Atrina Rigida Research Articles

Identification of pen shells (Bivalvia: Ostreida: Pinnidae) collected off northern Iloilo, Philippines using their morphological characters

This study describes morphological characteristics of pen shells collected along the coast of northern Iloilo, Philippines. A total of 380 pen shell individuals with intact shells and their adductor muscles were carefully examined and measured. Eleven species were studied, namely: Atrina pectinata (Linnaeus, 1767), Atrina inflata (Dillwyn, 1817), Atrina vexillum (Born, 1778), Atrina seminuda (Lamarck, 1819), Atrina rigida ([Lightfoot], 1786), Pinna bicolor Gmelin, 1791, Pinna atropurpurea Sowerby, 1825, Pinna deltodes Menke, 1843, Pinna muricata Linnaeus, 1758, Pinna incurva Gmelin, 1791, and Pinna nobilis Linnaeus, 1758. The species were identified based on the nine characteristics of the valve for the genus Pinna and eight for the genus Atrina. These characteristics were then correlated with their adductor muscles’ morphology. Analysis of variance revealed that the three most dominant species investigated under genus Atrina, namely Atrina pectinata, Atrina inflata, and Atrina vexillum, were found to be significantly different in the eight characters of the valve (P < 0.05). Moreover, three representative Pinna species, Pinna bicolor, Pinna atropurpurea, and Pinna deltodes, were significantly (P < 0.05) different based on shell width, dorsal posterior shell margin, sulcus width, and dorsal posterior margin to dorsal nacreous margin. The observed high correlation (P < 0.05) between adductor muscle properties and different shell length characteristics for five dominant species can be used as a predictor of growth and suggests that the increase in the size of adductor muscle correlates to the increase in the size of the mentioned shell characteristics. No previous study of this kind was conducted in the Philippines. This work provides relevant information for related biological research on other pen shell species and for the management of pen shell resources in northern Iloilo and, possibly, other regions in the world.

Comparative nanoindentation study of biogenic and geological calcite

Biogenic minerals are often reported to be harder and tougher than their geological counterparts. However, quantitative comparison of their mechanical properties, particularly fracture toughness, is still limited. Here we provide a systematic comparison of geological and biogenic calcite (mollusk shell Atrina rigida prisms and Placuna placenta laths) through nanoindentation under both dry and 90% relative humidity conditions. Berkovich nanoindentation is used to reveal the mechanical anisotropy of geological calcite when loaded on different crystallographic planes, i.e., reduced modulus Er{104} ≥ Er{108} > Er{001} and hardness H{001} ≥ H{104} ≥ H{108}, and biogenic calcite has comparable modulus but increased hardness than geological calcite. Based on conical nanoindentation, we elucidate that plastic deformation is activated in geological calcite at the low-load regime (<20 mN), involving r{104} and f{012} dislocation slips as well as e{018} twinning, while cleavage fracture dominates under higher loads by cracking along {104} planes. In comparison, biogenic calcite tends to undergo fracture, while the intercrystalline organic interfaces contribute to damage confinement. In addition, increased humidity does not show a significant influence on the properties of geological calcite and the single-crystal A. rigida prisms, however, the laminate composite of P. placenta laths (layer thickness, ∼250–300 nm) exhibits increased toughness and decreased hardness and modulus. We believe the results of this study can provide a benchmark for future investigations on biominerals and bio-inspired materials.

Intrinsic Mechanical Properties of Individual Biogenic Mineral Units in Biomineralized Skeletons.

A bioinspired study on replicating the superior damage tolerance of bioceramic composites requires a detailed understanding of the intrinsic properties of biogenic mineral units. Here, we investigate and compare the intrinsic properties of biogenic calcite (Atrina rigida) and aragonite (Sinanodonta woodiana) by conducting microbending experiments on the separated prismatic building blocks. Analyzed bending results indicate that the biogenic calcite has a higher modulus (36.24 ± 14.4 GPa for A. rigida vs. 29.9 ± 10.5 GPa for S. woodiana) and strength (446.5 ± 141.5 MPa for A. rigida vs. 338.6 ± 63.2 MPa for S. woodiana) than the biogenic aragonite, while the nanoindentation results indicate the opposite trend. Further systematic fractographic analysis suggests that the biogenic calcite fractures like amorphous glass, while the biogenic aragonite resembles polycrystalline ceramics. These contradictory behaviors of biogenic calcite and aragonite under tension-dominated (microbending) and indentation loading conditions are attributed to their different intrinsic structures, i.e., intracrystalline organic inclusions in single-crystal calcite vs. interlocked nanograins in polycrystalline aragonite.

