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Large Sulfur Isotope Fractionation Does Not Require Disproportionation

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Science  01 Jul 2011:
Vol. 333, Issue 6038, pp. 74-77
DOI: 10.1126/science.1205103

Abstract

The composition of sulfur isotopes in sedimentary sulfides and sulfates traces the sulfur cycle throughout Earth’s history. In particular, depletions of sulfur-34 (34S) in sulfide relative to sulfate exceeding 47 per mil (‰) often serve as a proxy for the disproportionation of intermediate sulfur species in addition to sulfate reduction. Here, we demonstrate that a pure, actively growing culture of a marine sulfate-reducing bacterium can deplete 34S by up to 66‰ during sulfate reduction alone and in the absence of an extracellular oxidative sulfur cycle. Therefore, similar magnitudes of sulfur isotope fractionation in sedimentary rocks do not unambiguously record the presence of other sulfur-based metabolisms or the stepwise oxygenation of Earth’s surface environment during the Proterozoic.

Dissimilatory microbial sulfate reduction (MSR) uses sulfate (SO42–) as an electron acceptor and simple organic compounds or hydrogen as electron donors, producing sulfide that is depleted in heavy isotopes of sulfur (33S, 34S, and 36S) relative to the starting sulfate. For more than 2.5 billion years of Earth history, this biological process has controlled the partitioning of sulfur isotopes between sedimentary sulfides and sulfates, leaving a sedimentary S isotope record that is commonly used to track the geochemical cycling of sulfur, the oceanic budgets of oxidants, the evolution of microbial metabolisms, and the levels of atmospheric oxygen throughout geologic history (13).

All interpretations of this geobiological record have drawn heavily on more than five decades of systematic studies of sulfur isotope effects produced by MSR under controlled laboratory conditions. Although previous environmental studies and models predicted that MSR alone could produce a sulfur isotope offset between sulfides and sulfates (δ34Ssulfate-sulfide) (4) as large as ~75 per mil (‰) (58), growth and chemical conditions that lead toward such large offsets remain poorly understood. On the other hand, laboratory culture studies of MSR have not reported sulfur isotope effects larger than 47‰ under chemically and biologically defined reproducible conditions (Fig. 1). Large δ34Ssulfate-sulfide values commonly measured in nature (Fig. 1) were thus attributed to a combination of MSR, extracellular oxidative recycling of sulfur by abiotic or microbial processes, and microbial sulfur disproportionation (MSD) (2, 9). This oxidative recycling model, applied to the temporal record of δ34Ssulfate-sulfide values, is used to track the evolution of more oxidized conditions and the progressive oxygenation of Earth’s surface environment (2, 1013).

Fig. 1

Fractionations of 34S reported by studies of environmental samples (δ34Ssulfate-sulfide or 34ε) and pure cultures of 44 different sulfate-reducing microbes (34ε). Isotope fractionations in environmental samples were estimated using dissolved sulfate and sedimentary or dissolved sulfide or were calculated from the concentration and the isotope composition of pore water sulfate. Each point in culture studies represents a different growth condition. Dashed black lines indicate the expanded range of 34S fractionations by MSR (this study). The gray shaded area outlines the equilibrium isotope effects between sulfate and sulfide with varying temperatures (0 to ~40°C). Complete lists of references, data, and criteria are available in tables S1 and S2.

We isolated a sulfate-reducing δ-proteobacterium [Desulfovibrio sp., (DMSS-1)] from marine coastal sediments from Cape Cod, Massachusetts (14), where δ34Ssulfate-sulfide exceeds 50‰ (15). This microbe couples the reduction of sulfate with the incomplete oxidation of various organic substrates including fructose and glucose (14), producing a wide range of isotope enrichment factors (34ε) from 6.1 to 65.6‰ (Fig. 1). Fractionations by DMSS-1 expand the range of 34ε and the triple isotope fractionation coefficient for 32S/33S/34S (33λ) (4) values produced by MSR, accommodating most S isotope fractionations observed in modern environments (Figs. 1 and 2 and Table 1). Very slow growth of DMSS-1 in batch cultures grown on glucose as an electron donor and carbon source produces 34ε values larger than 47‰ (Table 1) (16): During the early exponential growth, the 34ε values exceed 60‰, decreasing down to 44‰ at the very end of experiment (Table 1). Decreasing 34ε values are accompanied by an increasing growth rate and cell yields, as well as a different reaction stoichiometry (16). Because DMSS-1 can ferment glucose in the absence of sulfate, different stoichiometries during the early and the late exponential growth suggest that DMSS-1 ferments some glucose during the early exponential growth (16). Conditions producing large and constant 34ε values (>47‰) can be maintained indefinitely in continuous-flow cultures, where sulfate and glucose are the only available oxidant and reductant, respectively (Table 1). The production of large 34ε values cannot be attributed to MSD coupled with the extracellular oxidation of sulfide to intermediate sulfur species (for example, thiosulfate, sulfur, or sulfite), because other potential oxidants are absent from the defined culture medium (16). By proving that MSR alone can generate sulfides extremely depleted in 34S, even in the absence of extracellular oxidative recycling, our findings bridge the long-standing discrepancy between the ranges of sulfur isotope effects observed in laboratory cultures and geologic environments (Fig. 1).

