The majority of invertebrate skeletal tissues are composed of the most stable crystalline polymorphs of CaCO3, calcite, and/or aragonite. Here we describe a composite skeletal tissue from an ascidian ...in which amorphous and crystalline calcium carbonate coexist in well-defined domains separated by an organic sheath. Each biogenic mineral phase has a characteristic Mg content (5.9 and 1.7 mol %, respectively) and concentration of intramineral proteins (0.05 and 0.01 wt %, respectively). Macromolecular extracts from various biogenic amorphous calcium carbonate (ACC) skeletons are typically glycoproteins, rich in glutamic acid and hydroxyamino acids. The proteins from the crystalline calcitic phases are aspartate-rich. Macromolecules extracted from biogenic ACC induced the formation of stabilized ACC and/or inhibited crystallization of calcite in vitro. The yield of the synthetic ACC was 15−20%. The presence of Mg facilitated the stabilization of ACC: the protein content in synthetic ACC was 0.12 wt % in the absence of Mg and 0.07 wt % in the presence of Mg (the Mg content in the precipitate was 8.5 mol %). In contrast, the macromolecules extracted from the calcitic layer induced the formation of calcite crystals with modified morphology similar to that expressed by the original biogenic calcite. We suggest that specialized macromolecules and magnesium ions may cooperate in the stabilization of intrinsically unstable amorphous calcium carbonate and in the formation of complex ACC/calcite tissues in vivo.
A convenient tooth. Here we investigate how the different kinds of calcite crystals in a sea urchin tooth (left) work together as an effective grinding tool. The polycrystalline matrix has a higher ...elastic modulus and hardness than the single crystals, yet both work together to produce a flat grinding surface (right).
The sea urchin tooth is a remarkable grinding tool. Even though the tooth is composed almost entirely of calcite, it is used to grind holes into a rocky substrate itself often composed of calcite. ...Here, we use 3 complementary high-resolution tools to probe aspects of the structure of the grinding tip: X-ray photoelectron emission spectromicroscopy (X-PEEM), X-ray microdiffraction, and NanoSIMS. We confirm that the needles and plates are aligned and show here that even the high Mg polycrystalline matrix constituents are aligned with the other 2 structural elements when imaged at 20-nm resolution. Furthermore, we show that the entire tooth is composed of 2 cooriented polycrystalline blocks that differ in their orientations by only a few degrees. A unique feature of the grinding tip is that the structural elements from each coaligned block interdigitate. This interdigitation may influence the fracture process by creating a corrugated grinding surface. We also show that the overall Mg content of the tooth structural elements increases toward the grinding tip. This probably contributes to the increasing hardness of the tooth from the periphery to the tip. Clearly the formation of the tooth, and the tooth tip in particular, is amazingly well controlled. The improved understanding of these structural features could lead to the design of better mechanical grinding and cutting tools.
DNA from fossil human bones could provide invaluable information about population migrations, genetic relations between different groups and the spread of diseases. The use of ancient DNA from bones ...to study the genetics of past populations is, however, very often compromised by the altered and degraded state of preservation of the extracted material. The universally observed postmortem degradation, together with the real possibility of contamination with modern human DNA, makes the acquisition of reliable data, from humans in particular, very difficult. We demonstrate that relatively well preserved DNA is occluded within clusters of intergrown bone crystals that are resistant to disaggregation by the strong oxidant NaOCl. We obtained reproducible authentic sequences from both modern and ancient animal bones, including humans, from DNA extracts of crystal aggregates. The treatment with NaOCl also minimizes the possibility of modern DNA contamination. We thus demonstrate the presence of a privileged niche within fossil bone, which contains DNA in a better state of preservation than the DNA present in the total bone. This counterintuitive approach to extracting relatively well preserved DNA from bones significantly improves the chances of obtaining authentic ancient DNA sequences, especially from human bones.
Display omitted
Quantifying ion concentrations and mapping their intracellular distributions at high resolution can provide much insight into the formation of biomaterials. The key to achieving this ...goal is cryo-fixation, where the biological materials, tissues and associated solutions are rapidly frozen and preserved in a vitreous state. We developed a correlative cryo-Scanning Electron Microscopy (SEM)/Energy Dispersive Spectroscopy (EDS) protocol that provides quantitative elemental analysis correlated with spatial imaging of cryo-immobilized specimens. We report the accuracy and sensitivity of the cryo-EDS method, as well as insights we derive on biomineralization pathways in a foraminifer. Foraminifera are marine protozoans that produce Mg-containing calcitic shells and are major calcifying organisms in the oceans. We use the cryo-SEM/EDS correlative method to characterize unusual Mg and Ca-rich particles in the cytoplasm of a benthic foraminifer. The Mg/Ca ratio of these particles is consistently lower than that of seawater, the source solution for these ions. We infer that these particles are involved in Ca ion supply to the shell. We document the internal structure of the MgCa particles, which in some cases include a separate Si rich core phase. This approach to mapping ion distribution in cryo-preserved specimens may have broad applications to other mineralized biomaterials.
Ions are an integral part of life, and some ions play fundamental roles in cell metabolism. Determining the concentrations of ions in cells and between cells, as well as their distributions at high resolution can provide valuable insights into ion uptake, storage, functions and the formation of biomaterials. Here we present a new cryo-SEM/EDS protocol that allows the mapping of different ion distributions in solutions and biological samples that have been cryo-preserved. We demonstrate the value of this novel approach by characterizing a novel biogenic mineral phase rich in Mg found in foraminifera, single celled marine organisms. This method has wide applicability in biology, and especially in understanding the formation and function of mineral-containing hard tissues.