Strategies for simultaneous strengthening and toughening via nanoscopic intracrystalline defects in a biogenic ceramic

While many organisms synthesize robust skeletal composites consisting of spatially discrete organic and mineral (ceramic) phases, the intrinsic mechanical properties of the mineral phases are poorly understood. Using the shell of the marine bivalve Atrina rigida as a model system, and through a combination of multiscale structural and mechanical characterization in conjunction with theoretical and computational modeling, we uncover the underlying mechanical roles of a ubiquitous structural motif in biogenic calcite, their nanoscopic intracrystalline defects. These nanoscopic defects not only suppress the soft yielding of pure calcite through the classical precipitation strengthening mechanism, but also enhance energy dissipation through controlled nano- and micro-fracture, where the defects’ size, geometry, orientation, and distribution facilitate and guide crack initialization and propagation. These nano- and micro-scale cracks are further confined by larger scale intercrystalline organic interfaces, enabling further improved damage tolerance.

Hard to Soft: Biogenic Absorbent Sponge-like Material from Waste Mussel Shells

Summary Mussel shells are a source of renewable CaCO3 and can be obtained from aquaculture waste streams. Valorization of waste shells relieves strain on landfills and aligns with the goals of a circular economy. Here, we present the preparation of an absorbent CaCO3 material, soft calcite (SC), prepared from waste blue mussel shells under mild conditions. Calcite is the only polymorph present in SC, despite the presence of aragonite in the shell. SC has a nest-like morphology, unlike the highly ordered calcite in the shell. SC is formed by the reassembly of calcite prisms and may be held together by the organic matrix “glue” from the original shell as evidenced by NMR spectroscopy. This inorganic sponge adsorbs dyes from aqueous solution and absorbs crude oil from seawater with good recyclability. Our results demonstrate how a natural structure can be modified in a sustainable fashion to yield a higher-value material.

A Facilitation Cascade Enhances Local Biodiversity in Seagrass Beds

Invertebrate diversity can be a key driver of ecosystem functioning, yet understanding what factors influence local biodiversity remains uncertain. In many marine and terrestrial systems, facilitation cascades where primary foundation and/or autogenic ecosystem engineering species promote the settlement and survival of a secondary foundation/engineering species have been shown to enhance local biodiversity and ecosystem functioning. We experimentally tested if a facilitation cascade occurs among eelgrass (Zostera marina), pen clams (Atrina rigida), and community diversity in temperate seagrass beds in North Carolina, U.S.A., and if this sequence of direct positive interactions created feedbacks that affected various metrics of seagrass ecosystem function and structure. Using a combination of surveys and transplant experiments, we found that pen clam density and survivorship was significantly greater in seagrass beds, indicating that eelgrass facilitates pen clams. Pen clams in turn enhanced local diversity and increased both the abundance and species richness of organisms (specifically, macroalgae and fouling invertebrate fauna)—the effect of which scaled with increasing clam density. However, we failed to detect an impact of pen clams on other seagrass functions and hypothesize that functioning may more likely be enhanced in scenarios where secondary foundation species specifically increase the diversity of key functional groups such as epiphyte grazers and/or when bivalves are infaunal rather than epifaunal. Our findings add to the growing amount of literature that demonstrates that secondary foundation species are important drivers of local biodiversity in marine ecosystems. Further experimentation is needed that directly examines (i) the role of functional versus overall diversity on seagrass functions and (ii) the relative importance of life-history strategy in determining when and where engineering bivalves increase biodiversity and/or functioning of seagrass beds.

