The earliest wood and its hydraulic properties documented in c. 407‐million‐year‐old fossils using synchrotron microtomography

We document xylem structure and hydraulic properties in the earliest woody plant A rmoricaphyton chateaupannense gen. nov. & sp. nov. based on c. 407‐million‐year‐old fossils from the Armorican Massif, western France. The plant was small, and the woody axes were narrow and permineralized in pyrite (FeS₂). We used standard palaeobotanical methods and employed propagation phase contrast X‐ray synchrotron microtomography (PPC‐SRμCT) to create three‐dimensional images of the wood and to evaluate its properties. The xylem comprised tracheids and rays, which developed from a cambium. Tracheids possessed an early extinct type of scalariform bordered pitting known as P‐type. Our observations indicate that wood evolved initially in plants of small stature that were members of Euphyllophytina, a clade that includes living seed plants, horsetails and ferns. Hydraulic properties were calculated from measurements taken from the PPC‐SRμCT images. The specific hydraulic conductivity of the xylem area was calculated as 8.7 kg m^?1 s^?1 and the mean cell thickness‐to‐span ratio (t/b)² of tracheids was 0.0372. The results show that the wood was suited to high conductive performance with low mechanical resistance to hydraulic tension. We argue that axis rigidity in the earliest woody plants initially evolved through the development of low‐density woods. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 175 , 423–437.     

Spatially Resolved Sulphur K-edge XANES Spectroscopy for in situ Characterization of the Fungus–plant Interaction Puccinia triticina and Wheat Leaves

An innovative application of X‐ray absorption near edge structure (XANES) spectroscopy for the characterization of interactions of biotrophic plant pathogens with their hosts as exemplified by Puccinia triticina colonizing wheat leaves is described. Spatially resolved, synchrotron radiation‐based XANES spectroscopy was used for the detection of changes in sulphur metabolism induced by leaf rust infections. A significant accumulation of sulphate occurred at the site of the sporulating urediniosori of P. triticina. Compared with non‐infected leaf areas, minor changes in the spectra were observed for the non‐visibly colonized tissue neighbouring the rust sori. As the spectra for isolated urediniospores and the healthy leaf areas did not match the spectra of the urediniosori, a significant impact of the biotrophic pathogen on sulphur metabolism of wheat has been demonstrated. Spatially resolved XANES spectroscopy will extend the range of qualitative and quantitative methods for in situ investigations of host–pathogen interactions, thus contributing to enlarge our knowledge about the metabolism of diseased plants.     

Showcasing the application of synchrotron‐based X‐ray computed tomography in host–pathogen interactions: The role of wheat rachilla and rachis nodes in Type‐II resistance to Fusarium graminearum

Fusarium head blight, caused primarily by Fusarium graminearum (Fg), is one of the most devastating diseases of wheat. Host resistance in wheat is classified into five types (Type‐I to Type‐V), and a majority of moderately resistant genotypes carry Type‐II resistance (resistance to pathogen spread in the rachis) alleles, mainly from the Chinese cultivar Sumai 3. Histopathological studies in the past failed to identify the key tissue in the spike conferring resistance to pathogen spread, and most of the studies used destructive techniques, potentially damaging the tissue(s) under study. In the present study, nondestructive synchrotron‐based phase contrast X‐ray imaging and computed tomography techniques were used to confirm the part of the wheat spike conferring Type‐II resistance to Fg spread, thus showcasing the application of synchrotron‐based techniques to image host–pathogen interactions. Seven wheat genotypes of moderate resistance to Fusarium head blight were studied for changes in the void space volume fraction and grayscale/voxel intensity following Fg inoculation. Cell‐wall biopolymeric compounds were quantified using Fourier‐transform midinfrared spectroscopy for all genotype‐treatment combinations. The study revealed that the rachilla and rachis nodes together are structurally important in conferring Type‐II resistance. The structural reinforcement was not necessarily observed from lignin deposition but rather from an unknown mechanism.     

