The rumen microbial metagenome associated with high methane production in cattle.
BACKGROUND
Methane represents 16 % of total anthropogenic greenhouse gas emissions. It has been estimated that ruminant livestock produce ca. 29 % of this methane. As individual animals produce consistently different quantities of methane, understanding the basis for these differences may lead to new opportunities for mitigating ruminal methane emissions. Metagenomics is a powerful new tool for understanding the composition and function of complex microbial communities. Here we have applied metagenomics to the rumen microbial community to identify differences in the microbiota and metagenome that lead to high- and low-methane-emitting cattle phenotypes.


METHODS
Four pairs of beef cattle were selected for extreme high and low methane emissions from 72 animals, matched for breed (Aberdeen-Angus or Limousin cross) and diet (high or medium concentrate). Community analysis was carried out by qPCR of 16S and 18S rRNA genes and by alignment of Illumina HiSeq reads to the GREENGENES database. Total genomic reads were aligned to the KEGG genes databasefor functional analysis.


RESULTS
Deep sequencing produced on average 11.3 Gb per sample. 16S rRNA gene abundances indicated that archaea, predominantly Methanobrevibacter, were 2.5× more numerous (P = 0.026) in high emitters, whereas among bacteria Proteobacteria, predominantly Succinivibrionaceae, were 4-fold less abundant (2.7 vs. 11.2 %; P = 0.002). KEGG analysis revealed that archaeal genes leading directly or indirectly to methane production were 2.7-fold more abundant in high emitters. Genes less abundant in high emitters included acetate kinase, electron transport complex proteins RnfC and RnfD and glucose-6-phosphate isomerase. Sequence data were assembled de novo and over 1.5 million proteins were annotated on the subsequent metagenome scaffolds. Less than half of the predicted genes matched matched a domain within Pfam. Amongst 2774 identified proteins of the 20 KEGG orthologues that correlated with methane emissions, only 16 showed 100 % identity with a publicly available protein sequence.


CONCLUSIONS
The abundance of archaeal genes in ruminal digesta correlated strongly with differing methane emissions from individual animals, a finding useful for genetic screening purposes. Lower emissions were accompanied by higher Succinovibrionaceae abundance and changes in acetate and hydrogen production leading to less methanogenesis, as similarly postulated for Australian macropods. Large numbers of predicted protein sequences differed between high- and low-methane-emitting cattle. Ninety-nine percent were unknown, indicating a fertile area for future exploitation.
Rowett Institute of Nutrition and Health, University of Aberdeen, Bucksburn, Aberdeen, AB21 9SB, UK. john.wallace@abdn.ac.uk.
R John
RJ
Wallace
SRUC, West Mains Road, Edinburgh, EH9 3JG, UK. John.Rooke@sruc.ac.uk.
John A
JA
Rooke
Rowett Institute of Nutrition and Health, University of Aberdeen, Bucksburn, Aberdeen, AB21 9SB, UK. n.mckain@abdn.ac.uk.
Nest
N
McKain
SRUC, West Mains Road, Edinburgh, EH9 3JG, UK. Carol-Anne.Duthie@sruc.ac.uk.
Carol-Anne
CA
Duthie
SRUC, West Mains Road, Edinburgh, EH9 3JG, UK. Jimmy.Hyslop@sac.co.uk.
Jimmy J
JJ
Hyslop
SRUC, West Mains Road, Edinburgh, EH9 3JG, UK. Dave.Ross@sruc.ac.uk.
David W
DW
Ross
SRUC, West Mains Road, Edinburgh, EH9 3JG, UK. Tony.Waterhouse@sruc.ac.uk.
Anthony
A
Waterhouse
Edinburgh Genomics, The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush, Edinburgh, EH25 9RG, UK. mick.watson@roslin.ed.ac.uk.
Mick
M
Watson
SRUC, West Mains Road, Edinburgh, EH9 3JG, UK. rainer.roehe@sruc.ac.uk.
Rainer
R
Roehe
D000818:Animals; D001105:Archaea / Q000145:classification / Q000235:genetics; D001315:Australia; D001419:Bacteria / Q000145:classification / Q000235:genetics; D002417:Cattle; D054892:Metagenome / Q000235:genetics*; D056186:Metagenomics; D008697:Methane / Q000096:biosynthesis* / Q000378:metabolism; D064307:Microbiota / Q000235:genetics*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012417:Rumen / Q000378:metabolism / Q000382:microbiology*
D012336:RNA, Ribosomal, 16S; D008697:Methane