Microbial production of isotopically light iron(II) in a modern chemically precipitated sediment and implications for isotopic variations in ancient rocks

The inventories and Fe isotope composition of aqueous Fe(II) and solid‐phase Fe compounds were quantified in neutral‐pH, chemically precipitated sediments downstream of the Iron Mountain acid mine drainage site in northern California, USA. The sediments contain high concentrations of amorphous Fe(III) oxyhydroxides [Fe(III)_am] that allow dissimilatory iron reduction (DIR) to predominate over Fe–S interactions in Fe redox transformation, as indicated by the very low abundance of Cr(II)‐extractable reduced inorganic sulfur compared with dilute HCl‐extractable Fe. δ⁵⁶Fe values for bulk HCl‐ and HF‐extractable Fe were ≈ 0. These near‐zero bulk δ⁵⁶Fe values, together with the very low abundance of dissolved Fe in the overlying water column, suggest that the pyrite Fe source had near‐zero δ⁵⁶Fe values, and that complete oxidation of Fe(II) took place prior to deposition of the Fe(III) oxide‐rich sediment. Sediment core analyses and incubation experiments demonstrated the production of millimolar quantities of isotopically light (δ⁵⁶Fe ≈ ?1.5 to ?0.5‰) aqueous Fe(II) coupled to partial reduction of Fe(III)_am by DIR. Trends in the Fe isotope composition of solid‐associated Fe(II) and residual Fe(III)_am are consistent with experiments with synthetic Fe(III) oxides, and collectively suggest an equilibrium Fe isotope fractionation between aqueous Fe(II) and Fe(III)_am of approximately ?2‰. These Fe(III) oxide‐rich sediments provide a model for early diagenetic processes that are likely to have taken place in Archean and Paleoproterozoic marine sediments that served as precursors for banded iron formations. Our results suggest pathways whereby DIR could have led to the formation of large quantities of low‐δ⁵⁶Fe minerals during BIF genesis.     

Iron isotope fractionation during microbial dissimilatory iron oxide reduction in simulated Archaean seawater

The largest Fe isotope excursion yet measured in marine sedimentary rocks occurs in shales, carbonates, and banded iron formations of Neoarchaean and Paleoproterozoic age. The results of field and laboratory studies suggest a potential role for microbial dissimilatory iron reduction (DIR) in producing this excursion. However, most experimental studies of Fe isotope fractionation during DIR have been conducted in simple geochemical systems, using pure Fe(III) oxide substrates that are not direct analogues to phases likely to have been present in Precambrian marine environments. In this study, Fe isotope fractionation was investigated during microbial reduction of an amorphous Fe(III) oxide-silica coprecipitate in anoxic, high-silica, low-sulphate artificial Archaean seawater at 30 °C to determine if such conditions alter the extent of reduction or isotopic fractionations relative to those observed in simple systems. The Fe(III)-Si coprecipitate was highly reducible (c. 80% reduction) in the presence of excess acetate. The coprecipitate did not undergo phase conversion (e.g. to green rust, magnetite or siderite) during reduction. Iron isotope fractionations suggest that rapid and near-complete isotope exchange took place among all Fe(II) and Fe(III) components, in contrast to previous work on goethite and hematite, where exchange was limited to the outer few atom layers of the substrate. Large quantities of low-δ(56)Fe Fe(II) (aqueous and solid phase) were produced during reduction of the Fe(III)-Si coprecipitate. These findings shed new light on DIR as a mechanism for producing Fe isotope variations observed in Neoarchaean and Paleoproterozoic marine sedimentary rocks.     

