Establishment of dominance relationships in gilthead sea bream Sparus aurata juveniles during feeding: effects on feeding behaviour, feed utilization and fish health

In gilthead sea bream Sparus aurata held in groups of two, five or 10 fish, social hierarchies were observed. Subordinate S. aurata were characterized by elevation of basal levels of plasma cortisol, together with a reduced immunological potential. Subordinate fish also showed lower feed intake, feed utilization and lower growth. Fatty acid composition was also affected by social status, with a lower content of saturated acids, oleic and eicosapentaenoic, in muscle and liver of fish considered as subordinate. Results show that social hierarchy acts as a stressor in S . aurata .     

Cellular responses in the skin of the gilthead sea bream Sparus aurata L. and the sea bass Dicentrarchus labrax (L.) exposed to high ammonia

Adult gilthead sea bream Sparus aurata and sea bass Dicentrarchus labrax were exposed for 24 and 48 h, respectively, to two concentrations of ammonia each (mean values of 3·34 and 13·10 mg l(-1) TA-N in S. aurata; 2·99 and 11·90 mg l(-1) TA-N in D. labrax). Light microscopy and computerized morphometry were used to evaluate ammonia-induced alterations in skin structure during exposure and following recovery in normal water. In S. aurata, ammonia exposure induced a concentration-dependent increase in the number (hyperplasia) of neutral mucous cells (mc), with peak values at 24 h recovery after exposure. An increase in the dispersion of melanosomes in skin melanocytes was also observed in the dermis and occasionally in the epidermis of S. aurata, with peak values at 24 h of ammonia exposure. Exposure of D. labrax to ammonia had, likewise, concentration-dependent effects on mucous secretion. Of the two types of mc in this species, there was an increase in the number of the neutral mc and a reduction in the much more numerous acid mc, with peak values at 24 and 48 h, respectively, of ammonia exposure. The more intense mucous secretion in D. labrax compared to S. aurata could be related to the lower tolerance to ammonia in D. labrax, as reported elsewhere. Finally, the increase in melanosome dispersion was less evident in D. labrax, due to highly variable control values. These morphological alterations to the skin could be useful indicators of non-specific stress in cultured fishes.     

Feeding habits of the gilthead seabream (Sparus aurata) from the Ria Formosa (southern Portugal) as compared to the black seabream (Spondyliosoma cantharus) and the annular seabream (Diplodus annularis)

The feeding habits of Sparus aurata L., Diplodus annularis L. and Spondyliosoma cantharus L. in the Ria Formosa (southern Portugal) lagoon system were studied using three simple methods (frequency of occurrence, numeric percentage and percentage weight) and a composite index [index of relative importance (IRI)]. The Ivlev index was used to evaluate diet selectivity, while the Schoener overlap index was used to compare diets, and diet diversity was characterized by the Simpson index. The diets of the three species consist of a wide variety of food organisms, nevertheless S. aurata seems to be the most specialized. No significant dietary overlap was found, with S. aurata preferentially selecting gastropods and bivalves, while S. cantharus preferentially selected a wide variety of crustaceans and D. sargus a wider array, including crustaceans, gastropods and bivalves.     

A New Species of Acrodipsas Sands (Lepidoptera: Lycaenidae) from Southern New South Wales and the Australian Capital Territory

Acrodipsas aurata sp.n. from the Blue Mountains, the southern highlands of New South Wales and the Australian Capital Territory is described, figured and contrasted with the polymorphic A. cuprea (Sands) with which it has been previously considered to be con-specific. A. aurata is assigned to the myrmecophila species-group, based on leg morphology. Nine species are now recognised in the genus Acrodipsas Sands.     

Proteins from fish eggs that protect DNA from acid precipitation and inhibit DNA synthesis

We partially characterized proteins that inhibit DNA acid precipitation from various fish eggs (Sparus aurata, Dicentrarchus labrax, Mugil cephalus and Zeus faber). The active proteins were purified by acetone fractionation. The activity was found to be heat resistant. Of bivalent cations tested only Co(2+) and Cu(2+) exerted a profound promoting effect in the activity from all fish. The protein fraction from Sparus aurata inhibited DNA synthesis in PCR performed by different DNA polymerases. The possible role of DNA protective proteins in fish egg physiology is discussed.     

