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Mollusca is a morphologically diverse phylum, exhibiting an immense variety of calcium carbonate structures. Proteomic studies of adult shells often report high levels of rapidly-evolving, novel shell matrix proteins (SMPs), which are hypothesized to drive shell diversification. However, relatively little is known about the phylogenetic distribution of SMPs, or about the function of individual SMPs in shell construction. To understand how SMPs contribute to shell diversification a thorough characterization of SMPs is required. Here, we build tools and a foundational understanding of SMPs in the marine gastropod species Crepidula fornicata and Crepidula atrasolea because they are genetically-enabled mollusc model organisms. First, we established a staging system of shell development in C. atrasolea for the first time. Next, we leveraged previous findings in C. fornicata combined with phylogenomic analyses of 95 metazoan species to determine the evolutionary lineage of its adult SMP repertoire. We found that 55% of C. fornicatas SMPs belong to molluscan orthogroups, with 27% restricted to Gastropoda, and only 5% restricted at the species level. The low percentage of species-restricted SMPs underscores the importance of broad-taxon sampling and orthology inference approaches when determining homology of SMPs. From our transcriptome analysis, we found that the majority of C. fornicata SMPs that were found conserved in C. atrasolea were expressed in both larval and adult stages. We then selected a subset of SMPs of varying evolutionary ages for spatial-temporal analysis using in situ hybridization chain reaction (HCR) during larval shell development in C. atrasolea. Out of the 18 SMPs analyzed, 12 were detected in the larval shell field. These results suggest overlapping larval vs. adult SMP repertoires. Using multiplexed HCR, we observed five SMP expression patterns and three distinct cell populations within the shell field. These patterns support the idea that modular

Biominerals are organic-mineral composites formed by living organisms. They are the hardest and toughest tissues in those organisms, are often polycrystalline, and their mesostructure (which includes nano- and microscale crystallite size, shape, arrangement, and orientation) can vary dramatically. Marine biominerals may be aragonite, vaterite, or calcite, all calcium carbonate (CaCO3 ) polymorphs, differing in crystal structure. Unexpectedly, diverse CaCO3 biominerals such as coral skeletons and nacre share a similar characteristic: Adjacent crystals are slightly misoriented. This observation is documented quantitatively at the micro- and nanoscales, using polarization-dependent imaging contrast mapping (PIC mapping), and the slight misorientations are consistently between 1° and 40°. Nanoindentation shows that both polycrystalline biominerals and abiotic synthetic spherulites are tougher than single-crystalline geologic aragonite. Molecular dynamics (MD) simulations of bicrystals at the molecular scale reveal that aragonite, vaterite, and calcite exhibit toughness maxima when the bicrystals are misoriented by 10°, 20°, and 30°, respectively, demonstrating that slight misorientation alone can increase fracture toughness. Slight-misorientation-toughening can be harnessed for synthesis of bioinspired materials that only require one material, are not limited to specific top-down architecture, and are easily achieved by self-assembly of organic molecules (e.g., aspirin, chocolate), polymers, metals, and ceramics well beyond biominerals.

Structural characterization of biologically formed materials is essential for understanding biological phenomena and their enviro-nment, and for generating new bio-inspired engineering concepts. For example, nacre-the inner lining of some mollusk shells-encodes local environmental conditions throughout its formation and has exceptional strength due to its nanoscale brick-and-mortar structure. This layered structure, comprising alternating transparent aragonite (CaCO3) tablets and thinner organic polymer layers, also results in stunning interference colors. Existing methods of structural characterization of nacre rely on some form of cross-sectional analysis, such as scanning or transmission electron microscopy or polarization-dependent imaging contrast (PIC) mapping. However, these techniques are destructive and too time- and resource-intensive to analyze large sample areas. Here, we present an all-optical, rapid, and nondestructive imaging technique-hyperspectral interference tomography (HIT)-to spatially map the structural parameters of nacre and other disordered layered materials. We combined hyperspectral imaging with optical-interference modeling to infer the mean tablet thickness and its disorder in nacre across entire mollusk shells from red and rainbow abalone (Haliotis rufescens and Haliotis iris) at various stages of development. We observed that in red abalone, unexpectedly, nacre tablet thickness decreases with age of the mollusk, despite roughly similar appearance of nacre at all ages and positions in the shell. Our rapid, inexpensive, and nondestructive method can be readily applied to in-field studies.

