A US perspective on closing the carbon cycle to defossilize difficult-to-electrify segments of our economy.

Electrification to reduce or eliminate greenhouse gas emissions is essential to mitigate climate change. However, a substantial portion of our manufacturing and transportation infrastructure will be difficult to electrify and/or will continue to use carbon as a key component, including areas in aviation, heavy-duty and marine transportation, and the chemical industry. In this Roadmap, we explore how multidisciplinary approaches will enable us to close the carbon cycle and create a circular economy by defossilizing these difficult-to-electrify areas and those that will continue to need carbon. We discuss two approaches for this: developing carbon alternatives and improving our ability to reuse carbon, enabled by separations. Furthermore, we posit that co-design and use-driven fundamental science are essential to reach aggressive greenhouse gas reduction targets.

Capturing Rare-Earth Elements by Synthetic Aluminosilicate MCM-22: Mechanistic Understanding of Yb(III) Capture.

We studied the mechanism underlying the solid-phase adsorption of a heavy rare-earth element (HREE, Yb) from acidic solutions employing MCM-22 zeolite, serving as both a layered synthetic clay mimic and a new platform for the mechanistic study of HREE adsorption on aluminosilicate materials. Mechanistic studies revealed that the adsorption of Yb(III) at the surface adsorption site occurs primarily through the electrostatic interaction between the site and Yb(III) species. The dependence of Yb adsorption on the pH of the solution indicated the role of surface charge, and the content of framework Al suggested that the Brønsted acid sites (BAS) are involved in the adsorption of Yb(III) ions, which was further scrutinized by spectroscopic analysis and theoretical calculations. Our findings have illuminated the roles of surface sites in the solid-phase adsorption of HREEs from acidic solutions.

Interlayer spacing in pillared and grafted MCM-22 type silicas: density functional theory analysis versus experiment.

Pillaring of synthetic layered crystalline silicates and aluminosilicates provides a strategy to enhance their adsorption and separation performance, and can facilitate the understanding of such behavior in more complex natural clays. We perform the first-principles density functional theory calculations for the pillaring of the pure silica polymorph of an MCM-22 type molecular sieve. Starting with a precursor material MCM-22P with fully hydroxylated layers, a pillaring agent, (EtO)3SiR, can react with hydroxyl groups (-OH) on adjacent internal surfaces, 2(-OH) + (EtO)3SiR + H2O → (-O)2SiOHR + 3EtOH, to form a pillar bridging these surfaces, or with a single hydroxyl, -OH + (EtO)3SiR + 2H2O → (-O)Si(OH)2R + 3EtOH, grafting to one surface. For computational efficiency, we replace the experimental organic ligand, R, by a methyl group. We find that the interlayer spacing in MCM-22 is reduced by 2.66 Å relative to weakly bound layers in the precursor MCM-22P. Including (-O)2SiR bridges for 50% (100%) of the hydroxyl sites in MCM-22P increases the interlayer spacing relative to MCM-22 by 2.52 Å (2.46 Å). For comparison, we also analyze the system where all -OH groups in MCM-22P are replaced by non-bridging grafted (-O)Si(OH)2R which results in a smaller interlayer spacing expansion of 2.17 Å relative to MCM-22. Our results for the interlayer spacing in the pillared materials are compatible with experimental observations for a similar MCM-22 type material with low Al content (Si : Al = 51 : 1) of an expansion relative to MCM-22 of roughly 2.8 Å and 2.5 Å from our x-ray diffraction and scanning transmission electron microscopy analyses, respectively. The latter analysis reveals significant variation in individual layer spacings.

Biogeochemical fingerprinting of magnetotactic bacterial magnetite.

Biominerals are important archives of the presence of life and environmental processes in the geological record. However, ascribing a clear biogenic nature to minerals with nanometer-sized dimensions has proven challenging. Identifying hallmark features of biologically controlled mineralization is particularly important for the case of magnetite crystals, resembling those produced by magnetotactic bacteria (MTB), which have been used as evidence of early prokaryotic life on Earth and in meteorites. We show here that magnetite produced by MTB displays a clear coupled C-N signal that is absent in abiogenic and/or biomimetic (protein-mediated) nanometer-sized magnetite. We attribute the presence of this signal to intracrystalline organic components associated with proteins involved in magnetosome formation by MTB. These results demonstrate that we can assign a biogenic origin to nanometer-sized magnetite crystals, and potentially other biominerals of similar dimensions, using unique geochemical signatures directly measured at the nanoscale. This finding is significant for searching for the earliest presence of life in the Earth's geological record and prokaryotic life on other planets.

