Reproductive dynamics and nursery habitat preferences of two commercially important Indo-Pacific red snappers <Emphasis Type="Italic">Lutjanus erythropterus</Emphasis> and <Emphasis Type="Italic">L. malabaricus</Emphasis>

Red snappers were examined for reproductive biology and age-0 habitat preferences. Spawning in red snappers occurred throughout the year in northern Australia and eastern Indonesia; at least 10–30% of females and 40–80% of males were in ripe or spawning condition in most months. Northern Australian populations showed a spawning peak from July to December (L. erythropterus) and September to March (L. malabaricus). Eastern Indonesian L. malabaricus had a less defined pattern with two peaks: January–March and October. Size at first maturity was 240 mm for males and 250–300 mm for females. L ₅₀ estimates were similar between species in northern Australia: 270–280 mm (males) and 350–370 mm (females). Maximum batch fecundity was 676,100 oocytes for L. erythropterus and 997,000 oocytes for L. malabaricus. Higher mean abundances of age-0 L. erythropterus were found in silty and coarse sand/rubble estuarine habitats of northern Australia (456 ± 119 fish/km²) compared with sandy coastal habitats (5 ± 3 fish/km²). Most age-0 snapper caught at Sape (eastern Indonesia) were L. malabaricus (91%) with mean abundances of 312 ± 14 fish/km². The similarities in the reproductive characteristics of red snappers suggest that successful management approaches adopted in northern Australia should be considered in eastern Indonesia.     

Nucleotide sequence and mRNA expression analysis of <Emphasis Type="Italic">β</Emphasis>-actin gene in the orange-spotted grouper <Emphasis Type="Italic">Epinephelus coioides</Emphasis>

The full-length cDNA, encoding the orange-spotted grouper β-actin and spanning 1920 bp including a poly (A) tail, was cloned from its brain cDNA library. The open reading frame encodes a protein of 375 amino acids. Sequence analysis indicated that it contained the typical structural features of cytoplasmic actins, and showed higher homology with other vertebrate β-actin than any other members of the actin family. The partial genomic sequence indicated that the organization of the β-actin gene in the orange-spotted grouper might also be conserved. Northern blot analysis indicated that it was expressed at high levels in the brain, spleen, adipose tissue, ovary, and liver, but at low levels in the gill filament and heart, and at a very low level in the kidney. The expression of β-actin gene in the skeletal muscle was barely detectable. These results indicated that the expression of the orange-spotted grouper β-actin gene showed significant variation in different tissues. Therefore, caution should be taken when using β-actin gene as an internal control in the normalization of gene expression among tissues. Whereas, semi-quantitative RT-PCR analysis indicated that treatment with 17α–methyltestosterone (MT) had little effect on the mRNA expression of β-actin gene in the in vitro incubated hypothalamus, pituitary, and ovary fragments of the orange-spotted grouper, suggesting β-actin can be used as an internal control for RT-PCR analysis of MT effects on gene expression in these tissues.     

Comparison between α-LNA and DHA in early developmental stages of <Emphasis Type="Italic">Takifugu obscurus</Emphasis> and <Emphasis Type="Italic">Takifugu rubripes</Emphasis>

ABSTRACT:   Rearing experiments were conducted to investigate the essential fatty acid requirements in the early developmental stages of river puffer Takifugu obscurus and tiger puffer T. rubripes using two n-3 series unsaturated fatty acids, α-linolenic acid (18:3n-3, α-LNA) and docosahexaenoic acid (22:6n-3, DHA), under salinity of 30 and 18.5–20.3°C. River and tiger puffer larvae used in this study were 15 and 14 days old after hatching, and their average body weights were 30.1 and 20.8 mg, respectively. The results on fatty acid requirements of these two species were evaluated from fish growth, survival, fatty acid composition of the fish body and activity test results. The DHA groups of both river and tiger puffer exhibited better survival and weight gain. However, there was no difference in the mean final body weights of river puffer between two dietary groups. Also, the DHA group of tiger puffer showed better results in the recovery test from anesthetic condition than that obtained in the LNA group. In an examination of the fatty acid compositions of the whole body, the LNA group containing no dietary DHA resulted in 0.5% DHA in tiger puffer and 1.1% DHA in river puffer . These results suggest that α-LNA from Artemia converted to eicosapentaenoic acid (20:5n-3, EPA) and to DHA successively by their fatty acid metabolism. Symptoms following essential fatty acid deficiency were not observed in any experimental groups. As river puffer did not represent a significant difference in the dietary effects between α-LNA and DHA treatment groups, its essential fatty acid requirement was assumed to be somewhat closer to that of the freshwater fishes in comparison with that of marine fishes, including tiger puffer.     