Structural characterization of biologically formed materials is essential for understanding biological phenomena and their enviro-nment, and for generating new bio-inspired engineering concepts. For example, nacre-the inner lining of some mollusk shells-encodes local environmental conditions throughout its formation and has exceptional strength due to its nanoscale brick-and-mortar structure. This layered structure, comprising alternating transparent aragonite (CaCO3) tablets and thinner organic polymer layers, also results in stunning interference colors. Existing methods of structural characterization of nacre rely on some form of cross-sectional analysis, such as scanning or transmission electron microscopy or polarization-dependent imaging contrast (PIC) mapping. However, these techniques are destructive and too time- and resource-intensive to analyze large sample areas. Here, we present an all-optical, rapid, and nondestructive imaging technique-hyperspectral interference tomography (HIT)-to spatially map the structural parameters of nacre and other disordered layered materials. We combined hyperspectral imaging with optical-interference modeling to infer the mean tablet thickness and its disorder in nacre across entire mollusk shells from red and rainbow abalone (Haliotis rufescens and Haliotis iris) at various stages of development. We observed that in red abalone, unexpectedly, nacre tablet thickness decreases with age of the mollusk, despite roughly similar appearance of nacre at all ages and positions in the shell. Our rapid, inexpensive, and nondestructive method can be readily applied to in-field studies.

Anthropogenic warming and ocean acidification are predicted to negatively affect marine calcifiers. While negative effects of these stressors on physiology and shell calcification have been documented in many species, their effects on shell mineralogical composition remains poorly known, especially over longer time periods. Here, we quantify changes in the shell mineralogy of a foundation species, Mytilus californianus, under 60 y of ocean warming and acidification. Using historical data as a baseline and a resampling of present-day populations, we document a substantial increase in shell calcite and decrease in aragonite. These results indicate that ocean pH and saturation state, not temperature or salinity, play a strong role in mediating the shell mineralogy of this species and reveal long-term changes in this trait under ocean acidification.

Nature utilizes the available resources to construct lightweight, strong and tough materials under constrained environmental conditions. The impact surface of the fast-striking dactyl club from the mantis shrimp is an example of one such composite material; the shrimp has evolved the capability to localize damage and avoid catastrophic failure from high-speed collisions during its feeding activities. Here we report that the dactyl club of mantis shrimps contains an impact-resistant coating composed of densely packed (about 88 per cent by volume) ~65-nm bicontinuous nanoparticles of hydroxyapatite integrated within an organic matrix. These mesocrystalline hydroxyapatite nanoparticles are assembled from small, highly aligned nanocrystals. Under impacts of high strain rates (around 104 s-1), particles rotate and translate, whereas the nanocrystalline networks fracture at low-angle grain boundaries, form dislocations and undergo amorphization. The interpenetrating organic network provides additional toughening, as well as substantial damping, with a loss coefficient of around 0.02. An unusual combination of stiffness and damping is therefore achieved, outperforming many engineered materials.

Ocean acidification (OA) represents a serious challenge to marine ecosystems. Laboratory studies addressing OA indicate broadly negative effects for marine organisms, particularly those relying on calcification processes. Growing evidence also suggests OA combined with other environmental stressors may be even more deleterious. Scaling these laboratory studies to ecological performance in the field, where environmental heterogeneity may mediate responses, is a critical next step toward understanding OA impacts on natural communities. We leveraged an upwelling-driven pH mosaic along the California Current System to deconstruct the relative influences of pH, ocean temperature, and food availability on seasonal growth, condition and shell thickness of the ecologically dominant intertidal mussel Mytilus californianus. In 2011 and 2012, ecological performance of adult mussels from local and commonly sourced populations was measured at 8 rocky intertidal sites between central Oregon and southern California. Sites coincided with a large-scale network of intertidal pH sensors, allowing comparisons among pH and other environmental stressors. Adult California mussel growth and size varied latitudinally among sites and inter-annually, and mean shell thickness index and shell weight growth were reduced with low pH. Surprisingly, shell length growth and the ratio of tissue to shell weight were enhanced, not diminished as expected, by low pH. In contrast, and as expected, shell weight growth and shell thickness were both diminished by low pH, consistent with the idea that OA exposure can compromise shell-dependent defenses against predators or wave forces. We also found that adult mussel shell weight growth and relative tissue mass were negatively associated with increased pH variability. Including local pH conditions with previously documented influences of ocean temperature, food availability, aerial exposure, and origin site enhanced the explanatory power of models describing