Atom Probe Tomography Analysis of Mica.

Laser-assisted atom probe tomography (APT) is a relatively new, powerful technique for sub-nanometric mineral and biomineral analysis. However, the laser-assisted APT analysis of highly anisotropic and chemically diverse minerals, such as phyllosilicates, may prove especially challenging due to the complex interaction between the crystal structure and the laser pulse upon applying a high electric field. Micas are a representative group of nonswelling clay minerals of relevance to a number of scientific and technological fields. In this study, a Mg-rich biotite was analyzed by APT to generate preliminary data on nonisotropic minerals and to investigate the effect of the crystallographic orientation on mica chemical composition and structure estimation. The difference in results obtained for specimens extracted from the (001) and (hk0) mica surfaces indicate the importance of both experimental parameters and the crystallography. Anisotropy of mica has a strong influence on the physicochemical properties of the mineral during field evaporation and the interpretation of APT data. The promising results obtained in the present study open the way to future innovative APT applications on mica and clay minerals and contribute to the general discussion on the challenges for the analysis of geomaterials by atom probe tomography.

Metabolic engineering of an acid-tolerant yeast strain Pichia kudriavzevii for itaconic acid production.

Itaconic acid (IA), or 2-methylenesuccinic acid, has a broad spectrum of applications in the biopolymer industry owing to the presence of one vinyl bond and two acid groups in the structure. Its polymerization can follow a similar mechanism as acrylic acid but additional functionality can be incorporated into the extra beta acid group. Currently, the bio-based production of IA in industry relies on the fermentation of the filamentous fungus Aspergillus terreus. However, the difficulties associated with the fermentation undertaken by filamentous fungi together with the pathogenic potential of A. terreus pose a serious challenge for industrial-scale production. In recent years, there has been increasing interest in developing alternative production hosts for fermentation processes that are more homogenous in the production of organic acids. Pichia kudriavzevii is a non-conventional yeast with high acid tolerance to organic acids at low pH, which is a highly desirable trait by easing downstream processing. We introduced cis-aconitic acid decarboxylase gene (cad) from A. terreus (designated At_cad) into this yeast and established the initial titer of IA at 135 â€‹± â€‹5 â€‹mg/L. Subsequent overexpression of a native mitochondrial tricarboxylate transporter (herein designated Pk_mttA) presumably delivered cis-aconitate efficiently to the cytosol and doubled the IA production. By introducing the newly invented CRISPR-Cas9 system into P. kudriavzevii, we successfully knocked out both copies of the gene encoding isocitrate dehydrogenase (ICD), aiming to increase the availability of cis-aconitate. The resulting P. kudriavzevii strain, devoid of ICD and overexpressing Pk_mttA and At_cad on its genome produced IA at 505 â€‹± â€‹17.7 â€‹mg/L in shake flasks, and 1232 â€‹± â€‹64 â€‹mg/L in fed-batch fermentation. Because the usage of an acid-tolerant species does not require pH adjustment during fermentation, this work demonstrates the great potential of engineering P. kudriavzevii as an industrial chassis for the production of organic acid.

Direct Observation of Early Stages of Growth of Multilayered DNA-Templated Au-Pd-Au Core-Shell Nanoparticles in Liquid Phase.

We report here on direct observation of early stages of formation of multilayered bimetallic Au-Pd core-shell nanocubes and Au-Pd-Au core-shell nanostars in liquid phase using low-dose in situ scanning transmission electron microscopy (S/TEM) with the continuous flow fluid cell. The reduction of Pd and formation of Au-Pd core-shell is achieved through the flow of the reducing agent. Initial rapid growth of Pd on Au along <111> direction is followed by a slower rearrangement of Pd shell. We propose the mechanism for the DNA-directed shape transformation of Au-Pd core-shell nanocubes to adopt a nanostar-like morphology in the presence of T30 DNA and discuss the observed nanoparticle motion in the confined volume of the fluid cell. The growth of Au shell over Au-Pd nanocube is initiated at the vertices of the nanocubes, leading to the preferential growth of the {111} facets and resulting in formation of nanostar-like particles. While the core-shell nanostructures formed in a fluid cell in situ under the low-dose imaging conditions closely resemble those obtained in solution syntheses, the reaction kinetics in the fluid cell is affected by the radiolysis of liquid reagents induced by the electron beam, altering the rate-determining reaction steps. We discuss details of the growth processes and propose the reaction mechanism in liquid phase in situ.

New approach to electron microscopy imaging of gel nanocomposites in situ.