Chromium geochemistry of the ca. 1.85 Ga Flin Flon paleosol

Fractionation of stable Cr isotopes has been measured in Archaean paleosols and marine sedimentary rocks and interpreted to record the terrestrial oxidation of Cr(III) to Cr(VI), providing possible indirect evidence for the emergence of oxygenic photosynthesis. However, these fractionations occur amidst evidence from other geochemical proxies for a pervasively anoxic atmosphere. This study examined the Cr geochemistry of the ca. 1.85 Ga Flin Flon paleosol, which developed under an atmosphere unambiguously oxidising enough to quantitatively convert Fe(II) to Fe(III) during pedogenesis. The paleosol shows an extreme range in Cr isotope composition of 2.76 ‰ δ^53/52Cr. The protolith greenstone (δ^53/52Cr: ?0.23 ‰), the deepest weathering horizon (δ^53/52Cr: ?0.15 to ?0.23 ‰) and a residual corestone in the upper paleosol (δ^53/52Cr: ?0.01 ‰) all exhibit Cr isotopic compositions comparable to unaltered igneous rocks. The most significant isotopic fractionation is preserved in the areas influenced by oxidative subaerial weathering (i.e. increase in Fe(III)/Fe(II)) and the greatest loss of mobile elements. The uppermost paleosol horizon is both Cr and Mn depleted and offset to significantly ⁵³Cr‐enriched compositions (δ^53/52Cr values between +1.50 and +2.38 ‰), which is not easily modelled with the oxidation of Cr(III) and loss of isotopically heavy Cr(VI). Instead, the currently preferred model for these data invokes the open‐system removal of isotopically light aqueous Cr(III) during either pedogenesis or subsequent hydrothermal/metamorphic alteration. The ⁵³Cr enrichment would then represent the preferential dissolution or complexation of isotopically light aqueous Cr(III) species (enhanced by lower pH conditions and possibly the presence of complexing ligands) and/or the residual signature from preferential adsorption of isotopically heavy Cr(III). Both scenarios would contradict the widely held assumption that only redox reactions of Cr can generate large magnitude isotopic fractionations and, if substantiated, non‐redox isotope effects would complicate the conclusive fingerprinting of ancient atmospheric O₂ from Cr isotope data alone.     

Microbial Fe(III) oxide reduction potential in Chocolate Pots hot spring,Yellowstone National Park

Chocolate Pots hot springs (CP) is a unique, circumneutral pH, iron‐rich, geothermal feature in Yellowstone National Park. Prior research at CP has focused on photosynthetically driven Fe(II) oxidation as a model for mineralization of microbial mats and deposition of Archean banded iron formations. However, geochemical and stable Fe isotopic data have suggested that dissimilatory microbial iron reduction (DIR) may be active within CP deposits. In this study, the potential for microbial reduction of native CP Fe(III) oxides was investigated, using a combination of cultivation dependent and independent approaches, to assess the potential involvement of DIR in Fe redox cycling and associated stable Fe isotope fractionation in the CP hot springs. Endogenous microbial communities were able to reduce native CP Fe(III) oxides, as documented by most probable number enumerations and enrichment culture studies. Enrichment cultures demonstrated sustained DIR driven by oxidation of acetate, lactate, and H₂. Inhibitor studies and molecular analyses indicate that sulfate reduction did not contribute to observed rates of DIR in the enrichment cultures through abiotic reaction pathways. Enrichment cultures produced isotopically light Fe(II) during DIR relative to the bulk solid‐phase Fe(III) oxides. Pyrosequencing of 16S rRNA genes from enrichment cultures showed dominant sequences closely affiliated with Geobacter metallireducens, a mesophilic Fe(III) oxide reducer. Shotgun metagenomic analysis of enrichment cultures confirmed the presence of a dominant G. metallireducens‐like population and other less dominant populations from the phylum Ignavibacteriae, which appear to be capable of DIR. Gene (protein) searches revealed the presence of heat‐shock proteins that may be involved in increased thermotolerance in the organisms present in the enrichments as well as porin–cytochrome complexes previously shown to be involved in extracellular electron transport. This analysis offers the first detailed insight into how DIR may impact the Fe geochemistry and isotope composition of a Fe‐rich, circumneutral pH geothermal environment.     

Isotopic and chemical investigations of Fe (III) reduction by Shewanella putrefaciens strain 200R

Experiments were conducted using the Fe⁺³‐reducing bacterium Shewanella putrefaciens strain 200R to determine the stable carbon isotope fractionation during dissimilatory Fe (III) reduction and associated lactate oxidation at circum‐neutral pH. Previous studies used equilibrium fractionation factors (~14.3‰) between bacterial biomass and synthesized fatty acids to identify the predominant carbon fixation pathways for some of the most frequently isolated microbes including Shewanella under anaerobic conditions. We investigated the carbon isotope disproportionation among organic carbon substrate (lactate), biomass and respired carbon dioxide at the lag to stationary phase of the growth curve. Ferric citrate and sodium lactate were used as electron acceptor and donor, respectively. Sodium bicarbonate or potassium phosphate was used as buffering agent. Iron (II), iron (III), dissolved inorganic carbon (DIC) and carbon isotope ratios were measured for both bicarbonate‐ and phosphate‐buffered systems. Carbon isotope ratio measurements were made on the respired CO₂ (as DIC) and microbial biomass for both buffering conditions. The fraction of lactate consumed was estimated using DIC as a proxy and was verified by direct measurement using HPLC. Our result showed that bicarbonate‐buffered system has an enhancing effect in the reduction process compared to the phosphate system. Both systems resulted in carbon isotope fractionations between the lactate substrate and DIC that could be modelled as a Rayleigh process. The biomass produced under both buffer conditions was depleted on average by ~2‰ relative to the substrate and enriched by ~5‰ relative to the DIC. This translates to an overall isotopic fractionation of 10–12‰ between the biomass and respired CO₂ in both buffering systems.     