Although calcareous anatomical structures have evolved in diverse animal groups, such structures have been unknown in insects. Here, we report the discovery of high-magnesium calcite [CaMg(CO3)2] armor overlaying the exoskeletons of major workers of the leaf-cutter ant Acromyrmex echinatior. Live-rearing and in vitro synthesis experiments indicate that the biomineral layer accumulates rapidly as ant workers mature, that the layer is continuously distributed, covering nearly the entire integument, and that the ant epicuticle catalyzes biomineral nucleation and growth. In situ nanoindentation demonstrates that the biomineral layer significantly hardens the exoskeleton. Increased survival of ant workers with biomineralized exoskeletons during aggressive encounters with other ants and reduced infection by entomopathogenic fungi demonstrate the protective role of the biomineral layer. The discovery of biogenic high-magnesium calcite in the relatively well-studied leaf-cutting ants suggests that calcareous biominerals enriched in magnesium may be more common in metazoans than previously recognized.

Although calcareous anatomical structures have evolved in diverse animal groups, such structures have been unknown in insects. Here, we report the discovery of high-magnesium calcite [CaMg(CO3)2] armor overlaying the exoskeletons of major workers of the leaf-cutter ant Acromyrmex echinatior. Live-rearing and in vitro synthesis experiments indicate that the biomineral layer accumulates rapidly as ant workers mature, that the layer is continuously distributed, covering nearly the entire integument, and that the ant epicuticle catalyzes biomineral nucleation and growth. In situ nanoindentation demonstrates that the biomineral layer significantly hardens the exoskeleton. Increased survival of ant workers with biomineralized exoskeletons during aggressive encounters with other ants and reduced infection by entomopathogenic fungi demonstrate the protective role of the biomineral layer. The discovery of biogenic high-magnesium calcite in the relatively well-studied leaf-cutting ants suggests that calcareous biominerals enriched in magnesium may be more common in metazoans than previously recognized.

Ocean acidification (OA) represents a serious challenge to marine ecosystems. Laboratory studies addressing OA indicate broadly negative effects for marine organisms, particularly those relying on calcification processes. Growing evidence also suggests OA combined with other environmental stressors may be even more deleterious. Scaling these laboratory studies to ecological performance in the field, where environmental heterogeneity may mediate responses, is a critical next step toward understanding OA impacts on natural communities. We leveraged an upwelling-driven pH mosaic along the California Current System to deconstruct the relative influences of pH, ocean temperature, and food availability on seasonal growth, condition and shell thickness of the ecologically dominant intertidal mussel Mytilus californianus. In 2011 and 2012, ecological performance of adult mussels from local and commonly sourced populations was measured at 8 rocky intertidal sites between central Oregon and southern California. Sites coincided with a large-scale network of intertidal pH sensors, allowing comparisons among pH and other environmental stressors. Adult California mussel growth and size varied latitudinally among sites and inter-annually, and mean shell thickness index and shell weight growth were reduced with low pH. Surprisingly, shell length growth and the ratio of tissue to shell weight were enhanced, not diminished as expected, by low pH. In contrast, and as expected, shell weight growth and shell thickness were both diminished by low pH, consistent with the idea that OA exposure can compromise shell-dependent defenses against predators or wave forces. We also found that adult mussel shell weight growth and relative tissue mass were negatively associated with increased pH variability. Including local pH conditions with previously documented influences of ocean temperature, food availability, aerial exposure, and origin site enhanced the explanatory power of models describing

Oysters are unusual among bivalves in that they possess chambers, often filled with soft, chalky calcite, that are irregularly scattered throughout the shell. Because the function of these so-called chalky deposits is still unclear, evaluating the growth and distribution of chalk is important for elucidating the ecological function of this unique shell trait. Specimens of the Pacific oyster Magallana gigas, an oyster well known for chalk expression, were grown in Bodega Harbor, Bodega Bay, CA. At the end of an 11 month growing period, specimens were culled and selected animals were submitted for x-ray computed-tomography imaging. Three-dimensional reconstructions of oyster shells were used to assess the overall distribution of chalk, and also to better understand the relationship between chalk and other structures within the shell. Results indicate that chalky deposits underly sculptural features on the shell exterior, such as external ridges and changes in growth direction, and also that there is a relationship between chalk formation and oyster processes of cementation. Overall, chalk is useful for a cementing lifestyle because it enables morphological plasticity needed to conform to irregular substrates, but also acts as a cheap building material to facilitate rapid growth.