Characterization of Au-nanocomposites is routinely done with scattering techniques where the structure and ordering of nanoparticles can be analyzed. Imaging of Poloxamer gel-based Au-nanocomposites is usually limited to cryo-TEM imaging of cryo-microtomed thin sections of the specimen. While this approach is applicable for imaging of the individual nanoparticles and gauging their size distribution, it requires altering the state of the specimen and is prone to artifacts associated with preparation protocols. Use of Scanning Transmission Electron Microscopy (S/TEM) with fluid cell in situ provides an opportunity to analysis of these complex materials in their hydrated state with nanometer resolution, yet dispensing dense gel-based samples onto electron-transparent substrates remains challenging. We show that Poloxamer gel-based Au nanocomposites exhibiting thermoreversible behavior can be imaged in a fully hydrated state using a commercially available fluid cell holder, and we describe a specimen preparation method for depositing femtoliter amounts of gel-based nanocomposites directly onto the 50 nm-thick SiN window membranes. Ultimately, fluid cell S/TEM in situ imaging approach offers a pathway to visualization of individual nanoparticles within a thick gel media while maintaining the hydrated state of the carrier polymeric matrix.

Salt Mediated Self-Assembly of Poly(ethylene glycol)-Functionalized Gold Nanorods.

Although challenging, assembling and orienting non-spherical nanomaterials into two- and three-dimensional (2D and 3D) ordered arrays can facilitate versatile collective properties by virtue of their shape-dependent properties that cannot be realized with their spherical counterparts. Here, we report on the self-assembly of gold nanorods (AuNRs) into 2D films at the vapor/liquid interface facilitated by grafting them with poly(ethylene glycol) (PEG). Using surface sensitive synchrotron grazing incidence small angle X-ray scattering (GISAXS) and specular X-ray reflectivity (XRR), we show that PEG-AuNRs in aqueous suspensions migrate to the vapor/liquid interface in the presence of salt, forming a uniform monolayer with planar-to-surface orientation. Furthermore, the 2D assembled PEG functionalized AuNRs exhibit short range order into rectangular symmetry with side-by-side and tail-to-tail nearest-neighbor packing. The effect of PEG chain length and salt concentration on the 2D assembly are also reported.

Influence of physical activity on tibial bone material properties in laying hens.

Laying hens develop a type of osteoporosis that arises from a loss of structural bone, resulting in high incidence of fractures. In this study, a comparison of bone material properties was made for lines of hens created by divergent selection to have high and low bone strength and housed in either individual cages, with restricted mobility, or in an aviary system, with opportunity for increased mobility. Improvement of bone biomechanics in the high line hens and in aviary housing was mainly due to increased bone mass, thicker cortical bone and more medullary bone. However, bone material properties such as cortical and medullary bone mineral composition and crystallinity as well as collagen maturity did not differ between lines. However, bone material properties of birds from the different type of housing were markedly different. The cortical bone in aviary birds had a lower degree of mineralization and bone mineral was less mature and less organized than in caged birds. These differences can be explained by increased bone turnover rates due to the higher physical activity of aviary birds that stimulates bone formation and bone remodeling. Multivariate statistical analyses shows that both cortical and medullary bone contribute to breaking strengthThe cortical thickness was the single most important contributor while its degree of mineralization and porosity had a smaller contribution. Bone properties had poorer correlations with mechanical properties in cage birds than in aviary birds presumably due to the greater number of structural defects of cortical bone in cage birds.

Off-axis electron holography of bacterial cells and magnetic nanoparticles in liquid.

The mapping of electrostatic potentials and magnetic fields in liquids using electron holography has been considered to be unrealistic. Here, we show that hydrated cells of Magnetospirillum magneticum strain AMB-1 and assemblies of magnetic nanoparticles can be studied using off-axis electron holography in a fluid cell specimen holder within the transmission electron microscope. Considering that the holographic object and reference wave both pass through liquid, the recorded electron holograms show sufficient interference fringe contrast to permit reconstruction of the phase shift of the electron wave and mapping of the magnetic induction from bacterial magnetite nanocrystals. We assess the challenges of performing in situ magnetization reversal experiments using a fluid cell specimen holder, discuss approaches for improving spatial resolution and specimen stability, and outline future perspectives for studying scientific phenomena, ranging from interparticle interactions in liquids and electrical double layers at solid-liquid interfaces to biomineralization and the mapping of electrostatic potentials associated with protein aggregation and folding.

Magnetic microbes: Bacterial magnetite biomineralization.