Purification and kinetic characterization of 6-phosphofructo-1-kinase from the liver of gilthead sea bream (Sparus aurata)

6-phosphofructo-1-kinase (PFK) was purified to homogeneity from liver of gilthead sea bream (Sparus aurata) and kinetic properties of the enzyme were determined. The native enzyme had an apparent molecular mass of 510 kDa and was composed of 86 kDa subunits, suggesting homohexameric structure. At pH 7, S. aurata liver PFK (PFKL) showed sigmoidal kinetics for fructose-6-phosphate (fru-6-P) and hyperbolic kinetics for ATP. Fructose-2,6-bisphosphate (fru-2,6-P2) converted saturation curves for fru-6-P to hyperbolic and activated PFKL synergistically with AMP. Fru-2,6-P2 counteracted the inhibition caused by ATP, ADP and citrate. Compared to the S. aurata muscle isozyme, PFKL had lower affinity for fru-6-P, higher cooperativity, hyperbolic kinetics in relation to ATP, increased susceptibility to inhibition by ATP, and was less affected by AMP, ADP and inhibition by 3-phosphoglycerate, phosphoenolpyruvate, 6-phosphogluconate or phosphocreatine. The effect of starvation-refeeding on PFKL expression was studied at the levels of enzyme activity and protein content in the liver of S. aurata. Our findings indicate that short-term recovery of PFKL activity after refeeding previously starved fish, may result from allosteric regulation by fru-2,6-P2, whereas combination of activation by fru-2,6-P2 and increase in protein content may determine the long-term recovery of the enzyme activity.     

The influence of certain aminoacids and vitamins on post-thaw fish sperm motility, viability and DNA fragmentation

During cryopreservation, dilution in the extender media reduces the seminal plasma constituents being cells more vulnerable to oxidative stress. Vitamins (C and E) and the amino acids taurine and hypotaurine are powerful antioxidants naturally present in seminal plasma. Whether their effect may improve sperm quality and reduce sperm DNA damage after cryopreservation in fish sperm still remains unclear. Thus, the aim of the present work was to analyse the effect of extender supplementation with several antioxidant components on post-thawed sperm motility, viability and DNA integrity of two commercial species, gilthead seabream (Sparus aurata) and European seabass (Dicentrarchus labrax). Sperm collected from ten to twelve individuals was cryopreserved in ten different extenders containing: taurine and hypotaurine (1 and 10mM), ascorbic acid (1 and 10mM), α-tocoferol (0.1 and 0.5 mM) or 1 ml/l of a commercial cell antioxidant supplement. Cell viability, motility and DNA fragmentation were determined in post-thawed samples. Addition of antioxidants (vitamins and amino acids) to D. labrax and S. aurata extenders did not significantly increase the parameters of motility (TM, PM, VCL, VSL and Lin) or viability, although 1mM taurine slightly increased the percentage of motile cells (TM) in S. aurata. DNA fragmentation (DNA in tail and Olive tail moment) in D. labrax sperm was higher in treatments containing vitamins than amino acids or control. However in S. aurata sperm, antioxidants especially taurine and hypotaurine, significantly reduced both DNA fragmentation parameters, protecting DNA against strand breaks. These results suggest a species-specific effect depending on the type of antioxidants used.     

A novel second myostatin gene is present in teleost fish.

We report on the isolation and characterisation of the complete cDNA sequence encoding a novel bone morphogenetic protein-like protein (sbMSTN-b) in the teleost fish Sparus aurata. The encoded protein is 68% identical to S. aurata MSTN at the amino acid level, and homologues were also found in Umbrina cirrosa and Tetraodon nigroviridis. Phylogenetic analysis suggests that the MSTN-b gene may be present in most, perhaps all, teleost fish species. RT-PCR on different tissues/stages indicates that MSTN-b is expressed almost exclusively in the central nervous system, starting from late larval stages. Quantitative analyses indicate an increase of sbMSTN-b expression in the brain associated with metamorphosis, at the same time as completion of nervous system differentiation.     

Chronic toxicity of ammonia to juvenile gilthead seabream Sparus aurata and related histopathological effects

Effect of 20 days exposure of juvenile gilthead seabream ( Sparus aurata ) to elevated levels of ammonia on growth and survival was examined in a continuous flow system. Suppressed growth and reduced survival were observed at concentrations of 8.2 and 13 mg l⁻¹ total ammonia-N (0.5 and 0.7 mg l⁻¹ un-ionized ammonia-N, respectively) and higher. The maximum acceptable toxic concentration (MATC) for growth was between 4.8 to 8.2 mg l⁻¹ total ammonia-N (0.3 and 0.5 mg l⁻¹ un-ionized ammonia-N, respectively). Fish exposed to high ammonia levels (13 mg l⁻¹ total ammonia-N, 0.7 mg l⁻¹ un-ionized ammonia-N) displayed clear signs of liver pathology. Existing evidence suggests that S. aurata is less sensitive to ammonia than other reported marine and freshwater fish.
Under certain conditions ammonia concentration in the intensive fish ponds in Eilat may exceed the no observed effect concentration for S. aurata .     

