The triplite–triploidite supergroup: structural modulation in wagnerite, discreditation of magniotriplite, and the new mineral hydroxylwagnerite

Electron-microprobe analysis, single-crystal X-ray diffraction with an area detector, and high-resolution transmission electron microscopy show that minerals related to wagnerite, triplite and triploidite, which are monoclinic Mg, Fe and Mn phosphates with the formula Me2+ 2PO4(F,OH), constitute a modulated series based on the average triplite structure. Modulation occurs along b and may be commensurate with (2b periodicity) or incommensurate but generally close to integer values (∼3b, ∼5b, ∼7b, ∼9b), i.e. close to polytypic behaviour. As a result, the Mg- and F-dominant minerals magniotriplite and wagnerite can no longer be considered polymorphs of Mg2PO4F, i.e., there is no basis for recognizing them as distinct species. Given that wagnerite has priority (1821 vs. 1951), the name magniotriplite should be discarded in favour of wagnerite. Hydroxylwagnerite, end-member Mg2PO4OH, occurs in pyrope megablasts along with talc, clinochlore, kyanite, rutile and secondary apatite in two samples from lenses of pyrope–kyanite–phengite–quartz-schist within metagranite in the coesite-bearing ultrahigh-pressure metamorphic unit of the Dora-Maira Massif, western Alps, Vallone di Gilba, Val Varaita, Piemonte, Italy. Electron microprobe analyses of holotype hydroxylwagnerite and of the crystal with the lowest F content gave in wt%: P2O5 44.14, 43.99; SiO2 0.28, 0.02; SO3 –, 0.01; TiO2 0.20, 0.16; Al2O3 0.06, 0.03; MgO 48.82, 49.12; FeO 0.33, 0.48; MnO 0.01, 0.02; CaO 0.12, 0.10; Na2O 0.01, –; F 5.58, 4.67; H2O (calc) 2.94, 3.36; –O = F 2.35, 1.97; Sum 100.14, 99.98, corresponding to (Mg1.954Fe0.007Ca0.003Ti0.004Al0.002Na0.001)Σ=1.971(P1.003Si0.008)Σ=1.011O4(OH0.526F0.474)Σ=1 and (Mg1.971Fe0.011Ca0.003Ti0.003Al0.001)Σ=1.989(P1.002Si0.001)Σ=1.003O4(OH0.603F0.397)Σ=1, respectively. Due to the paucity of material, H2O could not be measured, so OH was calculated from the deficit in F assuming stoichiometry, i.e., by assuming F + OH = 1 per formula unit. Holotype hydroxylwagnerite is optically biaxial (+), α 1.584(1), β 1.586(1), γ 1.587(1) (589 nm); 2V Z(meas.) = 43(2)°; orientation Y = b. Single-crystal X-ray diffraction gives monoclinic symmetry, space group P21/c, a = 9.646(3) Å, b = 12.7314(16) Å, c = 11.980(4) Å, β = 108.38(4) , V = 1396.2(8) Å3, Z = 16, i.e., hydroxylwagnerite is the OH-dominant analogue of wagnerite [β-Mg2PO4(OH)] and a high-pressure polymorph of althausite, holtedahlite, and α- and ε-Mg2PO4(OH). We suggest that the group of minerals related to wagnerite, triplite and triploidite constitutes a triplite–triploidite super-group that can be divided into F-dominant phosphates (triplite group), OH-dominant phosphates (triploidite group), O-dominant phosphates (staněkite group) and an OH-dominant arsenate (sarkinite). The distinction among the three groups and a potential fourth group is based only on chemical features, i.e., occupancy of anion or cation sites. The structures of these minerals are all based on the average triplite structure, with a modulation controlled by the ratio of Mg, Fe2+, Fe3+ and Mn2+ ionic radii to (O,OH,F) ionic radii.

Osteoclast-like cells on deproteinized bovine bone mineral and biphasic calcium phosphate: light and transmission electron microscopical observations.

