The mechanism of calcification and its relation to photosynthesis and respiration in the scleractinian coral <Emphasis Type="Italic"> Galaxea fascicularis</Emphasis>

The mechanism of calcification and its relation to photosynthesis and respiration were studied with Ca²⁺, pH and O₂ microsensors using the scleractinian coral Galaxea fascicularis. Gross photosynthesis (Pg), net photosynthesis (Pn) and dark respiration (DR) were measured on the surface of the coral. Light respiration (LR) was calculated from the difference between Pg and Pn. Pg was about seven times higher than Pn; thus, respiration consumes most of the O₂ produced by the algal symbiont's photosynthesis. The respiration rate in light was ca. 12 times higher than in the dark. The coupled Pg and LR caused an intense internal carbon and O₂ cycling. The resultant product of this cycle is metabolic energy (ATP). The measured ATP content was about 35% higher in light-incubated colonies than in dark-incubated ones. Direct measurements of Ca²⁺ and pH were made on the outer surface of the polyp, inside its coelenteron and under the calicoblastic layer. The effects on Ca²⁺ and pH dynamics of switching on and off the light were followed in these three compartments. Ca²⁺ concentrations decreased in light on the surface of the polyp and in the coelenteron. They increased when the light was switched off. The opposite effect was observed under the calicoblastic layer. In light, the level of Ca²⁺ was lower on the polyp surface than in the surrounding seawater, and even lower inside the coelenteron. The concentration of calcium under the calicoblastic layer was about 0.6 mM higher than in the surrounding seawater. Thus Ca²⁺ can diffuse from seawater to the coelenteron, but metabolic energy is needed for its transport across the calicoblastic layer to the skeleton. The pH under the calicoblastic layer was more alkaline compared with the polyp surface and inside the coelenteron. This rise in pH increased the supersaturation of aragonite from 3.2 in the dark to 25 in the light, and brought about more rapid precipitation of CaCO₃. When ruthenium red was added, Ca²⁺ and pH dynamics were inhibited under the calicoblastic layer. Ruthenium red is a specific inhibitor of Ca-ATPase. The results indicated that Ca-ATPase transports Ca²⁺ across the calicoblastic layer to the skeleton in exchange for H⁺. Addition of dichlorophenyldimethylurea completely inhibited photosynthesis. The calcium dynamics under the calicoblastic layer continued; however, the process was less regular. Initial rates were maintained. We conclude that light and not energy generation triggers calcium uptake; however, energy is also needed.     

Patterns of formation and the ultrastructure of the larval skeleton of Pocillopora damicornis

The growth and development of the tissues and skeleton of settled larvae of the reef coral Pocillopora damicornis (Linnaeus), collected in December 1983 from Ko Phuket, Thailand, were investigated using light microscopy, scanning electron microscopy and transmission electron microscopy. The rate of development of larval skeletons was very variable, preventing the chronological sequencing of skeletal growth. However, four growth stages in the development of a complete larval skeleton from first settlement were identificd: Stage 1, deposition of the first elements of the basal plate upon settlement; Stage 2, completion of basal plate, and deposition of skeletal spines and ridges in positions corresponding to the septal cycles; Stage 3, formation of the corallite wall and septal and costal cycles; Stage 4, the complete larval skeleton which represented the maximum growth attained eight days after settlement. The configuration of the larval tissues, particularly the aboral ectoderm, mirrored the four developmental stages. The deposition of the larval skeleton was correlated with the metamorphosis of the aboral ectoderm from a columnar to a squamous morphology. The basal plate of the larval skeleton had two layers of crystals orientated perpendicular to each other. The architecture of the complete larval skeleton is described and compared to that of the adult skeleton of P. damicornis. The results are discussed with respect to previous concepts of the formation of the larval skeleton of scleractinian corals and coral calcification.     