Species of terebratulide brachiopods have been largely characterized qualitatively on the basis of morphology. Furthermore, species-level morphological variability has rarely been analyzed within a quantitative framework. The objective of our research is to quantify morphological variation to test the validity of extant named species of terebratulide brachiopods, focusing on the lophophore-supporting structures-the "long loops." Long loops are the most distinctive and complex morphological feature in terebratellidine brachiopods and are considered to be phylogenetically and taxonomically informative. We studied eight species with problematic species identities in three genera distributed in the North Pacific: Laqueus, Terebratalia, and Dallinella. Given how geometrically complex long loops are, we generated 3D models from computed tomography (CT) scans of specimens of these eight species and analyzed them using 3D geometric morphometrics. Our goal was to determine ranges of variation and to test whether species are clearly distinguishable from one another in morphospace and statistically. Previous studies have suggested that some species might be overly split and are indistinguishable. Our results show that these extant species of terebratellidines can be reliably distinguished on the basis of quantitative loop morphometrics. Using 3D geometric morphometric methods, we demonstrate the utility of CT beyond purely descriptive imaging purposes in testing the morphometric validity of named species. It is crucial to treat species described and named from qualitative morphology as working hypotheses to be tested; many macroevolutionary studies depend upon the accurate assessment of species in order to identify and seek to explain macroevolutionary patterns. Our results provide quantitative documentation of the distinction of these species and thus engender greater confidence in their use to characterize macroevolutionary patterns among extant terebratellidine brachiopods.

Crystallization by particle attachment (CPA) of amorphous precursors has been demonstrated in modern biomineralized skeletons across a broad phylogenetic range of animals. Precisely the same precursors, hydrated (ACC-H2O) and anhydrous calcium carbonate (ACC), have been observed spectromicroscopically in echinoderms, mollusks, and cnidarians, phyla drawn from the 3 major clades of eumetazoans. Scanning electron microscopy (SEM) here also shows evidence of CPA in tunicate chordates. This is surprising, as species in these clades have no common ancestor that formed a mineralized skeleton and appear to have evolved carbonate biomineralization independently millions of years after their late Neoproterozoic divergence. Here we correlate the occurrence of CPA from ACC precursor particles with nanoparticulate fabric and then use the latter to investigate the antiquity of the former. SEM images of early biominerals from Ediacaran and Cambrian shelly fossils show that these early calcifiers used attachment of ACC particles to form their biominerals. The convergent evolution of biomineral CPA may have been dictated by the same thermodynamics and kinetics as we observe today.

Many marine calcifiers experience metabolic costs when exposed to experimental ocean acidification conditions, potentially limiting the energy available to support regulatory processes and behaviors. Decorator crabs expend energy on decoration camouflage and may face acute trade-offs under environmental stress. We hypothesized that under reduced pH conditions, decorator crabs will be energy limited and allocate energy towards growth and calcification at the expense of decoration behavior. Decorator crabs, Pelia tumida, were exposed to ambient (8.01) and reduced (7.74) pH conditions for five weeks. Half of the animals in each treatment were given sponge to decorate with. Animals were analyzed for changes in body mass, exoskeleton mineral content (Ca and Mg), organic content (a proxy for metabolism), and decoration behavior (sponge mass and percent cover). Overall, decorator crabs showed no signs of energy limitation under reduced pH conditions. Exoskeleton mineral content, body mass, and organic content of crabs remained the same across pH and decoration treatments, with no effect of reduced pH on decoration behavior. Despite being a relatively inactive, osmoconforming species, Pelia tumida is able to maintain multiple regulatory processes and behavior when exposed to environmental pH stress, which underscores the complexity of responses within Crustacea to ocean acidification conditions.

Crystallization by particle attachment (CPA) of amorphous precursors has been demonstrated in modern biomineralized skeletons across a broad phylogenetic range of animals. Precisely the same precursors, hydrated (ACC-H2O) and anhydrous calcium carbonate (ACC), have been observed spectromicroscopically in echinoderms, mollusks, and cnidarians, phyla drawn from the 3 major clades of eumetazoans. Scanning electron microscopy (SEM) here also shows evidence of CPA in tunicate chordates. This is surprising, as species in these clades have no common ancestor that formed a mineralized skeleton and appear to have evolved carbonate biomineralization independently millions of years after their late Neoproterozoic divergence. Here we correlate the occurrence of CPA from ACC precursor particles with nanoparticulate fabric and then use the latter to investigate the antiquity of the former. SEM images of early biominerals from Ediacaran and Cambrian shelly fossils show that these early calcifiers used attachment of ACC particles to form their biominerals. The convergent evolution of biomineral CPA may have been dictated by the same thermodynamics and kinetics as we observe today.