Spatial–temporal variations in the composition of two <Emphasis Type="Italic">Zostera</Emphasis> species in a seagrass bed: implications for population management of a commercially exploited grass shrimp

We examined spatial and temporal variations in the species composition of two seagrass species, Zostera marina and Z. caespitosa, in a lagoon facing the Sea of Okhotsk. We also considered how those variations affected habitat quality for motile epifauna, especially for a commercial shrimp. A long-interval comparison between 1996 and 2013 showed that seagrass species composition in the lagoon did not vary, while their relative abundances did. A survey in 2012 revealed that the abundance of Z. caespitosa was affected by the abundance of Z. marina, water depth, and location in the lagoon. Although these seagrass species have similar aboveground morphology, differences in their fine structures were detected. Diversities of motile animals inhabiting the seagrass species were the same. Differences in seagrass utilization were observed when we focused on a commercial shrimp, Pandalus latirostris. This shrimp always preferred higher densities of shoots irrespective of the species and seasonally changed their preference for the number of leaves per shoot. They were therefore more abundant in Z. caespitosa, which had those structural characteristics. The results suggest that the management of shrimp resources could be improved by flexibly changing protected areas in accordance with the dynamics of seagrass distribution in the lagoon.     

Effects of <Emphasis Type="Italic">Euphorbia hirta</Emphasis> plant leaf extract on growth performance,hematological and organosomatic indices of hybrid catfish, <Emphasis Type="Italic">Clarias macrocephalus </Emphasis>× <Emphasis Type="Italic">C. gariepinus</Emphasis>

This study was conducted to evaluate the effects of Euphorbia hirta leaf extract on the growth performance, hematological and organosomatic indices of hybrid catfish, Clarias macrocephalus?×?C. gariepinus. The fish were treated with 0 (control), 300, 500 and 800 mg/kg (ppm) for 90 days. The weight gain, average daily growth rate, and specific growth rate were at significantly higher levels in fish from all the treatment groups on days 75 and 90, while the feed efficiency and protein efficiency ratio were consistently higher in fish from all the treatment groups from day 60 up until day 90. The feed conversion rate significantly decreased from day 60 until day 90 in all treatment groups when compared with the control group, and the survival rate was significantly different from day 30 until day 90; a consistently higher rate was observed in fish fed 800 mg/kg. The highest viscerosomatic index and intraperitoneal fat were observed in the group fed 500 mg/kg (p?<?0.05). The hepatosomatic index was significantly increased alongside increasing levels of E. hirta extract. The total white blood cell count in the control group was significantly higher on day 30, but on day 90 all the treatment groups showed higher levels. Hematocrit percentage was significantly different on days 30 and 90. Lymphocyte, eosinophil and thrombocyte levels were shown to be significantly different (p?<?0.05) when different groups of fish were compared. In conclusion, 300 mg/kg of E. hirta leaf extract could improve growth performance, hematological and some organosomatic indices in hybrid catfish.     

Effect of nitrite exposure on oxygen-carrying capacity and gene expression of <Emphasis Type="Italic">NF-κB</Emphasis>/<Emphasis Type="Italic">HIF-1α</Emphasis> pathway in gill of bighead carp (<Emphasis Type="Italic">Aristichthys nobilis</Emphasis>)

