Jennyfer Miot

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.

Formation of cell-iron-mineral aggregates by phototrophic and nitrate-reducing anaerobic Fe(II)-oxidizing bacteria.

Microbial anaerobic Fe(II) oxidation at neutral pH produces poorly soluble Fe(III) which is expected to bind to cell surfaces causing cell encrustation and potentially impeding cell metabolism. The challenge for Fe(II)-oxidizing prokaryotes therefore is to avoid encrustation with Fe(III). Using different microscopic techniques we tracked Fe(III) minerals at the cell surface and within cells of phylogenetically distinct phototrophic and nitrate-reducing Fe(II)-oxidizing bacteria. While some strains successfully prevented encrustation others precipitated Fe(III) minerals at the cell surface and in the periplasm. Our results indicate differences in the cellular mechanisms of Fe(II) oxidation, transport of Fe(II)/Fe(III) ions, and Fe(III) mineral precipitation.

Transformation of vivianite by anaerobic nitrate‐reducing iron‐oxidizing bacteria

In phosphate-rich environments, vivianite (Fe(II)(3)(PO(4))(2), 8H(2)O) is an important sink for dissolved Fe(II) and is considered as a very stable mineral due to its low solubility at neutral pH. In the present study, we report the mineralogical transformation of vivianite in cultures of the nitrate-reducing iron-oxidizing bacterial strain BoFeN1 in the presence of dissolved Fe(II). Vivianite was first transformed into a greenish phase consisting mostly of an amorphous mixed valence Fe-phosphate. This precipitate became progressively orange and the final product of iron oxidation consisted of an amorphous Fe(III)-phosphate. The sub-micrometer analysis by scanning transmission X-ray microscopy of the iron redox state in samples collected at different stages of the culture indicated that iron was progressively oxidized at the contact of the bacteria and at a distance from the cells in extracellular minerals. Iron oxidation in the extracellular minerals was delayed by a few days compared with cell-associated Fe-minerals. This led to strong differences of Fe redox in between these two types of minerals and finally to local heterogeneities of redox within the sample. In the absence of dissolved Fe(II), vivianite was not significantly transformed by BoFeN1. Whereas Fe(II) oxidation at the cell contact is most probably directly catalyzed by the bacteria, vivianite transformation at a distance from the cells might result from oxidation by nitrite. In addition, processes leading to the export of Fe(III) from bacterial oxidation sites to extracellular minerals are discussed including some involving colloids observed by cryo-transmission electron microscopy in the culture medium.

Preservation of protein globules and peptidoglycan in the mineralized cell wall of nitrate-reducing, iron(II)-oxidizing bacteria: a cryo-electron microscopy study.

Iron-oxidizing bacteria are important actors of the geochemical cycle of iron in modern environments and may have played a key role all over Earths history. However, in order to better assess that role on the modern and the past Earth, there is a need for better understanding the mechanisms of bacterial iron oxidation and for defining potential biosignatures to be looked for in the geologic record. In this study, we investigated experimentally and at the nanometre scale the mineralization of iron-oxidizing bacteria with a combination of synchrotron-based scanning transmission X-ray microscopy (STXM), scanning transmission electron microscopy (STEM) and cryo-transmission electron microscopy (cryo-TEM). We show that the use of cryo-TEM instead of conventional microscopy provides detailed information of the successive iron biomineralization stages in anaerobic nitrate-reducing iron-oxidizing bacteria. These results suggest the existence of preferential Fe-binding and Fe-oxidizing sites on the outer face of the plasma membrane leading to the nucleation and growth of Fe minerals within the periplasm of these cells that eventually become completely encrusted. In contrast, the septa of dividing cells remain nonmineralized. In addition, the use of cryo-TEM offers a detailed view of the exceptional preservation of protein globules and the peptidoglycan within the Fe-mineralized cell walls of these bacteria. These organic molecules and ultrastructural details might be protected from further degradation by entrapment in the mineral matrix down to the nanometre scale. This is discussed in the light of previous studies on the properties of Fe-organic interactions and more generally on the fossilization of mineral-organic assemblies.