Benthic iron cycling in a high‐oxygen environment: Implications for interpreting the Archean sedimentary iron isotope record

The most notable trend in the sedimentary iron isotope record is a shift at the end of the Archean from highly variable δ⁵⁶Fe values with large negative excursions to less variable δ⁵⁶Fe values with more limited negative values. The mechanistic explanation behind this trend has been extensively debated, with two main competing hypotheses: (i) a shift in marine redox conditions and the transition to quantitative iron oxidation; and (ii) a decrease in the signature of microbial iron reduction in the sedimentary record because of increased bacterial sulfate reduction (BSR). Here, we provide new insights into this debate and attempt to assess these two hypotheses by analyzing the iron isotope composition of siderite concretions from the Carboniferous Mazon Creek fossil site. These concretions precipitated in an environment with water column oxygenation, extensive sediment pile dissimilatory iron reduction (DIR) but limited bacterial sulfate reduction (BSR). Most of the concretions have slightly positive iron isotope values, with a mean of 0.15‰ and limited iron isotope variability compared to the Archean sedimentary record. This limited variability in an environment with high DIR and low BSR suggests that these conditions alone are insufficient to explain Archean iron isotope compositions. Therefore, these results support the idea that the unusually variable and negative iron isotope values in the Archean are due to dissimilatory iron reduction (DIR) coupled with extensive water column iron cycling.     

Chemical and Biological Interactions during Nitrate and Goethite Reduction by Shewanella putrefaciens 200

Although previous research has demonstrated that NO₃⁻ inhibits microbial Fe(III) reduction in laboratory cultures and natural sediments, the mechanisms of this inhibition have not been fully studied in an environmentally relevant medium that utilizes solid-phase, iron oxide minerals as a Fe(III) source. To study the dynamics of Fe and NO₃⁻ biogeochemistry when ferric (hydr)oxides are used as the Fe(III) source, Shewanella putrefaciens 200 was incubated under anoxic conditions in a low-ionic-strength, artificial groundwater medium with various amounts of NO₃⁻ and synthetic, high-surface-area goethite. Results showed that the presence of NO₃⁻ inhibited microbial goethite reduction more severely than it inhibited microbial reduction of the aqueous or microcrystalline sources of Fe(III) used in other studies. More interestingly, the presence of goethite also resulted in a twofold decrease in the rate of NO₃⁻ reduction, a 10-fold decrease in the rate of NO₂⁻ reduction, and a 20-fold increase in the amounts of N₂O produced. Nitrogen stable isotope experiments that utilized δ¹⁵N values of N₂O to distinguish between chemical and biological reduction of NO₂⁻ revealed that the N₂O produced during NO₂⁻ or NO₃⁻ reduction in the presence of goethite was primarily of abiotic origin. These results indicate that concomitant microbial Fe(III) and NO₃⁻ reduction produces NO₂⁻ and Fe(II), which then abiotically react to reduce NO₂⁻ to N₂O with the subsequent oxidation of Fe(II) to Fe(III).     

