Phototrophic Fe(II) oxidation by Rhodopseudomonas palustris TIE-1 in organic and Fe(II)-rich conditions.

Rhodopseudomonas palustris TIE-1 grows photoautotrophically with Fe(II) as an electron donor and photoheterotrophically with a variety of organic substrates. However, it is unclear whether R. palustris TIE-1 conducts Fe(II) oxidation in conditions where organic substrates and Fe(II) are available simultaneously. In addition, the effect of organic co-substrates on Fe(II) oxidation rates or the identity of Fe(III) minerals formed is unknown. We incubated R. palustris TIE-1 with 2 mM Fe(II), amended with 0.6 mM organic co-substrate, and in the presence/absence of CO2 . We found that in the absence of CO2 , only the organic co-substrates acetate, lactate and pyruvate, but not Fe(II), were consumed. When CO2 was present, Fe(II) and all organic substrates were consumed. Acetate, butyrate and pyruvate were consumed before Fe(II) oxidation commenced, whereas lactate and glucose were consumed at the same time as Fe(II) oxidation proceeded. Lactate, pyruvate and glucose increased the Fe(II) oxidation rate significantly (by up to threefold in the case of lactate). 57 Fe Mössbauer spectroscopy revealed that short-range ordered Fe(III) oxyhydroxides were formed under all conditions. This study demonstrates phototrophic Fe(II) oxidation proceeds even in the presence of organic compounds, and that the simultaneous oxidation of organic substrates can stimulate Fe(II) oxidation.

Anaerobic Neutrophilic Pyrite Oxidation by a Chemolithoautotrophic Nitrate-Reducing Iron(II)-Oxidizing Culture Enriched from a Fractured Aquifer.

Neutrophilic microbial pyrite (FeS2) oxidation coupled to denitrification is thought to be an important natural nitrate attenuation pathway in nitrate-contaminated aquifers. However, the poor solubility of pyrite raises questions about its bioavailability and the mechanisms underlying its oxidation. Here, we investigated direct microbial pyrite oxidation by a neutrophilic chemolithoautotrophic nitrate-reducing Fe(II)-oxidizing culture enriched from a pyrite-rich aquifer. We used pyrite with natural abundance (NA) of Fe isotopes (NAFe-pyrite) and 57Fe-labeled siderite to evaluate whether the oxidation of the more soluble Fe(II)-carbonate (FeCO3) can indirectly drive abiotic pyrite oxidation. Our results showed that in setups where only pyrite was incubated with bacteria, direct microbial pyrite oxidation contributed ca. 26% to overall nitrate reduction. The rest was attributed to the oxidation of elemental sulfur (S0), present as a residue from pyrite synthesis. Pyrite oxidation was evidenced in the NAFe-pyrite/57Fe-siderite setups by maps of 56FeO and 32S obtained using a combination of SEM with nanoscale secondary ion MS (NanoSIMS), which showed the presence of 56Fe(III) (oxyhydr)oxides that could solely originate from 56FeS2. Based on the fit of a reaction model to the geochemical data and the Fe-isotope distributions from NanoSIMS, we conclude that anaerobic oxidation of pyrite by our neutrophilic enrichment culture was mainly driven by direct enzymatic activity of the cells. The contribution of abiotic pyrite oxidation by Fe3+ appeared to be negligible in our experimental setup.

Organic Matter from Redoximorphic Soils Accelerates and Sustains Microbial Fe(III) Reduction.

Microbial reduction of Fe(III) minerals is a prominent process in redoximorphic soils and is strongly affected by organic matter (OM). We herein determined the rate and extent of microbial reduction of ferrihydrite (Fh) with either adsorbed or coprecipitated OM by Geobacter sulfurreducens. We focused on OM-mediated effects on electron uptake and alterations in Fh crystallinity. The OM was obtained from anoxic soil columns (effluent OM, efOM) and included-unlike water-extractable OM-compounds released by microbial activity under anoxic conditions. We found that organic molecules in efOM had generally no or only very low electron-accepting capacity and were incorporated into the Fh aggregates when coprecipitated with Fh. Compared to OM-free Fh, adsorption of efOM to Fh decelerated the microbial Fe(III) reduction by passivating the Fh surface toward electron uptake. In contrast, coprecipitation of Fh with efOM accelerated the microbial reduction, likely because efOM disrupted the Fh structure, as noted by Mössbauer spectroscopy. Additionally, the adsorbed and coprecipitated efOM resulted in a more sustained Fe(III) reduction, potentially because efOM could have effectively scavenged biogenic Fe(II) and prevented the passivation of the Fh surface by the adsorbed Fe(II). Fe(III)-OM coprecipitates forming at anoxic-oxic interfaces are thus likely readily reducible by Fe(III)-reducing bacteria in redoximorphic soils.