Molecular cloning of hepatic glucose-6-phosphatase catalytic subunit from gilthead sea bream (Sparus aurata): response of its mRNA levels and glucokinase expression to refeeding and diet composition

To examine the relationship between structure and function of glucose-6-phosphatase (G6Pase) in fish, we undertook molecular cloning and modulation of G6Pase expression by starvation and refeeding on diets with different nutrient composition in the liver of the carnivorous fish, Sparus aurata. A cDNA encoding the full-length G6Pase catalytic subunit from the liver of S. aurata was isolated. This cDNA encodes a 350-amino acid protein, with low homology to the mammalian G6Pase, although it contains most of the key residues required for catalysis. Based on hydrophobicity and membrane structure prediction, we propose a model containing nine-transmembrane regions for S. aurata G6Pase. Northern blots showed that refeeding after a prolonged starvation rapidly reverses the glucose/glucose-6-phosphate substrate cycle flux in the fish liver through decreased G6Pase expression and strong glucokinase (GK) induction. The effect of refeeding different diets on G6Pase and GK expression, indicated that hepatic intermediary metabolism of fish fed diets with low protein/high carbohydrate diets is impelled towards utilization of dietary carbohydrates, by means of modulation of GK mRNA levels rather than G6Pase expression. These findings challenge the role attributed to dysregulation of G6Pase or GK expression in the low ability of carnivorous fish to metabolise glucose.     

Fructose 2,6-bisphosphate in liver of Sparus aurata: influence of nutritional state

1. Fructose 2,6-bisphosphate (fru-2,6-P2) has been measured in liver and muscle of gilthead sea bream fish, Sparus aurata. 2. The fru-2,6-P2 levels in liver depend on the diet given to the fish: in fish fed a high carbohydrate diet, the fru-2,6-P2 levels are higher than any one previously reported. These changes are associated with differences in the phosphofructokinase 2 activity. 3. Fru-2,6-P2 levels has also been measured in liver of Sparus aurata after different fasting periods. In starved fish, fru-2,6-P2 did not decrease as sharply as in rat. The values found in fish starved for 20 days were similar to those reported for rats that had been starved for 24 hr.     

Characterization of Pseudomonas spp. associated with spoilage of gilt-head sea bream stored under various conditions.

The population dynamics of pseudomonads in gilt-head sea bream Mediterranean fish (Sparus aurata) stored under different conditions were studied. Phenotypic analysis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of whole-cell proteins were performed to identify a total of 106 Pseudomonas strains isolated from S. aurata stored under different temperatures (at 0, 10, and 20 degrees C) and packaging conditions (air and a modified atmosphere of 40% CO(2)-30% N(2)-30% O(2)). Pseudomonas lundensis was the predominant species, followed by Pseudomonas fluorescens, while Pseudomonas fragi and Pseudomonas putida were detected less frequently. Fluorescent Pseudomonas strains dominated under air conditions, while proteolytic and less lipolytic strains dominated under modified-atmosphere packaging. Different storage conditions appear to govern the selection of pseudomonads in gilt-head sea bream fish.     

Identification of an osteopontin-like protein in fish associated with mineral formation

Fish has been recently recognized as a suitable vertebrate model and represents a promising alternative to mammals for studying mechanisms of tissue mineralization and unravelling specific questions related to vertebrate bone formation. The recently developed Sparus aurata (gilthead seabream) osteoblast-like cell line VSa16 was used to construct a cDNA subtractive library aimed at the identification of genes associated with fish tissue mineralization. Suppression subtractive hybridization, combined with mirror orientation selection, identified 194 cDNA clones representing 20 different genes up-regulated during the mineralization of the VSa16 extracellular matrix. One of these genes accounted for 69% of the total number of clones obtained and was later identified as theS. aurata osteopontin-like gene. The 2138-bp full-length S. aurata osteopontin-like cDNA was shown to encode a 374 amino-acid protein containing domains and motifs characteristic of osteopontins, such as an integrin receptor-binding RGD motif, a negatively charged domain and numerous post-translational modifications (e.g. phosphorylations and glycosylations). The common origin of mammalian osteopontin and fish osteopontin-like proteins was indicated through an in silico analysis of available sequences showing similar gene and protein structures and was further demonstrated by their specific expression in mineralized tissues and cell cultures. Accordingly, and given its proven association with mineral formation and its characteristic protein domains, we propose that the fish osteopontin-like protein may play a role in hard tissue mineralization, in a manner similar to osteopontin in higher vertebrates.     