Iron cycling in the anoxic cryo-ecosystem of Antarctic Lake Vida

Iron redox cycling in metal-rich, hypersaline, anoxic brines plays a central role in the biogeochemical evolution of life on Earth, and similar brines with the potential to harbor life are thought to exist elsewhere in the solar system. To investigate iron biogeochemical cycling in a terrestrial analog we determined the iron redox chemistry and isotopic signatures in the cryoencapsulated liquid brines found in frozen Lake Vida, East Antarctica. We used both in situ voltammetry and the spectrophotometric ferrozine method to determine iron speciation in Lake Vida brine (LVBr). Our results show that iron speciation in the anoxic LVBr was, unexpectedly, not free Fe(II). Iron isotope analysis revealed highly depleted values of ?2.5‰ for the ferric iron of LVBr that are similar to iron isotopic signatures of Fe(II) produced by dissimilatory iron reduction. The presence of Fe(III) in LVBr therefore indicates dynamic iron redox cycling beyond iron reduction. Furthermore, extremely low δ¹⁸O–SO₄ ^2? values (?9.7‰) support microbial iron-sulfur cycling reactions. In combination with evidence for chemodenitrification resulting in iron oxidation, we conclude that coupled abiotic and biotic redox reactions are driving the iron cycle in Lake Vida brine. Our findings challenge the current state of knowledge of anoxic brine chemistry and may serve as an analogue for icy brines found in the outer reaches of the solar system.     

In Situ Fe and S isotope analyses in pyrite from the 3.2 Ga Mendon Formation (Barberton Greenstone Belt,South Africa): Evidence for early microbial iron reduction

On the basis of phylogenetic studies and laboratory cultures, it has been proposed that the ability of microbes to metabolize iron has emerged prior to the Archaea/Bacteria split. However, no unambiguous geochemical data supporting this claim have been put forward in rocks older than 2.7–2.5 giga years (Gyr). In the present work, we report in situ Fe and S isotope composition of pyrite from 3.28‐ to 3.26‐Gyr‐old cherts from the upper Mendon Formation, South Africa. We identified three populations of microscopic pyrites showing a wide range of Fe isotope compositions, which cluster around two δ⁵⁶Fe values of ?1.8‰ and +1‰. These three pyrite groups can also be distinguished based on the pyrite crystallinity and the S isotope mass‐independent signatures. One pyrite group displays poorly crystallized pyrite minerals with positive Δ³³S values > +3‰, while the other groups display more variable and closer to 0‰ Δ³³S values with recrystallized pyrite rims. It is worth to note that all the pyrite groups display positive Δ³³S values in the pyrite core and similar trace element compositions. We therefore suggest that two of the pyrite groups have experienced late fluid circulations that have led to partial recrystallization and dilution of S isotope mass‐independent signature but not modification of the Fe isotope record. Considering the mineralogy and geochemistry of the pyrites and associated organic material, we conclude that this iron isotope systematic derives from microbial respiration of iron oxides during early diagenesis. Our data extend the geological record of dissimilatory iron reduction (DIR) back more than 560 million years (Myr) and confirm that micro‐organisms closely related to the last common ancestor had the ability to reduce Fe(III).     

Fe isotope fractionation caused by translocation of iron during growth of bean and oat as models of strategy I and II plants

Background

The determination of the plant-induced Fe-isotopic fractionation is a promising tool to better quantify their role in the geochemical Fe cycle and possibly to identify the physiological mechanisms of Fe uptake and translocation in plants. Here we explore the isotope fractionation caused by translocation of Fe during growth of bean and oat as representatives of strategy I and II plants.

Methods

Plants were grown on a nutrient solution supplemented with Fe(III)-EDTA and harvested at three different ages. We used the technique of multi-collector ICP-MS to resolve the small differences in the stable iron isotope compositions of plants.

Results

Total bean plants, regardless of their age, were found to be enriched in the light iron isotopes by ?1.2‰ relative to the growth solution throughout. During growth plants internally redistributed isotopes where young leaves increasingly accumulated the lighter isotopes whereas older leaves and the total roots were simultaneously depleted in light iron isotopes. Oat plants were also enriched in the light iron isotopes but during growth the initial isotope ratio maintained in all organs at all growth stages.

Conclusions

We conclude that isotope fractionation in bean as a representative of strategy I plants is a result of translocation or re-translocation processes. Furthermore we assume that both uptake and translocation of Fe in oat maintains the irons’ ferric state, or that Fe is always bound to high-mass ligands, so that isotope fractionation is virtually absent in these plants. However, in contrast to our previous study in which strategy II plants were grown on soil substrate, oat plants grown on Fe(III)-EDTA contain iron that enriches ⁵⁴Fe by 0.5 permil over ⁵⁶Fe. A possible explanation for the enrichment is the prevalence of a constitutive reductive uptake mechanism of iron in the nutrient solution used which is non-deficient in iron.     