Influence of Physical Perturbation on Fe(II) Supply in Coastal Marine Sediments.

Iron (Fe) biogeochemistry in marine sediments is driven by redox transformations creating Fe(II) and Fe(III) gradients. As sediments are physically mixed by wave action or bioturbation, Fe gradients re-establish regularly. In order to identify the response of dissolved Fe(II) (Fe2+) and Fe mineral phases toward mixing processes, we performed voltammetric microsensor measurements, sequential Fe extractions, and Mössbauer spectroscopy of 12 h light-dark cycle incubated marine coastal sediment. Fe2+ decreased during 7 days of undisturbed incubation from approximately 400 to 60 µM. In the first 2-4 days of incubation, Fe2+ accumulated up to 100 µM in the top 2 mm due to Fe(III) photoreduction. After physical perturbation at day 7, Fe2+ was re-mobilized reaching concentrations of 320 µM in 30 mm depth, which decreased to below detection limit within 2 days afterward. Mössbauer spectroscopy showed that the relative abundance of metastable iron-sulfur mineral phases (FeSx) increased during initial incubation and decreased together with pyrite (FeS2) after perturbation. We show that Fe2+ mobilization in marine sediments is stimulated by chemical changes caused by physical disturbances impacting the Fe redox distribution. Our study suggests that, in addition to microbial and abiotic Fe(III) reduction, including Fe(III) photoreduction, physical mixing processes induce chemical changes providing sediments and the inhabiting microbial community with Fe2+.

Contribution of Microaerophilic Iron(II)-Oxidizers to Iron(III) Mineral Formation.

Neutrophilic microbial aerobic oxidation of ferrous iron (Fe(II)) is restricted to pH-circumneutral environments characterized by low oxygen where microaerophilic Fe(II)-oxidizing microorganisms successfully compete with abiotic Fe(II) oxidation. However, accumulation of ferric (bio)minerals increases competition by stimulating abiotic surface-catalyzed heterogeneous Fe(II) oxidation. Here, we present an experimental approach that allows quantification of microbial and abiotic contribution to Fe(II) oxidation in the presence or initial absence of ferric (bio)minerals. We found that at 20 µM O2 and the initial absence of Fe(III) minerals, an iron(II)-oxidizing enrichment culture (99.6% similarity to Sideroxydans spp.) contributed 40% to the overall Fe(II) oxidation within approximately 26 h and oxidized up to 3.6 × 10-15 mol Fe(II) cell-1 h-1. Optimum O2 concentrations at which enzymatic Fe(II) oxidation can compete with abiotic Fe(II) oxidation ranged from 5 to 20 µM. Lower O2 levels limited biotic Fe(II) oxidation, while at higher O2 levels abiotic Fe(II) oxidation dominated. The presence of ferric (bio)minerals induced surface-catalytic heterogeneous abiotic Fe(II) oxidation and reduced the microbial contribution to Fe(II) oxidation from 40% to 10% at 10 µM O2. The obtained results will help to better assess the impact of microaerophilic Fe(II) oxidation on the biogeochemical iron cycle in a variety of environmental natural and anthropogenic settings.

Phototrophic Fe(II) oxidation benefits from light/dark cycles.

Phototrophic Fe(II)-oxidizers use Fe(II) as electron donor for CO2 fixation thus linking Fe(II) oxidation, ATP formation, and growth directly to the availability of sunlight. We compared the effect of short (10 h light/14 h dark) and long (2-3 days light/2-3 days dark) light/dark cycles to constant light conditions for the phototrophic Fe(II)-oxidizer Chlorobium ferrooxidans KoFox. Fe(II) oxidation was completed first in the setup with constant light (9 mM Fe(II) oxidised within 8.9 days) compared to the light/dark cycles but both short and long light/dark cycles showed faster maximum Fe(II) oxidation rates. In the short and long cycle, Fe(II) oxidation rates reached 3.5 ± 1.0 and 2.6 ± 0.3 mM/d, respectively, compared to 2.1 ± 0.3 mM/d in the constant light setup. Maximum Fe(II) oxidation was significantly faster in the short cycle compared to the constant light setup. Cell growth reached roughly equivalent cell numbers across all three light conditions (from 0.2-2.0 × 106 cells/mL to 1.1-1.4 × 108 cells/mL) and took place in both the light and dark phases of incubation. SEM images showed different mineral structures independent of the light setup and 57 Fe Mössbauer spectroscopy confirmed the formation of poorly crystalline Fe(III) oxyhydroxides (such as ferrihydrite) in all three setups. Our results suggest that periods of darkness have a significant impact on phototrophic Fe(II)-oxidizers and significantly influence rates of Fe(II) oxidation.

