--> Microbes and Microbial Diagenesis in Methane-rich Sediments, by W. Ussler III, S.J. Hallam, C.K. Paull, V.J. Orphan, W.S. Borowski, M.K. Thompson, and E.F. DeLong; #90035 (2004)

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MICROBES AND MICROBIAL DIAGENESIS IN METHANE-RICH SEDIMENTS

W. Ussler III1, S. J. Hallam1, C. K. Paullv, V. J. Orphan2, W. S. Borowski3, M. K. Thompson3, and E. F. DeLong1
1 Monterey Bay Aquarium Research Institute, Moss Landing, CA
2 California Institute of Technology, Pasadena, CA
3 Eastern Kentucky University, Richmond, KY

The sulfate-methane interface (SMI) is a fundamental biogeochemical boundary that occurs in methane-rich and gas-hydrate-bearing continental margin sediments worldwide. The SMI marks the transition between sulfate-bearing sediments in contact with seawater and sulfate-depleted sediments at depth. In methane-rich sediments that often overlie gas hydrate deposits, sulfate depletion occurs at shallow depths (<50 mbsf) and methane concentrations rapidly increase below the SMI. Dissolved inorganic carbon concentrations (DIC) and light carbon isotope anomalies are often sharply defined across the SMI implying that 13C-depleted methane has been oxidized by microorganisms enriching the DIC pool in light carbon.

Recent microbiological and stable isotope studies of methane-oxidizing archaea (MOA) and sulfate-reducing bacteria (SRB) in seafloor sediments associated with methane venting have shown that these microorganisms are intimately involved in the anaerobic oxidation of methane (e.g., Orphan et al., 2001; PNAS 99:7663-7668). However, the ecological zonation and abundance of MOA and SRB across the SMI and their role in controlling biogeochemical patterns in deep-sea sediments have not been clearly established.

In many stratigraphic sections the effects of microbial activity are entombed in the accumulating sedimentary record. Early stages of diagenesis that occur in methane-rich sediments are represented by isotopic changes in the carbon and sulfur systems that are consistent with microbially-mediated diagenesis and isotope fractionation. Thus, combined geochemical and microbiological studies of modern marine sediments offer the possibility of better understanding the role microbes have in sedimentary diagenesis and how their effects on the carbon and sulfur systems might be preserved in the geologic record.

Methods

As part of a geochemical survey of the Blake Ridge Depression conducted in July 2000 from the R/V Cape Hatteras (see Paull and Ussler, this volume), pore water was extracted from whole-round sections of 33 sediment cores and analyzed shipboard for sulfate, chloride, dissolved inorganic carbon, ammonium, and methane concentrations. Preserved gas samples were subsequently analyzed for methane and carbon dioxide 13C isotopic composition. Sediment subsamples were analyzed for percentage carbonate, 13C and 18O in carbonate, percentage sulfide sulfur and 34S sulfide.

Microbial community structure associated with a deep SMI (~12 mbsf) was examined in one of the piston cores collected during the July 2000 geochemical survey (PC-26). A nearby companion core (PC-28) was also collected and sectioned at higher spatial resolution across the SMI zone, which was identified using shipboard sulfate measurements. Sixteen 15-cm3 sediment subsamples of PC-26 and twenty 15-cm3 subsamples of PC-28 were collected shipboard immediately after recovery on deck at a systematic spacing along the core and placed into cryovials and frozen for later DNA analysis. Total genomic DNA extracted from sediment using established protocols was used to generate archaeal and bacterial small subunit ribosomal RNA (SSU rRNA) gene libraries. These libraries were screened using both restriction fragment length polymorphism (RFLP) and clone-library sequencing approaches. FISH (fluorescence in situ hybridization) was used to visualize and count specific MOA and SRB groups (i.e., ANME-1 and Desulfosarcina respectively).