OBJECTIVES The occurrence of multinucleated giant cells (MNGCs) on bone substitute materials has been recognized for a long time. However, there have been no studies linking material characteristics with morphology of the MNGCs. The aim was to analyze the qualitative differences of MNGCs on two commercially available calcium phosphate bone substitute materials retrieved from bone defects. MATERIAL AND METHODS Six defects were prepared bilaterally in the mandibular body of three mini pigs. The defects were randomly grafted with either deproteinized bovine bone mineral (DBBM) or biphasic calcium phosphate (BCP). After a healing period of four weeks, bone blocks were embedded in LR White resin. Three consecutive sections per defect were analyzed as follows: two with light microscopy using toluidine blue and tartrate-resistant acid phosphatase (TRAP) staining and one with transmission electron microscopy. RESULTS Multinucleated giant cells appeared on both biomaterials. On BCP, MNGCs had a flat morphology and were not observed in resorption lacunae. On DBBM, the MNGCs appeared more round and were often found in shallow concavities. MNGCs on both biomaterials demonstrated a varying degree of TRAP staining, with a tendency toward higher staining intensity of MNGCs on BCP. At the ultrastructural level, signs of superficial dissolution of BCP together with phagocytosis of minor fragments were observed. MNGCs on the surface of DBBM demonstrated sealing zones and ruffled borders, both features of mature osteoclasts. CONCLUSION MNGCs demonstrated distinctly different histological features depending on the bone substitute material used. Further research is warranted to understand the clinical implications of these morphological observations.

The mammalian central nervous synaptic cleft contains a high density of periodically organized complexes.

Cryo-electron microscopy of vitreous section makes it possible to observe cells and tissues at high resolution in a close-to-native state. The specimen remains hydrated; chemical fixation and staining are fully avoided. There is minimal molecular aggregation and the density observed in the image corresponds to the density in the object. Accordingly, organotypic hippocampal rat slices were vitrified under high pressure and controlled cryoprotection conditions, cryosectioned at a final thickness of approximately 70 nm and observed below -170 degrees C in a transmission electron microscope. The general aspect of the tissue compares with previous electron microscopy observations. The detailed analysis of the synapse reveals that the density of material in the synaptic cleft is high, even higher than in the cytoplasm, and that it is organized in 8.2-nm periodic transcleft complexes. Previously undescribed structures of presynaptic and postsynaptic elements are also described.

Visualization of cell microtubules in their native state.

BACKGROUND INFORMATION Over the past decades, cryo-electron microscopy of vitrified specimens has yielded a detailed understanding of the tubulin and microtubule structures of samples reassembled in vitro from purified components. However, our knowledge of microtubule structure in vivo remains limited by the chemical treatments commonly used to observe cellular architecture using electron microscopy. RESULTS We used cryo-electron microscopy and cryo-electron tomography of vitreous sections to investigate the ultrastructure of microtubules in their cellular context. Vitreous sections were obtained from organotypic slices of rat hippocampus and from Chinese-hamster ovary cells in culture. Microtubules revealed their protofilament ultrastructure, polarity and, in the most favourable cases, molecular details comparable with those visualized in three-dimensional reconstructions of microtubules reassembled in vitro from purified tubulin. The resolution of the tomograms was estimated to be approx. 4 nm, which enabled the detection of luminal particles of approx. 6 nm in diameter inside microtubules. CONCLUSIONS The present study provides a first step towards a description of microtubules, in addition to other macromolecular assemblies, in an unperturbed cellular context at the molecular level. As the resolution appears to be similar to that obtainable with plunge-frozen samples, it should allow for the in vivo identification of larger macromolecular assemblies in vitreous sections of whole cells and tissues.

2D and 3D crystallization of the wild-type IIC domain of the glucose PTS transporter from Escherichia coli.

The bacterial phosphoenolpyruvate: sugar phosphotransferase system serves the combined uptake and phosphorylation of carbohydrates. This structurally and functionally complex system is composed of several conserved functional units that, through a cascade of phosphorylated intermediates, catalyze the transfer of the phosphate moiety from phosphoenolpyruvate to the substrate, which is bound to the integral membrane domain IIC. The wild-type glucose-specific IIC domain (wt-IIC(glc)) of Escherichia coli was cloned, overexpressed and purified for biochemical and functional characterization. Size-exclusion chromatography and scintillation-proximity binding assays showed that purified wt-IIC(glc) was homogenous and able to bind glucose. Crystallization was pursued following two different approaches: (i) reconstitution of wt-IIC(glc) into a lipid bilayer by detergent removal through dialysis, which yielded tubular 2D crystals, and (ii) vapor-diffusion crystallization of detergent-solubilized wt-IIC(glc), which yielded rhombohedral 3D crystals. Analysis of the 2D crystals by cryo-electron microscopy and the 3D crystals by X-ray diffraction indicated resolutions of better than 6Å and 4Å, respectively. Furthermore, a complete X-ray diffraction data set could be collected and processed to 3.93Å resolution. These 2D and 3D crystals of wt-IIC(glc) lay the foundation for the determination of the first structure of a bacterial glucose-specific IIC domain.

