Synthesis and maturation processes in the exoskeleton of the vent worm Riftia pachyptila

To investigate tube-wall maturation in Riftia pachyptila, tube-secretion experiments were performed on live individuals in pressurized aquaria during “HOT 96” cruise (East Pacific Rise, February and March 1996). Two major biochemical components of the tube, i.e. chitin and proteins, were measured during the process of tube maturation. Mean chitin and protein contents were two-fold lower in fresh (i.e. newly secreted) tube-walls than in mature tube-walls. Chitin content is related to age, i.e. to the time elapsed since the fresh tube material was extruded, and can thus provide a marker for determining tube maturation status. In contrast, variations in protein content suggest a difference in the nature of the secretion products in the upper and basal regions of the exoskeleton. The tube-secretion experiments analyzed the protein profiles of both fresh-tube material and actively secreting tissues. A protein triplet with an apparent molecular weight of ∼28 kdaltons proved useful as a marker for tracing exoskeleton protein synthesis and secretion pathways. Received: 17 February 1999 / Accepted: 10 January 2000     

Bioavailability and metabolism of hexavalent chromium compounds †

Intratracheal instillation of ⁵¹CrCl₃ in anaesthetized rabbits resulted in partial absorption. In blood, the absorbed material was entirely confined to the plasma compartment. Only trace amounts were deposited in liver and kidney. By contrast, after similar application of Na, ⁵¹CrO₄ the bulk of blood radioactivity was present in red blood cells (RBC). Substantial deposition occurred in liver and kidneys. It is concluded that Cr(VI) may enter the body unreduced via the lung and is partially deposited in cells over a prolonged period of time.

Since chromium was accumulated in liver after administration of Cr(VI) we investigated the intracellular disposition of Cr(VI) in the isolated perfused liver. No significant sex differences in chromium distribution were observed. At the end of the experiments (1 h), 60% of the applied dose (312μg Cr/liver) was located in the cytosol, whilst 14% was in the mitochondria, 9% in the microsomal pellet and 2% was associated with the nuclei. Gel chromatography of the cytosolic compartment showed that the overwhelming part of chromium was eluted in fractions with an apparent molecular weight of 6,000 dalton. These fractions exhibited absorption maxima at 410nm and 548nm. It is concluded, that cytosolic reduction might be the main intracellular redox pathway for chromates. This view was confirmed by monitoring the reaction of Cr(VI) with GSH in vitro. GSH reduced Cr(VI) without further cofactors under formation of GSH‐chromium complexes, which possibly represent major intermediates in the metabolism of Cr(VI).     

Feeding and cleaning mechanisms in the suspension feeding bivalve Mytilus edulis

Feeding and pseudofeces formation were studied in intact Mytilus edulis clearing suspensions of graphite particles, and the processes were compared with activities observed in mussels with a severed adductor muscle. Most particulate material in the stomach was present in the suspended state. Ingestion of particles in suspension could take place concurrently with the production of pseudofeces. Severing of the adductor muscle resulted in profuse secretion of mucus that gradually subsided to a constant low level. Addition of graphite particles at concentrations that did not cause mucus secretion in the intact mussels strongly stimulated secretion in the operated mussels, resulting in the accumulation of highly viscid, mucusparticle aggregates. These aggregates were mechanically stable, in contrast to the fragile pseudofeces. It is indicated that normal feeding depends upon hydromechanical mechanisms that produce highly concentrated suspensions of particles for ingestion, and that mucociliary mechanisms serve to clean the gills and other organs of the mantle cavity for excess particulate material. It is further indicated that intact mussels secrete mucus only in the amounts needed to consolidate excess particulate material, and that lesions affect the normal balance between particles in suspension and mucus secretion by enhancing the sensitivity of the mechanisms that control mucus secretion.     

Volatile substances in the pygidial secretion of gyrinid beetles (Coleoptera: Gyrinidae)

Summary Gyrinid beetles are common in freshwater habitats. They have paired pygidial glands with a secretion that contains high molecular weight substances rendering them toxic for predatory fish. In this paper we report on a laboratory study on volatile components released by three different gyrinid species when irritated. The volatile pattern had a clear difference between the different species.Gyrinus substriatus andG. aeratus, both produced 3-methyl-1-butanal and 3-methyl-1-butanol when irritated, but in quite different amounts. The third tested species,G. minutus, did not produce any substances above the detection level. It is suggested that the volatile compounds may be part of the beetles' communication and/or defence system.     

