Timothy G. Ferdelman

Prokaryotic cells of the deep sub-seafloor biosphere identified as living bacteria

Chemical analyses of the pore waters from hundreds of deep ocean sediment cores have over decades provided evidence for ongoing processes that require biological catalysis by prokaryotes. This sub-seafloor activity of microorganisms may influence the surface Earth by changing the chemistry of the ocean and by triggering the emission of methane, with consequences for the marine carbon cycle and even the global climate. Despite the fact that only about 1% of the total marine primary production of organic carbon is available for deep-sea microorganisms, sub-seafloor sediments harbour over half of all prokaryotic cells on Earth. This estimation has been calculated from numerous microscopic cell counts in sediment cores of the Ocean Drilling Program. Because these counts cannot differentiate between dead and alive cells, the population size of living microorganisms is unknown. Here, using ribosomal RNA as a target for the technique known as catalysed reporter deposition-fluorescence in situ hybridization (CARD-FISH), we provide direct quantification of live cells as defined by the presence of ribosomes. We show that a large fraction of the sub-seafloor prokaryotes is alive, even in very old (16 million yr) and deep (> 400 m) sediments. All detectable living cells belong to the Bacteria and have turnover times of 0.25–22 yr, comparable to surface sediments.

Deep sub-seafloor prokaryotes stimulated at interfaces over geological time

The sub-seafloor biosphere is the largest prokaryotic habitat on Earth but also a habitat with the lowest metabolic rates. Modelled activity rates are very low, indicating that most prokaryotes may be inactive or have extraordinarily slow metabolism. Here we present results from two Pacific Ocean sites, margin and open ocean, both of which have deep, subsurface stimulation of prokaryotic processes associated with geochemical and/or sedimentary interfaces. At 90 m depth in the margin site, stimulation was such that prokaryote numbers were higher (about 13-fold) and activity rates higher than or similar to near-surface values. Analysis of high-molecular-mass DNA confirmed the presence of viable prokaryotes and showed changes in biodiversity with depth that were coupled to geochemistry, including a marked community change at the 90-m interface. At the open ocean site, increases in numbers of prokaryotes at depth were more restricted but also corresponded to increased activity; however, this time they were associated with repeating layers of diatom-rich sediments (about 9 Myr old). These results show that deep sedimentary prokaryotes can have high activity, have changing diversity associated with interfaces and are active over geological timescales.

Zero-valent sulphur is a key intermediate in marine methane oxidation

Emissions of methane, a potent greenhouse gas, from marine sediments are controlled by anaerobic oxidation of methane coupled primarily to sulphate reduction (AOM). Sulphate-coupled AOM is believed to be mediated by a consortium of methanotrophic archaea (ANME) and sulphate-reducing Deltaproteobacteria but the underlying mechanism has not yet been resolved. Here we show that zero-valent sulphur compounds (S0) are formed during AOM through a new pathway for dissimilatory sulphate reduction performed by the methanotrophic archaea. Hence, AOM might not be an obligate syntrophic process but may be carried out by the ANME alone. Furthermore, we show that the produced S0—in the form of disulphide—is disproportionated by the Deltaproteobacteria associated with the ANME. Our observations expand the diversity of known microbially mediated sulphur transformations and have significant implications for our understanding of the biogeochemical carbon and sulphur cycles.

Subseafloor sedimentary life in the South Pacific Gyre

The low-productivity South Pacific Gyre (SPG) is Earths largest oceanic province. Its sediment accumulates extraordinarily slowly (0.1–1 m per million years). This sediment contains a living community that is characterized by very low biomass and very low metabolic activity. At every depth in cored SPG sediment, mean cell abundances are 3 to 4 orders of magnitude lower than at the same depths in all previously explored subseafloor communities. The net rate of respiration by the subseafloor sedimentary community at each SPG site is 1 to 3 orders of magnitude lower than the rates at previously explored sites. Because of the low respiration rates and the thinness of the sediment, interstitial waters are oxic throughout the sediment column in most of this region. Consequently, the sedimentary community of the SPG is predominantly aerobic, unlike previously explored subseafloor communities. Generation of H2 by radiolysis of water is a significant electron-donor source for this community. The per-cell respiration rates of this community are about 2 orders of magnitude higher (in oxidation/reduction equivalents) than in previously explored anaerobic subseafloor communities. Respiration rates and cell concentrations in subseafloor sediment throughout almost half of the world ocean may approach those in SPG sediment.

