Calcite and aragonite distributions in the skeletons of bimineralic bryozoans as revealed by Raman spectroscopy

Abstract. Bryozoans are among a diverse range of invertebrates capable of secreting calcium carbonate skeletons. Relatively little is known about biomineralization in bryozoans, despite the importance of understanding biomineralization processes for nanotechnology and the threats imposed by ocean acidification on organisms having calcareous skeletons. Ten species of cheilostome bryozoans that are reported to have bimineralic skeletons of calcite and aragonite are studied here using Raman spectroscopy. This technique allowed identification of the two mineral phases at submicron spatial resolution, allowing the distributions of calcite and aragonite within bryozoan skeletons to be determined with unprecedented precision. Confirming previous findings based on the use of chemical stains, most of the bimineralic species analyzed exhibited a calcitic skeletal framework, composed of basal, vertical, and inner frontal walls, having aragonite deposited subsequently onto the outer surfaces of the frontal walls. In one species ( Odontionella cyclops ), aragonite formed the superstructure above the autozooids, and in two others, traces of aragonite were detected on the undersides of the frontal shields. Using Raman spectroscopy, it was possible for the first time to determine the mineralogy of small-scale structures, including orificial rims, condyles and hinge teeth, avicularian pivotal bars and rostra, and ascopore rims and sieve plates. Even when surrounded by aragonitic frontal shields, these structures were found typically to be calcitic, the two exceptions being the aragonitic avicularia of Stylopoma inchoans and O. cyclops . Unexpectedly, the first-formed part of the basal wall at the distalmost growing edge of Pentapora foliacea was found to consist mainly of aragonite. This may point to a precursory phase of biomineralization comparable with the unusual mineralogies identified previously in the earliest-formed skeletons of members of some other invertebrate phyla.     

Skeletal Ultrastructure in the Cyclostome Bryozoan Hornera

Abstract Scanning electron microscopy of calcified walls in two species of the cyclostome bryozoan Hornera has revealed previously undescribed details of skeletal morphology and growth. The calcitic interior walls of both H. robusta MacGillivray and H. squamosa Hutton have a laminated structure. Walls are extended at distal growing edges where the formation of new crystallites is concentrated and wall fabric is nacreous or semi-nacreous. New crystallites are seeded on the surface of existing crystallites as six-sided rhombs. At the centres of the rhombs in H. robusta there are often three ‘spikes' which point towards alternate sides of the rhomb. Screw dislocations resulting in spiral overgrowths are also common at these distal wall edges. Wall thickening occurs further proximally where walls develop a regularly foliated structure of imbricated laths growing towards the colony base. Although often thought to be ubiquitous in cyclostomes, the division of walls into three layers (an inner, primary layer flanked on both sides by secondary layers) is absent in Hornera. Wall ultrastructure contrasts strongly with the lamellar–fibrous–lamellar structure recently described from cinctiporid cyclostomes. The c-axes of the crystallites are orientated perpendicular to the wall surface in Hornera, unlike cinctiporids in which they are orientated within the plane of the wall. Apparent similarities in ultrastructure suggest that Hornera may provide a good model for wall growth in extinct trepostome bryozoans.     

Evolution of larval size in cyclostome bryozoans

Cyclostomes are the only order of stenolaemate bryozoans living today. The non-feeding larvae of modern cyclostomes metamorphose on settlement to produce a calcified dome-shaped protoecium. Protoecial diameter provides a proxy for larval size. The sparse data available on living cyclostomes suggests that protoecial diameter is about one-and-a-half times greater than larval width. Here we use protoecial diameter to estimate larval sizes in fossil and Recent cyclostome species. A total of 233 protoecia were measured, 143 from Recent cyclostomes and 90 from fossil cyclostomes, of which 84 came from the Jurassic. Protoecial diameter ranged from 82.5 to 690 μm, with 89% of protoecia having diameters between 100 and 300 μm. A comparison of 30 Jurassic with 51 Recent taxa of tubuliporine cyclostomes showed a significant difference in size frequency. Although the Recent taxa have a larger size range (83–465 μm) than the Jurassic taxa (125–249 μm), Recent species have a lower mode (125–150 μm) than the Jurassic species (175–200 μm). Most Jurassic cyclostomes may therefore have had larger larvae than their extant relatives. Reduction in larval size may be a component of the previously hypothesized reduction in overall body size resulting from competitive displacement by cheilostome bryozoans.     