Macromolecular interaction studies on FF1-3 of human CA150 and STAT1-importin alpha5 using biophysical and biochemical methods

Macromolecular interactions, such as protein-protein interactions and protein-DNA interactions, play important roles in executing biological functions in cells. However the complexity of such interactions often makes it very challenging to elucidate the structural details of these subjects. In this thesis, two different research strategies were applied on two different two macromolecular systems: X-ray crystallography on three tandem FF domains of transcription regulator CA150 and electron microscopy on STAT1-importin α5 complex. The results from these studies provide novel insights into the function-structure relationships of transcription coupled RNA splicing mediated by CA150 and the nuclear import process of the JAK-STAT signaling pathway. ^ The first project aimed at the protein-protein interaction module FF domain, which often occurs as tandem repeats. Crystallographic structure of the first three FF domains of human CA150 was determined to 2.7 Å resolution. This is the only crystal structure of an FF domain and the only structure on tandem FF domains to date. It revealed a striking connectivity between an FF domain and the next. Peptide binding assay with the potential binding ligand of FF domains was performed using fluorescence polarization. Furthermore, for the first time, FF domains were found to potentially interact with DNA. DNA binding assays were also performed and the results were supportive to this newly proposed functionality of an FF domain. ^ The second project aimed at understanding the molecular mechanism of the nuclear import process of transcription factor STAT1. The first structural model of pSTAT1-importin α5 complex in solution was built from the images of negative staining electron microscopy. Two STAT1 molecules were observed to interact with one molecule of importin α5 in an asymmetric manner. This seems to imply that STAT1 interacts with importin α5 with a novel mechanism that is different from canonical importin α-cargo interactions. Further in vitro binding assays were performed to obtain more details on the pSTAT1-importin α5 interaction. ^

Magnetic properties of Cenozoic MORB and Cretaceous MORB from DSDP and ODP holes

The fact that the natural remanent magnetization (NRM) intensity of mid-oceanic-ridge basalt (MORB) samples shows systematic variations as a function of age has long been recognized: maximum as well as average intensities are generally high for very young samples, falling off rather rapidly to less than half the recent values in samples between 10 and 30 Ma, whereupon they slowly rise in the early Tertiary and Cretaceous to values that approach those of the very young samples. NRM intensities measured in this study follow the same trends as those observed in previous publications. In this study, we take a statistical approach and examine whether this pattern can be explained by variations in one or more of all previously proposed mechanisms: chemical composition of the magnetic minerals, abundance of these magnetization carriers, vectorial superposition of parallel or antiparallel components of magnetization, magnetic grain or domain size patterns, low-temperature oxidation to titanomaghemite, or geomagnetic field behavior. We find that the samples do not show any compositional, petrological, rock-magnetic, or paleomagnetic patterns that can explain the trends. Geomagnetic field intensity is the only effect that cannot be directly tested on the same samples, but it shows a similar pattern as our measured NRM intensities. We therefore conclude that the geomagnetic field strength was, on-average, significantly greater during the Cretaceous than during the Oligocene and Miocene.

Hydrate, ice and quartz content of eight holes of Leg 204

The state of preservation of natural gas hydrate samples, recovered from 6 sites drilled during ODP Leg 204 at southern summit of Hydrate Ridge, Oregon Margin, has been investigated by X-ray diffraction (XRD) and cryo-scanning-electron-microscopy (cryo-SEM) techniques. A detailed characterization of the state of decomposition of gas hydrates is necessary since no pressurized autoclave tools were used for sampling and partial dissociation must have occurred during recovery prior to the quench and storage in liquid nitrogen. Samples from 16 distinct horizons have been investigated by synchrotron X-ray diffraction measurements at HASYLAB/ Hamburg. A full profile fitting analysis ("Rietveld method") of synchrotron XRD data provides quantitative phase determinations of the major sample constituents such as gas hydrate structure I (sI), hexagonal ice (Ih) and quartz. The ice content (Ih) in each sample is related to frozen water composed of both original existing pore water and the water from decomposed hydrates. Hydrate contents as measured by diffraction vary between 0 and 68 wt.% in the samples we measured. Samples with low hydrate content usually show micro-structural features in cryo-SEM ascribed to extensive decomposition. Comparing the appearance of hydrates at different scales, the grade of preservation seems to be primarily correlated with the contiguous volume of the original existing hydrate; the dissociation front appears to be indicated by micrometer-sized pores in a dense ice matrix.