Yuchanyan Cave in Daoxian County, Hunan Province (People's Republic of China), yielded fragmentary remains of 2 or more ceramic vessels, in addition to large amounts of ash, a rich animal bone ...assemblage, cobble and flake artifacts, bone tools, and shell tools. The artifacts indicate that the cave was a Late Paleolithic foragers' camp. Here we report on the radiocarbon ages of the sediments based on analyses of charcoal and bone collagen. The best-preserved charcoal and bone samples were identified by prescreening in the field and laboratory. The dates range from around 21,000 to 13,800 cal BP. We show that the age of the ancient pottery ranges between 18,300 and 15,430 cal BP. Charcoal and bone collagen samples located above and below one of the fragments produced dates of around 18,000. These ceramic potsherds therefore provide some of the earliest evidence for pottery making in China.
Display omitted
Fish represent the most diverse and numerous of the vertebrate clades. In contrast to the bones of all tetrapods and evolutionarily primitive fish, many of the evolutionarily more ...advanced fish have bones that do not contain osteocytes. Here we use a variety of imaging techniques to show that anosteocytic fish bone is composed of a sequence of planar layers containing mainly aligned collagen fibrils, in which the prevailing principal orientation progressively spirals. When the sequence of fibril orientations completes a rotation of around 180°, a thin layer of poorly oriented fibrils is present between it and the next layer. The thick layer of aligned fibrils and the thin layer of non-aligned fibrils constitute a lamella. Although both basic components of mammalian lamellar bone are found here as well, the arrangement is unique, and we therefore call this structure lamellated bone. We further show that the lamellae of anosteocytic fish bone contain an array of dense, small-diameter (1–4μm) bundles of hypomineralized collagen fibrils that are oriented mostly orthogonal to the lamellar plane. Results of mechanical tests conducted on beams from anosteocytic fish bone and human cortical bone show that the fish bones are less stiff but much tougher than the human bones. We propose that the unique lamellar structure and the orthogonal hypomineralized collagen bundles are responsible for the unusual mechanical properties and mineral distribution in anosteocytic fish bone.
The crossed lamellar microstructure is the most commonly formed shell structure in mollusks. Although the mechanical properties and the architecture of the crossed lamellar microstructure have been ...extensively studied, its formation mechanisms are largely unknown. In this study, we obtained information on the formation processes of the aragonite crystals in the crossed lamellar microstructure using two converging approaches: cryo-scanning electron microscopy performed on the forming shell layer and in vitro crystallization experiments with extracted matrix macromolecules. We show that deposition of the first crystalline particles occurs in a thin granular layer on the growing surface of the mature crossed lamellar microstructure. The granules subsequently grow to form the lamellar prisms. In vitro, the ensemble of organic macromolecules extracted from the crossed lamellar layers induced the formation of only aragonite crystals from solutions containing Ca/Mg ion ratio of 1/2.5 within 6 h, and that the morphology and the crystallographic orientation of the synthetic crystals are the same as those of the biogenic crystals. In the absence of matrix macromolecules, both calcite and aragonite crystals formed. These observations provide insights into the early crystal nucleation and growth events of aragonite crystals in the formation of the crossed lamellar microstructure.
Combe-Grenal site (Southwest France) was excavated by F. Bordes between 1953 and 1965. He found several human remains in Mousterian levels 60, 39, 35 and especially 25, corresponding to MIS 4 ...(~75–70/60 ky BP) and with Quina Mousterian lithics. One of the fossils found in level 25 is Combe-Grenal IV, consisting of a fragment of the left corpus of a juvenile mandible. This fragment displays initial juvenile periodontitis, and the two preserved teeth (LLP4 and LLM1) show moderate attrition and dental calculus. The SEM tartar analysis demonstrates the presence of cocci and filamentous types of bacteria, the former being more prevalent. This result is quite different from those obtained for the two adult Neanderthals Kebara 2 and Subalyuk 1, where more filamentous bacteria appear, especially in the Subalyuk 1 sample from Central Europe. These findings agree with the available biomedical data on periodontitis and tartar development in extant individuals, despite the different environmental conditions and diets documented by numerous archeological, taphonomical and geological data available on Neanderthals and present-day populations. New metagenomic analyses are extending this information, and despite the inherent difficulties, they will open important perspectives in studying this ancient human pathology.
During mollusk shell formation, the mineral phase forms within an organic matrix composed of β-chitin, silk-like proteins, and acidic glycoproteins rich in aspartic acid. The matrix is widely assumed ...to play an important role in controlling mineralization. Thus, understanding its structure is of prime importance. Cryo-transmission electron microscopy (Cryo-TEM) studies of the matrix of the bivalve Atrina embedded in vitrified ice show that the interlamellar sheets are composed mainly of highly ordered and aligned β-chitin fibrils. The silk, which is quantitatively an important component of the matrix, could not be imaged within the sheets. Organic material was, however, observed between sheets. We infer that this is the location of the silk. As this material reveals no regular structure, we suggest that at least prior to mineralization the silk is in the form of a hydrated gel. This is supported by cryo-TEM structural observations of an artificial assembly of β-chitin with and without silk. This view of the nacreous organic matrix significantly changes previous models of the matrix structure and hence hypotheses pertaining to the mechanisms by which mineral formation occurs.
PDF