Osteologic development of the viscerocranial skeleton in sea bream: alternative ossification strategies in teleost fish

The ontogeny of the viscerocranial skeleton of sea bream Sparus aurata larvae was studied from 1 to 90 days post-hatching. In the smallest specimens analysed at 2·7 mm L _N no cephalic elements were present and at 3·1 mm L _N the following cartilaginous structures were visible: trabecula cranii, auditory capsule, Meckel's cartilage, quadrate, hyosymplectic cartilage, sclerotic, hypohyal, ceratohyal epihyal cartilage, interhyal, hypobranchial 1 and ceratobranchial 1. The only structure ossified at this size is the maxillary and the next ossified structures to appear are the preopercle and opercle at about 3·7 mm L _N. The last bones to appear are infraorbital 2 and 6 at 15·1 mm L _S. The first cartilaginous elements and structures to ossify in S. aurata appear to be related with functional requirements, so that structures involved directly in feeding and breathing generally appear and ossify before those that are not. The ontogeny of different regional structures revealed that generally the dermal bones ossify before the cartilage replacement bones. Comparison of S. aurata viscerocranial skeleton ontogeny with that of phylogenetically distant fish demonstrates that different ossification strategies exist in higher and lower teleost fish.     

Peptides from Liza aurata: Natural Source Attenuate Paracetamol Induced Nephrotoxicity by Modulation of the Inflammatory Response and DNA Damage

International Journal of Peptide Research and Therapeutics - The present study investigates the nephroprotective and the molecular mechanisms effects of Liza aurata protein hydrolysates (LAPHs)...     

Leptotheca sparidarum n. sp. (Myxosporea: Bivalvulida), a parasite from cultured common dentex (Dentex dentex L.) and gilthead sea bream (Sparus aurata L.) (Teleostei: Sparidae)

A new myxosporean, Leptotheca sparidarum n. sp., is described from the trunk kidney of two sparid fish, Sparus aurata and Dentex dentex, in several culture facilities from the Western Mediterranean coasts. It is distinguished from all the previously described species by spore morphometrics. Spores with two equal polar capsules and one binucleated sporoplasm. Spore measurements from D. dentex were 5-7.1 micron long x 8.8-12.3 micron thick x 5.88-6.18 micron wide; polar capsules 2.6-3.5 micron in diam. Spore measurements from S. aurata were 5.1-8.24 micron long x 9.41-11.76 micron thick; polar capsules 2.4-3.2 micron in diam. No significant differences were detected between either host. Prevalence could reach 21.4% in D. dentex and 19% in some stock of S. aurata. Renal tubules were the typical site of infection, which was also found in ureters and glomeruli, but seldom in the epiepithelial position of the gut. Spores were formed in disporous sporoblasts, and spore maturation seemed to proceed from the host epithelium towards the lumen. Trophozoites were attached to the host epithelial cells of renal tubules by pseudopodial-like projections, which were inserted into gaps between epithelial cells. Also, cell junctions were observed between primary cells and between neighboring spores.     

Molecular differentiation of three loach species (Pisces, Cobitidae) based on the nuclear 5S rDNA marker

The diversity of the 5S rDNA fragment among three loach species: Cobitis taenia, C. elongatoides and Sabanejewia aurata was investigated using universal PCR primers for this gene. Three amplification products were obtained: 220 bp length for C. taenia and C. elongatoides, and 330 bp for S. aurata. Two amplicons with the same length (in Cobitis) were digested with TaqI restriction endonuclease. This enzyme found one restriction site T/CGA in the C. elongatoides fragment, while in the case of C. taenia no cleavage effect was observed. On this basis we constructed an easy and cheap method for loach species discrimination. It seems adequate for effective support of conservation initiatives for endangered loaches.     

Ultrastructural and biochemical analysis of epidermal xanthophores and dermal chromatophores of the teleost Sparus aurata

We have studied the pigmentary system of the teleost Sparus aurata skin by electron microscopy and chromatographic analysis. Under electron microscopy, we found the dermis to contain the three major types of recognized chromatophores: melanophores, xanthophores and iridophores. Melanophores were more abundant in the dorsal region, whereas the iridophores were more abundant in the ventral region. The most important discovery was that of epidermal xanthophores. Epidermal xanthophores were the only chromatophores in the epidermis, something only found in S aurata and in a teleost species living in the Antartic sea. In contrast, the biochemical analysis did not establish any special characteristics: we found pteridine and flavin pigments located mostly in the pigmented dorsal region. Riboflavin and pterin were two of the most abundant coloured pigment types, but other colourless pigments such as xanthopterin and isoxanthopterin were also detected.