Survey of Bivalve Molluscs forBonamiaspp. and Other Parasitic Pathogens in Florida East Coast Lagoons

A 2007 study of Suminoe (Crassostrea ariakensis) and eastern (Crassostrea virginica) oysters exposed to Indian River Lagoon (IRL), Florida waters revealed the presence of Bonamia spp. in exposed oysters. In 2010, a survey of oysters and other bivalves (n = 721, 18 species) was conducted in the southern IRL (10 sites) and in the more southerly Lake Worth Lagoon (LWL, 3 sites). Polymerase chain reaction (PCR) was used to assess the prevalence of three oyster parasites, Bonamia spp., Perkinsus marinus, and Haplosporidium nelsoni, in collected samples. Histology, fluorescent in situ hybridization (FISH) and standard H&E staining, was used to confirm PCR-positive Bonamia results. Bonamia spp. was detected in eight species (Brachidontes exustus, Crassostrea virginica, Geukensia demissa, Ischadium recurvum, Isognomon alatus, Isognomon bicolor, Mercenaria campechiensis, and Ostrea stentina). The prevalence was 17.3% in the IRL and 9.9% in LWL with 66% of PCR-positive samples subsequently identified as either Bonamia exitiosa or Bonamia perspora. The presence of Bonamia spp. was confirmed by FISH in C. virginica, O. stentina, I. alatus, I. bicolor, and G. demissa, although only C. virginica and O. stentina showed evidence of infection. The presence of P. marinus was detected in eight species (Atrina rigida, B. exustus, C. virginica, G. demissa, I. recurvum, I. alatus, I. bicolor, and O. stentina). The prevalence was 12.4% in the IRL and 7.7% in LWL. The presence of MSX was not detected in any of the collected bivalves. This study confirms the presence of Bonamia spp. and P. marinus in Florida east coast waters and widens the described geographic range for Bonamia spp., and provides further evidence that MSX is unlikely to be found in Florida waters

Effect of pH Change on Exoskeletons of Selected Saltwater Organisms Which Rely on Calcium Fixation

The projections for rising atmospheric carbon dioxide concentrations indicate that the pH levels of the ocean surface could decrease by 0.3-0.4 units by the end of the 21st century. The objective of this research was to evaluate the effect of pH on the exoskeletons of six aquatic organisms commonly found in South Florida coastal waters. The exoskeleton samples studied were from the common nutmeg (Cancellaria reticulate), lettered olive (Oliva sayana), stiff pen shell (Atrina rigida), kitten’s paw (Plicatulidae), fan coral (Gorgonia ventalina), and common slipper shell (Crepidula fornicate). The exoskeleton samples were exposed to saltwater (34% salinity) at pH levels ranging from 8.3 to 6.0 for 5 days. The changes in the masses of the samples before and after exposure were compared. We normalized the data in reference to the observations at pH=8.3, which is the current pH level of the ocean surface. The fan coral had the highest percent mass loss, which increased with decreasing pH until pH 6.5. Exoskeleton samples from the stiff pen shell were not affected by the pH changes. Exoskeletons from the common nutmeg, lettered olive, and common slipper shell had similar responses to pH changes with relatively small changes in mass. Exoskeletons from the kitten’s paw had significant mass loss at lower pH conditions. The research results show that as ocean acidification increases, the exoskeletons of marine organisms will be affected. Some organisms, such as the fan coral and kitten’s paw, may lose their exoskeletons. Other organisms that rely on the marine organisms with exoskeletons for shelter and food could also be affected.

Life history affects how species experience succession in pen shell metacommunities

In nature, very few species are common and broadly distributed. Most species are rare and occupy few sites; this pattern is ubiquitous across habitats and taxa. In spatially structured communities (metacommunities), regional distribution and local abundance may change as the relative effects of within-habitat processes (e.g., species interactions) and among-habitat processes (e.g., dispersal) may vary through succession. A field experiment with the marine benthic inhabitants of pen shells (Atrina rigida) tested how common and rare species respond to succession and metacommunity size. I followed community development through time and partitioned species into sessile and motile based on their natural history. Rare species drive diversity patterns and are influenced by metacommunity size: there are strong abundance-distribution differences between common and rare species in large metacommunities, but motile species show lower rates of change than sessile species. In small metacommunities both common and rare species have similar changes through time; the dichotomous distinction of common and rare species is not present. Edge effects in metacommunities affect species' changes in distribution and abundance. In large metacommunities diversity is higher in edge habitats relative to small metacommunities during early succession. However, edge effects benefit motile species over time in small metacommunities showing a rapid increase in diversity. Individual mobility is sensitive to regional community size and allows individuals to sort among different communities. In contrast, sessile species do not show this edge effect. Metacommunity theory is a useful framework for understanding spatially structured communities, but the natural history of coexisting species cannot be ignored.