Limited reduction of ferrihydrite encrusted by goethite in freshwater sediment

Many physical and chemical processes control the extent of Fe(III) oxyhydroxide reduction by dissimilatory Fe(III)‐reducing bacteria. The surface precipitation of secondary Fe minerals on Fe(III) oxyhydroxides limits the extent of microbial Fe(III) reduction, but this phenomenon has not yet been observed in nature. This paper reports the observation of secondary Fe‐mineral (goethite) encrustation on ferrihydrite surface within freshwater sediment up to 10 cm deep. The sediment surface was characterized by the predominance of ferrihydrites with biogenic stalks and sheaths. An Fe(II)‐oxidizing bacterium (Gallionellaceae) was detected by 16S rRNA gene analysis at sediment depths of 1 and 2 cm. Fe²⁺ concentration in the sediment pore water was relatively higher at 2–4 cm depths. The 16S rRNA genes affiliated with dissimilatory Fe(III)‐reducing bacteria were detected at 1, 2, and 4 cm depths. The results of the Fe K‐edge extended X‐ray absorption fine structure (EXAFS) analysis suggested the presence of goethite and siderite at depths below 3 cm. However, the change in the Fe‐mineral composition was restricted to sediment depths between 3 and 4 cm, despite the presence of abundant ferrihydrite at depths below 4 cm. An increase in CH₄ concentration was observed at deeper than 6 cm. Stable isotopic analysis of CH₄ in the pore water indicated that acetoclastic CH₄ occurred at depths below 7 cm. Transmission electron microscope observations suggested the presence of goethite and siderite on stalks and sheaths at depths below 3 cm. Results from conversion electron yield EXAFS analysis suggested that goethite dominated at 10 cm depth, thereby indicating that ferrihydrite was encrusted by goethite at this depth. Moreover, the incomplete reduction of ferrihydrite below depths of 4 cm was not due to the lack of organic carbon, but was possibly due to the surface encrustation of goethite on ferrihydrite.     

Oxygen isotope analysis of bacterial and fungal manganese oxidation

The ability of micro‐organisms to oxidize manganese (Mn) from Mn(II) to Mn(III/IV) oxides transcends boundaries of biological clade or domain. Many bacteria and fungi oxidize Mn(II) to Mn(III/IV) oxides directly through enzymatic activity or indirectly through the production of reactive oxygen species. Here, we determine the oxygen isotope fractionation factors associated with Mn(II) oxidation via various biotic (bacteria and fungi) and abiotic Mn(II) reaction pathways. As oxygen in Mn(III/IV) oxides may be derived from precursor water and molecular oxygen, we use a twofold approach to determine the isotope fractionation with respect to each oxygen source. Using both ¹⁸O‐labeled water and closed‐system Rayleigh distillation approaches, we constrain the kinetic isotope fractionation factors associated with O atom incorporation during Mn(II) oxidation to ?17.3‰ to ?25.9‰ for O₂ and ?1.9‰ to +1.8‰ for water. Results demonstrate that stable oxygen isotopes of Mn(III/IV) oxides have potential to distinguish between two main classes of biotic Mn(II) oxidation: direct enzymatic oxidation in which O₂ is the oxidant and indirect enzymatic oxidation in which superoxide is the oxidant. The fraction of Mn(III/IV) oxide‐associated oxygen derived from water varies significantly (38%–62%) among these bio‐oxides with only weak relationship to Mn oxidation state, suggesting Mn(III) disproportionation may account for differences in the fraction of mineral‐bound oxygen from water and O₂. Additionally, direct incorporation of molecular O₂ suggests that Mn(III/IV) oxides contain a yet untapped proxy of of environmental O₂, a parameter reflecting the integrated influence of global respiration, photorespiration, and several other biogeochemical reactions of global significance.     

Pigment production and isotopic fractionations in continuous culture: okenone producing purple sulfur bacteria Part II