Animation of micro-computed tomography of bioerosion trace Entobia megastoma made by East Australian bioeroding sponge Cliona celata

Endolithic bioerosion is difficult to analyse and to describe, and it usually requires damaging of the sample material. Sponge erosion (Entobia) may be one of the most difficult to evaluate as it is simultaneously macroscopically inhomogeneous and microstructurally intricate. We studied the bioerosion traces of the two Australian sponges Cliona celata Grant, 1826 (sensu Schönberg 2000) and Cliona orientalis Thiele, 1900 with a newly available radiographic technology: high resolution X-ray micro-computed tomography (MCT). MCT allows non-destructive visualisation of live and dead structures in three dimensions and was compared to traditional microscopic methods. MCT and microscopy showed that C. celata bioerosion was more intense in the centre and branched out in the periphery. In contrast, C. orientalis produced a dense, even trace meshwork and caused an overall more intense erosion pattern than C. celata. Extended pioneering filaments were not usually found at the margins of the studied sponge erosion, but branches ended abruptly or tapered to points. Results obtained with MCT were similar in quality to observations from transparent optical spar under the dissecting microscope. Microstructures could not be resolved as well as with e.g. scanning electron microscopy (SEM). Even though sponge scars and sponge chips were easily recognisable on maximum magnification MCT images, they lacked the detail that is available from SEM. Other drawbacks of MCT involve high costs and presently limited access. Even though MCT cannot presently replace traditional techniques such as corrosion casts viewed by SEM, we obtained valuable information. Especially for the possibility to measure endolithic pore volumes, we regard MCT as a very promising tool that will continue to be optimised. A combination of different methods will produce the best results in the study of Entobia.

Peak areas and heights for minerals of ODP Leg 113-696B (Table 1)

Seismic data acquired over the eastern shelf and margin of the South Orkney microcontinent, Antarctica, have shown a high-amplitude reflection lying at a sub-bottom two-way traveltime (TWT) of 0.5-0.8 s. There appear to be two causes for the reflection which apply in different parts of the shelf. The more widespread cause of the reflection is a break-up unconformity associated with the opening of Jane Basin to the east. This is clearly seen where reflections in the underlying sequence are discordant. In contrast, in Eotvos Basin and the southeastern part of Bouguer Basin, the high-amplitude reflection in places cuts across bedding and is interpreted to be caused by silica diagenesis. A post-cruise analysis of core samples from Site 696 in Eotvos Basin by X-ray diffraction (XRD) and scanning electron microscopy (SEM) revealed the presence of a silica diagenetic front at 520-530 mbsf. The position of the unconformity at this site is uncertain, but probably coincides with a change of detrital input near 548 mbsf. Fluctuations of physical properties related to the depth of the diagenetic front are difficult to separate from those related to the variation of detrital composition over the same depth interval. Correlation of the drilling record with the seismic record is difficult but with a synthetic seismogram it is demonstrated that diagenesis is the probable cause of the high-amplitude reflection. In Bouguer Basin at Site 695 the depth of the high-amplitude reflection was not reached by drilling; however, the reflection is probably also caused by silica diagenesis because of the biogenic silica-rich composition of the sediments cored. The estimated temperatures and ages of the sediments at the depths of the high-amplitude reflections at Sites 695 and 696 compare favorably with similar data from other diagenetic fronts of the world. The high-amplitude reflection in Bouguer Basin is commonly of inverse polarity, possibly caused either by interference between reflections from several closely-spaced reflecting layers, such as chert horizons, or by free gas trapped near the diagenetic front.

Ba, Al and SO4 concentrations in sediments and pore fluid of ODP Leg 127/128 samples

The barium distribution in sediments and pore fluids from five sites drilled in the Japan Sea have been used to illustrate the geochemical behavior of this element as it pertains paleoproductivity reconstructions, diagenetic remobilization, and barite precipitation in authigenic fronts. Sites where sulfate is depleted in the pore fluids also show high concentrations of dissolved barium, reflecting dissolution of biogenic barite. The high rate of sedimentation at Sites 798 and 799 results in a rapid sulfate depletion, which in turn leads to barite dissolution and reprecipitation in diagenetic fronts. The dissolved barium distribution at these sites has been used to quantify the rate of barite dissolution; we estimate a first-order rate constant for barite dissolution to be 2*10**-6/s at Site 799 and 2*10**-7/s at Site 798. Authigenic barite has been documented in sediments from Site 799 at 323 meters below seafloor by scanning electron microscopy and X-ray fluorescence analysis. These results indicate barite precipitation in a diagenetic front near the zone of sulfate depletion by upward migration of dissolved barium and downward diffusion of sulfate. Barite precipitation has also been inferred at Sites 796 and 798 based on sedimentary and dissolved barium distributions. Sulfate is not depleted in the pore fluids of Site 794. The lack of diagenetic remobilization of biogenic barium at this site preserves the high barium signal associated with the high-productivity sequences deposited during the late Miocene to Pliocene. Significantly, the organic carbon distribution does not indicate high accumulation rates during the periods of high opal and barium deposition. Instead, higher organic carbon accumulations are recorded in the Quaternary and middle Miocene sequences; intervals that are also characterized by deposition of siliciclastic turbidites. The presence of a terrestrial component in the organic carbon record renders barium a more useful indicator than organic carbon for paleoproductivity reconstructions in this marginal sea.