Species of terebratulide brachiopods have been largely characterized qualitatively on the basis of morphology. Furthermore, species-level morphological variability has rarely been analyzed within a quantitative framework. The objective of our research is to quantify morphological variation to test the validity of extant named species of terebratulide brachiopods, focusing on the lophophore-supporting structures-the "long loops." Long loops are the most distinctive and complex morphological feature in terebratellidine brachiopods and are considered to be phylogenetically and taxonomically informative. We studied eight species with problematic species identities in three genera distributed in the North Pacific: Laqueus, Terebratalia, and Dallinella. Given how geometrically complex long loops are, we generated 3D models from computed tomography (CT) scans of specimens of these eight species and analyzed them using 3D geometric morphometrics. Our goal was to determine ranges of variation and to test whether species are clearly distinguishable from one another in morphospace and statistically. Previous studies have suggested that some species might be overly split and are indistinguishable. Our results show that these extant species of terebratellidines can be reliably distinguished on the basis of quantitative loop morphometrics. Using 3D geometric morphometric methods, we demonstrate the utility of CT beyond purely descriptive imaging purposes in testing the morphometric validity of named species. It is crucial to treat species described and named from qualitative morphology as working hypotheses to be tested; many macroevolutionary studies depend upon the accurate assessment of species in order to identify and seek to explain macroevolutionary patterns. Our results provide quantitative documentation of the distinction of these species and thus engender greater confidence in their use to characterize macroevolutionary patterns among extant terebratellidine brachiopods.

The eastern oyster (Crassostrea virginica) is a foundation species providing significant ecosystem services. However, the roles of oyster microbiomes have not been integrated into any of the services, particularly nitrogen removal through denitrification. We investigated the composition and denitrification potential of oyster microbiomes with an approach that combined 16S rRNA gene analysis, metabolic inference, qPCR of the nitrous oxide reductase gene (nosZ), and N2 flux measurements. Microbiomes of the oyster digestive gland, the oyster shell, and sediments adjacent to the oyster reef were examined based on next generation sequencing (NGS) of 16S rRNA gene amplicons. Denitrification potentials of the microbiomes were determined by metabolic inferences using a customized denitrification gene and genome database with the paprica (PAthway PRediction by phylogenetIC plAcement) bioinformatics pipeline. Denitrification genes examined included nitrite reductase (nirS and nirK) and nitrous oxide reductase (nosZ), which was further subdivided by genotype into clade I (nosZI) or clade II (nosZII). Continuous flow through experiments measuring N2 fluxes were conducted with the oysters, shells, and sediments to compare denitrification activities. Paprica properly classified the composition of microbiomes, showing similar classification results from Silva, Greengenes and RDP databases. Microbiomes of the oyster digestive glands and shells were quite different from each other and from the sediments. The relative abundance of denitrifying bacteria inferred by paprica was higher in oysters and shells than in sediments suggesting that oysters act as hotspots for denitrification in the marine environment. Similarly, the inferred nosZI gene abundances were also higher in the oyster and shell microbiomes than in the sediment microbiome. Gene abundances for nosZI were verified with qPCR of nosZI genes, which showed a significant positive correlation (F1,7 = 14.7, p = 6.0x10-3, R2 = 0.

The eastern oyster (Crassostrea virginica) is a foundation species providing significant ecosystem services. However, the roles of oyster microbiomes have not been integrated into any of the services, particularly nitrogen removal through denitrification. We investigated the composition and denitrification potential of oyster microbiomes with an approach that combined 16S rRNA gene analysis, metabolic inference, qPCR of the nitrous oxide reductase gene (nosZ), and N2 flux measurements. Microbiomes of the oyster digestive gland, the oyster shell, and sediments adjacent to the oyster reef were examined based on next generation sequencing (NGS) of 16S rRNA gene amplicons. Denitrification potentials of the microbiomes were determined by metabolic inferences using a customized denitrification gene and genome database with the paprica (PAthway PRediction by phylogenetIC plAcement) bioinformatics pipeline. Denitrification genes examined included nitrite reductase (nirS and nirK) and nitrous oxide reductase (nosZ), which was further subdivided by genotype into clade I (nosZI) or clade II (nosZII). Continuous flow through experiments measuring N2 fluxes were conducted with the oysters, shells, and sediments to compare denitrification activities. Paprica properly classified the composition of microbiomes, showing similar classification results from Silva, Greengenes and RDP databases. Microbiomes of the oyster digestive glands and shells were quite different from each other and from the sediments. The relative abundance of denitrifying bacteria inferred by paprica was higher in oysters and shells than in sediments suggesting that oysters act as hotspots for denitrification in the marine environment. Similarly, the inferred nosZI gene abundances were also higher in the oyster and shell microbiomes than in the sediment microbiome. Gene abundances for nosZI were verified with qPCR of nosZI genes, which showed a significant positive correlation (F1,7 = 14.7, p = 6.0x10-3, R2 = 0.