Okenone is a carotenoid pigment unique to certain members of Chromatiaceae, the dominant family of purple sulfur bacteria (PSB) found in euxinic photic zones. Diagenetic alteration of okenone produces okenane, the only recognized molecular fossil unique to PSB. The in vivo concentrations of okenone and bacteriochlorophyll a (Bchl a) on a per cell basis were monitored and quantified as a function of light intensity in continuous cultures of the purple sulfur bacterium Marichromatium purpuratum (Mpurp1591). We show that okenone‐producing PSB have constant bacteriochlorophyll to carotenoid ratios in light‐harvesting antenna complexes. The in vivo concentrations of Bchl a, 0.151 ± 0.012 fmol cell^?1, and okenone, 0.103 ± 0.012 fmol cell^?1, were not dependent on average light intensity (10–225 Lux) at both steady and non‐steady states. This observation revealed that in autotrophic continuous cultures of Mpurp1591, there was a constant ratio for okenone to Bchl a of 1:1.5. Okenone was therefore constitutively produced in planktonic cultures of PSB, regardless of light intensity. This confirms the legitimacy of okenone as a signature for autotrophic planktonic PSB and by extrapolation water column euxinia. We measured the δ¹³C, δ¹⁵N, and δ³⁴S bulk biomass values from cells collected daily and determined the isotopic fractionations of Mpurp1591. There was no statistical relationship in the bulk isotope measurements or stable isotope fractionations to light intensity or cell density under steady and non‐steady‐state conditions. The carbon isotope fractionation between okenone and Bchl a with respect to overall bulk biomass (¹³ε_{pigment – biomass}) was 2.2 ± 0.4‰ and ?4.1 ± 0.9‰, respectively. The carbon isotopic fractionation () for the production of pigments in PSB is more variable than previously thought with our reported values for okenone at ?15.5 ± 1.2‰ and ?21.8 ± 1.7‰ for Bchl a.     

Patterns of sulfur isotope fractionation during microbial sulfate reduction

Studies of microbial sulfate reduction have suggested that the magnitude of sulfur isotope fractionation varies with sulfate concentration. Small apparent sulfur isotope fractionations preserved in Archean rocks have been interpreted as suggesting Archean sulfate concentrations of <200 μm , while larger fractionations thereafter have been interpreted to require higher concentrations. In this work, we demonstrate that fractionation imposed by sulfate reduction can be a function of concentration over a millimolar range, but that nature of this relationship depends on the organism studied. Two sulfate‐reducing bacteria grown in continuous culture with sulfate concentrations ranging from 0.1 to 6 mm showed markedly different relationships between sulfate concentration and isotope fractionation. Desulfovibrio vulgaris str. Hildenborough showed a large and relatively constant isotope fractionation (³⁴ε_SO4‐H2S ? 25‰), while fractionation by Desulfovibrio alaskensis G20 strongly correlated with sulfate concentration over the same range. Both data sets can be modeled as Michaelis–Menten (MM)‐type relationships but with very different MM constants, suggesting that the fractionations imposed by these organisms are highly dependent on strain‐specific factors. These data reveal complexity in the sulfate concentration–fractionation relationship. Fractionation during MSR relates to sulfate concentration but also to strain‐specific physiological parameters such as the affinity for sulfate and electron donors. Previous studies have suggested that the sulfate concentration–fractionation relationship is best described with a MM fit. We present a simple model in which the MM fit with sulfate concentration and hyperbolic fit with growth rate emerge from simple physiological assumptions. As both environmental and biological factors influence the fractionation recorded in geological samples, understanding their relationship is critical to interpreting the sulfur isotope record. As the uptake machinery for both sulfate and electrons has been subject to selective pressure over Earth history, its evolution may complicate efforts to uniquely reconstruct ambient sulfate concentrations from a single sulfur isotopic composition.     

Effect of Oxidation Rate and Fe(II) State on Microbial Nitrate-Dependent Fe(III) Mineral Formation

A nitrate-dependent Fe(II)-oxidizing bacterium was isolated and used to evaluate whether Fe(II) chemical form or oxidation rate had an effect on the mineralogy of biogenic Fe(III) (hydr)oxides resulting from nitrate-dependent Fe(II) oxidation. The isolate (designated FW33AN) had 99% 16S rRNA sequence similarity to Klebsiella oxytoca. FW33AN produced Fe(III) (hydr)oxides by oxidation of soluble Fe(II) [Fe(II)_sol] or FeS under nitrate-reducing conditions. Based on X-ray diffraction (XRD) analysis, Fe(III) (hydr)oxide produced by oxidation of FeS was shown to be amorphous, while oxidation of Fe(II)_sol yielded goethite. The rate of Fe(II) oxidation was then manipulated by incubating various cell concentrations of FW33AN with Fe(II)_sol and nitrate. Characterization of products revealed that as Fe(II) oxidation rates slowed, a stronger goethite signal was observed by XRD and a larger proportion of Fe(III) was in the crystalline fraction. Since the mineralogy of Fe(III) (hydr)oxides may control the extent of subsequent Fe(III) reduction, the variables we identify here may have an effect on the biogeochemical cycling of Fe in anoxic ecosystems.     