Crystal lattice tilting in prismatic calcite

We analyzed the calcitic prismatic layers in Atrina rigida (Ar), Haliotis iris (Hi), Haliotis laevigata (HL), Haliotis rufescens (Hrf), Mytilus californianus (Mc), Pinctada fucata (Pf), Pinctada margaritifera (Pm) shells, and the aragonitic prismatic layer in the Nautilus pompilius (Np) shell. Dramatic structural differences were observed across species, with 100-μm wide single-crystalline prisms in Hi, HL and Hrf, 1-μm wide needle-shaped calcite prisms in Mc, 1-μm wide spherulitic aragonite prisms in Np, 20-μm wide single-crystalline calcite prisms in Ar, and 20-μm wide polycrystalline calcite prisms in Pf and Pm. The calcite prisms in Pf and Pm are subdivided into sub-prismatic domains of orientations, and within each of these domains the calcite crystal lattice tilts gradually over long distances, on the order of 100 μm, with an angle spread of crystal orientation of 10-20°. Furthermore, prisms in Pf and Pm are harder than in any other calcite prisms analyzed, their nanoparticles are smaller, and the angle spread is strongly correlated with hardness in all shells that form calcitic prismatic layers. One can hypothesize a causal relationship of these correlated parameters: greater angle spread may confer greater hardness and resistance to wear, thus providing Pf and Pm with a structural advantage in their environment. This is the first structure-property relationship thus far hypothesized in mollusk shell prisms.

Evaluation of strengthening mechanisms in calcite single crystals from mollusk shells

Biogenic single-crystal calcite is often reported to be harder and tougher than geologic calcite in the form of Iceland spar. However, the mechanistic origins of the superior mechanical properties of the biogenic materials are still debated. We investigate the hardness and modulus of biogenic calcite from the prismatic layer of the mollusk Atrina rigida compared with a pure geologic calcite, Iceland spar. On the {001} face, biogenic calcite is found to be 50–70% harder than geologic calcite. This range is due to the fact that changes in azimuthal angle of the indenter tip lead to a hardness variation of ∼20% in A. rigida but only ∼7% in Iceland spar. The higher hardness and increased anisotropy of biogenic calcite could be accounted for by hardening mechanisms based on hindered dislocation motion rather than crack deflection.

The effect of magnesium substitution on the hardness of synthetic and biogenic calcite

Abstract

Mollusk Shell Nacre Ultrastructure Correlates with Environmental Temperature and Pressure

Nacre, or mother-of-pearl, the tough, iridescent biomineral lining the inner side of some mollusk shells, has alternating biogenic aragonite (calcium carbonate, CaCO(3)) tablet layers and organic sheets. Nacre has been common in the shells of mollusks since the Ordovician (450 million years ago) and is abundant and well-preserved in the fossil record, e.g., in ammonites. Therefore, if any measurable physical aspect of the nacre structure was correlated with environmental temperatures, one could obtain a structural paleothermometer of ancient climates. Using X-ray absorption near-edge structure (XANES) spectroscopy, Photoelectron emission spectromicroscopy (PEEM), and X-ray linear dichroism we acquired polarization-dependent imaging contrast (PIC) maps of pristine nacre in cross-section. The new PIC-map data reveal that the nacre ultrastructure (nacre tablet width, thickness, and angle spread) is species-specific in at least eight mollusk species from completely different environments: Nautilus pompilius, Haliotis iris, Haliotis rufescens, Bathymodiolus azoricus, Atrina rigida, Lasmigona complanata, Pinctada margaritifera, and Mytilus californianus. Nacre species-specificity is interpreted as a result of adaptation to diverging environments. We found strong correlation between nacre crystal misorientations and environmental temperature, further supported by secondary ion mass spectrometry measurements of in situ δ(18)O in the nacre of one shell. This has far-reaching implications: nacre texture may be used as a paleothermometer of ancient climate, spanning 450 million years of Earth's history.