Scanning transmission X-ray microscopy study of microbial calcification

Calcium phosphates and calcium carbonates are among the most prevalent minerals involved in microbial fossilization. Characterization of both the organic and mineral components in biomineralized samples is, however, usually difficult at the appropriate spatial resolution (i.e. at the submicrometer scale). Scanning transmission X‐ray microscopy (STXM) was used to measure C K‐edge, P L‐edge, and Ca L‐edge near‐edge X‐ray absorption fine structure (NEXAFS) spectra of some calcium‐containing minerals common in biomineralization processes and to study the experimental biomineralization by the model microorganism, Caulobacter crescentus. We show that the Ca L_2,3‐edges for hydroxyapatite, calcite, vaterite, and aragonite are unique and can be used as probes to detect these different mineral phases. Using these results, we showed that C. crescentus cells, when cultured in the presence of high calcium concentration, precipitated carbonate hydroxyapatite. In parallel, we detected proteins, polysaccharides, and nucleic acids in the mineralizing bacteria at the single‐cell scale. Finally, we discussed the utility of STXM for the study of natural fossilized microbial systems.     

Na‐Ion Intercalation and Charge Storage Mechanism in 2D Vanadium Carbide

2D vanadium carbide MXene containing surface functional groups (denoted as V₂CT_x , where T_x are surface functional groups) is synthesized and studied as anode material for Na‐ion batteries. V₂CT_x anode exhibits reversible charge storage with good cycling stability and high rate capability through electrochemical test. The charge storage mechanism of V₂CT_x material during Na⁺ intercalation/deintercalation and the redox reaction of vanadium are studied using a combination of synchrotron based X‐ray diffraction, hard X‐ray absorption near edge spectroscopy (XANES), and soft X‐ray absorption spectroscopy (sXAS). Experimental evidence of a major contribution of redox reaction of vanadium to the charge storage and the reversible capacity of V₂CT_x during sodiation/desodiation process are provided through V K ‐edge XANES and V L _2,3‐edge sXAS results. A correlation between the CO₃^2? content and the Na⁺ intercalation/deintercalation states in the V₂CT_x electrode observed from C and O K ‐edge in sXAS results implies that some additional charge storage reactions may take place between the Na⁺‐intercalated V₂CT_x and the carbonate‐based nonaqueous electrolyte. The results of this study provide valuable information for the further studies on V₂CT_x as anode material for Na‐ion batteries and capacitors.     

Three‐dimensional rendering of otolith growth using phase contrast synchrotron tomography

A three‐dimensional computer reconstruction of a plaice Pleuronectes platessa otolith is presented from data acquired by the Diamond Light synchrotron, beamline I12, X‐ray source, a high energy (53–150 keV) source particularly well suited to the study of dense objects. The data allowed non‐destructive rendering of otolith structure, and for the first time allows otolith annuli (internal ring structures) to be analysed in X‐ray tomographic images.     

Structural and functional characterization of solute binding proteins for aromatic compounds derived from lignin: p‐Coumaric acid and related aromatic acids

Lignin comprises 15–25% of plant biomass and represents a major environmental carbon source for utilization by soil microorganisms. Access to this energy resource requires the action of fungal and bacterial enzymes to break down the lignin polymer into a complex assortment of aromatic compounds that can be transported into the cells. To improve our understanding of the utilization of lignin by microorganisms, we characterized the molecular properties of solute binding proteins of ATP‐binding cassette transporter proteins that interact with these compounds. A combination of functional screens and structural studies characterized the binding specificity of the solute binding proteins for aromatic compounds derived from lignin such as p‐coumarate, 3‐phenylpropionic acid and compounds with more complex ring substitutions. A ligand screen based on thermal stabilization identified several binding protein clusters that exhibit preferences based on the size or number of aromatic ring substituents. Multiple X‐ray crystal structures of protein–ligand complexes for these clusters identified the molecular basis of the binding specificity for the lignin‐derived aromatic compounds. The screens and structural data provide new functional assignments for these solute‐binding proteins which can be used to infer their transport specificity. This knowledge of the functional roles and molecular binding specificity of these proteins will support the identification of the specific enzymes and regulatory proteins of peripheral pathways that funnel these compounds to central metabolic pathways and will improve the predictive power of sequence‐based functional annotation methods for this family of proteins.Proteins 2013; 81:1709–1726. © 2013 Wiley Periodicals, Inc.     