In vitro establishment of continuous cell cultures and cell lines from ten colonial cnidarians

Continuous cell cultures from the ten taxa of sedentary colonial marine cnidariansStylophora pistillata, Porites lutea, Favia favus (Anthozoa, Hexacorallia, Madeporaria),Parerythropodium fulvum fulvum, Dendronephthya hemprichi, Nephthya sp.Heteroxenia fuscescence (Anthozoa, Octocorallia, Alcyonacea),Clathraria rubrinoides, Plexaura A (Anthozoa, Octocorallia, Gorgonacea) andMillepora dichotoma (Hydrozoa) were established in vitro. Primary cultures of various cell types (5 to 20 m) were obtained from colony fragments and/or planula larvae (collected in 1993 from coral reefs at Eilat, Red Sea and from the San Blas islands, Panama) using three dissociation approaches: a mechanical approach, chemical approach and a novel approach, spontaneous dissociation. Cells were cultured in a modified Leibowitz L15 medium, with 5 to 10% heat-inactivated fetal bovine serum, diluted in seawater. Cell proliferation was observed in primary cultures within 7 to 20 d following dissociation. When developed to secondary cultures, spindle-shaped cells (5 to 10 m) gradually replaced all other cell types seen in primary cultures ofS. pistillata, C. rubrinoides, P.f. fulvum andM. dichotoma. The spindle-shaped cells differentiated to several cell types when the diluted medium was replaced with a concentrated one, or when microbial contamination occurred. Secondary cultures of spindle-shaped cells from three species (S. pistillata, C. rubrinoides andM. dichotoma) were cloned. They gave rise to continuously proliferating cell lines (NIO-SPP-1, NIO-CR-1 and NIO-MD-1, respectively). A clone fromPlexaura planula, originated from one rounded cell, started to differentiate and gave rise to a heterogeneous culture. Samples of the above four cultures were frozen for future work. Rounded cells of various sizes (5 to 30 m) dominated secondary cell cultures ofH. fuscescence, D. hemprichi, Nephthya sp.,P. f. fulvum andF. favus. Several cell cultures fromP. lutea developed flattened cell interconnected by ectoplasmic networks characteristic of the eukaryotic unicellular organisms of the phylum Labyrinthulomycota. We propose that this culture methodology may be used as a ubiquitous protocol for producing tissue cultures from other cnidarians. The established cell lines may be used in a variety of disciplines in cnidarian biology and ecology.     

The biology of colonial hydroids. II. The morphology and ultrastructure of the medusa of Eirene viridula (Thecata-Leptomedusa: Campanulinidae)

The free-swimming medusae of Eirene viridula are typical marine hydroids, with distinct subdivisions into body regions, which were studied by comparison of light and electron microscopy. Particular attention is given to the epithelio-muscular and digestive muscle cells. The epidermis — apart from a few nerve cells and cnidocytes-consists of epitheliomuscular cells. At their bases these cells form a musculature and at the apex many secretory vesicles. In the velum, large expansions of the intercellular space are found. The gastrodermis — apart from the mucous and zymogen cells in the manubrium — consists of digestive muscle cells, the structure of which corresponds in principle to that of the epitheliomuscular cells. In its apex, this type of cells contains both normal coated vesicles and discoidal coated vesicles. In the solid tentacles, the digestive muscle cells form a turgescent vesicular tissue. The central nervous system consists of the exumbrellar, subumbrellar, and gastrodermal nerve rings. They are composed of ganglion cells, their processes, and intraepithelial flagella. The static organs are purely ectodermal statocysts which have only one lithocyte and only one sensory cell.     

Scanning and transmission electron microscope study of specialized mantle structures in dorid nudibranchs (Gastropoda: Opisthobranchia: Anthobranchia)

Mantle surfaces of the dorid nudibranchs Rostanga arbutus and Jorunna sp. were examined using scanning and transmission electron microscopy, and two specialized structures are described. These are caryophyllidia and mantle rim organs; the latter being described for the first time. Caryophyllidia occur in large numbers (several thousand) uniformly distributed over the entire upper surface, while the mantle rim organs, numbering only 40 to 60, are restricted to the upper mantle margin. Caryophyllidia are minute (40 to 50 m diam), erect tubercles supported internally by 4 to 7, vertical, calcareous spicules which emerge in a crown surrounding an apical knob. Caryophyllidia display a high level of spicular organization and incorporate a complex muscle system at their base. The apical knob is formed from specialized epidermis capping a sub-epithelial ganglion. The highly organized structure of the caryophyllidium indicates its potential importance as a new character in dorid taxonomy and phylogeny. Mantle rim organs (80 to 300 m diam) contain large numbers of vacuolated cells and cells containing pellet-shaped nodules.     