Stabilization of mineral-associated organic carbon in Pleistocene permafrost.

Ice-rich Pleistocene-age permafrost is particularly vulnerable to rapid thaw, which may quickly expose a large pool of sedimentary organic matter (OM) to microbial degradation and lead to emissions of climate-sensitive greenhouse gases. Protective physico-chemical mechanisms may, however, restrict microbial accessibility and reduce OM decomposition; mechanisms that may be influenced by changing environmental conditions during sediment deposition. Here we study different OM fractions in Siberian permafrost deposited during colder and warmer periods of the past 55,000 years. Among known stabilization mechanisms, the occlusion of OM in aggregates is of minor importance, while 33-74% of the organic carbon is associated with small, <6.3 µm mineral particles. Preservation of carbon in mineral-associated OM is enhanced by reactive iron minerals particularly during cold and dry climate, reflected by low microbial CO2 production in incubation experiments. Warmer and wetter conditions reduce OM stabilization, shown by more decomposed mineral-associated OM and up to 30% higher CO2 production. This shows that considering the stability and bioavailability of Pleistocene-age permafrost carbon is important for predicting future climate-carbon feedback.

Influence of Fe(III) source, light quality, photon flux and presence of oxygen on photoreduction of Fe(III)-organic complexes - Implications for light-influenced coastal freshwater and marine sediments.

Iron(III) photoreduction is an important source of Fe(II) in illuminated aquatic and sedimentary environments. Under oxic conditions, the Fe(II) can be re-oxidized by oxygen (O2) forming reactive O-species such as hydrogen peroxide (H2O2) which further react with Fe(II) thus enhancing Fe(II) oxidation rates. However, it is unknown by aquatic sediments how the parameters wavelength of radiation, photon flux, origin of Fe(III) source and presence or absence of O2 influence the extent of Fe(II) and H2O2 turnover. We studied this using batch experiments with different Fe(III)-organic complexes mimicking sedimentary conditions. We found that wavelengths <500 nm are necessary to initiate Fe(III) photoreduction and that the photon flux, wavelength and identity of Fe(III)-complexing organic acids control the kinetics of Fe(III) photoreduction. The formation of photo-susceptible Fe(III)-organic complexes did not depend on whether the Fe(III) source was biogenically produced, poorly-crystalline Fe(III) oxyhydroxides or chemically synthesized ferrihydrite. Oxic conditions caused chemical re-oxidation of Fe(II) and accumulation of H2O2. The photon flux, wavelength and availability of Fe(III)-complexing organic molecules are critical for the balance between concurrent Fe(III) photoreduction and abiotic Fe(II) oxidation and may even lead to a steady-state concentration of Fe(II) in the micromolar range. These results help understand and predict Fe(III) photoreduction dynamics and in-situ formation of Fe(II) in oxic or anoxic, illuminated and organic-rich environments.

Growth of microaerophilic Fe(II)-oxidizing bacteria using Fe(II) produced by Fe(III) photoreduction.

Iron(II) (Fe(II)) can be formed by abiotic Fe(III) photoreduction, particularly when Fe(III) is organically complexed. Light-influenced environments often overlap or even coincide with oxic or microoxic geochemical conditions, for example, in sediments. So far, it is unknown whether microaerophilic Fe(II)-oxidizing bacteria are able to use the Fe(II) produced by Fe(III) photoreduction as electron donor. Here, we present an adaption of the established agar-stabilized gradient tube approach in comparison with liquid cultures for the cultivation of microaerophilic Fe(II)-oxidizing microorganisms by using a ferrihydrite-citrate mixture undergoing Fe(III) photoreduction as Fe(II) source. We quantified oxygen and Fe(II) gradients with amperometric and voltammetric microelectrodes and evaluated microbial growth by qPCR of 16S rRNA genes. We showed that gradients of dissolved Fe(II) (maximum Fe(II) concentration of 1.25 mM) formed in the gradient tubes when incubated in blue or UV light (400-530 nm or 350-400 nm). Various microaerophilic Fe(II)-oxidizing bacteria (Curvibacter sp. and Gallionella sp.) grew by oxidizing Fe(II) that was produced in situ by Fe(III) photoreduction. Best growth for these species, based on highest gene copy numbers, was observed in incubations using UV light in both liquid culture and gradient tubes containing 8 mM ferrihydrite-citrate mixtures (1:1), due to continuous light-induced Fe(II) formation. Microaerophilic Fe(II)-oxidizing bacteria contributed up to 40% to the overall Fe(II) oxidation within 24 h of incubation in UV light. Our results highlight the potential importance of Fe(III) photoreduction as a source of Fe(II) for Fe(II)-oxidizing bacteria by providing Fe(II) in illuminated environments, even under microoxic conditions.