Contrasting calcium localization and speciation in leaves of the Medicago truncatula mutant cod5 analyzed via synchrotron X–ray techniques

Oxalate‐producing plants accumulate calcium oxalate crystals (CaOx_(c)) in the range of 3–80% w/w of their dry weight, reducing calcium (Ca) bioavailability. The calcium oxalate deficient 5 (cod5) mutant of Medicago truncatula has been previously shown to contain similar Ca concentrations to wild‐type (WT) plants, but lower oxalate and CaOx_(c) concentrations. We imaged the Ca distribution in WT and cod5 leaflets via synchrotron X–ray fluorescence mapping (SXRF). We observed a difference in the Ca distribution between cod5 and WT leaflets, manifested as an abundance of Ca in the interveinal areas and a lack of Ca along the secondary veins in cod5, i.e. the opposite of what is observed in WT. X–ray microdiffraction (μXRD) of M. truncatula leaves confirmed that crystalline CaOx_(c) (whewellite; CaC₂O₄·H₂O) was present in the WT only, in cells sheathing the secondary veins. Together with μXRD, microbeam Ca K–edge X–ray absorption near‐edge structure spectroscopy (μXANES) indicated that, among the forms of CaOx, i.e. crystalline or amorphous, only amorphous CaOx was present in cod5. These results demonstrate that deletion of COD5 changes both Ca localization and the form of CaOx within leaflets.     

The origin of novel features by changes in developmental mechanisms: ontogeny and three‐dimensional microanatomy of polyodontode scales of two early osteichthyans

Recent advances in synchrotron imaging allow us to study the three‐dimensional (3D) histology of vertebrate fossils, including microfossils (e.g. teeth and scales) of early jawed vertebrates. These microfossils can often be scanned at submicron resolution (<1 µm) because of their small size. The resulting voxel (3D pixel) stacks can be processed into virtual thin sections revealing almost every internal detail of the samples, comparable to traditional thin sections. In addition, 3D models of the internal microanatomical structures, such as embedded odontodes and vasculature, can be assembled and examined in situ. Scales of two early osteichthyans, Psarolepis romeri from the Early Devonian of China and Andreolepis hedei from the Late Silurian of Sweden, were scanned using propagation phase‐contrast synchrotron X‐ray microtomography (PPC‐SRµCT), and 3D models of internal canal systems and buried odontodes were created from the scans. Based on these new data, we review the evolutionary origin of cosmine and its associated pore‐canal system, which has been long recognized as a synapomorphy of sarcopterygians. The first odontode that appeared during growth shows almost identical morphology in the two scales, but the second odontode of the Psarolepis scale shows a distinctive morphology with several pores on the surface. It is suggested that a shift from ridge‐like odontode to pore‐bearing odontode was the key step in the origin of cosmine, which was then elaborated further in more‐derived sarcopterygians. We perform a detailed comparison between the two scales and propose a primary homology framework to generate microanatomical characters, which can be used in the phylogenetic analysis of early osteichthyans when more 3D data become available. Our results highlight the importance of 3D data for the study of histology and ontogeny of the dermal skeleton of early jawed vertebrates, especially scales of the polyodontode type. The traditional microvertebrate collection is not only useful for biostratigraphic studies, but also preserves invaluable biological information about the growth of vertebrate hard tissues. Today, we are only beginning to understand the biological meaning of the new 3D data. The increasing availability of such data will enable, and indeed require, a complete revision of traditional palaeohistological studies on early vertebrates.     