Fig. 2

(A) Comparison of 34ε and 33λ values in this study to the values reported by previous culture studies (23, 34) and to the range (34ε, 33λ) from coexisting dissolved sulfide and sulfate in modern euxinic environments [Green Lake (Fayetteville, New York), Lago di Cadagno (Ticino, Switzerland), and Cariaco Basin (Venezuela)] (8, 27, 35). (B) δ34S and Δ33S (4) of seawater sulfate proxies (circles) from 2 to 0.2 billion years ago standardized to the Vienna Canyon Diablo Troilite (11, 32). Shown here is the predicted range of δ34S and Δ33S values of seawater sulfate based on the steady-state global sulfur-cycle model including only MSR without MSD, constrained by the previous range of 34ε and 33λ values for MSR (11) (dashed outline) or the expanded range reported in this study (solid outline).

Table 1

Sulfur isotope effects during the growth of DMSS-1 on glucose. Solid horizontal lines indicate separate experiments. For 8.5 mM glucose/21 mM sulfate, we carried out the experiment two times using different inocula. Individual numbers within solid lines represent cultures inoculated at the same time and grown simultaneously in identical growth media, but in separate bottles. Calculated 34ε values vary depending on the growth stage. The largest and the smallest 34ε values were always found during the early and late exponential growth, respectively. Errors were propagated from analytic uncertainties of isotope analysis and sulfide concentration measurement. f, fraction of remaining sulfate.

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One may ask whether large sulfur isotope effects produced by a single sedimentary microbe are truly representative of the natural environment. However, it is similarly unclear whether previous culture studies of MSR are representative of conditions conducive to large isotope fractionations in the environment, because most of these studies investigated MSR during growth on simple organic acids (e.g., lactate) and hydrogen and only rarely attained very slow growth rates and cell-specific sulfate reduction rates (csSRRs) (16). Here, slow growth rates and csSRRs are attained by growing DMSS-1 on glucose. This is not a conventional substrate for sulfate-reducing microbes, but it is a common building block in the biofilms (17) and storage polymers of some sulfate reducers (18), as well as a common monosaccharide in the ocean (19).

Sulfate-reducing microbes accomplish the eight-electron reduction of sulfate to sulfide in a stepwise and reversible manner (7, 20). The overall isotope effects between sulfate and sulfide thus depend both on the ratio between forward and backward fluxes at each intermediate step and on the isotope effect intrinsic to each transfer flux (20, 21). The largest isotope effect is expected when the sulfate-reduction pathway operates in a highly reversible manner, leading to near equilibrium conditions (7). Accordingly, the largest 34ε values produced by MSR should approach the equilibrium isotope effect between dissolved sulfate and sulfide, calculated to 68 ± 2‰ at 20°C (22). This upper boundary is also supported by the relation among multiple isotopes of sulfur. As 34ε approaches the equilibrium value, so does 33λ (Fig. 2A), whereas smaller 34ε values are associated with smaller 33λ values, as expected for the kinetic isotope effects in multiple-isotope systems (23). Our estimate for the largest 34ε value is close to the apparent upper boundary for the values of δ34Ssulfate-sulfide in nature (Fig. 1). In contrast, models that include oxidative recycling do not necessarily limit the values of δ34Ssulfate-sulfide to the equilibrium value [figure 2 in (9)].