Crystallographic orientations of structural elements in skeletons of Syringoporicae (tabulate corals,Carboniferous): implications for biomineralization processes in Palaeozoic corals

The crystallographic orientation of structural elements in skeletons of representatives of Carboniferous Syringoporicae (Auloporida) has been analysed by scanning electron microscopy (SEM), petrographic microscopy and electron backscatter diffraction (EBSD) on specimens from the Iberian Peninsula. The skeletons of the tabulate corals of the Syringoporicae consist of biogenic calcite crystals, and their microstructure is composed of lamellae, fibres and granules, or of a combination of these. Independent of the microstructure, the c‐axis is oriented towards the lumen, quasi‐perpendicular to the growth direction of the skeleton (perpendicular to the morphological axis lamellae, parallel to fibres). Most phaceloid taxa have a turbostratic distribution, as a biogenic response to prevent the cleavage of crystals. Cerioid and some phaceloid corals, whose microstructure is conditioned by wall elements, do not exhibit turbostratic distribution. Wall elements are determined by the biology of each taxon. Holacanth septal spines are composed of fibres arranged in a cone‐shape structure, sometimes clamped to the external part of the corallite and show a complex crystallography. Monacanth septal spines are spindle shaped and composed of bundles of fibres. Tabulae are composed of lamellae. Their development and crystallographic orientation depends on the position of the epithelium in each case. Shared walls are formed by a combination of the walls of two independent corallites with a median lamina, composed of granules; these have a crystallographic orientation between that of the two corallites. The growth of the microstructure is derived by a coordinated stepping mode of growth, similar to other groups of organisms such as molluscs and scleractinians. The nucleation and formation of packages of co‐oriented microcrystals suggest a growth mode similar to mineral bridges with a competitive growth mode between each crystal. The growth pattern of corallites suggests that the growth direction is divided into two main components: a horizontal growth direction towards the lumen and a vertical direction towards the top.     

Breaking the long-standing morphological paradigm: Individual prisms in the pearl oyster shell grow perpendicular to the c-axis of calcite

Cross-sections of calcitic prismatic layers in mollusk shells, cut perpendicular to growth direction, reveal well-defined polygonal shapes of individual “grains” clearly visible by light and electron microscopy. For several kinds of shells, it was shown that the average number of edges in an individual prism approaches six during the growth process. Taking into account the rhombohedral symmetry of calcite, often presented in hexagonal axes, all this led to the long-standing opinion that calcitic prisms grow along the c-axis of calcite. In this paper, using X-ray diffraction and electron backscatter diffraction (EBSD), we unambiguously show that calcitic prisms in pearl oyster Pinctada margaritifera predominantly grow perpendicular to the c-axis. The obtained results imply that the hexagon-like habitus of growing crystallites may be not necessarily connected to calcite crystallography and, therefore, other factors should be taken into consideration. We analyze this phenomenon by comparing the organic contents in Pinctada margaritifera and Pinna nobilis shells, the later revealing regular growth of calcitic prisms along the c-axis.     

Late Campanian to late Maastrichtian bryozoans encrusting on belemnite rostra from the Aktolagay Plateau in western Kazakhstan

Twenty species belonging to fifteen genera of cyclostome and cheilostome bryozoans encrusting belemnite rostra are described from the late Campanian to Maastrichtian of the Aktolagay Plateau, in western Kazakhstan. Due to the moderate to poor preservation of the material, only four cheilostome species are identified down to the species level: Wilbertopora? besoktiensis (Voigt, 1967), ‘Aechmellina’ stenostoma Voigt, 1930, and two new species, ‘Aechmellina’ viskovae and Cheethamia aktolagayensis. All remaining species are left in open nomenclature. Type material of Wilbertopora? besoktiensis from the early Maastrichtian of the Mangyshlak Peninsula in Kazakhstan, has been re-examined. Palaeobiogeographical and implications are discussed. Cheilostomes slightly dominated over cyclostomes in the Aktolagay Plateau fauna encrusting on belemnites in terms of diversity. The dominant colony forms observed were spots and sheets.     