XANES Investigation of Dynamic Phase Transition in Olivine Cathode for Li‐Ion Batteries

Dynamic phase transformation in olivine LiFePO₄ involving formation of one or more intermediate or metastable phases is revealed by an in situ time‐resolved X‐ray absorption near edge structure (XANES) technique. The XANES spectra measured during relaxation immediately after the application of relatively high overpotentials, where metastable phases are expected, show a continuous shift of the Fe K‐edge toward higher energy. Surprisingly, the Fe K‐edge relaxes to higher energies after current interrupt regardless of whether the cell is being charged or discharged. This relaxation phenomenon is superimposed upon larger shifts in K‐edge due to changes in Fe^2+/Fe³⁺ ratio due to charging and discharging, and implies an intermediate phase of larger Fe? O bond length than any of the known crystalline phases. No intermediate crystalline phases are observed by X‐ray diffraction (XRD). A metastable amorphous phase formed during dynamic cycling and which structurally relaxes to the equilibrium crystalline phases over a time scale of about 10 min after cessation of charging/discharging current is consistent with the experimental observations.     

Genomic conflict drives patterns of X‐linked population structure in Drosophila neotestacea

Intragenomic conflict has the potential to cause widespread changes in patterns of genetic diversity and genome evolution. In this study, we investigate the consequences of sex‐ratio (SR) drive on the population genetic patterns of the X‐chromosome in Drosophila neotestacea. An SR X‐chromosome prevents the maturation of Y‐bearing sperm during male spermatogenesis and thus is transmitted to ~100% of the offspring, nearly all of which are daughters. Selection on the rest of the genome to suppress SR can be strong, and the resulting conflict over the offspring sex ratio can result in the accumulation of multiple loci on the X‐chromosome that are necessary for the expression of drive. We surveyed variation at 12 random X‐linked microsatellites across 16 populations of D. neotestacea that range in SR frequency from 0% to 30%. First, every locus was differentiated between SR and wild‐type chromosomes, and this drives genetic structure at the X‐chromosome. Once the association with SR is accounted for, the patterns of differentiation among populations are similar to the autosomes. Second, within wild‐type chromosomes, the relative heterozygosity is reduced in populations with an increased prevalence of drive, and the heterozygosity of SR chromosomes is higher than expected based on its prevalence. The combination of the relatively high prevalence of SR drive and the structuring of polymorphism between the SR and wild‐type chromosomes suggests that genetic conflict because of SR drive has had significant consequences on the patterns of X‐linked polymorphism and thus also probably affects the tempo of X‐chromosome evolution in D. neotestacea.     

Occasional recombination of a selfish X‐chromosome may permit its persistence at high frequencies in the wild

The sex‐ratio X‐chromosome (SR) is a selfish chromosome that promotes its own transmission to the next generation by destroying Y‐bearing sperm in the testes of carrier males. In some natural populations of the fly Drosophila neotestacea, up to 30% of the X‐chromosomes are SR chromosomes. To investigate the molecular evolutionary history and consequences of SR, we sequenced SR and standard (ST) males at 11 X‐linked loci that span the ST X‐chromosome and at seven arbitrarily chosen autosomal loci from a sample of D. neotestacea males from throughout the species range. We found that the evolutionary relationship between ST and SR varies among individual markers, but genetic differentiation between SR and ST is chromosome‐wide and likely due to large chromosomal inversions that suppress recombination. However, SR does not consist of a single multilocus haplotype: we find evidence for gene flow between ST and SR at every locus assayed. Furthermore, we do not find long‐distance linkage disequilibrium within SR chromosomes, suggesting that recombination occurs in females homozygous for SR. Finally, polymorphism on SR is reduced compared to that on ST, and loci displaying signatures of selection on ST do not show similar patterns on SR. Thus, even if selection is less effective on SR, our results suggest that gene flow with ST and recombination between SR chromosomes may prevent the accumulation of deleterious mutations and allow its long‐term persistence at relatively high frequencies.     

