Martin Obst

Extracellular iron biomineralization by photoautotrophic iron-oxidizing bacteria.

ABSTRACT Iron oxidation at neutral pH by the phototrophic anaerobic iron-oxidizing bacterium Rhodobacter sp. strain SW2 leads to the formation of iron-rich minerals. These minerals consist mainly of nano-goethite (α-FeOOH), which precipitates exclusively outside cells, mostly on polymer fibers emerging from the cells. Scanning transmission X-ray microscopy analyses performed at the C K-edge suggest that these fibers are composed of a mixture of lipids and polysaccharides or of lipopolysaccharides. The iron and the organic carbon contents of these fibers are linearly correlated at the 25-nm scale, which in addition to their texture suggests that these fibers act as a template for mineral precipitation, followed by limited crystal growth. Moreover, we evidence a gradient of the iron oxidation state along the mineralized fibers at the submicrometer scale. Fe minerals on these fibers contain a higher proportion of Fe(III) at cell contact, and the proportion of Fe(II) increases at a distance from the cells. All together, these results demonstrate the primordial role of organic polymers in iron biomineralization and provide first evidence for the existence of a redox gradient around these nonencrusting, Fe-oxidizing bacteria.

Green Rust Formation during Fe(II) Oxidation by the Nitrate-Reducing Acidovorax sp Strain BoFeN1

Green rust (GR) as highly reactive iron mineral potentially plays a key role for the fate of (in)organic contaminants, such as chromium or arsenic, and nitroaromatic compounds functioning both as sorbent and reductant. GR forms as corrosion product of steel but is also naturally present in hydromorphic soils and sediments forming as metastable intermediate during microbial Fe(III) reduction. Although already suggested to form during microbial Fe(II) oxidation, clear evidence for GR formation during microbial Fe(II) oxidation was lacking. In the present study, powder XRD, synchrotron-based XAS, Mössbauer spectroscopy, and TEM demonstrated unambiguously the formation of GR as an intermediate product during Fe(II) oxidation by the nitrate-reducing Fe(II)-oxidizer Acidovorax sp. strain BoFeN1. The spatial distribution and Fe redox-state of the precipitates associated with the cells were visualized by STXM. It showed the presence of extracellular Fe(III), which can be explained by Fe(III) export from the cells or extracellular Fe(II) oxidation by an oxidant diffusing from the cells. Moreover, GR can be oxidized by nitrate/nitrite and is known as a catalyst for oxidation of dissolved Fe(II) by nitrite/nitrate and may thus contribute to the production of extracellular Fe(III). As a result, strain BoFeN1 may contribute to Fe(II) oxidation and nitrate reduction both by an direct enzymatic pathway and an indirect GR-mediated process.

ARE PICOPLANKTON RESPONSIBLE FOR CALCITE PRECIPITATION IN LAKES

Abstract Deposits of lacustrine calcite are important records of environmental changes. In order to interpret these archives, knowledge about the origin of the calcite is essential. It has been accepted that calcite precipitation can be induced by bacteria and algae. However, the detailed mechanisms are still unclear. This review summarizes what is known about the interactions between calcite precipitation and the autotrophic picoplankton. We consider findings from both field and laboratory studies. Field studies show that calcite precipitation in oligotrophic lakes is strongly linked with picocyanobacteria blooms. Laboratory experiments led to the formulation of the mechanism of precipitation induced by microalga. Experiments also showed that precipitation induced by picocyanobacteria is influenced by various factors including the uptake of inorganic carbon and the structure of the cell walls. Recent studies indicate that the influence of environmental conditions like the composition of lake water has to be taken into account as well. We conclude that in situ observations of precipitation processes at picoplankton cells under controlled conditions are needed to improve our understanding of mineral bacteria interaction.

