Caroline Schmidt

The interplay of microbially mediated and abiotic reactions in the biogeochemical Fe cycle

Many iron (Fe) redox processes that were previously assumed to be purely abiotic, such as photochemical Fe reactions, are now known to also be microbially mediated. Owing to this overlap, discerning whether biotic or abiotic processes control Fe redox chemistry is a major challenge for geomicrobiologists and biogeochemists alike. Therefore, to understand the network of reactions within the biogeochemical Fe cycle, it is necessary to determine which abiotic or microbially mediated reactions are dominant under various environmental conditions. In this Review, we discuss the major microbially mediated and abiotic reactions in the biogeochemical Fe cycle and provide an integrated overview of biotic and chemically mediated redox transformations.

Microbial Iron(II) Oxidation in Littoral Freshwater Lake Sediment: The Potential for Competition between Phototrophic vs. Nitrate-Reducing Iron(II)-Oxidizers

The distribution of neutrophilic microbial iron oxidation is mainly determined by local gradients of oxygen, light, nitrate and ferrous iron. In the anoxic top part of littoral freshwater lake sediment, nitrate-reducing and phototrophic Fe(II)-oxidizers compete for the same e− donor; reduced iron. It is not yet understood how these microbes co-exist in the sediment and what role they play in the Fe cycle. We show that both metabolic types of anaerobic Fe(II)-oxidizing microorganisms are present in the same sediment layer directly beneath the oxic-anoxic sediment interface. The photoferrotrophic most probable number counted 3.4·105 cells·g−1 and the autotrophic and mixotrophic nitrate-reducing Fe(II)-oxidizers totaled 1.8·104 and 4.5·104 cells·g−1 dry weight sediment, respectively. To distinguish between the two microbial Fe(II) oxidation processes and assess their individual contribution to the sedimentary Fe cycle, littoral lake sediment was incubated in microcosm experiments. Nitrate-reducing Fe(II)-oxidizing bacteria exhibited a higher maximum Fe(II) oxidation rate per cell, in both pure cultures and microcosms, than photoferrotrophs. In microcosms, photoferrotrophs instantly started oxidizing Fe(II), whilst nitrate-reducing Fe(II)-oxidizers showed a significant lag-phase during which they probably use organics as e− donor before initiating Fe(II) oxidation. This suggests that they will be outcompeted by phototrophic Fe(II)-oxidizers during optimal light conditions; as phototrophs deplete Fe(II) before nitrate-reducing Fe(II)-oxidizers start Fe(II) oxidation. Thus, the co-existence of the two anaerobic Fe(II)-oxidizers may be possible due to a niche space separation in time by the day-night cycle, where nitrate-reducing Fe(II)-oxidizers oxidize Fe(II) during darkness and phototrophs play a dominant role in Fe(II) oxidation during daylight. Furthermore, metabolic flexibility of Fe(II)-oxidizing microbes may play a paramount role in the conservation of the sedimentary Fe cycle.

Coexistence of Microaerophilic, Nitrate-Reducing, and Phototrophic Fe(II) Oxidizers and Fe(III) Reducers in Coastal Marine Sediment

ABSTRACT Iron is abundant in sediments, where it can be biogeochemically cycled between its divalent and trivalent redox states. The neutrophilic microbiological Fe cycle involves Fe(III)-reducing and three different physiological groups of Fe(II)-oxidizing microorganisms, i.e., microaerophilic, anoxygenic phototrophic, and nitrate-reducing Fe(II) oxidizers. However, it is unknown whether all three groups coexist in one habitat and how they are spatially distributed in relation to gradients of O2, light, nitrate, and Fe(II). We examined two coastal marine sediments in Aarhus Bay, Denmark, by cultivation and most probable number (MPN) studies for Fe(II) oxidizers and Fe(III) reducers and by quantitative-PCR (qPCR) assays for microaerophilic Fe(II) oxidizers. Our results demonstrate the coexistence of all three metabolic types of Fe(II) oxidizers and Fe(III) reducers. In qPCR, microaerophilic Fe(II) oxidizers (Zetaproteobacteria) were present with up to 3.2 × 106 cells g dry sediment−1. In MPNs, nitrate-reducing Fe(II) oxidizers, anoxygenic phototrophic Fe(II) oxidizers, and Fe(III) reducers reached cell numbers of up to 3.5 × 104, 3.1 × 102, and 4.4 × 104 g dry sediment−1, respectively. O2 and light penetrated only a few millimeters, but the depth distribution of the different iron metabolizers did not correlate with the profile of O2, Fe(II), or light. Instead, abundances were homogeneous within the upper 3 cm of the sediment, probably due to wave-induced sediment reworking and bioturbation. In microaerophilic Fe(II)-oxidizing enrichment cultures, strains belonging to the Zetaproteobacteria were identified. Photoferrotrophic enrichments contained strains related to Chlorobium and Rhodobacter; the nitrate-reducing Fe(II) enrichments contained strains related to Hoeflea and Denitromonas. This study shows the coexistence of all three types of Fe(II) oxidizers in two near-shore marine environments and the potential for competition and interrelationships between them.