Nitrite (NO₂^?) contamination of water can severely impact the health of aquatic life and is a major concern for commercial aquaculture. In order to study the effect of nitrite on Aristichthys nobilis, we investigated the oxygen-carrying capacity, NF-κB/HIF-1α pathway, and the gill tissue structure under nitrite stress. In the current study, bighead carp (initial weight 180.05?±?0.092 g) were exposed to nitrite (48.634 mg/L) for 96 h and then for 96 h recovery test. After nitrite exposure for 6 h, hypoxia-inducible factor-1α (HIF-1α) and nuclear factor-κB (NF-κB) mRNA expression increased significantly in the gill of bighead carp (P?<?0.05). After nitrite exposure for 12 h, hemoglobin (Hb) and methemoglobin reductase (MHBR) content in blood decreased significantly (P?<?0.05); TLR4 mRNA expression increased significantly (P?<?0.05). After nitrite exposure for 24 h, methemoglobin (MetHb) content increased significantly (P?<?0.05). After recovery test, all the indicators except TLR4 mRNA expression level recovered to initial level. In conclusion, nitrite exposure can affect hemoglobin dynamics, as oxidization of nitrite by hemoglobin results in the reduction of Hb to MetHb leading to hypoxia and nitrite exposure can also result into gill tissue damage. In the face of nitrite exposure, NF-κB and HIF-1α mRNA expression level increased immediately to protect the body from oxidative damage and eased hypoxic condition caused by nitrite. It was also observed that nitrite damage is recoverable in Aristichthys nobilis, but it may be need more than 96 h.     

Protective effect of squilla chitosan–silver nanoparticles for <Emphasis Type="Italic">Dicentrarchus labrax</Emphasis> larvae infected with <Emphasis Type="Italic">Vibrio anguillarum</Emphasis>

Antimicrobial nanoparticle therapy was proposed as an alternative strategy to reduce the use of antibiotics in larval-rearing systems. Antibacterial potential of the prepared squilla chitosan–silver nanoparticles and its protective effect on Dicentrarchus labrax (sea bass) larvae in the early stages were studied against Vibrio angularium. Different concentrations of squilla chitosan (Csq) and squilla chitosan–silver nanoparticles (Csq–AgNps) (1, 2, 5, 10, and 20 %) were, in vitro, tested against V.anguillarum and expressed as a role of Log₁₀ mean. Sea bass larvae were treated using: 10 % Csq and 5 % Csq–AgNps as effective inhibitory concentrations against the pathogen either encapsulated during the feeding regime or added directly to the model system via the water from the onset of 4 weeks. The long-term administration of Csq–AgNps through enriched food for both non-infected and infected systems had survival % of 74.5 ± 1.5 and 72.5 ± 2.5, respectively. Larval clinical observations using Csq–AgNps were studied compared with the two controls. The current study found that 5 % encapsulated Csq–AgNps was enough to suppress infection and considered as an alternative to antibiotics in controlling virulent fish pathogens.     

Age,growth and mortality estimates for populations of red snappers <Emphasis Type="Italic">Lutjanus erythropterus</Emphasis> and <Emphasis Type="Italic">L. malabaricus</Emphasis> from northern Australia and eastern Indonesia

Lutjanus erythropterus and L. malabaricus were examined for life-history differences among northern Australian and eastern Indonesian populations. Formation of opaque growth increments in otoliths began during April to September for northern Australian fishes and September to April for eastern Indonesian fishes. Maximum observed ages were greater than previously reported: 42 and 48 years for L. erythropterus and L. malabaricus, respectively. Eastern Indonesian red snappers grew faster than northern Australian fish. Growth was similar for northern Australian L. erythropterus populations. However, Kupang and Sape populations of L. erythropterus in eastern Indonesian showed different growth from the Arafura North population, which was similar to northern Australian fish. There were growth differences among northern Australian populations of L. malabaricus. Arafura South and Timor populations were similar, but differed from Groote and Weipa populations. There were no significant differences in growth among populations of L. malabaricus in eastern Indonesia. Total mortality estimates were similar between northern Australian and eastern Indonesian fish: 0.09 and 0.16 year⁻¹ for L. erythropterus and 0.11 and 0.14 year⁻¹ for L. malabaricus, respectively. Life-history characteristics of these red snappers are typical of other tropical snappers: slow-growing and long-lived with low mortality. However, population growth differences suggest that their management should be based on biological information from distinct stocks.     