Marine organisms have developed a wide variety of protective strategies to thrive in their native environments. These biological materials, although formed from simple biopolymer and biomineral constituents, take on many intricate and effective designs. The specific environmental conditions that shape all marine organisms have helped modify these materials into their current forms: complete hydration, and variation in hydrostatic pressure, temperature, salinity, as well as motion from currents and swells. These conditions vary throughout the ocean, being more consistent in the pelagic and deep benthic zones while experiencing more variability in the nearshore and shallows (e.g. intertidal zones, shallow bays and lagoons, salt marshes and mangrove forests). Of note, many marine organisms are capable of migrating between these zones. In this review, the basic building blocks of these structural biological materials and a variety of protective strategies in marine organisms are discussed with a focus on their structure and mechanical properties. Finally, the bioinspired potential of these biological materials is discussed.

Plankton, corals, and other organisms produce calcium carbonate skeletons that are integral to their survival, form a key component of the global carbon cycle, and record an archive of past oceanographic conditions in their geochemistry. A key aspect of the formation of these biominerals is the interaction between organic templating structures and mineral precipitation processes. Laboratory-based studies have shown that these atomic-scale processes can profoundly influence the architecture and composition of minerals, but their importance in calcifying organisms is poorly understood because it is difficult to measure the chemistry of in vivo biomineral interfaces at spatially relevant scales. Understanding the role of templates in biomineral nucleation, and their importance in skeletal geochemistry requires an integrated, multiscale approach, which can place atom-scale observations of organic-mineral interfaces within a broader structural and geochemical context. Here we map the chemistry of an embedded organic template structure within a carbonate skeleton of the foraminifera Orbulina universa using both atom probe tomography (APT), a 3D chemical imaging technique with Ångström-level spatial resolution, and time-of-flight secondary ionization mass spectrometry (ToF-SIMS), a 2D chemical imaging technique with submicron resolution. We quantitatively link these observations, revealing that the organic template in O. universa is uniquely enriched in both Na and Mg, and contributes to intraskeletal chemical heterogeneity. Our APT analyses reveal the cation composition of the organic surface, offering evidence to suggest that cations other than Ca2+, previously considered passive spectator ions in biomineral templating, may be important in defining the energetics of carbonate nucleation on organic templates.

In this paper, we describe 485 Oliva spp. shell beads recovered from four archaeological cave sites Jerimalai, Lene Hara, Matja Kuru 1, and Matja Kuru 2, located in Timor-Leste, Island Southeast Asia. While Pleistocene-aged examples of modified marine shells used for personal ornamentation are common in African and Eurasian assemblages, they are exceedingly rare in Southeast Asia, leading some researchers to suggest that these Modern Human societies were less complex than those found further west. In Timor-Leste, the lowest Oliva bead to be recovered was directly dated to ca. 37,000 cal. BP, making it the oldest piece of personal ornamentation in Southeast Asia. Morphometric, taphonomic, use wear, and residue analyses of these beads alongside modern reference specimens, and experimentally made examples indicate that the Oliva shells were modified to be strung consecutively (as in a necklace), and while their mode of production changed remarkably little over the thousands of years they were utilised, an increase in their deposition around 6,000 cal. BP suggests that there was a change in their use coinciding with sea-level stabilisation. These tiny beads demonstrate that early Island Southeast Asian societies produced the same kinds of symbolic material culture we have come to expect from the more intensively studied African/Eurasian region, and that limited sampling and poor recovery methods have biased our perspectives of this region.