Occurrence of a crinotoxin and hemagglutinin in the skin mucus of the moray eel Lycodontis nudivomer

The tropical Indo-Pacific moray eel Lycodontis nudivomer possesses a mucous skin secretion which shows hemolytic, toxic and hemagglutinating activities. The active components were extracted from the mucous secretion with 0.02 M phosphate buffer (pH 7.5) and purified by DEAE-cellulose and Sephadex G-200 column chromatography. The component(s) responsible for the hemolytic-toxic activities differ(s) from that (those) causing the hemagglutinating activity. The hemolysin was unstable in the presence of heat, acidic and alkaline media, and several organic solvents. It was undialyzable throught a cellophane membrane, and showed an absorption maxium at 280 nm in the phosphate buffer (pH 7.5). The hemolytic activity was completely lost when treated with trypsin, indicating that the hemolysin is a protein. Histochemical examination on the epidermis of L. nudivomer revealed the presence of clubbed peculiar cells, in addition to common mucous cells, which we speculate produced the hemolytic-toxic component(s). Eight other species of morays were found to possess clubbed peculiar cells similar to those of L. nudivomer in their epidermis. There are marked differences, however, in the abundance of these cells among these muraenids.     

Dragon’s blood secretion and its ecological significance

Dragon’s blood is the name given to a red exudate produced by some plant species belonging to the genera Daemonorops, Dracaena, Croton and Pterocarpus. These are endemic to various parts of the globe. It is classified as a resin or latex depending on its mode of secretion and its chemical composition, which is species specific. This red substance functions in defence and is produced (a) constitutively and stored in preformed anatomical structures, or (b) by induction in response to traumatic events, such as mechanical injury, pathogen attack or invasion by insects. Apart from its defensive role in plants, dragon’s blood is also a valuable natural resource renowned since antiquity for its diverse medicinal properties and uses in art. Despite the great importance of dragon’s blood, our knowledge of the biological basis for its secretion is still incomplete. This review summarizes recent advances in the study of the anatomical basis for its secretion, and discusses its classification and ecological function. Bringing some clarity to these issues may also help in the commercial sourcing of dragon’s blood.     

Assessment of the role of thymol in combating chromium (VI)-induced oxidative stress in isolated rat erythrocytes in vitro

Thymol, the main phenolic compound in Thymus vulgaris, has been shown to have various biological effects. The main objective of this study was to investigate the efficacy of thymol on counteracting hexavalent chromium-induced oxidative damage in rat erythrocytes in vitro. The radical scavenging activity of thymol was examined using the 1, 1-diphenyl-2-picrylhydrazyl assay. Erythrocytes resistance to oxidative damage, lipid peroxidation, osmotic pressure, hemolysis as well as morphological alterations were evaluated in the presence of 2.5 µg thymol mL^?1 with or without 5 µmol hexavalent chromium mL^?1 of the incubation media. Results from the 1,1-diphenyl-2-picrylhydrazyl assay denoted good radical scavenging activity of thymol. Thymol caused a significant increase in superoxide dismutase and catalase activities and reduced glutathione content in erythrocytes intoxicated with hexavalent chromium. In contrast, the presence of thymol resulted in markedly less-elevated malondialdehyde levels, hemolysis, and destabilization of erythrocytes exposed to hexavalent chromium. Microscopically, thymol markedly reduced hexavalent chromium-induced morphological alterations in rat red blood cells. Conclusively, thymol counteracted hexavalent chromium-induced oxidative damage in rat erythrocytes.     