Distribution patterns of settlement-stage juveniles of <Emphasis Type="Italic">Girella punctata</Emphasis> and <Emphasis Type="Italic">Girella leonina</Emphasis> on the rocky coast of the Kanto–Izu region,Japan

The early life history of girellid fishes in Japanese waters is unclear, and little is known about their species-specific reproductive strategies. We examined seasonal changes of distribution patterns for settlement-stage juveniles of Girella punctata and Girella leonina on the rocky shore in the regions of Kanto and Izu, Japan, to infer the influence of the Kuroshio Current on their reproduction. We collected 813 settlement-stage juveniles mainly in Sagami Bay and genetically identified the species. The juveniles of G. punctata were collected on the rocky shore in Sagami Bay during April to August, with the abundant catch in May and June. Thus, we infer that juvenile G. punctata ubiquitously inhabit the rocky shore in the area in spring and summer. By contrast, juveniles of G. leonina were rarely collected in Sagami Bay, with a total catch of only 66. Notably, no juveniles were collected during the wintertime in Sagami Bay, although an abundant catch of G. leonina had been previously reported for Sagami Nada off Sagami Bay during January to March. This clear-cut difference between the areas likely reflects the difference in proximity to the path of the Kuroshio Current. We expect that the Kuroshio Current strongly influences the reproductive success of G. leonina.     

Segregation of microsatellite loci in second generation hybrid striped bass (<Emphasis Type="Italic">Morone saxatilis</Emphasis> × <Emphasis Type="Italic">Morone chrysops</Emphasis>)

Within the framework of a larger project aimed to assess the potential of second generation hybrid striped bass for German aquaculture the genotypic segregation of five microsatellite loci was analysed in two progeny lots (n = 74 and 76, respectively). There was no consistent correlation between microsatellite genotypes and phenotypic category (white bass, hybrid, or striped bass). None of the individuals expressed neither only white bass nor only striped bass genotypes at all five loci. On the other hand, only hybrid genotypes at all five loci were detected in three individuals of lot 1 and four individuals of lot 2. Single loci tests for conformity of microsatellite genotypic segregation with Mendelian rules revealed significant deviations (P < 0.05) in four cases for lot 1 and in three cases for lot 2. If pooled over all five loci, both lots displayed highly significant deviations (P < 0.01) with an excess of hybrid genotypes and a deficiency of white bass genotypes. It is concluded that stabilizing selection performed on hybrid genotypes might be a suitable approach for practical aquaculture in Europe if the goal is to become independent of first generation hybrid fry supply and/or if establishing domesticated brood stocks of both parental species is impossible. However, more detailed studies on the characteristics and performance of multiple hybrid generations are needed.     

Optimum time for weaning South African <Emphasis Type="Italic">Scylla serrata</Emphasis> (Forskål) larvae from rotifers to <Emphasis Type="Italic">Artemia</Emphasis>

To determine the optimum time at which to wean Scylla serrata larvae from rotifers onto Artemia two experiments were conducted, approximately 1 month apart, using larvae from two different female crabs. In the first experiment, the larvae in three treatment groups, with nine replicates each, were fed rotifers for the first 8 days after hatching. Artemia were introduced on days after hatch (DAH) 0 – during the first zoeal instar (treatment R + A); on DAH 4 – during the second zoeal instar (treatment R4A); on DAH 8 – during the third zoeal instar (treatment R8A). In a control (ROT) larvae were fed with rotifers exclusively for 18 days until the completion of metamorphosis to megalopa. In the second experiment, the same four feeding schedules as in experiment 1 were used with an additional group of larvae (treatment AC) that were fed only on Artemia throughout the rearing period. Similar results were recorded in the two experiments. Larvae in treatments R + A and R4A performed significantly better than those in treatments R8A, ROT and AC. This was particularly evident when examining the proportion of zoeae which successfully completed metamorphosis to megalopa. Poor performance of larvae in treatments AC and ROT implied that rotifers are needed as a first food, but that rotifers alone do not fill the nutritional requirements of S. serrata larvae. Poor performance of larvae in treatment R8A suggested that the diet should be supplemented with Artemia before the end of the zoea 3 stage.     