Marine organisms have developed a wide variety of protective strategies to thrive in their native environments. These biological materials, although formed from simple biopolymer and biomineral constituents, take on many intricate and effective designs. The specific environmental conditions that shape all marine organisms have helped modify these materials into their current forms: complete hydration, and variation in hydrostatic pressure, temperature, salinity, as well as motion from currents and swells. These conditions vary throughout the ocean, being more consistent in the pelagic and deep benthic zones while experiencing more variability in the nearshore and shallows (e.g. intertidal zones, shallow bays and lagoons, salt marshes and mangrove forests). Of note, many marine organisms are capable of migrating between these zones. In this review, the basic building blocks of these structural biological materials and a variety of protective strategies in marine organisms are discussed with a focus on their structure and mechanical properties. Finally, the bioinspired potential of these biological materials is discussed.

Plankton, corals, and other organisms produce calcium carbonate skeletons that are integral to their survival, form a key component of the global carbon cycle, and record an archive of past oceanographic conditions in their geochemistry. A key aspect of the formation of these biominerals is the interaction between organic templating structures and mineral precipitation processes. Laboratory-based studies have shown that these atomic-scale processes can profoundly influence the architecture and composition of minerals, but their importance in calcifying organisms is poorly understood because it is difficult to measure the chemistry of in vivo biomineral interfaces at spatially relevant scales. Understanding the role of templates in biomineral nucleation, and their importance in skeletal geochemistry requires an integrated, multiscale approach, which can place atom-scale observations of organic-mineral interfaces within a broader structural and geochemical context. Here we map the chemistry of an embedded organic template structure within a carbonate skeleton of the foraminifera Orbulina universa using both atom probe tomography (APT), a 3D chemical imaging technique with Ångström-level spatial resolution, and time-of-flight secondary ionization mass spectrometry (ToF-SIMS), a 2D chemical imaging technique with submicron resolution. We quantitatively link these observations, revealing that the organic template in O. universa is uniquely enriched in both Na and Mg, and contributes to intraskeletal chemical heterogeneity. Our APT analyses reveal the cation composition of the organic surface, offering evidence to suggest that cations other than Ca2+, previously considered passive spectator ions in biomineral templating, may be important in defining the energetics of carbonate nucleation on organic templates.

Seawater pH and the availability of carbonate ions are decreasing due to anthropogenic carbon dioxide emissions, posing challenges for calcifying marine species. Marine mussels are of particular concern given their role as foundation species worldwide. Here, we document shell growth and calcification patterns in Mytilus californianus, the California mussel, over millennial and decadal scales. By comparing shell thickness across the largest modern shells, the largest mussels collected in the 1960s-1970s and shells from two Native American midden sites (∼1000-2420 years BP), we found that modern shells are thinner overall, thinner per age category and thinner per unit length. Thus, the largest individuals of this species are calcifying less now than in the past. Comparisons of shell thickness in smaller individuals over the past 10-40 years, however, do not show significant shell thinning. Given our sampling strategy, these results are unlikely to simply reflect within-site variability or preservation effects. Review of environmental and biotic drivers known to affect shell calcification suggests declining ocean pH as a likely explanation for the observed shell thinning. Further future decreases in shell thickness could have significant negative impacts on M. californianus survival and, in turn, negatively impact the species-rich complex that occupies mussel beds.

The anticipated effects of CO2-induced ocean acidification on marine calcifiers are generally negative, and include dissolution of calcified elements and reduced calcification rates. Such negative effects are not typical of crustaceans for which comparatively little ocean acidification research has been conducted. Crustaceans, however, depend on their calcified exoskeleton for many critical functions. Here, we conducted a short-term study on a common caridean shrimp, Lysmata californica, to determine the effect of CO2-driven reduction in seawater pH on exoskeleton growth, structure, and mineralization and animal cryptic coloration. Shrimp exposed to ambient (7.99 ± 0.04) and reduced pH (7.53 ± 0.06) for 21 days showed no differences in exoskeleton growth (percent increase in carapace length), but the calcium weight percent of their cuticle increased significantly in reduced pH conditions, resulting in a greater Ca:Mg ratio. Cuticle thickness did not change, indicating an increase in the mineral to matrix ratio, which may have mechanical consequences for exoskeleton function. Furthermore, there was a 5-fold decrease in animal transparency, but no change in overall shrimp coloration (red). These results suggest that even short-term exposure to CO2-induced pH reduction can significantly affect exoskeleton mineralization and shrimp biophotonics, with potential impacts on crypsis, physical defense, and predator avoidance.