Magnetotactic bacteria are a diverse group of prokaryotes with the ability to orient and migrate along the magnetic field lines in search for a preferred oxygen concentration in chemically stratified water columns and sediments. These microorganisms produce magnetosomes, the intracellular nanometer-sized magnetic crystals surrounded by a phospholipid bilayer membrane, typically organized in chains. Magnetosomes have nearly perfect crystal structures with narrow size distribution and species-specific morphologies, leading to well-defined magnetic properties. As a result, the magnetite biomineralization in these organisms is of fundamental interest to diverse disciplines, from biotechnology to astrobiology. This article highlights recent advances in the understanding of the bacterial magnetite biomineralization.

Direct Visualization of the Hydration Layer on Alumina Nanoparticles with the Fluid Cell STEM in situ.

Rheological behavior of aqueous suspensions containing nanometer-sized powders is of relevance to many branches of industry. Unusually high viscosities observed for suspensions of nanoparticles compared to those of micron size powders cannot be explained by current viscosity models. Formation of so-called hydration layer on alumina nanoparticles in water was hypothesized, but never observed experimentally. We report here on the direct visualization of aqueous suspensions of alumina with the fluid cell in situ. We observe the hydration layer formed over the particle aggregates and show that such hydrated aggregates constitute new particle assemblies and affect the flow behavior of the suspensions. We discuss how these hydrated nanoclusters alter the effective solid content and the viscosity of nanostructured suspensions. Our findings elucidate the source of high viscosity observed for nanoparticle suspensions and are of direct relevance to many industrial sectors including materials, food, cosmetics, pharmaceutical among others employing colloidal slurries with nanometer-scale particles.

Correlative electron and fluorescence microscopy of magnetotactic bacteria in liquid: toward in vivo imaging.

Magnetotactic bacteria biomineralize ordered chains of uniform, membrane-bound magnetite or greigite nanocrystals that exhibit nearly perfect crystal structures and species-specific morphologies. Transmission electron microscopy (TEM) is a critical technique for providing information regarding the organization of cellular and magnetite structures in these microorganisms. However, conventional TEM can only be used to image air-dried or vitrified bacteria removed from their natural environment. Here we present a correlative scanning TEM (STEM) and fluorescence microscopy technique for imaging viable cells of Magnetospirillum magneticum strain AMB-1 in liquid using an in situ fluid cell TEM holder. Fluorescently labeled cells were immobilized on microchip window surfaces and visualized in a fluid cell with STEM, followed by correlative fluorescence imaging to verify their membrane integrity. Notably, the post-STEM fluorescence imaging indicated that the bacterial cell wall membrane did not sustain radiation damage during STEM imaging at low electron dose conditions. We investigated the effects of radiation damage and sample preparation on the bacteria viability and found that approximately 50% of the bacterial membranes remained intact after an hour in the fluid cell, decreasing to ~30% after two hours. These results represent a first step toward in vivo studies of magnetite biomineralization in magnetotactic bacteria.

Nucleation of iron oxide nanoparticles mediated by Mms6 protein in situ.

Biomineralization proteins are widely used as templating agents in biomimetic synthesis of a variety of organic-inorganic nanostructures. However, the role of the protein in controlling the nucleation and growth of biomimetic particles is not well understood, because the mechanism of the bioinspired reaction is often deduced from ex situ analysis of the resultant nanoscale mineral phase. Here we report the direct visualization of biomimetic iron oxide nanoparticle nucleation mediated by an acidic bacterial recombinant protein, Mms6, during an in situ reaction induced by the controlled addition of sodium hydroxide to solution-phase Mms6 protein micelles incubated with ferric chloride. Using in situ liquid cell scanning transmission electron microscopy we observe the liquid iron prenucleation phase and nascent amorphous nanoparticles forming preferentially on the surface of protein micelles. Our results provide insight into the early steps of protein-mediated biomimetic nucleation of iron oxide and point to the importance of an extended protein surface during nanoparticle formation.

Stomatobaculum longum gen. nov., sp. nov., an obligately anaerobic bacterium from the human oral cavity.

