The Håkon Mosby Mud Volcano (HMMV) is an active methane driven cold seep at the bottom of the Barents Sea (Fig. 1). This cold seep harbors a highly diverse microbial community that uses the reverse methanogenesis pathway to anaerobically oxidize methane [1]. The energy released is used by the prokaryotic community. To understand the diversity and metabolic capacity of the prokaryotic community we created a metagenomic library using 454 GS FLX technology. We combined taxonomic binning of sequences with metabolic annotation to analyze the metabolic capacity of different taxa found in the HMMV microbial community.

1. Figure 1. Location of the HMMV sampling site
Metagenome pyrosequencing of Håkon Mosby Mud Volcano sediments
Othilde Elise Håvelsrud1, 2, 3, Thomas H.A. Haverkamp3,4, Tom Kristensen2, Anne Gunn Rike1and Kjetill S. Jakobsen3,4
1 Norwegian Geotechnical Institute, Oslo, Norway. 2 Department of Molecular Biosciences, University of Oslo, Oslo, Norway. 3 Microbial Evolution Research
Group, University of Oslo, Oslo, Norway. 4 Centre for Evolutionary and Ecological Synthesis, University of Oslo, Oslo, Norway.
Methods
A sediment sample (2-20cm bsf) obtained from the
northern outer rim of the HMMV, was used for DNA
extraction.
Extracted DNA was sequenced following the standard
GS FLX protocols using the 70x75 format
PicoTiterPlateTM on a GS FLX instrument at Roche
Penzburg.
Analysis of the metagenome was done as shown in the
flowchart (Fig. 2) using MG-RAST [2], KAAS [3] and
MEGAN [4].
References
[1] Niemann et al. (2006). Novel microbial communities
of the Håkon Mosby mud volcano and their role as a
methane sink. Nature 443: 854-858.
[2] Meyer F, et al. (2008). The metagenomics RAST
server - a public resource for the automatic phylogenetic
and functional analysis of metagenomes. BMC
Bioinformatics 9: Article No.: 386.
[3] Moriya Y et al. (2007). KAAS: an automatic genome
annotation and pathway reconstruction server. Nucleic
Acids Res 35: W182-185.
[4] Huson et al. (2009). Methods for comparative
metagenomics. BMC Bioinformatics 10 Suppl 1: S12.
Contact information
e-mail: thhaverk@bio.uio.no
webpage T. Haverkamp:
http://www.cees.uio.no/about/staff/frida/407495.xml
Figure 2. Flowchart to generate taxon specific
annotation of metagenomic reads
Results
The HMMV metagenome has 392.628 reads, average
length: 222 bp; average GC content 44 %.
68% of the HMMV metagenome reads were assigned to
three dominating taxa: Proteobacteria, Archaea and
Bacteriodetes (Fig. 3). KAAS annotated reads showed
metabolic capacity differences between these groups for:
Anaerobic oxidation of methane (Fig.4); Methane
metabolism (Fig.5); Sulfur metabolism (Fig.6) and
Nitrogen metabolism (Fig.7).
Results
Conclusions
The dominating taxa within the HMMV prokaryotic
community belong to Proteobacteria, Bacteroidetes and
Archaea.
The combination of taxonomic binning and subsequent
annotation of the binned reads, as performed here is
particularly powerful in revealing taxon- or group-
specific metabolism.
Essential steps in anaerobic oxidation of methane are
only found among archaeal reads.
The Proteobacteria have the capacity for aerobic
oxidation of methane.
Only among proteobacterial reads a full pathway for
sulfate reduction could be detected.
The Bacteriodetes at the HMMV are not involved in
direct methane metabolism.
Introduction
The Håkon Mosby Mud Volcano (HMMV) is an active
methane driven cold seep at the bottom of the Barents
Sea (Fig. 1). This cold seep harbors a highly diverse
microbial community that uses the reverse
methanogenesis pathway to anaerobically oxidize
methane [1]. The energy released is used by the
prokaryotic community.
To understand the diversity and metabolic capacity of
the prokaryotic community we created a metagenomic
library using 454 GS FLX technology. We combined
taxonomic binning of sequences with metabolic
annotation to analyze the metabolic capacity of different
taxa found in the HMMV microbial community.
Acknowledgements
The VISTA program for financial support.
Jochen M. Knies (NGU, Trondheim, Norway) for
providing samples.
The crew of RV Håkon Mosby (IMR, Bergen, Norway).
Roche Penzburg for 454 sequencing.
Lex Nederbragt for help with bioinformatic analyses
(NSC, Oslo, Norway).
Figure 3. A) Taxonomic assignment of HMMV
metagenome reads using MEGAN. B)
Proteobacterial taxa. C) Archaeal taxa.
Figure 4. KEGG map folate biosynthesis,
including the reverse methanogenesis pathway.
Figure 5. KEGG map of methane metabolism.
Figure 6. KEGG map of sulfur metabolism
Figure 7. KEGG map of nitrogen metabolism