Carbonate mineralogy of a tropical bryozoan biota and its vulnerability to ocean acidification

Decreasing pH levels in the world’s oceans are widely recognized as a threat to marine life. Bryozoans are among several phyla that produce calcium carbonate skeletons potentially affected by ocean acidification (OA). Depending on species, bryozoan skeletons can consist of calcite, aragonite or have a bimineralic combination of these two minerals. Aragonite is generally more soluble in seawater than calcite, making aragonitic species more vulnerable to OA. Here, for the first time we use Raman spectroscopy to determine the mineral composition of a tropical bryozoan biota. Compared with bryozoan biotas from higher latitudes in which calcite predominates, aragonite was found to occur in a much higher proportion of the 22 cheilostome bryozoan species collected from the shorelines of Penang and Langkawi in Malaysia, where 46% of species are calcitic, 41% aragonitic and 13% bimineralic. All but one of the aragonitic or bimineralic species belong to the ascophorans, whereas calcitic skeletons characterized most of the anascans, many of which are primitive ‘weedy’ malacostegines. These results suggest a relatively high vulnerability of tropical bryozoan faunas to OA, with the weedier taxa likely to be least impacted.     

Computer-integrated polarisation (CIP) in the analysis of fossils: a case of study in a Palaeozoic coral (Sinopora,Syringoporicae, Carboniferous)

Computer-integrated polarisation (CIP) method has been applied satisfactorily in the study of fossils skeletons of Sinopora (tabulate coral, Auloporida and Carboniferous). A previous characterisation of sample by scanning electron microscopy, atomic force microscopy and cathodoluminescence (CL) with the purpose of distinguishing the diagenetical alteration was done. Subsequently, a crystallographic comparison between CIP and electron-backscattering diffraction has been made getting a very good correlation between both methods. The CIP method allows obtaining c-axis orientation images, pole figures, and measure and mapping the misorientation of uniaxial biominerals in recent and fossil skeletons. This technique can only be used in uniaxial biominerals (calcite, quartz and hydroxylapatite), limiting its use for biaxial or bimineralic and polimineralic biominerals. CIP method has good spatial resolution (limited by camera); in our example 90 nm. The main advantage of this technique, versus other with similar properties, is the fast acquisition of data in low and high magnifications. This method does not require special treatment of samples and can be very useful for the analysis of microstructures in thin and ultra-thin sections. CIP method detects diagenetic alterations in fossil skeletons by modifications in the inner arrangement of biominerals, which combined with CL offers valuable geochemical and crystallographic information.     

a new ichnogenus for etchings made by cheilostome bryozoans into calcareous substrates

Some encrusting cheilostome bryozoans etch a pattern of small pits into hard calcareous substrates, especially calcitic and aragonitic shells of molluscs. These patterns, herein described as Leptichnus ichnogen. nov., comprise pits which are sub-circular to elongate in cross section and are found in either uniserial ( L. dromeus isp. nov.) or multiserial arrangements ( L. peristroma isp. nov., the type species). Each pit corresponds to the location of a single zooid in the bryozoan colony. The oldest known Leptichnus is Late Cretaceous (Maastrichtian), the trace fossil first becomes common in the Cenozoic, and at least nine modern cheilostome genera produce incipient Leptichnus. Leptichnus can be the only evidence remaining of encrusting cheilostomes following taphonomic or diagenetic loss of their calcareous skeletons. The mechanism by which bryozoans etch into their calcareous substrates is unknown but is almost certain to be chemical and necessitates having windows in the basal walls of the zooids which permit contact with the substratum beneath. Etching may result in better adherence to the substrate, giving protection from abrasion and bioerosion.     

Comparative analysis of the nervous system structure of polymorphic zooids in marine bryozoans

The nervous system structure was compared for the first time in avicularia and vibracula in seven species of the cheilostome bryozoans from six families by immunohistochemical methods and confocal scanning microscopy. Regardless of significant differences in heterozooid shape, size, and position in a colony, their muscular and nervous systems have a common structure, which suggests their parallel evolution.     