Models of S isotope effects produced by a combination of MSR and MSD predict that the largest δ34Ssulfate-sulfide should occur in areas of intense sulfur redox cycling and substantial MSD. Although these processes demonstrably occur in modern coastal environments (9, 24), large (>47‰) δ34Ssulfate-sulfide values are not reported frequently in these settings (Fig. 1). Large δ34Ssulfate-sulfide values are instead commonly reported from deep-sea sediments, where the extremely slow microbial metabolisms are attributed to the limited availability and the poor reactivity of organic substrates (25, 26). The diversity of microbes and growth conditions that generate natural δ34Ssulfate-sulfide values larger than 47‰ remains to be determined, but the overall scarcity of sulfides exhibiting δ34Ssulfate-sulfide > 47‰ over geologic history (2) thus might simply be related to the lack of preservation of deep-sea sediments. Euxinic basins also exhibit a wide range of δ34Ssulfate-sulfide values that commonly exceed 47‰ and 33λ values larger than those previously attributed to MSR (Fig. 2A). These δ34Ssulfate-sulfide and 33λ values have been used as an indication of the contribution of MSD to the cycling of sulfur in modern euxinic environments (27). However, the combined isotopic signatures of 34ε and 33λ produced by DMSS-1 can explain nearly all observations from modern euxinic settings (Fig. 2A) and demonstrate that neither 34ε nor 33λ unambiguously indicates MSD in modern environments.

The earliest values of δ34Ssulfate-sulfide larger than 50‰ occur in a 1.2-billion-year-old nonmarine environment (13) and may have become more widespread in marine environments after ~700 million years ago (28) or even later (10, 12). Given the assumption that 34ε values larger than 47‰ do not occur during MSR alone, this temporal trend was attributed to various mechanisms including the growth of the marine sulfate reservoir (29), the increasing importance of disproportionation (11), the progressive oxygenation of the oceans (2), and the advent of bioturbating organisms close to the Precambrian/Cambrian boundary (30). However, given that DMSS-1 in the presence of just 2 mM sulfate produces 34ε values of 61.1‰ (Table 1) [i.e., outside the limit previously attributed to MSR alone (2, 11)], MSR alone could have produced similar S isotope signatures after a moderate increase in the size of marine sulfate reservoir (2 mM) during the mid-Mesoproterozoic (31).

In addition to 34S (2), the 33S record has been used to show the substantial contribution of MSD to the global sulfur cycle as early as 1300 million years ago (11, 32). This constraint is based on model estimates for sulfur isotope compositions (δ34S and Δ33S) of proxies for seawater sulfate (Fig. 2B). However, the input parameters for this model include the relatively small ranges of 34ε and 33λ values from previous laboratory experiments (11). When the same box model is solved with new expanded ranges of 34ε and 33λ values produced by DMSS-1, all but one sample of Phanerozoic and Proterozoic sedimentary sulfates (11, 32) are consistent with the global sulfur cycle including only MSR without MSD (Fig. 2B). Therefore, 33S isotope signatures in sedimentary records do not clearly indicate sulfur disproportionation in the ancient oceans (2, 9, 11).

Because the fractionation of sulfur isotopes between sulfate and sulfide can exceed 50‰, even if sulfide is not reoxidized outside of the cell and at an environmental scale, more Proterozoic samples exhibiting a large δ34Ssulfate-sulfide value may be found. Any temporal changes in the sulfur isotope record during this time could reflect the changing nature of organic material that fueled sulfate reduction, rather than measure the extent of oxygenated areas in oceans. The relative contributions of MSR alone and of the environmental-scale oxidative recycling toward large present and past natural fractionations of S isotope ratios now remain to be evaluated.

Supporting Online Material

www.sciencemag.org/cgi/content/full/333/6038/74/DC1

Materials and Methods

SOM Text

Fig. S1

Tables S1 to S5

References (36133)

References and Notes

  1. δ34Ssulfate-sulfide is the depletion of 34S in sulfide relative to sulfate source (e.g., seawater sulfate for marine environments) (≈δ34Ssulfate – δ34Ssulfide). δxS values are defined as δxS = 1000·[(xS/32S)sample/(xS/32S)standard – 1], where x is 33 or 34. Δ33S = δ33S – 1000·[(δ34S/1000 + 1)0.515 – 1]. xε = 1000·[1-(xS/32S)sulfide/(xS/32S)sulfate] and 33λ = ln[(33S/32S)sulfide/(33S/32S)sulfate]/ln[(34S/32S)sulfide/(34S/32S)sulfate], where (xS/32S)sulfide and (xS/32S)sulfate are the instantaneous S isotope ratios of sulfide and the remaining sulfate, respectively.
  2. Materials and methods are available as supporting material on Science Online.
  3. The range reflects values derived by either extrapolation from high temperature (>200°C) or theoretical calculations (33), assuming pH = 7. The equilibrium isotope fractionation factor between aqueous H2S and HS is ~6‰.
  4. Acknowledgments: We thank W. Olszewski, K. Donovan, S. Templer, J. Seewald, S. Sylva, B. Weiss, L. Kump, J. Grabenstatter, B. Bannon, P. Hedman, and Y. J. Joo. This work was partially supported by NASA Astrobiology Institute (#NNA08CN84A).
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