Ecosystem functioning from a geomicrobiological perspective a conceptual framework for biogeochemical iron cycling

Environmental context. Microbial ecosystems are characterised by the interplay of various microorganisms with their biotic and abiotic environment. Biogeochemical niches host adapted microbial communities that are in constant competition for substrates and nutrients. Their natural distribution, interactions and responses to fluctuating environmental conditions are often impossible to simulate in laboratory studies. Using biogeochemical iron redox cycling as an example, we suggest the application of a conceptual framework to improve our understanding of the principal functioning of (geo)microbial ecosystems.

Metabolic Flexibility and Substrate Preference by the Fe(II)-Oxidizing Purple Non-Sulphur Bacterium Rhodopseudomonas palustris Strain TIE-1

It is unknown to which extent phototrophic Fe(II) oxidation takes place in the simultaneous presence of organic electron donors (e.g., acetate/lactate). Therefore, the photoferrotrophic strain Rhodopseudomonas palustris TIE-1 was inoculated with various combinations of Fe(II), acetate and lactate to understand metabolic substrate preference. When acetate was provided together with Fe(II), TIE-1 consumed acetate first before Fe(II). When provided lactate plus Fe(II), TIE-1 consumed both substrates in parallel. When all three substrates were provided in one culture, TIE-1 used all substrates simultaneously. Our study suggests that the availability of alternative electron donors in addition to ferrous iron limits phototrophic iron oxidation.

High spatial resolution of distribution and interconnections between Fe‐ and N‐redox processes in profundal lake sediments

The Fe and N biogeochemical cycles play key roles in freshwater environments. We aimed to determine the spatial positioning and interconnections of the N and Fe cycles in profundal lake sediments. The gradients of O2, NO3(-), NH4(+), pH, Eh, Fe(II) and Fe(III) were determined and the distribution of microorganisms was assessed by most probable numbers and quantitative polymerase chain reaction. The redox zones could be divided into an oxic zone (0-8 mm), where microaerophiles (Gallionellaceae) were most abundant at a depth of 7 mm. This was followed by a denitrification zone (6-12 mm), where NO3(-)-dependent Fe(II) oxidizers and organoheterotrophic denitrifiers both reduce nitrate. Lastly, an iron redox transition zone was identified at 12.5-22.5 mm. Fe(III) was most abundant above this zone while Fe(II) was most abundant beneath. The high abundance of poorly crystalline iron suggested iron cycling. The Fe and N cycles are biologically connected through nitrate-reducing Fe(II) oxidizers and chemically by NOx(-) species formed during denitrification, which can chemically oxidize Fe(II). This study combines high resolution chemical, molecular and microbiological data to pinpoint sedimentary redox zones in which Fe is cycled between Fe(II) and Fe(III) and where Fe and N-redox processes interact.

Anaerobic microbial Fe(II) oxidation and Fe(III) reduction in coastal marine sediments controlled by organic carbon content

Coastal marine sediments contain varying concentrations of iron, oxygen, nitrate and organic carbon. It is unknown how organic carbon content influences the activity of nitrate-reducing and phototrophic Fe(II)-oxidizers and microbial Fe-redox cycling in such sediments. Therefore, microcosms were prepared with two coastal marine sediments (Kalø Vig and Norsminde Fjord at Aarhus Bay, Denmark) varying in TOC from 0.4 to 3.0 wt%. The microcosms were incubated under light/dark conditions with/without addition of nitrate and/or Fe(II). Although most probable number (MPN) counts of phototrophic Fe(II)-oxidizers were five times lower in the low-TOC sediment, phototrophic Fe(II) oxidation rates were higher compared with the high-TOC sediment. Fe(III)-amended microcosms showed that this lower net Fe(II) oxidation in the high-TOC sediment is caused by concurrent bacterial Fe(III) reduction. In contrast, MPN counts of nitrate-reducing Fe(II)-oxidizers and net rates of nitrate-reducing Fe(II) oxidation were comparable in low- and high-TOC sediments. However, the ratio of nitratereduced :iron(II)oxidized was higher in the high-TOC sediment, suggesting that a part of the nitrate was reduced by mixotrophic nitrate-reducing Fe(II)-oxidizers and chemoorganoheterotrophic nitrate-reducers. Our results demonstrate that dynamic microbial Fe cycling occurs in these sediments and that the extent of Fe cycling is dependent on organic carbon content.