Skeletal Ultrastructures in some Cerioporine Cyclostome Bryozoans

Abstract The ultrastructure of the calcareous skeleton is described in nine species of Recent cyclostome bryozoans belonging to the suborder Cerioporina. Two species of Heteropora have interior zooecial walls comprising a granular precursory layer followed by a thick layer of transverse fibres and a subordinate foliated fabric with, in mature proximal walls, a semi-nacreous layer. The remaining seven species have interior walls with no transverse fibres and instead predominantly comprise a distally-imbricated, regularly foliated fabric overlying a granular precursory layer. Older, proximal surfaces often have abundant screw dislocations, but true semi-nacre is absent. Basal walls comprise an outer finely granular precursory fabric and planar spherulitic layer, succeeded by the same ultrastructural succession seen in the interior zooecial walls of the respective groups. Exterior walled diaphragms, peristomes and gonozooids similarly comprise an external fabric of planar spherulitic calcite, lined internally by the predominant fabric seen in the interior walls. Ultrastructurally, therefore, cerioporines may be split into two groups with different fabric suites, the first resembling cinctiporids and many tubuliporines in having interior walls with fabrics of transverse fibres, foliated crystallites and semi-nacre; and the second resembling the rectangulates Lichenopora and Disporella in having interior walls comprising only the foliated fabric. These findings support the close phylogenetic relationship between cerioporines and other cyclostomes but suggest that the cerioporines may constitute either a diphyletic or a paraphyletic group.     

Morphological aspects of the mineral phase of the bone

By means of light, scanning and transmission electron microscopy structure of the mineral phase of human compact bone have been studied. Mechanical properties of deorganified bone samples have been determined. High compressive strength may be in connection with interactions of plate-like mineral particles at the level of hydrate shell of the bone mineral crystallites.     

Biomineralization in bryozoans: present,past and future

Many animal phyla have the physiological ability to produce biomineralized skeletons with functional roles that have been shaped by natural selection for more than 500 million years. Among these are bryozoans, a moderately diverse phylum of aquatic invertebrates with a rich fossil record and importance today as bioconstructors in some shallow‐water marine habitats. Biomineralizational patterns and, especially, processes are poorly understood in bryozoans but are conventionally believed to be similar to those of the related lophotrochozoan phyla Brachiopoda and Mollusca. However, bryozoan skeletons are more intricate than those of these two phyla. Calcareous skeletons have been acquired independently in two bryozoan clades – Stenolaemata in the Ordovician and Cheilostomata in the Jurassic – providing an evolutionary replicate. This review aims to highlight the importance of biomineralization in bryozoans and focuses on their skeletal ultrastructures, mineralogy and chemistry, the roles of organic components, the evolutionary history of bimineralization in bryozoans with respect to changes in seawater chemistry, and the impact of contemporary global changes, especially ocean acidification, on bryozoan skeletons. Bryozoan skeletons are constructed from three different wall types (exterior, interior and compound) differing in the presence/absence and location of organic cuticular layers. Skeletal ultrastructures can be classified into wall‐parallel (i.e. laminated) and wall‐perpendicular (i.e. prismatic) fabrics, the latter apparently found in only one of the two biomineralizing clades (Cheilostomata), which is also the only clade to biomineralize aragonite. A plethora of ultrastructural fabrics can be recognized and most occur in combination with other fabrics to constitute a fabric suite. The proportion of aragonitic and bimineralic bryozoans, as well as the Mg content of bryozoan skeletons, show a latitudinal increase into the warmer waters of the tropics. Responses of bryozoan mineralogy and skeletal thickness to oscillations between calcite and aragonite seas through geological time are equivocal. Field and laboratory studies of living bryozoans have shown that predicted future changes in pH (ocean acidification) combined with global warming are likely to have detrimental effects on calcification, growth rate and production of polymorphic zooids for defence and reproduction, although some species exhibit reasonable levels of resilience. Some key questions about bryozoan biomineralization that need to be addressed are identified.     

Mineralogy of Arctic bryozoan skeletons in a global context

Bryozoans are major carbonate producers in some ancient and Recent benthic environments, including parts of the Arctic Ocean. Seventy-six species of bryozoans from within the Arctic Circle have been studied using XRD to determine their carbonate mineralogies and the Mg content of the calcite. The majority of species were found to be calcitic, only four having bimineralic skeletons that combined calcite and aragonite, and none being entirely aragonitic. In almost all species, the calcite was of the low- (<4 mol% MgCO₃) or intermediate-Mg (4–11.99 mol% MgCO₃) varieties. Previous regional studies of bryozoan biomineralogy have found higher proportions of bimineralic and/or aragonitic species in New Zealand and the Mediterranean, with a greater number of calcitic species employing intermediate- and high-Mg calcite. The Antarctic bryozoan fauna, however, has a similar mineralogical composition to the Arctic. The lesser solubility of low-Mg calcite compared to both Mg calcite and aragonite in cold polar waters is most likely responsible for this latitudinal pattern. However, it is unknown to what extent environmental factors drive the pattern directly through eliciting an ecophenotypic response from the bryozoans concerned or the pattern reflects genetic adaptations by particular bryozoan clades.     