3‐D analysis of bacterial cell‐(iron)mineral aggregates formed during Fe(II) oxidation by the nitrate‐reducing Acidovorax sp. strain BoFeN1 using complementary microscopy tomography approaches

The formation of cell‐(iron)mineral aggregates as a consequence of bacterial iron oxidation is an environmentally widespread process with a number of implications for processes such as sorption and coprecipitation of contaminants and nutrients. Whereas the overall appearance of such aggregates is easily accessible using 2‐D microscopy techniques, the 3‐D and internal structure remain obscure. In this study, we examined the 3‐D structure of cell‐(iron)mineral aggregates formed during Fe(II) oxidation by the nitrate‐reducing Acidovorax sp. strain BoFeN1 using a combination of advanced 3‐D microscopy techniques. We obtained 3‐D structural and chemical information on different cellular encrustation patterns at high spatial resolution (4–200 nm, depending on the method): more specifically, (1) cells free of iron minerals, (2) periplasm filled with iron minerals, (3) spike‐ or platelet‐shaped iron mineral structures, (4) bulky structures on the cell surface, (5) extracellular iron mineral shell structures, (6) cells with iron mineral filled cytoplasm, and (7) agglomerations of extracellular globular structures. In addition to structural information, chemical nanotomography suggests a dominant role of extracellular polymeric substances (EPS) in controlling the formation of cell‐(iron)mineral aggregates. Furthermore, samples in their hydrated state showed cell‐(iron)mineral aggregates in pristine conditions free of preparation (i.e., drying/dehydration) artifacts. All these results were obtained using 3‐D microscopy techniques such as focused ion beam (FIB)/scanning electron microscopy (SEM) tomography, transmission electron microscopy (TEM) tomography, scanning transmission (soft) X‐ray microscopy (STXM) tomography, and confocal laser scanning microscopy (CLSM). It turned out that, due to the various different contrast mechanisms of the individual approaches, and due to the required sample preparation steps, only the combination of these techniques was able to provide a comprehensive understanding of structure and composition of the various Fe‐precipitates and their association with bacterial cells and EPS.     

Deciphering the Reaction Mechanism of Lithium–Sulfur Batteries by In Situ/Operando Synchrotron‐Based Characterization Techniques

Lithium–sulfur (Li–S) batteries have received extensive attention as one of the most promising next‐generation energy storage systems, mainly because of their high theoretical energy density and low cost. However, the practical application of Li–S batteries has been hindered by technical obstacles arising from the polysulfide shuttle effect and poor electronic conductivity of sulfur and discharge products. Therefore, it is of profound significance for understanding the underlying reaction mechanism of Li–S batteries to circumvent these problems and improve the overall battery performance. Advanced characterization techniques, especially synchrotron‐based X‐ray techniques, have been widely applied to the mechanistic understanding of Li–S batteries. Specifically, in situ/operando synchrotron‐based techniques allows chemical and structural evolution to be directly observed under real operation conditions. Here, recent progress in the understanding of the operating principles of Li–S batteries based on in situ/operando synchrotron‐based techniques, including X‐ray absorption spectroscopy, X‐ray diffraction, and X‐ray microscopy, is reviewed. The aim of this progress report is to provide a comprehensive treatise on in situ/operando synchrotron‐based techniques for mechanism understanding of Li–S batteries, and thereby provide guidance for optimizing their overall electrochemical performances.     