Results and Discussion

In PC-26 sulfate concentrations decreased linearly with depth (r2=0.991) until depletion at 12.36 mbsf. Except for slight increases in methane concentration at 0.76 mbsf (~27 µM) and at 6.73 mbsf (38 µM), samples from PC-26 above 12 mbsf had typical background methane concentrations (~2 µM). However, below 12 mbsf methane concentration increased rapidly to 1,500 µM at 13.18 mbsf. Sulfate depletion and the sharp rise in methane concentration indicate that the SMI is a zone at least 50 cm thick centered at 12.11 mbsf. Methane 13C values in PC-26 and a nearby companion core PC-28 below the SMI zone were tightly clustered (-98.26+1.35‰, PDB, n=8) and isotopically lighter than samples from above (-89.0+8.28‰, PDB, n=11). These isotopic values indicate that the methane contained in these cores is microbial in origin. Maximum DIC concentrations in PC-26 (~9 mM) were found between 4 and 7 mbsf, and declined smoothly towards seawater-like values (~3 mM) above and below this depth. A spike in DIC concentration (9.3 mM) occurred near the center of the SMI zone at 12.11 mbsf in PC-26. Stable isotopic values of bulk sedimentary carbonate throughout PC-26 and PC-28 (13C=0.22+0.37‰, PDB; 18O=-0.47+0.88‰, PDB, n=43) resemble those for pelagic carbonates, indicating that limited amounts of methane-derived authigenic carbonates have formed. Sulfide sulfur is distinctly more abundant in sediment within the SMI zone (~0.35% S) compared to background sediments between the seafloor and 10 mbsf (0.05-0.2% S) and has a very large 34S enrichment (up to 34S~+25‰, CDT) relative to background sediments (34S~-35‰, CDT), indicating that a large transfer of sulfate to sulfide sulfur has occurred within the SMI zone.

In PC-26, SSU rRNA libraries at 0.76 mbsf and 11.82 mbsf were found to contain ribosomal sequences corresponding to MOA, including ANME-1 and ANME-2 and Desulfosarcina-type of SRB. In addition to MOA, the 0.76-mbsf library contained ribotypes affiliated with the methanogenic genus, Methanosaeta. Although methane was detected at both depths, only the deeper sample was associated with the geochemically-defined SMI. MOA community structure varied between the two libraries. The 0.76-mbsf library contained both ANME-1 and ANME-2 ribotypes, whereas the 11.82-mbsf library was dominated by a novel ribotype, ANME-X, phylogenetically bracketed by the ANME-1 and ANME-2 groups. FISH experiments with SRB- and MOA-specific rRNA-targeted probes revealed that ANME-X forms aggregates with SRB similar to those formed by ANME-2.

To better resolve microbial community structure across the SMI, archaeal SSU rRNA libraries from 11.42, 11.67, 12.07, 12.32, 12.57, and 12.82-mbsf depths were constructed and screened. MOA were detected in all samples between 11.67 and 12.57 mbsf, but not in samples from 11.42 and 12.82 mbsf. However, MOA community structure based on clone representation varied among these depth intervals. The 11.67-mbsf library contained ANME-1, ANME-2, and ANME-X ribotypes. In contrast, deeper libraries (between 11.82 and 12.57 mbsf) were dominated by ANME-X ribotypes.

Analysis of the SSU rRNA libraries in PC-26 indicates that MOA exist in a 0.9 meter interval spanning the SMI, and are not detected immediately above or below this interval. Thus, within our ability to resolve biogeochemical changes and microbial distributions, the SMI in sediments from the Blake Ridge Depression is a zone of rapid transition at least 0.5 meters thick that separates sulfate-bearing sediments above from methane-rich sediments below and contains evidence for MOA and of focused microbial metabolic activity. These microbes and their metabolic effects on the biogeochemistry of marine sediments can produce diagnostic diagenetic effects in sediments overlying methane-rich and gas-hydrate-bearing sedimentary sections that should be preserved in the geologic record.