Purification,characterization and toxicity of a mannose-binding lectin from the seeds of Treculia africana plant

In this study, a mannose-specific, homodimeric lectin from the seeds of Treculia africana was purified, characterized and its adverse effects were investigated in mice. The purification protocol involved anionic exchange chromatography on DEAE-Cellulose followed by gel filtration on Sephadex G-100. The hemagglutinating activity of lectin towards human erythrocytes was sensitive to inhibition by D-mannose. Treatment of the protein with EDTA exerted no inhibitory effect; however, analysis of metal content by atomic absorption spectroscopy revealed the presence of Cu²⁺, Fe³⁺, and Mg²⁺. The results obtained showed that the lectin possesses maximum hemagglutinating activity towards human erythrocytes activity over the pH range 3–7.2 and is relatively thermostable up to 50°C. Periodic acid Schiff's (PAS) reagent staining showed that the protein was non-glycosylated while its amino acid composition analysis revealed that the protein contained 155 residues per subunit. The subunit had a minimal molecular weight of 22,139 Daltons, while the native molecular weight was estimated to be 41,000 Daltons. The lectin was found to be moderately toxic to mice with an LD₅₀ of 47.21 µg g^?1 body weight while, histopathological analysis showed no treatment related effects in any of the organs examined.     

Feeding of calanoid copepods in relation to Phaeocystis pouchetii blooms in the German Wadden Sea area off Sylt

Surface sediments from the Svartnes basin (195 m deep) in Balsfjorden, northern Norway (ca. 70°N), were partially characterized to assess the nature and origin of the organic material present and its potential nutritive value for sediment-ingesting animals. Seasonal analyses were carried out on material collected between May 1979 and August 1980 for total organic matter, organic C and N, acid-extractable amino acids and lipids extractable with chloroform:methanol. Little or no seasonal variation was seen in any of the parameters analysed. The mean apparent organic content was 9.3% of sediment dry weight, the organic C content was 2.38%, the organic N content was 0.26% and the C:N ratio was 9.1:1. Acid hydrolysis of sediment yielded 575 mg of amino acids and 41 mg of NH₃ per 100 g sediment dry weight, the composition of the amino acids being similar to that of nutritionally highquality animal or microbial protein. Glucoseamine was not detected in acid hydrolysates of sediment, consistent with the absence of chitin. Chloroform:methanol extraction yielded 133 mg of material per 100 g sediment dry weight, 62% of which was accounted for by saponifiable lipids (fatty acids) and non-saponifiable lipids present in approximately equal amounts. Fatty alcohols accounted for 30% of the non-saponifiable lipids and phytol accounted for 40% of the fatty alcohols. Small amounts of very long-chain fatty alcohols characteristic of terrestrial plants were present, but long-chain monounsaturated fatty alcohols characteristic of marine zooplankton were essentially absent. Very small quantities of long-chain ketones characteristic of marine coccolithophores were detected. The major fatty acids present in sediments were 16:0, 14:0, 16:1 (n-7) and 18:1 (n-9), and 11% of the total fatty acids were comprised of a mixture of odd-numbered straight-chain and branched-chain moieties characteristic of micro-organisms. The data point to small amounts of material characteristic of marine and terrestrial photosynthetic organisms being present in sediments at any given time, whereas material characteristic of marine zooplankton is not present. The presence of material characteristic of micro-organisms is consistent with conversion of sedimenting material into a pool of sediment micro-organisms. Although the biological availability to sediment-ingesting organisms of the esterified fatty acids and the acid-extractable amino acids in sediments remains to be assessed, the maximum amounts available are equivalent to approximately 60 mg of polar lipid and 575 mg of protein per 100 g sediment dry weight. These amounts are small in relation to the production rates in the photic zone of the fjord, implying that the bulk of the energy flow in Balsfjorden occurs in its relatively short pelagic food chains.     

Assessing feeding of a carnivorous copepod using species-specific PCR

The polymerase chain reaction (PCR) offers a sensitive and selective way to detect trace amounts of biological remnants. Here, we show that this simple molecular technique can be applied to identify prey copepods in the fecal pellets of carnivorous zooplankton. Using variation in the mitochondrial cytochrome C oxidase subunit I (mtCOI) sequence, we developed a species-specific oligonucleotide PCR primer (COI-2026) for Calanus helgolandicus. In a touch-down PCR, Calanus DNA was amplified from pellets collected from freshly incubated individuals of the carnivorous copepod Pareuchaeta norvegica. Positive results could easily be detected by agarose gel electrophoresis.Electronic Supplementary Material Supplementary material is available in the online version of this article at Communicated by M. Kühl, Helsingør     

Catfish epidermal secretions in response to threat or injury

The Arabian Gulf catfish (Arius thalassinus, Rüppell) secrete a thick layer of proteinaceous material when threatened or injured. The chemical and pharmacological properties of this material indicate that it may promote the healing of wounds. These studies suggest that the abrupt secretion of mucous-like gel in response to a perceived hazard is an externally expressed defense response; the secretion may have a primary function in healing injury beyond the commonly postulated roles of deterring predators or facilitating escape.     