Micro‐ to nanostructure and geochemistry of extant crinoidal echinoderm skeletons

This paper reports the results of micro‐ to nanostructural and geochemical analyses of calcitic skeletons from extant deep‐sea stalked crinoids. Fine‐scale (SEM, FESEM, AFM) observations show that the crinoid skeleton is composed of carbonate nanograins, about 20–100 nm in diameter, which are partly separated by what appears to be a few nm thick organic layers. Sub‐micrometre‐scale geochemical mapping of crinoid ossicles using a NanoSIMS ion microprobe, combined with synchrotron high‐spatial‐resolution X‐ray micro‐fluorescence (μ‐XRF) maps and X‐ray absorption near‐edge structure spectroscopy (XANES) show that high Mg concentration in the central region of the stereom bars correlates with the distribution of S‐sulphate, which is often associated with sulphated polysaccharides in biocarbonates. These data are consistent with biomineralization models suggesting a close association between organic components (including sulphated polysaccharides) and Mg ions. Additionally, geochemical analyses (NanoSIMS, energy dispersive spectroscopy) reveal that significant variations in Mg occur at many levels: within a single stereom trabecula, within a single ossicle and within a skeleton of a single animal. Together, these data suggest that physiological factors play an important role in controlling Mg content in crinoid skeletons and that great care should be taken when using their skeletons to reconstruct, for example, palaeotemperatures and Mg/Ca palaeo‐variations of the ocean.     

Enhancement of the Amazonian Açaí Waste Fibers through Variations of Alkali Pretreatment Parameters

The açaí fruit depulping produces large amounts of long lignocellulosic fiber bundles that are disposed in the environment. Chemical pretreatments may improve açaí fibers favoring their usage in advanced materials. This work aimed to define optimal alkali reaction parameters to improve the properties of açaí fibers. Two NaOH concentrations (5 % and 10 %) and two reaction temperatures (80 °C and 100 °C) were tested. The raw and treated fibers were analyzed by scanning electron microscopy, Fourier transformed infrared spectroscopy, X‐ray diffraction, and thermal analyses. All the alkali pretreatments separated fibers from the bundles, unblocked pit channels by removing silicon structures, exposed the inner lignin, partially removed non‐cellulosic compounds, and raised the cellulose crystalline index. The highest temperature and NaOH content resulted in better cleaning and isolation of the fibers, while milder conditions better preserved the cellulose crystalline structure and thermal stability.     

Differential cadmium and zinc distribution in relation to their physiological impact in the leaves of the accumulating Zygophyllum fabago L.

Cadmium and zinc share many similar physiochemical properties, but their compartmentation, complexation and impact on other mineral element distribution in plant tissues may drastically differ. In this study, we address the impact of 10 μm Cd or 50 μm Zn treatments on ion distribution in leaves of a metallicolous population of the non‐hyperaccumulating species Zygophyllum fabago at tissue and cell level, and the consequences on the plant response through a combined physiological, proteomic and metabolite approach. Micro‐proton‐induced X‐ray emission and laser ablation inductively coupled mass spectrometry analyses indicated hot spots of Cd concentrations in the vicinity of vascular bundles in response to Cd treatment, essentially bound to S‐containing compounds as revealed by extended X‐ray absorption fine structure and non‐protein thiol compounds analyses. A preferential accumulation of Zn occurred in vascular bundle and spongy mesophyll in response to Zn treatment, and was mainly bound to O/N‐ligands. Leaf proteomics and physiological status evidenced a protection of photosynthetically active tissues and the maintenance of cell turgor through specific distribution and complexation of toxic ions, reallocation of some essential elements, synthesis of proteins involved in photosynthetic apparatus or C‐metabolism, and metabolite synthesis with some specificities regarding the considered heavy metal treatment.     

Synchrotron microcomputer tomography for the non‐destructive visualization of the fish skeleton

The objective of this study was to elucidate the potential of synchrotron radiation‐based microcomputer tomography as a non‐destructive method to visualize hard tissue in fish with high resolution. The high X‐ray contrast between mineralized tissue (bone and teeth) and soft tissue permits an easy differentiation. The nature of this method avoids preparation artefacts which may occur during the preparation for histology or scanning electron microscopy. The spatial resolution is of the order of a few μm, depending on the sample. Results obtained for the species medaka (Oryzias latipes) and zebrafish (Danio rerio) are presented.