Ultrastructure of the corallinaceae. I. The vegetative cells of Corallina officinalis and C. cuvierii

A technique utilizing combined fixation and gentle decalcification has been employed to study the ultrastructure of the vegetative cells of the articulated calcareous coralline algae Corallina officinalis Linnaeus and C. cuvierii Lamouroux (Rhodophyta: Cryptonemiales). The epidermal cells are distinctive, with many cell wall inggrowths which pass between the chloroplasts. It is suggested that these cells function as transfer cells. The epidermal cells contain no starch, although the chloroplasts have well developed photosynthetic lamellae. Damage to these epidermal cells leads to formation of new cells by renewed division of sub-epidermal meristematic cells. The outer cortical cells have few small vacuoles and many plastids, with an extensive photosynthetic lamellar system. Deeper into the thallus, the vacuoles increase in size and free cytoplasmic starch grains occur. The medullary cells have a very large vacuole and in older tissue often appear dead. The long genicular cells have calcareous walls at either end while the wall in the middle of these cells is non-calcareous and has an inner fibrillar layer and a thin outer cuticle. In partially decalcified material, the orientation of the CaCO₃ (calcite) crystals next to the cells can clearly be seen. Immediately next to the cell the crystals are fairly small and arranged at right angles to the plasmalemma. Further away from the cell the crystal size is larger and their orientation is more random. The crystals are surrounded by organic material, and the possible rôle of this material in calcification in coralline algae is discussed.     

A chromatophore system in the hermatypic,deep-water coral Leptoseris fragilis (Anthozoa: Hexacorallia)

Ultrastructural evidence is presented of a chromatophoresystem in the zooxanthellae containing hermatypic, deep-water coral Leptoseris fragilis (Milne Edwards and Haime). It consists of multilobed cells which mainly occupy the intercellular space of the oral gastrodermis. The cellular processes are filled with electron-dense granules up to 1-m-long and 0.5-m-wide. Within the cytoplasm an elaborate system of microtubules is established. The ramifications of the pigment cells, containing the pigment granules, form a dense and nearly continuous layer close to the overlying zooxanthellae. It is speculated that host pigments may transform the violet portion of the incident light into longer wavelengths, thus increasing the photosynthetic efficiency of the zooxanthellae.     

Isolation and characterization of granules from the kidney of the bivalveMercenaria mercenaria

Intracellular and extracellular granules are found in most bivalve kidneys. We examined the composition of kidney granules from the infaunal bivalveMercenaria mercenaria (L.) collected in 1985 from Cataumet Bay, Bourne, and Waquoit Bay, Falmouth, Massachusetts, USA. Small granules are numerically dominant, but large granules dominate the samples on the basis of weight. Large granules (estimated diam. >15 m) are composed primarily of metals (28% by weight) with Mn (8%), Ca (8%), Zn (4%) and Fe (4%) as the major contributors. Non-metal elements (P, C, H, N, S) together constituted 21% of the granule dry weight, though organic content (0.02% protein, 0.3% carbohydrate and 0.5% lipid) was low. Small kidney granules (10 m diam.) and digestive gland granules fromM. mercenaria had significantly higher C, H, and N contents than larger kidney granules. Our results, taken together with those from other bivalve species, suggest that kidney granule formation and subsequent increase in size is a continuous process of lysosomal maturation, residual body release and extracellular accumulation of predominately inorganic elements.     