Chromium (VI) removal kinetics by magnetite-coated sand: Small-scale flow-through column experiments.

Magnetite nanoparticles are promising materials for treating toxic Cr(VI), but safe handling is challenging due to their small size. We prepared flow-through columns containing 10% or 100% (v/v) magnetite-coated sand. Cr(VI) removal efficiency was determined for different Cr(VI) concentrations (0.1 or 1.0 mM), neutral or alkaline pH, and oxic/anoxic conditions. We formulated a reactive-transport model that accurately predicted total Cr removal, accounting for reversible and irreversible (chemi)sorption reactions. Our results show that the material removes and irreversibly sequesters Cr(VI). For the concentration range used 10% and 100% (v/v) -packed columns removed > 99% and 72% of influent Cr(VI), respectively. Two distinct parameter sets were necessary to fit the identical model formulation to the 10 or 100% (v/v) columns (e.g., maximum sorption capacities (qmax) of 1.37 µmol Cr/g sand and 2.48 µmol Cr/g, respectively), which we attributed to abrasion-driven magnetite micro-particle detachment during packing yielding an increase in reactive surface area. Furthermore, experiments under oxic conditions showed that, even when handled in the presence of O2, the magnetite-coated sand maintained a high removal capacity (47%). Our coupled experimental and modelling analyses indicates that magnetite-coated sand is a promising and suitable medium for treating Cr(VI)-contaminated water in fixed-bed reactors or permeable reactive barriers.

Potentially bioavailable iron produced through benthic cycling in glaciated Arctic fjords of Svalbard.

The Arctic has the highest warming rates on Earth. Glaciated fjord ecosystems, which are hotspots of carbon cycling and burial, are extremely sensitive to this warming. Glaciers are important for the transport of iron from land to sea and supply this essential nutrient to phytoplankton in high-latitude marine ecosystems. However, up to 95% of the glacially-sourced iron settles to sediments close to the glacial source. Our data show that while 0.6-12% of the total glacially-sourced iron is potentially bioavailable, biogeochemical cycling in Arctic fjord sediments converts the glacially-derived iron into more labile phases, generating up to a 9-fold increase in the amount of potentially bioavailable iron. Arctic fjord sediments are thus an important source of potentially bioavailable iron. However, our data suggests that as glaciers retreat onto land the flux of iron to the sediment-water interface may be reduced. Glacial retreat therefore likely impacts iron cycling in coastal marine ecosystems.

Aggregation-dependent electron transfer via redox-active biochar particles stimulate microbial ferrihydrite reduction.

Microbial Fe(III) reduction plays an important role for biogeochemical carbon and iron cycling in sediments and soils. Biochar is used as a soil amendment to increase fertility and lower N2O/CO2 emissions. It is redox-active and can stimulate microbial Fe(III) mineral reduction. It is currently unknown, however, how the aggregation of cells and Fe(III) minerals with biochar particles influence microbial Fe(III) reduction. Therefore, we determined rates and extent of ferrihydrite (Fh) reduction in S. oneidensis MR-1 cell suspensions with different particles sizes of wood-derived Swiss biochar and KonTiki biochar at different biochar/Fh ratios. We found that at small biochar particle size and high biochar/Fh ratios, the biochar, MR-1 cells and Fh closely aggregated, therefore addition of biochar stimulated electron transfer and microbial Fh reduction. In contrast, large biochar particles and low biochar/Fh ratios inhibited the electron transfer and Fe(III) reduction due to the lack of effective aggregation. These results suggest that for stimulating Fh reduction, a certain biochar particle size and biochar/Fh ratio is necessary leading to a close aggregation of all phases. This aggregation favors electron transfer from cells to Fh via redox cycling of the electron donating and accepting functional groups of biochar and via direct electron transfer through conductive biochar carbon matrices. These findings improve our understanding of electron transfer between microorganisms and Fe(III) minerals via redox-active biochar and help to evaluate the impact of biochar on electron transfer processes in the environment.