Endosymbiotic sulphate-reducing and sulphide-oxidizing bacteria in an oligochaete worm

Stable associations of more than one species of symbiont within a single host cell or tissue are assumed to be rare in metazoans because competition for space and resources between symbionts can be detrimental to the host. In animals with multiple endosymbionts, such as mussels from deep-sea hydrothermal vents and reef-building corals, the costs of competition between the symbionts are outweighed by the ecological and physiological flexibility gained by the hosts. A further option for the coexistence of multiple symbionts within a host is if these benefit directly from one another, but such symbioses have not been previously described. Here we show that in the gutless marine oligochaete Olavius algarvensis, endosymbiotic sulphate-reducing bacteria produce sulphide that can serve as an energy source for sulphide-oxidizing symbionts of the host. Thus, these symbionts do not compete for resources but rather share a mutalistic relationship with each other in an endosymbiotic sulphur cycle, in addition to their symbiotic relationship with the oligochaete host.

Sulfate reduction and methanogenesis in a Thioploca-dominated sediment off the coast of Chile

Continental shelf sediments of the central Chile upwelling area are dominated by the presence of dense mats of the filamentous, sulfur-depositing bacterium Thioploca spp. We examined rates and pathways of S and methane cycling in these sediments along a transect from the Bay of Concepcion to the continental slope. Sulfate reduction rates (170–4670 nmol cm−3 d−1) were equal to or exceeded rates reported for other subtidal marine sediments. Elemental S and pyrite were the dominant end-products of sulfate reduction in Thioploca mats on the continental shelf, whereas, in the highly-reducing, Beggiatoa-dominated sediments of the nearby Bay of Concepcion, acid-volatile S was the principal end-product. Dissolved organic C values were lowest at the stations with the highest sulfate reduction rates and increased offshore. Sediment porewater methane concentrations in all surface sediments were low (<12 nmol cm−3), and methane production rates at the station most dominated by Thioploca were extremely low ( <0.5 nmol cm−3 d−1). Low methane production rates and concentrations were matched by low methane oxidation rates (<0.1 nmol cm−3 d−1). Radio-tracer studies showed that methane production was almost exclusively from methylamines, substrates which are noncompetitive with sulfate reduction, rather than from acetate or CO2/H2. Bacterial MPN (most probable number) counts also indicated the presence of a methylotrophic population of methanogens. Surprisingly, high numbers of autotrophic acetogenic bacteria were found, suggesting that the bacterial population involved in anaerobic DOC degradation is more complex than expected. In spite of the high sulfate reduction rates, sulfide concentrations in the shelf and slope were low or undetectable (<0.5 μM), and sulfate concentrations were never depleted below bottom water levels down to depths of 25–30 cm. Calculations suggest that Thioploca were oxidizing a maximum of 35% of sulfide production—not enough to prevent sulfate depletion. Either other sulfide oxidizers were also present or transient hydrodynamic conditions coupled with bioturbation resulted in oxidation of the sediments.