The changes in the biochemical compositions and enzymatic activities of rotifer (<Emphasis Type="Italic">Brachionus plicatilis</Emphasis>, Müller) and <Emphasis Type="Italic">Artemia</Emphasis> during the enrichment and starvation periods

The changes in the biochemical compositions and enzymatic activities of rotifer (Brachionus plicatilis) and Artemia, enriched and stored at 4°C temperature, were determined. The total starvation period was 16 h and samples were taken at the end of the 8th and 16th hours. In present study, the rotifer and nauplii catabolized a large proportion of the protein during the enrichment period. Lipid contents of both live preys increased during the enrichment period and decreased in nauplii and metanauplii throughout the starvation period but lipid content of the rotifer remained relatively constant during the starvation period. The changes observed in the amino acid compositions of Artemia and the rotifer were statistically significant (P < 0.05). The conspicuous decline the essential amino acid (EAA) and nonessential amino acid (NEAA) content of the rotifer was observed during the enrichment period. However, the essential amino acid (EAA) and nonessential amino acid (NEAA) contents of Artemia nauplii increased during the enrichment period. The unenriched and enriched rotifers contained more monounsaturated fatty acid (MUFAs) than polyunsaturated fatty acid (PUFAs) and saturated fatty acids (SFA). However, Artemia contained more PUFAs than MUFAs and SFA during the experimental period. A sharp increase in the amounts of docosahexaenoic acid (DHA) during the enrichment of the rotifer and Artemia nauplii was observed. However, the amount of DHA throughout the starvation period decreased in Artemia metanauplii but not in Artemia nauplii. Significant differences in tryptic, leucine aminopeptidase N (LAP), and alkaline phosphatase (AP) enzyme activities of Artemia and rotifer were observed during the enrichment and starvation period (P < 0.05). The digestive enzymes derived from live food to fish larvae provided the highest contribution at the end of the enrichment period. In conclusion, the results of the study provide important contributions to determine the most suitable live food offering time for marine fish larvae. Rotifer should be offered to fish larvae at the end of the enrichment period, Artemia nauplii just after hatching and before being stored at 4°C, and Artemia metanauplii at the end of the enrichment and throughout the starvation period.     

Species differences and effects of soft coral extracts from <Emphasis Type="Italic">Sinnularia maximus</Emphasis> on the expression of cytochrome P4501A and 2N in butterflyfishes (<Emphasis Type="Italic">Chaetodon</Emphasis> spp.)

Cytochrome P450 (CYP) has been shown to confer resistance in numerous terrestrial insects that consume potentially toxic secondary metabolites in plants, but fewer studies have examined the role of critical biotransformation enzymes in allowing marine organisms to consume chemically defended foods. This study examined the expression of CYP1A and CYP2N mRNAs in several butterflyfish species, which can feed on numerous chemically defended soft and hard corals. In addition, the effect of an extract from a soft coral (Sinnularia maxima) on expression of hepatic CYP1A and CYP2 mRNAs was also examined. Fish were fed extracts on days 1, 3 and 5, and expression was examined on day 5. Phylogenetic analyses of the CYP1A cDNA from 12 species of butterflyfish (DNA, amino acid) indicate well-separated groupings according to their feeding strategies. The non-coralline feeding Chaetodon xanthurus exhibited a 7-fold higher basal expression of CYP2N8 relative to the other species studied. Although induction of CYP2N7 expression was observed in C. punctatofasciatus, CYP1A and CYP2N was largely unaffected or diminished by extract treatment in the other species of butterflyfish. These results indicated groupings of feeding strategy with CYP1A phylogeny in Chaetodon, but generally unaltered expression of CYP1A and CYP2N following dietary treatment with an extract from a chemically defended soft coral suggesting an inconclusive role of these isoforms in the detoxification of chemicals in these extracts.     

Experimental infection of <Emphasis Type="Italic">Betanodavirus</Emphasis> in freshwater fish <Emphasis Type="Italic">Gambusia affinis</Emphasis> (Baird and Girard, 1853)—a potential infection model for viral encephalopathy and retinopathy

Betanodaviruses are the causative agents of the disease known as viral nervous necrosis (VNN) or viral encephalopathy and retinopathy (VER) in a variety of marine and freshwater fish species. The aim of this study was to demonstrate experimental infection of an isolate of betanodavirus (RGNNV genotype) in freshwater fish, Gambusia affinis, for elucidation of transmission mechanism and potential use as a laboratory model. Morbidity and mortality rate was significantly higher by injection route of infection as compared to immersion by bath and resembled the natural infection of juvenile marine fish. The fish in disease affected group showed severe neurological disorders accompanied by extensive vacuolar degeneration and mild to moderate neuronal necrosis of the brain in comparison to control. Amplification of ~?427 bp product in the variable region of the coat protein gene of betanodavirus was achieved by RT-PCR with 100% sequence homology to RGNNV genotype.     