Cooperative Modulation of Mineral Growth by Prismatic-Associated Asprich Sequences and Mg(II)

Cooperative effects of magnesium ions and acidic polypeptides originating from a family of proteins known as Asprich (mollusk Atrina rigida) were studied. In our previous studies, these two acidic polypeptides were found to be effective in controlling the morphology of the calcium carbonate mineral, the main inorganic constituent of prismatic layer of the mollusk shell. Since these Asprich sequences are believed to contain a putative magnesium binding domain, the morphology-controlling effects were further investigated with the addition of magnesium ions. The mineral morphology was dramatically changed by the combined influence of each polypeptides and the magnesium ions, substantiating the recognized importance of magnesium in the formation of calcium carbonate-based biominerals.

Oligomer formation, metalation, and the existence of aggregation-prone and mobile sequences within the intracrystalline protein family, Asprich

The formation of crack propagation—resistant single crystal calcite in the prismatic layer of the mollusk shell involves the participation of a number of different proteins, some of which form intracrystalline organic inclusions. One protein family, Asprich (Atrina rigida), participate in the ACC formation/transformation process and become occluded within calcite. However, these two phenomena are poorly understood. Here, we experimentally establish that the Asprich “3” protein oligomerizes in solution over a wide pH range and that this oligomerization process is enhanced by the presence of Ca2+ ions, which form complexes with this protein as verified by ESI-MS. Bioinformatics analyses confirm that intrinsic disorder and unstable interactive domains constitute the entire length of each Asprich sequence. In addition, an amyloid- or prion-like aggregation-prone region was identified within the highly conserved 12 AA N-terminal sequence, KPVFKRSLSDPS. Together, these bioinformatics findings suggest that different sequence elements of the Asprich family contribute to the observed oligomerization process. Solution NMR studies of Asprich “3” protein assemblies document that portions of the 12 AA N-terminal domain are mobile within these assemblies. This mobility is driven by Pro imido ring cis-trans isomerization at P11, and this induces multiple conformational states for V3, R6, and L8 which lie upstream of this Pro residue. This Asprich “3” Pro-induced conformational exchange phenomena parallels other findings obtained for self-assembling nacre layer- and tooth enamel-specific biomineralization proteins. In conclusion, we have identified the presence of self-association, unusual sequence behavior, and ion clustering phenomena in Asprich “3” and we believe that these are key factors in the ability of Asprich proteins to control ACC formation and form organic inclusions that resist crack propagation.

Transmembrane myosin chitin synthase involved in mollusc shell formation produced in Dictyostelium is active

Several mollusc shells contain chitin, which is formed by a transmembrane myosin motor enzyme. This protein could be involved in sensing mechanical and structural changes of the forming, mineralizing extracellular matrix. Here we report the heterologous expression of the transmembrane myosin chitin synthase Ar-CS1 of the bivalve mollusc Atrina rigida (2286 amino acid residues, M.W. 264 kDa/monomer) in Dictyostelium discoideum, a model organism for myosin motor proteins. Confocal laser scanning immunofluorescence microscopy (CLSM), chitin binding GFP detection of chitin on cells and released to the cell culture medium, and a radiochemical activity assay of membrane extracts revealed expression and enzymatic activity of the mollusc chitin synthase in transgenic slime mold cells. First high-resolution atomic force microscopy (AFM) images of Ar-CS1 transformed cellulose synthase deficient D. discoideum dcsA − cell lines are shown.

Calcite Prisms from Mollusk Shells (Atrina Rigida): Swiss‐cheese‐like Organic–Inorganic Single‐crystal Composites

AbstractBiogenic single‐crystal composites, such as sea urchin spines and calcitic prisms from mollusk shells, contain organic macromolecules inside of inorganic single‐crystal matrices. The nanoscale internal structure of these materials, however, is poorly understood, especially how the biomacromolecules are distributed within the crystals without significantly disrupting the crystalline lattice. Here, annular dark‐field scanning transmission electron microscopy and electron tomography reveal, in three dimensions, how biomacromolecules are distributed within the calcitic prisms from Atrina rigida shells. Disk‐like nanopatches, whose scattering intensity is consistent with organic inclusions, are observed to be anisotropically arranged within a continuous, single‐crystalline calcite matrix. These nanopatches are preferentially aligned with the (000l) planes of calcite. Along the crystallographic c‐axis, there are alternating organic‐rich and ‐poor regions on a length scale of tens of nanometers, while, in the ab plane, the distribution of nanopatches is more random and uniform. The structural features elucidated in this work have relevance to understanding the structure–property relationships and formation mechanisms of biominerals, as well as to the development of bio‐inspired strategies to extrinsically tune the properties of single‐crystals.