A basidiomycete isolated from the skeleton of<Emphasis Type="Italic"> Pocillopora damicornis</Emphasis> (Scleractinia) selectively stimulates short-term survival of coral skeletogenic cells

Endolithic fungi bore through the extracellular calcium carbonate skeleton of reef-building scleractinian corals, both healthy and dead, and effect net erosion of coral reefs. Potential fungal interactions with coral tissue were investigated using an in vitro approach suggested by earlier observations of skeletal repair cones at the site of fungal perforation in Porites sp. A fungal strain was isolated from the skeleton of a long-term culture of healthy, tissue-covered, Pocillopora damicornis Linnaeus colonies maintained in a recirculating system in Monaco. As coral soft tissue spontaneously dissociated in vitro, the skeleton became exposed and hyaline hyphae emerged radially from 15% of the total clipped branches. In this study, which was performed between January 2001 and March 2003, 35 skeleton–hypha explants were embedded in agar-based solid medium, yielding 60% hyphal growth. A fungal strain (F19-3-1) of the dominant (80%) morphology was isolated and propagated in agar-based solid medium. The strain was identified by 18S and 26S rDNA gene sequence analysis as a basidiomycete in the genus Cryptococcus. Co-cultures were used to provide experimental exposure of coral soft tissue to the fungus. The fungus extended the survival of coral cells by 2 days, selectively maintaining skeletogenic cell types. This effect may be interpreted as stimulation by the fungus of a short-term coral defense response.Communicated by J.P. Grassle, New Brunswick     

Studies on reef corals. III. Fine structural changes of calicoblast cells in Pocillopora damicornis during settling and calcification

The site of reef-coral calcification has been studied in the branching coral Pocillopora damicornis Lamarck. Electron microscopy and X-ray microprobe analysis were performed on the calicoblast epidermis of newly settled larval stages and of adult coral. During settling, the heterogeneous columnar cell composition of the planktonic larva epidermis is replaced by a simple epithelium consisting of a single cell type, the calicoblast cell. Metamorphosis appears tightly linked to settling, with cell changes occurring within hours after attachment, and is marked by the appearance of a new secretory cell. The calicoblast cell of the adult coral is extremely flattened, and interdigitates extensively with adjacent calicoblast cells. This cell possesses a featureless plasma membrane lacking microvilli or flagella. It characteristically contains large membrane-bound vesicles with homogeneously fine granular contents. Preliminary microprobe analysis indicated a higher calcium content in these vesicles than in surrounding tissue; however, not in concentrations suggesting calcium-carbonate precipitation. They may represent sites of organic matrix synthesis. The calicoblast epidermis is separated from the underlying coral skeleton by a narrow gap. This gap appeared devoid of substructure, either organic or inorganic. The coral soft tissues are attached to the skeleton by mesogleal attachment processes, the desmoidal processes. These consist of a complex fibrous network originating in the mesoglea, and inserting onto the skeleton via specialized attachment regions consisting of electron-opaque membranous plaques. Skeletogenesis in reef-corals probably occurs extra-cellularly, external to the calicoblast epidermis, by simple overgrowth of the skeleton.     

Skeleton and sclerite formation in the precious red coral Corallium rubrum

The carbonate skeleton of the gorgonian coral Corallium rubrum (L.) is composed of both a skeletal axis and numerous sclerites scattered in the mesoglea. Studies carried out on these skeletal elements and their associated tisues using microscopy and X-ray microanalysis, suggest a close relationship between the process of sclerite formation and skeletogenesis. The skeleton is surrounded by an axial epithelium composed of a single cell type. These cells associate intimately with mesogleal sclerites and scleroblasts, incorporating them into a nascent skeleton at the branch tip. Subsequent (sub-apical) growth appears to occur solely through the agency of the axis epithelial cells that serve to physically separate mesogleal sclerites and scleroblasts from contact with the axis. The epithelium is associated with the production of layered calcite crystals and irregular protuberances that constitute the mature, calcareous skeleton. Free sclerites in the mesoglea appear to be the product of multiple cells that are cytologically indistinguishable from those in the axis epithelium. Like the axis, sclerites are produced as layers of calcite crystals with irregular protuberances. The protuberances differ only slightly from those of the axis, and the skeleton is mineralogically indistinguishable from the sclerites. Thus, the skeleton of red coral is not primarily the product of fused sclerites. Instead, we suggest that the axis epithelium treats the incipient skeleton as if it were the core of a single sclerite, and conversely, that the mesogleal scleroblasts of C. rubrum constitute a fragmented axis epithelium.     