The impact of net-isolated polyculture of tilapia (<Emphasis Type="Italic">Oreochromis niloticus</Emphasis>) on plankton community in saline–alkaline pond of shrimp (<Emphasis Type="Italic">Penaeus vannamei</Emphasis>)

We used 12 land-based experimental enclosures (6 m × 5 m) in a saline–alkaline pond of shrimp (Penaeus vannamei) to determine the impact of net-isolated polyculture of tilapia (Oreochromis niloticus) on plankton communities for 40 days. Tilapias were stocked in net cages suspended in enclosures, in polyculture ponds including tilapia and shrimp. Four tilapia biomass were tested: 0, 39, 115 and 227 g m⁻². Shrimp stocking biomass were 0.7 g m⁻² in all treatments. There were three replicates in each treatment. Our results showed that the presence of tilapia significantly reduced phytoplankton biomass directly through predation and indirectly through top-down effect. The stocking of tilapia reduced zooplankton biomass, particularly rotifer biomass. However, copepod biomass was not been significantly affected. So, net-isolated polyculture of tilapia can thus have a strong impact on phytoplankton allowing the co-existence of large numbers of copepods with planktivorous fish and improving the water quality of shrimp ponds.     

Talking to the dead: using <Emphasis Type="Italic">Post</Emphasis>-<Emphasis Type="Italic">mortem</Emphasis> data in the assessment of stress in tiger sharks (<Emphasis Type="Italic">Galeocerdo cuvier</Emphasis>) (Péron and Lesueur, 1822)

Sharks are very sensitive to stress and prone to a high mortality rate after capture. Since approximately 50 million of sharks are caught as bycatch every year, and current recommendations to reduce the impact of commercial fishing strongly support immediate release, it is imperative to better understand post-release mortality caused by the stress of capture and handling. Blood samples allow the assessment of stress levels which are valuable tools to reduce mortality in commercial, recreational and scientific fishing, being essential for the improvement in those conservation measures. Biochemical analyses are widely used for sharks as stress indicators, with secondary plasma parameters (lactate, glucose and ions) being the most often employed assays. However, it is virtually impossible to determine baseline plasma parameters in free-ranging sharks, since blood withdrawal involves animal capture and restrain, which are stressful procedures. This study aims at analyzing secondary parameters of five healthy tiger sharks captured with circular hooks and handlines in Fernando de Noronha (Northeastern Brazil) and comparing them with secondary parameters of three dead tiger sharks caught off Recife (also Northeastern Brazil). The results showed that the analysis of some plasma constituents in dead animals may be an efficient tool to assess stress and lethality. However, traditional parameters such as glucose and calcium, need to be used with caution. The results also demonstrated the extreme importance of urea and phosphorus for assessing stress response and mortality in tiger sharks, both parameters frequently neglected and of utmost importance for shark’s homeostasis.     

Purification and characterization of α<Subscript>1</Subscript>-proteinase inhibitor and antithrombin III: major serpins of rainbow trout (<Emphasis Type="Italic">Oncorhynchuss mykiss</Emphasis>) and carp (<Emphasis Type="Italic">Cyprinus carpio</Emphasis>) blood plasma

The main serine proteinase inhibitors of rainbow trout (Oncorhynchuss mykiss) and common carp (Cyprinus carpio) blood plasma were isolated and purified. The investigated inhibitors, α₁-proteinase inhibitor (α₁-PI) and antithrombin III (AT III), act by forming stable complexes with target proteinases. The association rate constants k _on for the interaction of fish plasma inhibitors with several serine proteinases have been determined: k _on for both carp and rainbow trout α₁-PI were >10⁷ M⁻¹ s⁻¹ for human neutrophil elastase, and in the case of bovine trypsin and chymotrypsin k _on values were 2.0–5.2 × 10⁶ M⁻¹ s⁻¹. Association rate constants k _on for the interaction of carp and rainbow trout AT III with bovine trypsin and thrombin were about 1.3 × 10⁴–7.9 × 10⁵ M⁻¹ s⁻¹ without and >10⁷ M⁻¹ s⁻¹ in presence of heparin; so antithrombins require the presence of heparin to become effective proteinase inhibitors. The high degree of homology of the estimated amino acid sequences of fish inhibitors reactive site loops confirms their similarity with other proteinase inhibitors from the serpin family.