Potential role of nitrite for abiotic Fe(II) oxidation and cell encrustation during nitrate reduction by denitrifying bacteria

ABSTRACT Microorganisms have been observed to oxidize Fe(II) at neutral pH under anoxic and microoxic conditions. While most of the mixotrophic nitrate-reducing Fe(II)-oxidizing bacteria become encrusted with Fe(III)-rich minerals, photoautotrophic and microaerophilic Fe(II) oxidizers avoid cell encrustation. The Fe(II) oxidation mechanisms and the reasons for encrustation remain largely unresolved. Here we used cultivation-based methods and electron microscopy to compare two previously described nitrate-reducing Fe(II) oxidizers ( Acidovorax sp. strain BoFeN1 and Pseudogulbenkiania sp. strain 2002) and two heterotrophic nitrate reducers (Paracoccus denitrificans ATCC 19367 and P. denitrificans Pd 1222). All four strains oxidized ∼8 mM Fe(II) within 5 days in the presence of 5 mM acetate and accumulated nitrite (maximum concentrations of 0.8 to 1.0 mM) in the culture media. Iron(III) minerals, mainly goethite, formed and precipitated extracellularly in close proximity to the cell surface. Interestingly, mineral formation was also observed within the periplasm and cytoplasm; intracellular mineralization is expected to be physiologically disadvantageous, yet acetate consumption continued to be observed even at an advanced stage of Fe(II) oxidation. Extracellular polymeric substances (EPS) were detected by lectin staining with fluorescence microscopy, particularly in the presence of Fe(II), suggesting that EPS production is a response to Fe(II) toxicity or a strategy to decrease encrustation. Based on the data presented here, we propose a nitrite-driven, indirect mechanism of cell encrustation whereby nitrite forms during heterotrophic denitrification and abiotically oxidizes Fe(II). This work adds to the known assemblage of Fe(II)-oxidizing bacteria in nature and complicates our ability to delineate microbial Fe(II) oxidation in ancient microbes preserved as fossils in the geological record.

TEM-specimen preparation of cell/mineral interfaces by Focused Ion Beam milling

Abstract Picocyanobacteria were found to play an important role in calcite precipitation in oligotrophic lakes. In this study, investigations on the interface between cyanobacteria and attached biogenic calcite crystals have been performed to gain further insights into the mechanisms of nucleation of these precipitates. Ultramicrotomy, the conventional preparation technique of thin sections for Transmission Electron Microscopy (TEM) investigations, often fails when working on heterogeneous samples containing soft organic material and hard minerals. Thus, in this study the thin sections were prepared using Focused Ion Beam (FIB) milling. This approach is usually applied in material sciences but until recently was not very common in environmental research. Different analytical TEM methods like Electron Spectroscopic Imaging (ESI) and Electron Energy Loss Spectrometry (EELS) were used to test the suitability of FIB-milling for the preparation of organic/inorganic interface specimens. With this approach we were able to analyze both organic and the inorganic phases of the same sample. Elemental maps of the samples were also calculated. By analyzing the structure of the C K-absorption edge, the different bonding forms of the organic carbon cell and the inorganic carbon of the crystal could be clearly distinguished

Dissimilatory Reduction and Transformation of Ferrihydrite-Humic Acid Coprecipitates

Organic matter (OM) is present in most terrestrial environments and is often found coprecipitated with ferrihydrite (Fh). Sorption or coprecipitation of OM with Fe oxides has been proposed to be an important mechanism for long-term C preservation. However, little is known about the impact of coprecipitated OM on reductive dissolution and transformation of Fe(III) (oxyhydr)oxides. Thus, we study the effect of humic acid (HA) coprecipitation on Fh reduction and secondary mineral formation by the dissimilatory Fe(III)-reducing bacterium Shewanella putrefaciens strain CN32. Despite similar crystal structure for all coprecipitates investigated, resembling 2-line Fh, the presence of coprecipitated HA resulted in lower specific surface areas. In terms of reactivity, coprecipitated HA resulted in slower Fh bioreduction rates at low C/Fe ratios (i.e., C/Fe ≤ 0.8), while high C/Fe ratios (i.e., C/Fe ≥ 1.8) enhanced the extent of bioreduction compared to pure Fh. The coprecipitated HA also altered the secondary Fe mineralization pathway by inhibiting goethite formation, reducing the amount of magnetite formation, and increasing the formation of a green rust-like phase. This study indicates that coprecipitated OM may influence the rates, pathway, and mineralogy of biogeochemical Fe cycling and anaerobic Fe respiration within soils.

