Roni J. Kingsley

MULTIMINERAL CALCAREOUS DEPOSITS IN THE MARINE ALGA ACETABULARIA ACETABULUM (CHLOROPHYTA; DASYCLADACEAE)

The ultrastructure and mineralogy of cultured and wild specimens of the green algae Acetabularia acetabulum were studied. Sites of calcification were clarified using TEM and SEM. Minerology was determined via electron diffraction, energy‐dispersive x‐ray microanalysis, and Fourier transform infrared spectral analyses of the calcareous deposits. These deposits were found covering the surface and within the cell wall of the cultured juveniles and adult wild specimens of A. acetabulum. Deposits within and directly over the wall were amorphous calcium carbonate (ACC) in granular form. In‐wall and outer cell wall ACC appear to develop independently from each other. The formation of the in‐wall ACC may be mediated by the wall structure and/or chemistry, whereas mucin may be the factor mediating formation of outer cell wall ACC. In contrast to the stable in‐wall ACC, outer cell wall ACC could be transitory, transforming into aragonite. In cultured cells, a small amount of monohydrocalcite crystals and calcite were found. It is possible that these were transformation products of ACC. In wild specimens, whewellite (calcium oxalate monohydrate) was present inside and outside of the cap wall. It is unclear if whewellite is present within the cytoplasm. Local and temporal differences in the microenvironments for the multiphase mineralization are suggested.

The dynamics of spicule calcification in whole colonies of the gorgonian Leptogorgia virgulata (Lamarck) (Coelenterata: Gorgonacea)

The dynamics of calcification was examined in fractionated segments throughout entire colonies of the gorgonian Leptogorgia virgulata (Lamarck). The uptake of 45Ca, 3H-aspartic acid and 3H-thymidine was evaluated in isolated fractions including tissues, spicules and axes. Polyp number and distribution, axis weight and spicule type and size were also examined. The highest rates of uptake for all labeled materials were in the branch tips, indicating that the greatest degree of organic matrix formation, subsequent spicule formation and cell division occur in this apical region of the colony. Ca uptake decreased from the branch tips to the mid-portions of colonies, followed by an increase toward the bases. This latter rise in uptake is due to the increase in size and density of spicules which causes an increase in isotopic exchange. At branch junctions, the predominant cellular activity appears to be spicule formation vs. cell division. Axes of apical regions of branches displayed high rates of Ca uptake supporting its active role in the calcification process. The number and distribution of polyps, as well as spicule types, was variable among colonies indicating individual variability and/or the impact of varying environmental conditions. Data indicate that, although spicule formation occurs at its highest rate at branch tips, it occurs throughout the remainder of the colony as well.

Seasonal localization of a collagenous protein in the organic matrix of spicules from the gorgonian Leptogorgia virgulata (Cnidaria: Gorgonacea)

Spicules of the gorgonian Leptogorgia virgulata possess an insoluble matrix fraction that is predominantly collagenous in summer months. This collagenous component is largely absent in winter months. Using an antibody directed against the 140 kD collagenous protein (CP) of the insoluble matrix, immuno-gold labelling was employed to localized this protein at the transmission electron-microscopy level throughout the year, and in different areas of the gorgonian colonies. Within the tip regions, the 140 kD CP varied throughout the year in the spicules, electron-dense bodies (EDBs) of scleroblasts, polyp vesicles, desmocytes and axes. In the mid and base regions, the 140 kD CP varied throughout the year in the spicules, EDBs and lysosomes of scleroblasts, desmocytes and axes. This variation in the location and density of the label suggests a dynamic annual cycling of the collagenous component of the insoluble matrix. EDBs may transport a collagenous component of the matrix to the spicule-forming vacuole. A component of the 140 kD CP may be transported and/or degraded by polyp vesicles and lysosomes, respectively. The pattern of labelling of the axial region suggests that translocation and storage of a component of the collagenous protein may occur. Environmental factors may be responsible for the triggering of matrix cycling.

Formation and morphology of epidermal sclerites from a deep-sea hydrothermal vent solenogaster (Helicoradomenia sp., Solenogastres, Mollusca)

The deep-sea hydrothermal vent solenogaster Helicoradomenia is covered with calcium carbonate sclerites. Light and electron microscopy reveal varying morphologies of these sclerites. Many sclerites have hollow tips and/or are pitted and etched. Bacteria are found on and in sclerites. Initial sclerite formation occurs in an extracellular crystalline chamber formed by the invagination of a cuboidal basal cell of the columnar microvillus mantle epithelium. As the sclerite grows, it fills the crystalline chamber resulting in direct contact with the microvilli of both the basal cell and neighboring secondary sclerite-forming cells. These cells shape a collar around the base of the growing sclerite. As growth continues, the sclerite-forming cells stretch around the sclerite forming a sheath in which the base of the sclerite resides. Mature sclerites grow through the cuticle into the external environment. The erosion pattern of sclerites reveals a less stable inner medullary region and a harder outer cortical region. This points to a secondary character state, where foremost hollow acicular sclerites develop into solid sclerites. This is in agreement with the systematic position of the genus Helicoradomenia within Simrothiellidae, a taxon typically with hollow sclerites.

Calcification in Octocorals

The octocorals are colonial cnidarians, many of which contain calcareous spicules in the mesoglea and, in some species, in the axis as well. The octocorals are excellent organisms for probing the mechanisms of biomineralization and demineralization, and the processes of calcification of spicules have been investigated extensively in recent years, especially in Leptogorgia virgulata. In this review, octocoral spicule formation is discussed relative to (1) the characteristics of scleroblasts, the spicule-forming cells, (2) calcium transport by the colonies, (3) rate of calcification, (4) role of carbonic anhydrase, (5) hormonal control of calcification, and (6) organic matrices and their possible functions in spicule formation.

Ca-binding glycoprotein from the spicules of the octocoral Leptogorgia virgulata

Abstract 1. 1. A Ca-binding component (CaBC) was isolated from the soluble matrix of spicules of the gorgonian Leptogargia virgulata. 2. 2. The CaBC is a glycoprotein with apparent mol. wt between 70,000 and 100,000, and contains very high levels of aspartate. 3. 3. The CaBC probably plays an important role in the formation of the coral spicules.

Investigating Bacterial Endosymbiosis in Leptogorgia virgulata Using a Molecular and SEM Approach

The common sea-whip, Leptogorgia virgulata is reported to be asymbiotic in terms of endosymbiotic dinoflagellates. Though we tend to believe that L. virgulata uses possible bacterial symbionts as a mechanism to reduce acidic environments within the gorgonian tissue to form calcium carbonate spicules. This study investigates urease production by bacteria within L. virgulata to reduce acidity. The goal of the study is to characterize possible bacterial symbionts and using SEM determine locality within L. virgulata tissue. Possible microbe symbionts were isolated from L. virgulata tissue through a series of dilutions where bacteria colonies unique to surface bacteria were cultured and tested for urease production. Bacteria that were positive for urease production were then prepared for SEM analysis and a series of magnification SEM references for cultured bacteria were compared to freeze fractured L. virgulata SEM prepared samples to determine possible microbe symbiont location in the gorgonian tissue. Possible bacterial endosymbionts were characterized using univ518r primers typical for identifying universal bacteria. Preliminary molecular data suggests there are unique microbes inhabiting L. virgulata tissue, though their location is still being investigated. Microsc Microanal 15(Suppl 2), 2009 Copyright 2009 Microscopy Society of America doi: 10.1017/S1431927609098675 836