Isotopic discrimination and kinetic parameters of RubisCO from the marine bloom‐forming diatom,Skeletonema costatum

The cosmopolitan, bloom‐forming diatom, Skeletonema costatum, is a prominent primary producer in coastal oceans, fixing CO₂ with ribulose 1,5‐bisphosphate carboxylase/oxygenase (RubisCO) that is phylogenetically distinct from terrestrial plant RubisCO. RubisCOs are subdivided into groups based on sequence similarity of their large subunits (IA–ID, II, and III). ID is present in several major oceanic primary producers, including diatoms such as S. costatum, coccolithophores, and some dinoflagellates, and differs substantially in amino acid sequence from the well‐studied IB enzymes present in most cyanobacteria and in green algae and plants. Despite this sequence divergence, and differences in isotopic discrimination apparent in other RubisCO enzymes, stable carbon isotope compositions of diatoms and other marine phytoplankton are generally interpreted assuming enzymatic isotopic discrimination similar to spinach RubisCO (IB). To interpret phytoplankton δ¹³C values, S. costatum RubisCO was characterized via sequence analysis, and measurement of its K_CO2 and V_max, and degree of isotopic discrimination. The sequence of this enzyme placed it among other diatom ID RubisCOs. Michaelis‐Menten parameters were similar to other ID enzymes (K_CO2 = 48.9 ± 2.8 μm ; V_max = 165.1 ± 6.3 nmol min^?1 mg^?1). However, isotopic discrimination (ε = [¹²k/¹³k ? 1] × 1000) was low (18.5‰; 17.0–19.9, 95% CI) when compared to IA and IB RubisCOs (22–29‰), though not as low as ID from coccolithophore, Emiliania huxleyi (11.1‰). Variability in ε‐values among RubisCOs from primary producers is likely reflected in δ¹³C values of oceanic biomass. Currently, δ¹³C variability is ascribed to physical or chemical factors (e.g. illumination, nutrient availability) and physiological responses to these factors (e.g. carbon‐concentrating mechanisms). Estimating the importance of these factors from δ¹³C measurements requires an accurate ε‐value, and a mass‐balance model using the ε‐value for S. costatum RubisCO is presented. Clearly, appropriate ε‐values must be included in interpreting δ¹³C values of environmental samples.     

Diversity decoupled from sulfur isotope fractionation in a sulfate‐reducing microbial community

The extent of fractionation of sulfur isotopes by sulfate‐reducing microbes is dictated by genomic and environmental factors. A greater understanding of species‐specific fractionations may better inform interpretation of sulfur isotopes preserved in the rock record. To examine whether gene diversity influences net isotopic fractionation in situ, we assessed environmental chemistry, sulfate reduction rates, diversity of putative sulfur‐metabolizing organisms by 16S rRNA and dissimilatory sulfite reductase (dsrB) gene amplicon sequencing, and net fractionation of sulfur isotopes along a sediment transect of a hypersaline Arctic spring. In situ sulfate reduction rates yielded minimum cell‐specific sulfate reduction rates < 0.3 × 10^?15 moles cell^?1 day^?1. Neither 16S rRNA nor dsrB diversity indices correlated with relatively constant (38‰–45‰) net isotope fractionation (ε³⁴S_{sulfide‐sulfate}). Measured ε³⁴S values could be reproduced in a mechanistic fractionation model if 1%–2% of the microbial community (10%–60% of Deltaproteobacteria) were engaged in sulfate respiration, indicating heterogeneous respiratory activity within sulfate‐reducing populations. This model indicated enzymatic kinetic diversity of Apr was more likely to correlate with sulfur fractionation than DsrB. We propose that, above a threshold Shannon diversity value of 0.8 for dsrB, the influence of the specific composition of the microbial community responsible for generating an isotope signal is overprinted by the control exerted by environmental variables on microbial physiology.     