A strictly anaerobic Gram-stain-variable but positive by structure, non-spore-forming bacterium designated Lachnospiraceae bacterium ACC2 strain DSM 24645(T) was isolated from human subgingival dental plaque. Bacterial cells were 4-40 µm long non-motile rods, often swollen and forming curved filaments up to 200 µm. Cells contained intracellular, poorly crystalline, nanometre-sized iron- and sulfur-rich particles. The micro-organism was able to grow on yeast extract, trypticase peptone, milk, some sugars and organic acids. The major metabolic end-products of glucose fermentation were butyrate, lactate, isovalerate and acetate. The growth temperature and pH ranges were 30-42 °C and 4.9-7.5, respectively. Major fatty acids were C14 : 0, C14 : 0 DMA (dimethyl aldehyde), C16 : 0, C16 : 1ω7c DMA. The whole-cell hydrolysate contained meso-diaminopimelic acid, indicating peptidoglycan type A1γ. The DNA G+C content was calculated to be 55.05 mol% from the whole-genome sequence and 55.3 mol% as determined by HPLC. There were no predicted genes responsible for biosynthesis of respiratory lipoquinones, mycolic acids and lipopolysaccharides. Genes associated with synthesis of teichoic and lipoteichoic acids, diaminopimelic acid, polar lipids and polyamines were present. According to the 16S rRNA gene sequence phylogeny, strain DSM 24645(T) formed, together with several uncultured oral clones, a separate branch within the family Lachnospiraceae, with the highest sequence similarity to the type strain of Moryella indoligenes at 94.2 %. Based on distinct phenotypic and genotypic characteristics, we suggest that strain DSM 24645(T) represents a novel species in a new genus, for which the name Stomatobaculum longum gen. nov., sp. nov. is proposed. The type strain of Stomatobaculum longum is DSM 24645(T) ( = HM-480(T); deposited in BEI Resources, an NIH collection managed by the ATCC).

Self-assembly and biphasic iron-binding characteristics of Mms6, a bacterial protein that promotes the formation of superparamagnetic magnetite nanoparticles of uniform size and shape.

Highly ordered mineralized structures created by living organisms are often hierarchical in structure with fundamental structural elements at nanometer scales. Proteins have been found responsible for forming many of these structures, but the mechanisms by which these biomineralization proteins function are generally poorly understood. To better understand its role in biomineralization, the magnetotactic bacterial protein, Mms6, which promotes the formation in vitro of superparamagnetic magnetite nanoparticles of uniform size and shape, was studied for its structure and function. Mms6 is shown to have two phases of iron binding: one high affinity and stoichiometric and the other low affinity, high capacity, and cooperative with respect to iron. The protein is amphipathic with a hydrophobic N-terminal domain and hydrophilic C-terminal domain. It self-assembles to form a micelle, with most particles consisting of 20-40 monomers, with the hydrophilic C-termini exposed on the outside. Studies of proteins with mutated C-terminal domains show that the C-terminal domain contributes to the stability of this multisubunit particle and binds iron by a mechanism that is sensitive to the arrangement of carboxyl/hydroxyl groups in this domain.

A cultured greigite-producing magnetotactic bacterium in a novel group of sulfate-reducing bacteria.

Magnetotactic bacteria contain magnetosomes--intracellular, membrane-bounded, magnetic nanocrystals of magnetite (Fe(3)O(4)) or greigite (Fe(3)S(4))--that cause the bacteria to swim along geomagnetic field lines. We isolated a greigite-producing magnetotactic bacterium from a brackish spring in Death Valley National Park, California, USA, strain BW-1, that is able to biomineralize greigite and magnetite depending on culture conditions. A phylogenetic comparison of BW-1 and similar uncultured greigite- and/or magnetite-producing magnetotactic bacteria from freshwater to hypersaline habitats shows that these organisms represent a previously unknown group of sulfate-reducing bacteria in the Deltaproteobacteria. Genomic analysis of BW-1 reveals the presence of two different magnetosome gene clusters, suggesting that one may be responsible for greigite biomineralization and the other for magnetite.

Isolation of obligately alkaliphilic magnetotactic bacteria from extremely alkaline environments.

Large numbers of magnetotactic bacteria were discovered in mud and water samples collected from a number of highly alkaline aquatic environments with pH values of ≈ 9.5. These bacteria were helical in morphology and biomineralized chains of bullet-shaped crystals of magnetite and were present in all the highly alkaline sites sampled. Three strains from different sites were isolated and cultured and grew optimally at pH 9.0-9.5 but not at 8.0 and below, demonstrating that these organisms truly require highly alkaline conditions and are not simply surviving/growing in neutral pH micro-niches in their natural habitats. All strains grew anaerobically through the reduction of sulfate as a terminal electron acceptor and phylogenetic analysis, based on 16S rRNA gene sequences, as well as some physiological features, showed that they could represent strains of Desulfonatronum thiodismutans, a known alkaliphilic bacterium that does not biomineralize magnetosomes. Our results show that some magnetotactic bacteria can be considered extremophilic and greatly extend the known ecology of magnetotactic bacteria and the conditions under which they can biomineralize magnetite. Moreover, our results show that this type of magnetotactic bacterium is common in highly alkaline environments. Our findings also greatly influence the interpretation of the presence of nanometer-sized magnetite crystals, so-called magnetofossils, in highly alkaline environments.