Sulfate reduction and methane oxidation in continental margin sediments influenced by irrigation (South-East Atlantic off Namibia)

Abstract Sulfate reduction rates (SRR) and concentrations of SO 4 2− , H 2 S, pyrite sulfur, total sulfur, CH 4 , and organic carbon were measured with high depth resolution through the entire length of the SO 4 2− -zone and well into the CH 4 -zone at two continental slope stations in the eastern South Atlantic (Benguela upwelling area). The sediments were characterized by a high organic carbon content of approx. 7.5% at GeoB 3703 and 3.7% at GeoB 3714. At GeoB 3703 SO 4 2− concentrations decreased linearly with depth to about 40 μM at the sulfate-methane transition zone (SMT) at 3.5 m, while at GeoB 3714, SO 4 2− remained at sea water concentration in the top 2 m of the sediment and then decreased linearly to about 70 μM at the SMT at 6 m. Direct rate measurements of SRR ( 35 SO 4 2− ) showed that the highest SRR occurred within the surface 3–5 cm with peak rates of up to 20 and 7 nmol SO 4 2− cm −3 day −1 at GeoB 3703 and GeoB 3714, respectively. SRR decreased quasi-exponentially with depth at GeoB 3703 and the cumulative SRR over the length of the SO 4 2− zone resulted in an areal SRR (SRR area ) of 1114–3493 μmol m −2 day −1 (median value: 2221 μmol m −2 day −1 ) at GeoB 3703 with more than 80% of the total sulfate reduction proceeding in the top 30 cm sediment. At GeoB 3714 SRR exhibited more scatter with a cumulative SRR area of 398–1983 μmol m −2 day −1 (median value: 1251 μmol m −2 day −1 ) and with >60% of the total sulfate reduction occurring below a depth of 30 cm due partially to a deeply buried zone of sulfate reduction located between 3 and 5 m depths. SRR peaks were also observed in SMT of both cores, ostensibly associated with methane oxidation, but with rates about 10 times lower than at the surface. Modeled SRR balanced both methane oxidation rates and measured SRR within the SMT, but severely underestimated by up to 89% the total SRR area that were obtained from direct measurements. Modeled and measured SRR were reconciled by including solute transport by irrigation described by a non-local pore water exchange function (α) which had values of up to 0.3 year −1 in the top sediment, and decreased exponentially to zero (i.e., no irrigation) at 2–3 meters (i.e., above SMT). These results suggested that co-existing sulfate reduction processes and linear SO 4 2− -gradients can be maintained by a non-local transport mechanism such as irrigation, by which pore water in tubes or burrows is exchanged with bottom waters by activities of tube-dwelling animals, or some similar physical transport phenomenon (i.e., bubble ebullition). Further support for an irrigation mechanism was found in the observations of open tubes of up to 8 mm (ID) at depths down to 6 m, which also contained fecal pellets, indicating that these tubes were or had been inhabited.

A constant flux of diverse thermophilic bacteria into the cold Arctic seabed.

Monitoring Massive Microbial Dispersal Quantifying the relative influence of present-day environmental conditions and geological history on the spatial distribution of species represents a major challenge in microbial ecology. Ecological approaches to distinguish between these two biogeographic controls are limited by environmental variability both in space and through time (see the Perspective by Patterson). Using a 1.5-million-year fossil record of marine diatoms, Cermeño and Falkowski (p. 1539) show that, even at the largest (global) spatial scale, the dispersal of marine diatoms is not very limited. Environmental factors are the primary control shaping the global biogeography of marine diatom morphospecies. Thermophilic microorganisms are routinely detected in permanently cold environments from deep sea sediments to polar soils. Hubert et al. (p. 1541) provide a quantitative analysis of a potentially large-scale dispersion of thermophilic bacteria in the ocean. Approximately 108 thermophilic spores are deposited each year on every square meter of Arctic sediment. Spore-forming bacteria adapted to the hot subsurface biosphere are continually deposited in polar marine sediments. Microorganisms have been repeatedly discovered in environments that do not support their metabolic activity. Identifying and quantifying these misplaced organisms can reveal dispersal mechanisms that shape natural microbial diversity. Using endospore germination experiments, we estimated a stable supply of thermophilic bacteria into permanently cold Arctic marine sediment at a rate exceeding 108 spores per square meter per year. These metabolically and phylogenetically diverse Firmicutes show no detectable activity at cold in situ temperatures but rapidly mineralize organic matter by hydrolysis, fermentation, and sulfate reduction upon induction at 50°C. The closest relatives to these bacteria come from warm subsurface petroleum reservoir and ocean crust ecosystems, suggesting that seabed fluid flow from these environments is delivering thermophiles to the cold ocean. These transport pathways may broadly influence microbial community composition in the marine environment.