Physiological and immunological conditions of wild populations of <Emphasis Type="Italic">Farfantepenaeus duorarum </Emphasis>from the campeche sound (Crustacea,Penaeidae)

To assess the health status of aquatic ecosystems, specialized tools and strategies are needed to study the changes induced in oceanic systems by human activity. The Gulf of Mexico has a tremendous ecological importance because of its biological diversity. The present paper defines baseline levels of blood metabolites and immune components for wild tropical populations of F. duorarum: an important shrimp species of the Gulf of Mexico. Osmotic pressure (OP), osmotic capability (OC), oxy hemocyanin (OxyHc), glucose, digestive gland glycogen, proteins, cholesterol, lactate, acylglycerols (AG) were used as indicators of physiological and nutritional status; phenoloxidase activity, quantification of hemocytes, as well as respiratory burst, were used to assess the immunological status. There were no differences between sexes in either live weight or physiological and nutritional conditions. Only total blood protein, oxy hemocyanin, and digestive gland glycogen showed normally distributed data. All the remaining blood parameters were right- or left-skewed. A median live weight of 9.5 g was registered for the sampled population. Median values of 955 and −91.85 mOsm/kg (OP and OC, respectively) were obtained, indicating that shrimp were slightly hypo-osmotic in relation to sea water. A proportion of 27% of OxyHc of the total protein (OxyHc/Prot) was calculated for the F. duorarum population. ProPO and PO showed no normal distribution and were not different between sexes. A right-skewed distribution was observed. Total hemocytes, hyaline (HC) and granular cells (GC) showed differences between sexes, with higher values in females than males. A similar proportion of HC (69%) and GC (30%) of total cells were observed between sexes. Basal and activated respiratory burst showed differences between sexes with higher values for females than for males. The current results provide indicators of the nutritional and immune status of F. duorarum that can be used to detect changes in trophic relations and health at a population level.     

Blooms of surf-zone diatoms along the coast of the Olympic Peninsula,Washington. X. Chemical composition of the surf diatom Chaetoceros armatum and its major herbivore,the Pacific razor clam Siliqua patula

The surf diatom Chaetoceros armatum T. West, collected from its natural habitat along the Washington coast (USA), had a large inorganic component in the form of a clay coat (consisting of clay minerals, illite and montmorillonite) surrounding the cell chains: 63% of the dry weight of the natural material collected in November was inorganic. The organic fraction was composed of 67.6% lipid, 29.7% protein, and 1.3% carbohydrate. Culture cells were likewise high in protein and lipid and low in carbohydrate. Traces of chitin found in field samples were probably a contaminant, since chitin was absent from cultured cells. This diatom species serves as a major food source throughout the year for the Pacific razor clam Siliqua patula Dixon, which inhabits these same beaches. Protein constituted 47% and lipid 42% of the dry weight of razor clam tissue. The fatty acid distribution in the diatom lipid resembled that previously reported for other diatom species; similarly, the fatty acid distribution of the clam lipid was similar to that previously reported for other bivalve molluscs. The clam fatty acids differed in chain length and degree of saturation from those extracted from its food source, indicating an active fatty acid metabolism in the clam.Contribution No. 995 from the Department of Oceanography, University of Washington, Seattle, Washington 98195, USA.     