Physical properties, grain size distribution and clay mineralogy of ODP Leg 117 sites

The fabric of sediments recovered at sites drilled on the Indus Fan, Owen Ridge, and Oman margin during Ocean Drilling Program Leg 117 was examined by scanning electron microscopy to document changes that accompany sediment burial. Two sediment types were studied: (1) biogenic sediments consisting of a variety of marly nannofossil and nannofossil oozes and chalks and (2) terrigenous sediments consisting of fine-grained turbidites deposited in association with the Indus Fan. Biogenic sediments were examined with samples from the seafloor to depths of 306 m below seafloor (mbsf) on the Owen Ridge (Site 722) and 368 mbsf on the Oman margin (Sites 723 and 728). Over these depth ranges the biogenic sediments are characterized by a random arrangement of microfossils and display little chemical diagenetic alteration. The microfossils are dispersed within a fine-grained matrix that is predominantly microcrystalline carbonate particles on the Owen Ridge and clay and organic matter on the Oman margin. Sediments with abundant siliceous microfossils display distinct, open fabrics with high porosity. Porosity reduction resulting from gravitational compaction appears to be the primary process affecting fabric change in the biogenic sediment sections. Fabric of illite-rich clayey silts and silty claystones from the Indus Fan (Site 720) and Owen Ridge (Sites 722 and 731) was examined for a composite section extending from 45 to 985 mbsf. In this section fabric of the fine-grained turbidites changes from one with small flocculated clay domains, random particle arrangement, and high porosity to a fabric with larger domains, strong preferred particle orientation roughly parallel to bedding, and lower porosity. These changes are accomplished by a growth in domain size, primarily through increasing face-to-face contacts, and by particle reorientation which is characterized by a sharp increase in alignment with bedding between 200 and 400 mbsf. Despite extensive particle reorientation, flocculated clay fabric persists in the deepest samples examined, particularly adjacent to silt grains, and the sediments lack fissility. Fabric changes over the 45-985 mbsf interval occur in response to gravitational compaction. Porosity reduction and development of preferred particle orientation in the Indus Fan and Owen Ridge sections occur at greater depths than outlined in previous fabric models for terrigenous sediments as a consequence of a greater abundance of silt and a greater abundance of illite and chlorite clays.

Effective stress, porosity, p-wave velocity and mineral composition of ODP Hole 174A-1073A sediments

Porosity, permeability, and compressional (P-wave) velocity were measured as a function of stress on sediments from Ocean Drilling Program Site 1073, U.S. Mid-Atlantic continental slope. Thin sections, scanning electron microscopy, and X-ray diffraction analyses provided mineralogical characteristics of the samples. Uniaxial strain boundary conditions were imposed on the samples during consolidation tests with the maximum effective axial stress reaching 13 MPa. The maximum effective radial stress necessary to maintain uniaxial strain was 7.6 MPa. Over an effective axial stress interval of 0 to 5.2 MPa, Sample 174A-1073A-26X-2, 82-89 cm (226.65 meters below seafloor [mbsf]), exhibited the largest decrease in porosity (51% to 41%), whereas Sample 71X-1, 2-8 cm (644.70 mbsf), exhibited the smallest decrease in porosity (48% to 45%). All samples showed negligible porosity increases during unloading. The permeability (on the order of 1 x 10-17 m**2) of Sample 174A-1073A-71X-1, 2-8 cm, was twice that measured on Sample 8H-1, 23-26 cm (63.75 mbsf), even though the former was considerably deeper and older. The differences in porosity-stress behavior and permeability between shallow and deep samples is related to lithologic, mineralogic, and diagenetic differences between the sediments above and below the Pliocene-Pleistocene to Miocene unconformity. P-wave velocity for Samples 174A-1073A-41X-5, 97-103 cm (372.35 mbsf), and 71X-1, 2-8 cm, increased with decreasing porosity, but did not change significantly during unloading.