Interactions Between Fe(III)-Oxides and Fe(III)-Phyllosilicates During Microbial Reduction 1: Synthetic Sediments

Fe(III)-oxides and Fe(III)-bearing phyllosilicates are the two major iron sources utilized as electron acceptors by dissimilatory iron-reducing bacteria (DIRB) in anoxic soils and sediments. Although there have been many studies on microbial Fe(III)-oxide and Fe(III)-phyllosilicate reduction with both natural and specimen materials, no controlled experimental information is available on the interaction between these two phases when both are available for microbial reduction. In this study, the model DIRB Geobacter sulfurreducens was used to examine the pathways of Fe(III) reduction in Fe(III)-oxide stripped subsurface sediment that was coated with different amounts of synthetic high surface area (HSA) goethite. Cryogenic (12K) ⁵⁷Fe Mössbauer spectroscopy was used to determine changes in the relative abundances of Fe(III)-oxide, Fe(III)-phyllosilicate, and phyllosilicate-associated Fe(II) [Fe(II)-phyllosilicate] in bioreduced samples. Analogous Mössbauer analyses were performed on samples from abiotic Fe(II) sorption experiments in which sediments were exposed to a quantity of exogenous soluble Fe(II) (FeCl₂?2H₂O) comparable to the amount of Fe(II) produced during microbial reduction. A Fe partitioning model was developed to analyze the fate of Fe(II) and assess the potential for abiotic Fe(II)-catalyzed reduction of Fe(III)-phyllosilicates. The microbial reduction experiments indicated that although reduction of Fe(III)-oxide accounted for virtually all of the observed bulk Fe(III) reduction activity, there was no significant abiotic electron transfer between oxide-derived Fe(II) and Fe(III)-phyllosilicatesilicates, with 26–87% of biogenic Fe(II) appearing as sorbed Fe(II) in the Fe(II)-phyllosilicate pool. In contrast, the abiotic Fe(II) sorption experiments showed that 41 and 24% of the added Fe(II) engaged in electron transfer to Fe(III)-phyllosilicate surfaces in synthetic goethite-coated and uncoated sediment. Differences in the rate of Fe(II) addition and system redox potential may account for the microbial and abiotic reaction systems. Our experiments provide new insight into pathways for Fe(III) reduction in mixed Fe(III)-oxide/Fe(III)-phyllosilicate assemblages, and provide key mechanistic insight for interpreting microbial reduction experiments and field data from complex natural soils and sediments.     

Multiple sulphur isotopic interpretations of biosynthetic pathways: implications for biological signatures in the sulphur isotope record

Isotopic fractionations produced by biosynthetic processes are the result of networks of individual biochemical reactions that operate at differing efficiencies and with distinct fractionation factors. These reaction networks determine the magnitude and direction of the net isotopic fractionation associated with a given process. Here we examine the ways that biological reaction networks control mass‐dependent isotopic fractionations of multiple sulphur isotopes. We describe how material‐flow through some networks can produce characteristic multiple‐sulphur‐isotope signatures that differ from those produced by their constituent steps and demonstrate that experimental results with Archaeaglobus fulgidus can be evaluated using multiple sulphur isotopes in the context of previously published models for dissimilatory sulphate reduction. Our evaluation of these data is consistent with the interpretation that the dependence of sulphur isotope fractionation on external sulphate concentration is rooted in differences between the forward and reverse  ? adenosine‐5′‐phosphosulphate (APS) ?  steps. The framework provided by our analysis has the potential to evaluate the biosynthetic pathways that produce the isotopic fractionations, to isolate the primary sources of isotopic fractionations (sulphate reduction or disproportionation reactions) and to establish criteria to identify the signature of specific sulphur metabolisms in the geological record. The results highlight the new types of information that can be obtained by including measurements of δ³³S {δ³³S = [(³³S/³²S)_sample/(³³S/³²S)_reference ? 1]*1000} with measurements of δ³⁴S.     

Nitrogen isotope effects in the assimilation of inorganic nitrogenous compounds by marine diatoms

Nitrogen isotope fractionation in the assimilation of inorganic nitrogenous compounds was studied using marine diatoms (Phaeodactylum tricornutum and Chaetoceros sp.). The isotopic composition (δ¹⁵N) of the diatoms ranged from 7 to ‐18‰ relative to that of the nitrogen source, i.e., ammonium, nitrite, or nitrate. When the growth was light‐limited, the isotope fractionation in nitrate assimilation was inversely correlated with the growth rate. The highest fractionation factor of 1.016 was obtained when the growth rate was as low as 0.025 day^‐1. Fractionation was negligible when the growth, rate was higher than 1 day^‐1. A steady‐state kinetic model was applied to explain the isotope fractionation in nitrate assimilation. The nitrogen isotope fractionation primarily takes place at the step of N‐O bond breaking in nitrate reduction to nitrite. The extent of the isotope fractionation associated with the nitrate uptake is very small, and barely exceeds the limit of detection.