The color of the trophosome: elemental sulfur distribution in the endosymbionts of<Emphasis Type="Italic"> Riftia pachyptila</Emphasis> (Vestimentifera; Siboglinidae)

Riftia pachyptila Jones, 1981, lives in association with a chemoautotrophic, sulfide-oxidizing -Proteobacterium that occurs in a specialized organ, the trophosome. Ultrastructurally different bacterial subpopulations occur in different regions of the trophosome lobules (central rods, median small cocci, peripheral large cocci) and contain vesicles, which have been proposed to be sites of elemental sulfur storage. Differently colored trophosomes have been suggested to reflect different amounts of elemental sulfur in the tissue. In this study, the presence of elemental sulfur (S⁰) was confirmed in the vesicles of the symbionts of R. pachyptila by electron energy loss spectrography (EELS). The proportion of (two-dimensional) area occupied by sulfur vesicles in the cells was found to be strongly correlated with trophosome color, both in individuals with uniformly colored trophosomes and individuals that exhibited a gradual color change along the length of their trophosomes. Elemental sulfur content was highly variable between individuals from a single collection, suggesting a high degree of microhabitat heterogeneity within vestimentiferan aggregations.Communicated by O. Kinne, Oldendorf/Luhe     

An electron-microscope study of periostracum formation in some marine bivalves. II. The cells lining the periostracal groove

The histology and ultrastructure of the epithelial cells of the outer and middle mantle folds of the bivalves Mytilus edulis (L), Cardium edule (L), Macoma balthica (L) and Nucula sulcata Bronn are described. The cells lining the inner face of the outer fold exhibit a prominent granular endoplasmic reticulum, and Golgi complexes which are concerned with the elaboration of granules and vesicles eventually incorporated into the periostracum. A gradual reduction in the protein synthetic apparatus occurs towards the tip of the fold. Within the cells, it is proposed that the ovoid inclusion bodies are lysosomes and that they control the rate of secretion. The cells of the middle fold are cuboidal in appearance. Those of M. edulis and N. sulcata exhibit prominent granules, whereas those of C. edule and M. balthica possess vesicles. The cells of M. edulis differ from the others in possessing stout bundles of filaments, which occupy large areas of the cell and constitute a cell web. The cells of the epithelium in all cases do not appear to be implicated in periostracum formation.     

Effects of zinc on epidermal ultrastructure in the larva of Clupea harengus

In March, 1983 changes in epidermal ultrastructure were examined in Clupea harengus L. larvae hatched from eggs incubated in four zinc concentrations (0.5 2.0, 6.0 and 12.0 ppm). In addition to the outer and inner epidermal cell types described previously, a third type of cell is present. Ultrastructurally this resembles the epidermal chloride cells of Sardinops caerulea Girard, characterised by numerous mitochondria, extensive smooth endoplasmic reticulum and a free surface exposed to the environment. These cells occur on the yolk sac, head and in the regions of the trunk just above the yolk sac. In larvae treated in 12.0 ppm zinc, the cells contain fewer mitochondria with fewer cristae than those in the controls. Smooth endoplasmic reticulum is much reduced forming nodular masses and contains granular osmiophilic inclusions. In larvae hatching from eggs previously incubated in 6.0 and 12.0 ppm zinc, the epidermal cells contain more vesicles, intracellular spaces, and swollen mitochondria and show signs of necrosis. The viability of these larvae is discussed.     

Cuvierian tubules of the holothuroid Holothuria forskali (Echinodermata): a morphofunctional study