Humoral opsonin from the colonial ascidian Botryllus schlosseri as a member of the galectin family

The synthesis and secretion of the humoral opsonin from the colonial ascidian Botryllus schlosseri was studied by using specific polyclonal antibodies that recognise both of the bands obtained after SDS-PAGE of the purified lectin. The agglutinin was revealed on the surface of agglutinated yeast and red blood cells and in the cytoplasm of the phagocytes, either uniformly distributed or localised inside the cytoplasmic projections. A previous incubation of haemocytes with the agglutinin resulted in intense labelling of the surface that disappeared in the presence of D-galactose, whereas it was not affected by treatment with proteases. These results, together with the already published data, suggest that our molecule is a new member of the galectin family. Received: 16 July 1999 / Accepted: 7 February 2000     

Isolation of novel metal-binding proteins distinct from metallothionein from spotted seatrout (Cynoscion nebulosus) and Atlantic croaker (Micropogonias undulatus) ovaries

Metal-binding proteins were isolated from ovaries of the spotted seatroutCynoscion nebulosus and the Atlantic croakerMicropogonias undulatus collected in 1988 near Port Aransas, Texas, USA. Gel-filtration analysis of spotted seatrout trout ovarian cytosolic fraction on Sephadex G-75 revealed the presence of three zincbinding protein fractions. A major zinc/calcium-binding protein fraction had a low molecular weight (M _r)(6 000 to 10 000), similar to mammalian hepatic metallothionein (MT). All the metals were displaced from this fraction following saturation with exogenous cadmium. After exposure of Atlantic croaker to 2 mg cadmium l^–1 seawater for 2 mo, the majority of the cadmium in the ovarian cytosolic fraction was associated with a similar low molecular weight protein fraction. These proteins were further purified by heat treatment and sequential acetone precipitation. Three isoforms were isolated by reversephase high-performance liquid chromatography. All the isoforms were found to be distinct from mammalian MT, based on amino acid composition. The major isoform contained low amounts of cysteine (approximately 5 residues per molecule) and aromatic amino acids, compared to high amounts of cysteine (typically 17 to 20 residues/molecule) and a lack of aromatic amino acids for mammalian MT. All the ovarian protein isoforms contained more glutamate than mammalian MT. The spotted seatrout and Atlantic croaker ovarian isoforms showed a high degree of homology with metal-binding proteins isolated from mammalian gonadal tissues. The results suggest a physiological role for these metal-binding proteins in developing vertebrate ovaries as well as an involvement in the sequestration of cadmium following environmental exposure.     

Chitinolytic enzymes in the digestive system of marine fishes

Chitinase, exo-N-acetyl--D-glycosaminidase (NAGase) and lysozyme activities were assayed in the digestive tract of 6 species of marine fishes: Myxine glutinosa (cyclostome), Chimaera monstrosa (holocephalan), Squalus acanthias, Etmopterus spinax, Raja radiata (elasmobranchs) and Coryphaenoides rupestris (teleost). Strong chitinase activity was found in the gastric mucosa of the elasmobranchs (S. acanthias, E. spinax and R. radiata) and the teleost (Coryphaenoides rupestris). A remarkably high chitinase activity occurred in the pancreas of the stomachless holocephalan fish Chimaera monstrosa. NAGase activity was strong in the digestive tract of all species. It could be concluded that marine fishes with diets consisting largely of chitinous invertebrates may display high chitinase and NAGase activities in their digestive system; however, only low chitinase activity was found in the intestine of the cyclostome Myxine glutinosa. Coryphaenoides rupestris gastric mucosa chitinase had one optimum activity at pH 1.25, whereas S. acanthias chitinase had two optima, at pH 1.6 and 3.6. The NAGase pH-activity curves from S. acanthias and R. radiata gastric mucosa displayed similar optima, at pH 4.5 and 4.25 respectively. Chimaera monstrosa pancreatic chitinase had a very strong optimum around pH 8 to 10, and one less strong at pH 3. These enzyme activities could not be separated by gel filtration or isoelectric focusing. The pI (isoelectric point) was approximately 4.9 for both enzymes. The molecular weight of the C. monstrosa pancreatic chitinase was estimated to be approximately 43 000. Lysozyme activity was absent or extremely weak in the material studied.     