Secondary Mineral Formation During Ferrihydrite Reduction by Shewanella oneidensis MR-1 Depends on Incubation Vessel Orientation and Resulting Gradients of Cells, Fe2+ and Fe Minerals

In previous studies on microbial ferric iron (Fe(III)) reduction varying results regarding reduction rates and secondary mineral formation have been reported for almost identical conditions regarding temperature, pH, medium composition, Fe(III) mineral identity and bulk iron concentration. Here we show that in addition to physico-chemical parameters also geometric aspects, i.e., incubation orientation and dimension of cultivation vessels, influence the reduction rates and mineralogy. We incubated the Fe(III)-reducer Shewanella oneidensis MR-1 in test tubes at ferrihydrite (FH) concentrations of 1.3–50 mM either in vertical or horizontal orientation. Cells and minerals formed a pellet at the bottom of the tubes with different thicknesses at the same initial FH concentration depending on the incubation orientation. In vertically incubated tubes thick FH pellets were present at the bottom of the tubes and magnetite was formed in all setups with ≥2.5 mM initial FH. In tubes that were incubated horizontally no magnetite was formed in presence of <5 mM initial FH. Spatially resolved analysis of the supernatant and mineral sediment including voltammetric microelectrodes, Xray diffraction and Mössbauer spectroscopy revealed strong gradients of Fe2+ in both the aqueous supernatant and mineral pellets, whereas a heterogeneous distribution of cells and minerals in the sediment pellet was detected. The highest cell density and, consequently, the initiation of FH reduction was found at the mineral-supernatant interface. This study demonstrates that small changes in incubation conditions can significantly influence and even change the experimental results of geomicrobiological experiments.

A Revised Iron Extraction Protocol for Environmental Samples Rich in Nitrite and Carbonate

ABSTRACT Wet-chemical iron extraction is widely applied to quantify the mineral-bound ferriferous fraction of sediments and soils. As previously shown, this method is strongly affected by the composition of the soil/sediment. Samples enriched in mostly microbially produced nitrite require the removal of the nitrite-containing aqueous phase or the replacement of HCl with sulfamic acid (SA) as the extractant. In this study, we show that sedimentary carbonate buffers SA, inhibiting the stabilization of Fe(II) and effective extraction of iron. We therefore provide a revised extraction protocol which allows the preservation of low pH conditions, leading to efficient iron extraction in nitrite- and carbonate-enriched samples.

Microbially mediated coupling of Fe and N cycles by nitrate-reducing Fe(II)-oxidizing bacteria in littoral freshwater sediments

ABSTRACT Nitrate-reducing iron(II)-oxidizing bacteria have been known for approximately 20 years. There has been much debate as to what extent the reduction of nitrate and the oxidation of ferrous iron are coupled via enzymatic pathways or via abiotic processes induced by nitrite formed by heterotrophic denitrification. The aim of the present study was to assess the coupling of nitrate reduction and iron(II) oxidation by monitoring changes in substrate concentrations, as well as in the activity of nitrate-reducing bacteria in natural littoral freshwater sediment, in response to stimulation with nitrate and iron(II). In substrate-amended microcosms, we found that the biotic oxidation of ferrous iron depended on the simultaneous microbial reduction of nitrate. Additionally, the abiotic oxidation of ferrous iron by nitrite in sterilized sediment was not fast enough to explain the iron oxidation rates observed in microbially active sediment. Furthermore, the expression levels of genes coding for enzymes crucial for nitrate reduction were in some setups stimulated by the presence of ferrous iron. These results indicate that there is a direct influence of ferrous iron on bacterial denitrification and support the hypothesis that microbial nitrate reduction is stimulated by biotic iron(II) oxidation. IMPORTANCE The coupling of nitrate reduction and Fe(II) oxidation affects the environment at a local scale, e.g., by changing nutrient or heavy metal mobility in soils due to the formation of Fe(III) minerals, as well as at a global scale, e.g., by the formation of the primary greenhouse gas nitrous oxide. Although the coupling of nitrate reduction and Fe(II) oxidation was reported 20 years ago and has been studied intensively since then, the underlying mechanisms still remain unknown. One of the main knowledge gaps is the extent of enzymatic Fe(II) oxidation coupled to nitrate reduction, which has frequently been questioned in the literature. In the present study, we provide evidence for microbially mediated nitrate-reducing Fe(II) oxidation in freshwater sediments. This evidence is based on the rates of nitrate reduction and Fe(II) oxidation determined in microcosm incubations and on the effect of iron on the expression of genes required for denitrification.