Quiescent Cuvierian tubules of Holothuria forskali (Delle Chiaje, 1823) have a pseudostratified mesothelium made of ciliated cells that cover elongated cells containing lipoproteie granules. Together these cells form conspicuous transversal folds that penetrate the underlaying connective tissue. Circular and longitudinal muscle cells occur under the tips of the mesothelial folds. These cells are presumably myoepithelial cells; they divide the connective tissue into a thin outer layer (where collagen fibers are directed perpendicularly to the tubule long axis) and a thick inner layer (where collagen fibers are arranged in helices parallel to the tubular long axis). Nerve processes are associated with the longitudinal muscles; they also occur between the spirally arranged collagen fibers. A spiral nerve runs along the tubules; it innervates the circular muscles. The tubule lumen is narrow with a highly convoluted limiting epithelium. Epithelial cells are not ciliated; they have basal processes that contain enlarged spherules including mucosubstances and proteins. While having the same diameter as quiescent tubules, elongated tubules have a much thiner wall surrounding an enlarged cylindrical lumen. The mesothelium has no adluminal cells and consists of unfolded granular cells. In contact with a solid surface, granules become a sticky secretion. Longitudinal muscle fibers disintegrate and circular fibers form conspicuous rings along the tubules. Collagen helices still occur, but are considerably elongated and flattened. Inner epithelial cells dissociate from each other and typically lack basal processes and spherules, the latter being secreted into the lumen of elongating tubules. Discharge of Cuvierian tubules results from tubule expulsion and elongation. Tubule expulsion is triggered by the opening of the anus of a contracted holothuroid and results from the rupture of the cloaca with subsequent expulsion of coelomic fluid. Tubule elongation results both from the mechanical stimulation of the tubule sphincter by hydrostatic pressure and the entry of water into the tubule. Mechanical stimulation of the sphincter results in a rapid contraction of the circular muscles that causes the disintegration of adluminal cells, the unfolding of granular cells and the stretching of inner epithelial cells. Entry of water immediately follows sphincter stimulation and complete tubule elongation. Water pressure causes the irreversible elongation of collagen helices, which results both in a considerable thinning of the tubule wall and the formation of enlarged water-filled lumen. It is suggested that slippage of collagen fibers in elongating tubules is nervously mediated.     

Micromorphology and ultrastructure of Caribbean sclerosponges

Fine structural analysis of living tissue of the sclerosponges Ceratoporella nicholsoni (Hickson) and Stromatospongia norae Hartman, collected near Discovery Bay, Jamaica, between 1984 and 1986, was carried out using transmission and scanning electron microscopy (TEM and SEM). The thick dermal membrane of these sponges is covered by exopinacocytes having a T shape in sections perpendicular to the surface. A dense, complex glycocalyx is produced at the surface of these cells. Choanocyte chambers are diplodal and unusually small. The inhalant and exhalant canals of both species are characterized by the presence of valvules, made by transverse lamellipodial processes of the endopinacocytes lining them. An abundant and diversified bacterial community occupies almost 20% of the mesohyl. A single layer of active basopinacocytes lines the mesohyl at the interface between the living tissue and the aragonitic skeleton. Basopinacocytes are presumed to be precursors of the irregular fibrillar organic matrix found in the aragonitic skeleton. Sclerocytes and spongocytes are abundant in the vicinity of the siliceous spicules. Typical lophocytes releasing smooth collagen fibrils are common in the dermal membrane as well as in the choanosome where they can be grouped in bundles. Uniquely, C. nicholsoni elaborates rough intercellular fibrils characterized by periodically spaced thickenings. The endolithic algae Ostreobium sp. is present in the most apical zones of the aragonitic skeleton, but does not seem to interfere with its development. The striking micromorphological resemblances between both species are discussed and compared to demosponges.Contribution no. 472 from the Discovery Bay Marine Laboratory, University of the West Indies, Discovery Bay, Jamaica, West Indies     

Accumulation of copper within the “Hepatopancreatic” caeca of Corophium volutator (Crustacea: Amphipoda)

The concentrations of various metals in specimens of Corophium volutator (Pallas) which have been taken from a site that receives freshwater drainage with a high concentration of copper, zinc and manganese, are compared with those in individuals from a site with a normal concentration of these metals for coastal waters. The amphipods at the contaminated site contain 259 g g⁺¹ dry weight of copper, which is three times that of individuals from the uncontaminated site. The concentrations of the other metals do not differ substantially between the two populations. The hepatopancreatic caeca of the midgut of amphipods with a high concentration of copper have numerous granules of homogeneous electron-dense material which can aggregate within the multivesicular bodies. Some of the amphipods with a lower concentration of copper have similar granules, but these are fewer in number. X-ray microporbe analysis shows that these granules contain copper and sulphur, and occasionally calcium. Various tests on the granules in frozen section indicate that they are relatively insoluble in a wide range of reagents and that the copper is probably complexed with organic matter.     