Kinetics of osmoregulation in the crab Carcinus maenas

When shore crabs Carcinus maenas are transferred from 11 to 38 S at 11°C, new constant levels of hemolymph freezing points and of concentrations of Na, K, Ca, and Mg in the hemolymph are accomplished within 24h. From a decrease in serum protein and in serum free amino acids and an increase in the relative amounts of individual essential free amino acids in the body fluids of whole crabs, a participation of proteolytic activities and a transport of amino acids from hemolymph to cell is deduced; the stationary concentration of total intracellular free amino acids increases up to a nearly two-fold value, compared with the concentration in crabs remaining in diluted sea water. Also, the low molecular neutral sugars increase in whole crabs after high salinity stress, reaching values of more than two-fold initial concentrations within a period of 10 days. This increase is fully accounted for by a 6.7-fold increase in trehalose levels. The processes of increasing the concentrations of low molecular organic material seem to be slower than those of establishing new osmotic pressures in the hemolymph. The oxygen consumption decreases by 30 to 45% to new constant values within 8 to 12 h after the salinity change from 11 to 38, reflecting similar kinetics as the establishment of new osmotic pressures in the hemolymph.     

Food habits, gut morphology and pH, and assimilation efficiency of the zebraperch Hermosilla azurea, an herbivorous kyphosid fish of temperate marine waters

Adult zebraperch, Hermosilla azurea, were found to be functional herbivores in that animal matter constituted <0.01% of the total dry weight of stomach contents of fish collected off Santa Catalina Island in southern California waters. The diet of these fish consisted mainly of red algae (88.2% by dry wt) and also small amounts of brown (7.8%) and green (4.0%) algae. The most important dietary item, the filamentous red algae Polysiphonia spp., was found in >78% of the stomachs and comprised >60% of the contents by dry weight. The digestive tract was long, on average 4.0 times the standard length of the fish, and was composed of the stomach, pyloric caeca, intestine, hindgut chamber with a blind caecum, and rectum. The mean pH of the cardiac stomach was acidic (3.9), whereas that of the intestine was nearly neutral (6.9) and that of the hindgut and blind caecum slightly acidic (6.3 and 6.6, respectively). Algal foods are apparently digested by acid lysis in the stomach and by microbial fermentation in the hindgut. Zebraperch assimilated nutritional constituents from six species of algae with varying degrees of efficiency: carbon (73.7 to 89.7%), nitrogen (72.4 to 84.5%), and protein (71.9 to 94.9%). The fish assimilated these constituents as efficiently or more efficiently from three species of nondietary brown algae as from three species of dietary red and green algae. These results show that zebraperch, like their tropical and subtropical relatives (members of the genus Kyphosus), can digest a wide variety of algae including brown algae containing defensive secondary compounds. Received: 3 November 1997 / Accepted: 19 June 1998     

Blutgruppen-ähnliche substanzen in einigen marinen invertebraten. IV. H-und A-ähnliche substanzen,agglutinine und präzipitine: Ihre verteilung bei Cerianthus sp.

In tentacles and other parts of Cerianthus sp., besides a protease, a strong agglutinin has been found which agglutinates red cells of different origin and which shows an antigalactose specificity. In gonads and mantle mucus, a Blood-Group-H-like material could be demonstrated. The agglutinin is an acid protein, and can be precipitated by base polymers. For tentacles, a precipitin with anti-galactogen specificity is described and analysed. In the mantle, the commensal Phoronis australis was discovered which contained material similar to human blood, Group A.     

High molecular weight algal substances in the sea

Phytoplankton exudates into culture media and extracts from littoral algae are shown to contain significant quantities of extracellular materials having molecular weights in excess of 50,000. A recent development in experimental fluid mechanics (the reduction of turbulent friction coefficients of flowing solutions by dissolved macromolecules) has been utilized as a specific test for high molecular weight compounds. Phytoplankton species in all major algal groups could be found which exude high molecular weight compounds into their culture media; the red alga Porphyridium cruentum typically produced friction reductions as high as 60%. Seaweed samples in the Chlorophyta, Phaeophyta, and Rhodophyta showed major changes in turbulent friction coefficient; in some cases the friction was less than 1/2 that of pure seawater. All samples of the genus Porphyra and Gigartina produced substantial friction changes. From these experiments it is concluded that algal exudates can be a prominent source of high molecular weight compounds in the sea.