Oxic Fe(III) reduction could have generated Fe(II) in the photic zone of Precambrian seawater.

Many marine Precambrian iron formations (IF) record deep anoxic seawater enriched in Fe(II) (i.e. ferruginous) overlain by mildly oxygenated surface water. This is reflected by iron-rich sediments forming in deep basins, and relatively iron-poor sediments forming in shallow, sunlit waters. Such an iron gradient is often interpreted as a redox interface where dissolved Fe(II) was oxidized and precipitated as Fe(III)-bearing minerals. As such, sedimentary iron enrichments are proxy to the progressive oxidation of the oceans through geological time. However, this interpretation is founded on the assumption that Fe(II) could not persist within an oxygenated water column. Here, we cultivated cyanobacteria in an illuminated column supplied with Fe(II)-rich seawater medium in a laboratory-scale analog of a continental margin supporting IF deposition. We first observed Fe(II) oxidation with oxygen, then biologically-mediated reduction of Fe(III) (oxyhydr)oxides, which maintained a pool of Fe(II) in the presence of oxygen. Such steady-state iron redox cycling may have maintained dissolved, and hence mobile Fe(II) in oxygenated seawater above ferruginous deep basins such as those inferred for many Precambrian IF.

Sterilization impacts on marine sediment---Are we able to inactivate microorganisms in environmental samples?

To distinguish between biotic and abiotic processes in laboratory experiments with environmental samples, an effective sterilization method is required that prevents biological activity but does not change physico-geochemical properties of samples. We compared standard sterilization methods with respect to their impact on microbial abundance and activity. We exposed marine sediment to (i) autoclaving, (ii) gamma-radiation or (iii) sodium azide (NaN3) and determined how nucleic acids, microbial productivity, colony forming units (CFUs) and community composition of microorganisms, fungi, unicellular protists and protozoa were affected. In autoclaved and gamma-sterilized sediments, only few colonies formed within 16 days. After addition of NaN3 to the sediment, numerous CFUs (>50) but lower 3H-leucine incorporation rates, i.e. lower protein biosynthesis rates, were found compared to the other two sterilization techniques. Extractable RNA was detected immediately after all sterilization treatments (0.2-17.9 ng/g dry sediment) but decreased substantially by 84%-98% after 16 days of incubation. The total organic carbon content increased from 18 mg L-1 to 220 mg L-1 (autoclaving) and 150 mg L-1 (gamma-radiation) after sterilization. We compare advantages and disadvantages for each tested sterilization method and provide a helpful decision-making resource for choosing the appropriate sterilization technique for environmental studies, particularly for marine sediments.

Laboratory Simulation of an Iron(II)-rich Precambrian Marine Upwelling System to Explore the Growth of Photosynthetic Bacteria.

A conventional concept for the deposition of some Precambrian Banded Iron Formations (BIF) proceeds on the assumption that ferrous iron [Fe(II)] upwelling from hydrothermal sources in the Precambrian ocean was oxidized by molecular oxygen [O2] produced by cyanobacteria. The oldest BIFs, deposited prior to the Great Oxidation Event (GOE) at about 2.4 billion years (Gy) ago, could have formed by direct oxidation of Fe(II) by anoxygenic photoferrotrophs under anoxic conditions. As a method for testing the geochemical and mineralogical patterns that develop under different biological scenarios, we designed a 40 cm long vertical flow-through column to simulate an anoxic Fe(II)-rich marine upwelling system representative of an ancient ocean on a lab scale. The cylinder was packed with a porous glass bead matrix to stabilize the geochemical gradients, and liquid samples for iron quantification could be taken throughout the water column. Dissolved oxygen was detected non-invasively via optodes from the outside. Results from biotic experiments that involved upwelling fluxes of Fe(II) from the bottom, a distinct light gradient from top, and cyanobacteria present in the water column, show clear evidence for the formation of Fe(III) mineral precipitates and development of a chemocline between Fe(II) and O2. This column allows us to test hypotheses for the formation of the BIFs by culturing cyanobacteria (and in the future photoferrotrophs) under simulated marine Precambrian conditions. Furthermore we hypothesize that our column concept allows for the simulation of various chemical and physical environments - including shallow marine or lacustrine sediments.