Biomineralized α-Fe2O3: texture and electrochemical reaction with Li

Sustainable batteries call for the development of new eco-efficient processes for preparation of electrode materials based on low cost and abundant chemical elements. Here we report a method based on bacterial iron biomineralization for the synthesis of α-Fe2O3 and its subsequent use as a conversion-based electrode material in Li batteries. This high-yield synthesis approach enlists (1) the room temperature formation of γ-FeOOH via the use of an anaerobic Fe(II)-oxidizing bacterium Acidovorax sp. strain BoFeN1 and (2) the transformation of these BoFeN1/γ-FeOOH assemblies into an alveolar bacteria-free α-Fe2O3 material by a short heat treatment under air. As the γ-FeOOH precursor particles are precipitated between the two membranes of the bacterial cell wall (40 nm-thick space), the final material consists of highly monodisperse nanometric ([similar]40 × 15 nm) and oriented hematite crystals, assembled to form a hollow shell having the same size and shape as the initial bacteria (bacteriomorph). This double level of control (nanometric particle size and particle organization at the micrometric scale) provided powders exhibiting (1) enhanced electrochemical reversibility when fully reacted with Li and (2) an impressive high rate capability when compared to non-textured primary α-Fe2O3 particles of similar size. This bacterially induced eco-efficient and scalable synthesis method opens wide new avenues to be explored at the crossroads of biomineralization and electrochemistry for energy storage.

Fe biomineralization mirrors individual metabolic activity in a nitrate-dependent Fe(II)-oxidizer

Microbial biomineralization sometimes leads to periplasmic encrustation, which is predicted to enhance microorganism preservation in the fossil record. Mineral precipitation within the periplasm is, however, thought to induce death, as a result of permeability loss preventing nutrient and waste transit across the cell wall. This hypothesis had, however, never been investigated down to the single cell level. Here, we cultured the nitrate reducing Fe(II) oxidizing bacteria Acidovorax sp. strain BoFeN1 that have been previously shown to promote the precipitation of a diversity of Fe minerals (lepidocrocite, goethite, Fe phosphate) encrusting the periplasm. We investigated the connection of Fe biomineralization with carbon assimilation at the single cell level, using a combination of electron microscopy and Nano-Secondary Ion Mass Spectrometry. Our analyses revealed strong individual heterogeneities of Fe biomineralization. Noteworthy, a small proportion of cells remaining free of any precipitate persisted even at advanced stages of biomineralization. Using pulse chase experiments with 13C-acetate, we provide evidence of individual phenotypic heterogeneities of carbon assimilation, correlated with the level of Fe biomineralization. Whereas non- and moderately encrusted cells were able to assimilate acetate, higher levels of periplasmic encrustation prevented any carbon incorporation. Carbon assimilation only depended on the level of Fe encrustation and not on the nature of Fe minerals precipitated in the cell wall. Carbon assimilation decreased exponentially with increasing cell-associated Fe content. Persistence of a small proportion of non-mineralized and metabolically active cells might constitute a survival strategy in highly ferruginous environments. Eventually, our results suggest that periplasmic Fe biomineralization may provide a signature of individual metabolic status, which could be looked for in the fossil record and in modern environmental samples.

Preservation of Archaeal Surface Layer Structure During Mineralization

Proteinaceous surface layers (S-layers) are highly ordered, crystalline structures commonly found in prokaryotic cell envelopes that augment their structural stability and modify interactions with metals in the environment. While mineral formation associated with S-layers has previously been noted, the mechanisms were unconstrained. Using Sulfolobus acidocaldarius a hyperthermophilic archaeon native to metal-enriched environments and possessing a cell envelope composed only of a S-layer and a lipid cell membrane, we describe a passive process of iron phosphate nucleation and growth within the S-layer of cells and cell-free S-layer “ghosts” during incubation in a Fe-rich medium, independently of metabolic activity. This process followed five steps: (1) initial formation of mineral patches associated with S-layer; (2) patch expansion; (3) patch connection; (4) formation of a continuous mineral encrusted layer at the cell surface; (5) early stages of S-layer fossilization via growth of the extracellular mineralized layer and the mineralization of cytosolic face of the cell membrane. At more advanced stages of encrustation, encrusted outer membrane vesicles are formed, likely in an attempt to remove damaged S-layer proteins. The S-layer structure remains strikingly well preserved even upon the final step of encrustation, offering potential biosignatures to be looked for in the fossil record.