The microstructure of isotropic vapor-deposited carbon films

The structure of thin, vapor-deposited carbon films was characterized by transmission electron microscopy and electron diffraction. Selected area electron diffraction showed very weak and broad peaks, indicating that these carbons contain extremely small crystallites whose dimension in the crystallographic c-direction is about 8 to 10 a. The observed diffraction bands are (h, k, 1 = 0) type reflections, which suggests that individual crystallites consist of graphitic layer planes stacked in parallel groups but with no order between atoms in adjacent planes (turbostratic). The carbon films exhibit no preferred orientation, indicating that the small crystallites are randomly oriented in the film and that the films are therefore isotropic. The measured density (1.8 g/cm3) and the structure of the vapor-deposited carbons are accordingly similar to those of low-temperature isotropic (LTI) pyrolytic carbons.     

Investigation of the early mineralisation on collagen in dentine of rat incisors by quantitative electron spectroscopic diffraction (ESD)

The earliest crystallites in dentine appear as chains of dots in ultra-thin sections viewed by transmission electron microscopy. These dots rapidly coalesce along the longitudinal directions of the collagen microfibrils to form needle-like structures that coalesce preferentially in lateral directions to form ribbon-like or plate-like crystallites. This morphological interpretation is supported by line-scans of the corresponding zero-loss filtered electron spectroscopic diffraction patterns, which demonstrate the crystalline structure of the dentine mineral (apatite). The intensity ratio of the Debye-Scherrer rings of the characteristic Bragg-reflections (002 to 300, together with 1 or 2 unresolved reflections) shows a maximum in the region of early chain-like and needle-like crystallites, decreasing with maturation of the dentine mineral to the ribbon-plate-like crystallites. Detailed investigations using line-scans of the zero-loss filtered electron spectroscopic diffraction patterns through the dentine zone show that the intensity ratio found near the mineralisation front is repeated 3–5 times at distances of about 10–20 m. This may represent a circadian pattern of mineralisation corresponding to light microscopically visible incremental lines in dentine.     

Predation by Patiria miniata (Asteroidea) on bryozoans: Prey diversity may depend on the mechanism of succession

Summary In environments where frequent disturbances interrupt the successional process there will usually be many patches of habitat at intermediate stages of succession. It is then relevant to consider the factors which control local diversity during succession. In offshore kelp forests across the whole Southern Californian Bight settlement panels were rapidly colonised by two species of cyclostome bryozoans (Tubulipora tuba and T. pacifica); but cheilostome bryozoans eventually became dominant during succession because they were able to grow over Tubulipora spp. When abundant, Tubulipora spp. were apparently able to reduce the number of colonies, and hence the number of species, of cheilostome bryozoans settling on the panels. Thus the rapid colonisers may delay the process of succession and reduce the diversity of bryozoans during succession. In field and laboratory experiments we found that the asteroid Patiria miniata preys on Tubulipora spp. but not on cheilostome bryozoans. The predator speeds up the successional process and increases bryozoan diversity by reducing the cover of Tubulipora spp. Other ways in which predators may influence diversity during succession are discussed. The effect of predation may depend on the abundance of the prey and the mechanism of succession.     

Cenomanian cheilostome bryozoans from Devon,England

The Cenomanian witnessed a spectacular evolutionary radiation of cheilostome bryozoans, both in terms of species diversity and morphological disparity. However, Cenomanian cheilostome faunas are inadequately known. Twelve species of cheilostome bryozoans are here described from the Cenomanian Beer Head Limestone Formation of SE Devon, England, a nearshore facies of limestones and sands. Two of the cheilostome species are new – Wilbertopora manubriformis sp. nov. and Foratella cervisia sp. nov. – and five cannot be identified beyond genus level. Syntypes of Foratella forata (d’Orbigny, 1853), the Senonian type species of Foratella Canu, 1900, are illustrated using SEM and a lectotype is chosen. All of the species present are neocheilostomes, which were larval brooders. Compared with the non-brooding malacostegans that dominate pre-Cenomanian faunas, most species have avicularia and extensive frontal walls, features probably adaptive against small predators.     