The anticipated effects of CO2-induced ocean acidification on marine calcifiers are generally negative, and include dissolution of calcified elements and reduced calcification rates. Such negative effects are not typical of crustaceans for which comparatively little ocean acidification research has been conducted. Crustaceans, however, depend on their calcified exoskeleton for many critical functions. Here, we conducted a short-term study on a common caridean shrimp, Lysmata californica, to determine the effect of CO2-driven reduction in seawater pH on exoskeleton growth, structure, and mineralization and animal cryptic coloration. Shrimp exposed to ambient (7.99 ± 0.04) and reduced pH (7.53 ± 0.06) for 21 days showed no differences in exoskeleton growth (percent increase in carapace length), but the calcium weight percent of their cuticle increased significantly in reduced pH conditions, resulting in a greater Ca:Mg ratio. Cuticle thickness did not change, indicating an increase in the mineral to matrix ratio, which may have mechanical consequences for exoskeleton function. Furthermore, there was a 5-fold decrease in animal transparency, but no change in overall shrimp coloration (red). These results suggest that even short-term exposure to CO2-induced pH reduction can significantly affect exoskeleton mineralization and shrimp biophotonics, with potential impacts on crypsis, physical defense, and predator avoidance.

As the climate warms, species that cannot tolerate changing conditions will only persist if they undergo range shifts. Redistribution ability may be particularly variable for benthic marine species that disperse as pelagic larvae in ocean currents. The blue mussel, Mytilus edulis, has recently experienced a warming-related range contraction in the southeastern USA and may face limitations to northward range shifts within the Gulf of Maine where dominant coastal currents flow southward. Thus, blue mussels might be especially vulnerable to warming, and understanding dispersal patterns is crucial given the species' relatively long planktonic larval period (>1 month). To determine whether trace elemental "fingerprints" incorporated in mussel shells could be used to identify population sources (i.e. collection locations), we assessed the geographic variation in shell chemistry of blue mussels collected from seven populations between Cape Cod, Massachusetts and northern Maine. Across this ∼500 km of coastline, we were able to successfully predict population sources for over two-thirds of juvenile individuals, with almost 80% of juveniles classified within one site of their collection location and 97% correctly classified to region. These results indicate that significant differences in elemental signatures of mussel shells exist between open-coast sites separated by ∼50 km throughout the Gulf of Maine. Our findings suggest that elemental "fingerprinting" is a promising approach for predicting redistribution potential of the blue mussel, an ecologically and economically important species in the region.

As the climate warms, species that cannot tolerate changing conditions will only persist if they undergo range shifts. Redistribution ability may be particularly variable for benthic marine species that disperse as pelagic larvae in ocean currents. The blue mussel, Mytilus edulis, has recently experienced a warming-related range contraction in the southeastern USA and may face limitations to northward range shifts within the Gulf of Maine where dominant coastal currents flow southward. Thus, blue mussels might be especially vulnerable to warming, and understanding dispersal patterns is crucial given the species' relatively long planktonic larval period (>1 month). To determine whether trace elemental "fingerprints" incorporated in mussel shells could be used to identify population sources (i.e. collection locations), we assessed the geographic variation in shell chemistry of blue mussels collected from seven populations between Cape Cod, Massachusetts and northern Maine. Across this ∼500 km of coastline, we were able to successfully predict population sources for over two-thirds of juvenile individuals, with almost 80% of juveniles classified within one site of their collection location and 97% correctly classified to region. These results indicate that significant differences in elemental signatures of mussel shells exist between open-coast sites separated by ∼50 km throughout the Gulf of Maine. Our findings suggest that elemental "fingerprinting" is a promising approach for predicting redistribution potential of the blue mussel, an ecologically and economically important species in the region.

Engraved by J. Mulder from a specimen by Frederik Ruysch., A specimen jar with the severed arm on a human infant holding the egg of a sea turtle, the hatchling of which is popping it's head out (fig. 2); in the adjacent jar, a spotted 'Zeylanicus' fish and sea-nettles (fig. 2). Each jar is top with sea plants, shells and other marine life., Title derived from caption in text.

Engraved by J. Mulder from a specimen by Frederik Ruysch., A specimen jar with a large sea snail, 'Buccinum Guienense', (fig. 1); in the adjacent jar, a crocodile egg broken open so the fetus is visible (fig. 2). Each jar is top with sea plants, shells and other marine life., Title derived from caption in text.