Synchrotron X-ray studies of heavy metal mineral-microbe interactions

The availability of analytical methods that utilize the very intense and bright X-rays from synchrotron radiation sources has fundamentally changed the way in which geoscientists, environmental scientists and soil scientists study complex environmental samples and decipher the chemical and biological processes that impact the speciation, transport and potential bioavailability of environmental toxins (Brown et al. , 2006). Such samples are often mixtures of crystalline and amorphous phases in particle-sizes ranging from cm to nm, adsorbed metal ions and organic molecules, natural organic matter, microbial organisms, algae, plant materials and aqueous solutions. The processes that affect the chemical forms and environmental fate of contaminants in such mixtures range from surface adsorption, desorption, precipitation and dissolution reactions, often involving a combination of hydrolysis, ligand exchange and electron transfer, to biological interactions in which microbial organisms, algae or plants interact with mineral surfaces and environmental contaminants. These processes can result in: (1) the formation of biominerals, which can effectively sequester contaminants; (2) oxidation-reduction reactions of redox-sensitive contaminants, which can transform such contaminants into more (or less) toxic forms; and (3) mineral dissolution reactions, which can release heavy metal and metalloid contaminants. Determining the effects of these processes on environmental contaminants and characterizing the types and speciation of contaminants present in complex environmental samples are challenging tasks that require a variety of analytical methods. Such methods must be element-selective, sensitive enough to detect elemental concentrations at the ppm level, have spatial resolutions comparable to the spatial scales of elemental and structural heterogenieties of the samples, and be capable of providing molecular-scale structural and compositional information so that contaminant speciation can be defined quantitatively. Because many of the chemical and biological processes of interest in this context occur at environmental interfaces (e.g. mineral/water or mineral/microbe interfaces), it is also essential that some of the …

Nitrate-Dependent Iron Oxidation: A Potential Mars Metabolism

This work considers the hypothetical viability of microbial nitrate-dependent Fe2+ oxidation (NDFO) for supporting simple life in the context of the early Mars environment. This draws on knowledge built up over several decades of remote and in situ observation, as well as recent discoveries that have shaped current understanding of early Mars. Our current understanding is that certain early martian environments fulfill several of the key requirements for microbes with NDFO metabolism. First, abundant Fe2+ has been identified on Mars and provides evidence of an accessible electron donor; evidence of anoxia suggests that abiotic Fe2+ oxidation by molecular oxygen would not have interfered and competed with microbial iron metabolism in these environments. Second, nitrate, which can be used by some iron oxidizing microorganisms as an electron acceptor, has also been confirmed in modern aeolian and ancient sediment deposits on Mars. In addition to redox substrates, reservoirs of both organic and inorganic carbon are available for biosynthesis, and geochemical evidence suggests that lacustrine systems during the hydrologically active Noachian period (4.1–3.7 Ga) match the circumneutral pH requirements of nitrate-dependent iron-oxidizing microorganisms. As well as potentially acting as a primary producer in early martian lakes and fluvial systems, the light-independent nature of NDFO suggests that such microbes could have persisted in sub-surface aquifers long after the desiccation of the surface, provided that adequate carbon and nitrates sources were prevalent. Traces of NDFO microorganisms may be preserved in the rock record by biomineralization and cellular encrustation in zones of high Fe2+ concentrations. These processes could produce morphological biosignatures, preserve distinctive Fe-isotope variation patterns, and enhance preservation of biological organic compounds. Such biosignatures could be detectable by future missions to Mars with appropriate instrumentation.

Experimental maturation of Archaea encrusted by Fe-phosphates

Burial is generally detrimental to the preservation of biological signals. It has often been assumed that (bio)mineral-encrusted microorganisms are more resistant to burial-induced degradation than non-encrusted ones over geological timescales. For the present study, we submitted Sulfolobus acidocaldarius experimentally encrusted by amorphous Fe phosphates to constrained temperature conditions (150 °C) under pressure for 1 to 5 days, thereby simulating burial-induced processes. We document the molecular and mineralogical evolution of these assemblages down to the sub-micrometer scale using X-ray diffraction, scanning and transmission electron microscopies and synchrotron-based X-ray absorption near edge structure spectroscopy at the carbon K-edge. The present results demonstrate that the presence of Fe-phosphates enhances the chemical degradation of microbial organic matter. While Fe-phosphates remained amorphous in abiotic controls, crystalline lipscombite (FeIIxFeIII3−x(PO4)2(OH)3−x) entrapping organic matter formed in the presence of S. acidocaldarius cells. Lipscombite textures (framboidal vs. bipyramidal) appeared only controlled by the initial level of encrustation of the cells, suggesting that the initial organic matter to mineral ratio influences the competition between nucleation and crystal growth. Altogether these results highlight the important interplay between minerals and organic matter during fossilization, which should be taken into account when interpreting the fossil record.