Inferring larval type in fossil bryozoans

Pachut, J.F. & Fisherkeller, P. 2010: Inferring larval type in fossil bryozoans. Lethaia, Vol. 43, pp. 396–410. Larval type in extinct organisms might be recognizable because larvae of living marine invertebrates are approximately of the same size as the initial post‐larval organism. Two larval types typically occur. Planktotrophic larvae feed on other members of the plankton, potentially prolonging their larval existence and producing broad geographic distributions. Conversely, lecithotrophic larvae feed on yolk supplied by the fertilized egg, often settle quickly after release, and display more restricted distributions. However, some lecithotrophic bryozoans undergo embryonic fission forming multiple, small, polyembryonic larvae. The relationship between post‐larval size and larval type was evaluated in bryozoans by comparing the size of the ancestrula, the founding individual of a colony, to the sizes of extant planktotrophic, lecithotrophic and polyembryonic lecithotrophic larvae and ancestrulae. The sizes of larvae and ancestrulae in extant lecithotrophic and planktotrophic cheilostome (gymnolaemate) species are statistically the same. They are, however, statistically larger than the polyembryonic larvae of extant cyclostomes (stenolaemates). In turn, the sizes of cyclostome larvae are indistinguishable from the ancestrulae of extant and fossil cyclostomes, the ancestrulae of other fossil stenolaemate species measured from the literature, and the ancestrulae of three of four genera from North American Cincinnatian strata. Ancestrulae of a fourth genus, Dekayia, are the same size as cyclostome ancestrulae but are statistically smaller than the ancestrulae of other stenolaemates. With few exceptions, stenolaemates have statistically smaller larvae and ancestrulae than both lecithotrophic and planktotrophic cheilostomes. We infer that the sizes of fossil ancestrulae permit the discrimination of taxa that had polyembryonic lecithotrophic larvae from those possessing other larval types. This inference is strengthened, in several cases, by the co‐occurrence of brood chambers (gynozooecia) and restricted palaeobiogeographic distributions. The presence of cyclostomes in Early Ordovician strata suggests that polyembryony may have been acquired during the initial radiation of Class Stenolaemata. Polyembryony appears to be a monophyletic trait, but confirmation requires the demonstration that species of several stenolaemate suborders lacking skeletally expressed brood chambers possessed polyembryonic larvae. □Ancestrulae, evolution, fossil bryozoans, gynozooecia, larvae.     

Understanding the crystallographic and nanomechanical properties of bryozoans

This study examines how microscale differences in skeletal ultrastructure affect the crystallographic and nanomechanical properties of two related bryozoan species: (i) Hornera currieae, which is found at relatively quiescent depths of c. 1000 m, and (ii) Hornera robusta, which lives at depths of 50–400 m where it is exposed to currents and storm waves. Microstructural and Electron Backscatter Diffraction (EBSD) observations show that in both species the secondary walls are composed of low-Mg calcite crystallites that grow with their c-axes perpendicular to the wall. Branches in H. currieae develop a strong preferred orientation of the calcite c-axes, while in H. robusta the c-axes are more scattered. Microstructural observations suggest that the degree of scattering is controlled by the underlying morphology of the skeletons: in H. currieae the laminated branch walls are smooth and relatively uninterrupted, whereas the wall architecture of H. robusta is modified by numerous deflections, forming pustules and ridges associated with microscopic tubules. Modelling of the Young’s modulus and measurements of nanoindentation hardness indicate that the observed scattering of the crystallite c-axes affects the elastic modulus and nanohardness of the branches, and therefore controls the mechanical properties of the skeletal walls. At relatively high pressure in deep waters, the anisotropic skeletal architecture of H. currieae is aimed at concentrating elasticity normal to the skeleton wall. In comparison, in the relatively shallow and active hydrographic regime of the continental shelf, the elastically isotropic skeleton of H. robusta is designed to increase protection from external predators and stronger omni-directional currents.