Intrinsically Disordered Mollusk Shell Prismatic Protein That Modulates Calcium Carbonate Crystal Growth

The formation of calcite prism architecture in the prismatic layer of the mollusk shell involves the participation of a number of different proteins. One protein family, Asprich, has been identified as a participant in amorphous calcium carbonate stabilization and calcite architecture in the prismatic layer of the mollusk, Atrina rigida . However, the functional role(s) of this protein family are not fully understood due to the fact that insufficient quantities of these proteins are available for experimentation. To overcome this problem, we employed stepwise solid-phase synthesis to recreate one of the 10 members of the Asprich family, the 61 AA single chain protein, Asprich "3". We find that the Asprich "3" protein inhibits the formation of rhombohedral calcite crystals and induces the formation of round calcium carbonate deposits in vitro that contain calcite and amorphous calcium carbonate (ACC). This mineralization behavior does not occur under control conditions, and the formation of ACC and calcite is similar to that reported for the recombinant form of the Asprich "g" protein. Circular dichroism studies reveal that Asprich "3" is an intrinsically disordered protein, predominantly random coil (66%), with 20-30% β-strand content, a small percentage of β-turn, and little if any α-helical content. This protein is not extrinsically stabilized by Ca(II) ions but can be stabilized by 2,2,2-trifluoroethanol to form a structure consisting of turn-like and random coil characteristics. This finding suggests that Asprich "3" may require other extrinsic interactions (i.e., with mineral or ionic clusters or other macromolecules) to achieve folding. In conclusion, Asprich "3" possesses in vitro functional and structural qualities that are similar to other reported for other Asprich protein sequences.

Hydroxyapatite synthesis from biogenic calcite single crystals into phosphate solutions at ambient conditions

Hydroxyapatite is one of the most important bone substitute biomaterials. Here, it has been successfully overgrown on biogenic seed crystals at ambient conditions. Single crystals of calcite from Atrina rigida, Paracentrotus lividus and Heterocentrotus mammillatus have been soaked in phosphate solution with different concentrations and pHs for 2 months. X-ray powder diffraction, scanning electron microscopy and Fourier transform infrared spectroscopy have been used to characterize soaking precipitates. The results show that the conversion of calcite to hydroxyapatite occurs to an extent which depends on composition and morphology of seed crystals, and starting concentration and pH of phosphate solutions. In the same experimental conditions, synthetic calcite single crystals did not convert to hydroxyapatite. The morphological observations suggest for hydroxyapatite formation, a mechanism that involves a superficial dissolution of calcite and a subsequently overgrowth of hydroxyapatite. Moreover, the final architectural assembly of the hydroxyapatite crystals resembles the shape of the starting biogenic seed crystals.

Biomechanics of byssal threads outside the Mytilidae: Atrina rigida and Ctenoides mitis

The byssus is the set of proteinaceous threads widely used by bivalves to attach themselves to the substrate. Previous researchers have focused on a single byssate family, the Mytilidae. However, the properties of byssal threads from species outside this family are of interest - first, because evolutionary patterns are only detectable if species from a range of taxa are examined, and second, because recent biomimetic research efforts would benefit from a wider range of ;mussel glue' exemplars. In the present study, we measured the mechanical properties of the byssal threads of two species outside the Mytilidae, the pen shell Atrina rigida Lightfoot and the flame ;scallop' Ctenoides mitis Lamarck. The mechanical properties of their byssal threads were significantly different from those of mytilids. For instance, the byssal threads of both species were significantly weaker than mytilid threads. Atrina rigida threads were significantly less extensible than mytilid threads, while C. mitis threads exhibited the highest extensibility ever recorded for the distal region of byssal threads. However, there were also interesting similarities in material properties across taxonomic groups. For instance, the threads of A. rigida and Modiolus modiolus Linnaeus both exhibited a prominent double-yield behavior, high stiffness combined with low extensibility, and similar correlations between stiffness and other thread properties. These similarities suggest that the thread properties of some semi-infaunal species may have evolved convergently. Further research on these patterns, along with biochemical analysis of threads which exhibit unusual properties like double-yield behavior, promises to contribute to both evolutionary biology and materials engineering.