Magnetite impregnation effects on the sorbent properties of activated carbons and biochars

This paper discusses the sorbent properties of magnetic activated carbons and biochars produced by wet impregnation with iron oxides. The sorbents had magnetic susceptibilities consistent with theoretical predictions for carbon-magnetite composites. The high BET surface areas of the activated carbons were preserved in the synthesis, and enhanced for one low surface area biochar by dissolving carbonates. Magnetization decreased the point of zero charge. Organic compound sorption correlated strongly with BET surface areas for the pristine and magnetized materials, while metal cation sorption did not show such a correlation. Strong sorption of the hydrophobic organic contaminant phenanthrene to the activated carbon or biochar surfaces was maintained following magnetite impregnation, while phenol sorption was diminished, probably due to enhanced carbon oxidation. Copper, zinc and lead sorption to the activated carbons and biochars was unchanged or slightly enhanced by the magnetization, and iron oxides also contributed to the composite metal sorption capacity. While a magnetic biochar with 219 ± 3.7 m(2)/g surface area nearly reached the very strong organic pollutant binding capacity of the two magnetic activated carbons, a magnetic biochar with 68 ± 2.8 m(2)/g surface area was the best metal sorbent. Magnetic biochars thus hold promise as more sustainable alternatives to coal-derived magnetic activated carbons.

Linking environmental processes to the in situ functioning of microorganisms by high‐resolution secondary ion mass spectrometry (NanoSIMS) and scanning transmission X‐ray microscopy (STXM)

Environmental microbiology research increasingly focuses on the single microbial cell as the defining entity that drives environmental processes. The interactions of individual microbial cells with each other, the environment and with higher organisms shape microbial communities and control the functioning of whole ecosystems. A single-cell view of microorganisms in their natural environment requires analytical tools that measure both cell function and chemical speciation at the submicrometre scale. Here we review the technical capabilities and limitations of high-resolution secondary ion mass spectrometry (NanoSIMS) and scanning transmission (soft) X-ray microscopy (STXM) and give examples of their applications. Whereas NanoSIMS can be combined with isotope-labelling, thereby localizing the distribution of cellular activities (e.g. carbon/nitrogen fixation/turnover), STXM provides information on the location and chemical speciation of metabolites and products of redox reactions. We propose the combined use of both techniques and discuss the technical challenges of their joint application. Both techniques have the potential to enhance our understanding of cellular mechanisms and activities that contribute to microbially mediated processes, such as the biogeochemical cycling of elements, the transformation of contaminants and the precipitation of mineral phases.

Soft X-ray spectromicroscopy of nickel sorption in a natural river biofilm

Scanning transmission X-ray microscopy (STXM) at the C 1s, O 1s, Ni 2p, Ca 2p, Mn 2p, Fe 2p, Mg 1s, Al 1s and Si 1s edges was used to study Ni sorption in a complex natural river biofilm. The 10-week grown river biofilm was exposed to 10 mg L(-1) Ni(2+) (as NiCl(2)) for 24 h. The region of the biofilm examined was dominated by filamentous structures, which were interpreted as the discarded sheaths of filamentous bacteria, as well as a sparse distribution of rod-shaped bacteria. The region also contained discrete particles with spectra similar to those of muscovite, SiO(2) and CaCO(3). The Ni(II) ions were selectively adsorbed by the sheaths of the filamentous bacteria. The sheaths were observed to be metal rich with significant amounts of Ca, Fe and Mn, along with the Ni. In addition, the sheaths had a large silicate content but little organic material. The metal content of the rod-shaped bacterial cells was much lower. The Fe on the sheath was mainly in the Fe(III) oxidation state. Mn was found in II, III and IV oxidation states. The Ni was likely sorbed to Mn-Fe minerals on the sheath. These STXM results have probed nano-scale biogeochemistry associated with bacterial species in a complex, natural biofilm community. They have implications for selective Ni contamination of the food chain and for developing bioremediation strategies.

Helium Ion Microscopy (HIM) for the imaging of biological samples at sub-nanometer resolution

Scanning Electron Microscopy (SEM) has long been the standard in imaging the sub-micrometer surface ultrastructure of both hard and soft materials. In the case of biological samples, it has provided great insights into their physical architecture. However, three of the fundamental challenges in the SEM imaging of soft materials are that of limited imaging resolution at high magnification, charging caused by the insulating properties of most biological samples and the loss of subtle surface features by heavy metal coating. These challenges have recently been overcome with the development of the Helium Ion Microscope (HIM), which boasts advances in charge reduction, minimized sample damage, high surface contrast without the need for metal coating, increased depth of field, and 5 angstrom imaging resolution. We demonstrate the advantages of HIM for imaging biological surfaces as well as compare and contrast the effects of sample preparation techniques and their consequences on sub-nanometer ultrastructure.