.2%).…”