Sulfur enrichment of humic substances in a Delaware salt marsh sediment core

Abstract Humic sulfur, operationally defined as the sulfur extracted with humic substances in 0. l N NaOH solution, comprises up to 51% of the total sulfur inventory in a sediment core taken from a Delaware Spartina alterniflora marsh. Pyrite sulfur is the next largest fraction, except at the near-surface sediments, where zerovalent sulfur concentrations are significant. X-ray photoelectron spectroscopy indicates that the humic sulfur consists of sulfoxides or sulfones and, in a more reduced state, organic sulfides and/or organic polysulfides. A subsurface decrease in the humic acid C:S atomic ratio to 56 ± 2 suggests that the upper 4 cm of marsh sediment is the locus for humic sulfur formation. S. alterniflora detritus and microbial biomass cannot fully account for observed sulfur enrichment of humic C:S atomic ratios. Therefore, the enrichment of humic substances by sulfur is probably via reaction of reduced sulfur compounds with organic matter. A humic sulfur formation rate of 10.6 μmol S · cm−3 · a−1 is calculated for the surface sediments and leads to an areal production of 18 μmol S · cm−2 · a−1 of humic sulfur. Humic sulfur formation and preservation is enhanced by the limited availability of iron for the rapid precipitation of iron sulfide minerals and the apparent resistance of organic sulfur compounds towards reoxidation to sulfate, especially in the upper 9 cm of marsh sediment where inorganic sulfur compounds are rapidly oxidized.

Influence of water column dynamics on sulfide oxidation and other major biogeochemical processes in the chemocline of Mariager Fjord (Denmark)

Abstract Major electron donors (H2S, NH4+, Mn2+, Fe2+) and acceptors (O2, NO3−, Mn(IV), Fe(III)), process rates (35SO42− reduction, dark 14CO2 fixation) and vertical fluxes were investigated to quantify the dominant biogeochemical processes at the chemocline of a shallow brackish fjord. Under steady-state conditions, the upward fluxes of reductants and downward fluxes of oxidants in the water column were balanced. However, changes in the hydrographical conditions caused a transient nonsteady-state at the chemocline and had a great impact on process rates and the distribution of chemical species. Maxima of S0 (17.8 μmol l−1), thiosulfate (5.2 μmol l−1) and sulfite (1.1 μmol l−1) occurred at the chemocline, but were hardly detectable in the sulfidic deep water. The distribution of S0 suggested that the high concentration of S0 was (a) more likely due to a low turnover than a high formation rate and (b) was only transient, caused by chemocline perturbations. Kinetic calculations of chemical sulfide oxidation based on actual conditions in the chemocline revealed that under steady-state conditions with a narrow chemocline and low reactant concentrations, biological sulfide oxidation may account for more than 88% of the total sulfide oxidation. Under nonsteady-state conditions, where oxic and sulfidic water masses were recently mixed, resulting in an expanded chemocline, the proportion of chemical sulfide oxidation increased. The sulfide oxidation rate determined by incubation experiments was 0.216 μmol l−1 min−1, one of the highest reported for stratified basins and about 15 times faster than the initial rate for chemical oxidation. The conclusion of primarily biological sulfide oxidation was consistent with the observation of high rates of dark 14CO2 fixation (10.4 mmol m−2 day−1) in the lower part of the chemocline. However, rates of dark 14CO2 fixation were too high to be explained only by lithoautotrophic processes. CO2 fixation by growing populations of heterotrophic microorganisms may have additionally contributed to the observed rates.