An electron-microscope study of periostracum repair in Mytilus edulis

The ultrastructural details of changes that occur in the inner face of the outer fold of Mytilus edulis during periostracum repair are described. Initially, after the periostracum is slit, increased secretory activity occurs in the cells, apparently controlled by lysosomes. Proto-ostracal material deposited outside the cells seems to be due to Golgi secretory granules, which dissociate and then re-assemble outside the cells to form an organised layer. During the later stages of repair, autophagic vacuole formation increases in the cells, possibly because of increased demands for synthetic material. When substantial amounts of proto-ostracal material are secreted and the outer fold is sealed off from the external environment, the cells return to their normal condition. During the early stages of repair, amoebocytes appear to be implicated in the laying down of proto-ostracal material.     

A brominated secondary metabolite synthesized by the cyanobacterial symbiont of a marine sponge and accumulation of the crystalline metabolite in the sponge tissue

The dictyoceratid marine sponge Dysidea herbacea (Keller, 1889) is common in shallow waters of the tropical Pacific Ocean. Polybrominated biphenyl ethers such as 2-(2,4-dibromophenyl)-4,6-dibromophenol (1) are characteristic secondary metabolites of some specimens of this sponge and may represent as much as 12% of the dry weight. We have found 1 to be deposited as conspicuous crystals throughout the sponge tissue. The dominant prokaryotic endosymbiont in the mesohyl of the sponge is a filamentous cyanobacterium (Oscillatoria spongeliae), although a vacuole-containing, heterotrophic bacterium is also present. The cyanobacteria were separated from the sponge cells and heterotrophic bacteria by flow cytometry. Coupled gas chromatography—mass spectrometry and proton nuclear magnetic-resonance spectroscopy revealed that the major brominated Compound 1 isolated from the intact symbiotic association is found in the cyanobacteria and not in the sponge cells or heterotrophic bacteria. This suggests that the production of the compound is due to the cyanobacterium, and not to the sponge or symbiotic heterotrophic bacteria, as had been suggested earlier.     

Seasonal cycle of lysosomal enzyme activities in the mantle tissue and isolated cells from the musselMytilus edulis

InMytilus edulis L., gametogenesis takes place in the mantle at the expense of the connective storage tissue. There are two main types of storage cells: vesicular (VC) cells storing large amounts of glycogen and adipogranular (ADG) cells containing large numbers of protein granules, lipid droplets and lesser amounts of glycogen. One of the ways in which stored reserves can be mobilized for gamete formation is by controlled autophagy, in which the cellular constituents are degraded by lysosomes. Mussels were collected from the Menai Strait, North Wales, and monthly measurements made, over two years (1984–1986), of the activities of lysosomal acid hydrolases (acid phosphatase,-N-acetyl-D-glucosaminidase and-glucuronidase) and Cathepsins B and L in the mantle tissue, isolated ADG cells, low-density cells and, during spawning, in the mature oocytes of female mussels. The lysosomal proteinases, Cathepsins B, L and H, were further characterised by activation with thiol compounds and inhibition with thiol blockers and by leupeptin. Because of the low activity in the mantle tissue ofM. edulis, Cathepsin H was not assayed on a seasonal basis. There was a general increase in lysosomal enzyme activity during the winter, which can be related to increased autolysis in the storage cells and to the process of maturation in the developing oocytes. The activity of Cathepsin B was highest in the ADG and low-density cells, implying an important role in proteolysis within the ADG cells. By contrast, Cathepsin L displayed the highest activity in the mature oocytes, suggesting a major function of Cathepsin L in the development and maturation of the oocytes. Two different-glucosidase activities were measured in the monthly assays, one with a pH optimum of 4.5 (acid) and the other at pH 7.5 (neutral). Highest activities of the acid-glycosidase were found in the low-density cells, but there were no significant seasonal changes in the mantle tissue as a whole. Activities of the neutral-glucosidase were low in the ADG cells and mature oocytes, but showed high activities in the mantle tissue, with marked seasonal changes that corresponded to the mobilization of glycogen reserves in the VC cells.