Molecular cloning of sex hormone-binding globulin cDNA in carp, Cyprinus carpio

Two types of cDNAs encoding carp SHBGs were isolated and characterized. Northern blot analysis showed their mRNAs distributed mainly in liver, and mRNA levels were almost constant during annual reproductive cycle. This result suggests SHBG synthesis is independent of serum steroid concentration.     

Studies on the Toxicity of Shrimp (Penaeus vannamei) Fed Diets Dosed with Aflatoxin B1 to Humans

Abstract

The objective of this study was to determine the potential for transmission of aflatoxin B₁ to humans through consumption of shrimp contaminated with this toxin. Pacific white shrimp (Penaeus vannamei) were fed diets dosed with three levels of afaltoxin B₁ (300, 400, and 900 ppb) in an 8 week indoor aquarium feeding trial. The shrimp were analyzed for residues of the toxin in the feces, whole shrimp and tail muscle after concentrating the aflatoxin by monoclonal antibody affinity chromatography, followed by radioimmunoassay. Aflatoxin B₁ was not detected in shrimp feces, whole shrimp or tail muscle within detection limits of 2 ppb. Our study suggests that the possibilities for aflatoxin B₁ toxicity occurring in humans via consumption of shrimp fed on diets contaminated with this toxin are limited.     

Identification of a telomere sequence type in three sponge species (Porifera) by fluorescence <Emphasis Type="Italic">in situ</Emphasis> hybridization analysis

The telomere sequence type (TTAGGG)_n is known to be distributed in various phyla in the Animalia and in Mastigophora (Protista). However, the telomere type of Porifera (sponges), a phylum comprising the lowest multicellular animals, has not been reported. In this study, we examined the three sponge species Leucetta chagosensis, Halichondria japonica, and Halichondria panicea for the presence of the telomere type (TTAGGG)_n by fluorescence in situ hybridization (FISH). The oligonucleotide probe (TTAGGG)₇ clearly displayed signals on the interphase nuclei of all three sponges. In contrast, the (TTAGG)₇ probe, which has one base fewer than (TTAGGG)₇, did not display the signals. These results suggest that the telomeres of the three sponges consist of (TTAGGG)_n, which is identical to the sequence type found in many higher multicellular animals and in Mastigophora. Additionally, this is the first study to reveal a telomere sequence type for Porifera. Moreover, these results suggest that Porifera are phylogenetically related to Mastigophora, and supports the general theory that Porifera evolved from Mastigophora. Further, this study strongly suggests that the origin of the (TTAGGG)_n telomere sequence is to be found in a common ancestor of either the Bilateria and Porifera, or the Protista.     

Generation of reactive oxygen species by leukocytes of Prochilodus lineatus

Prochilodus lineatus (curimbatá), from the Procholodontidae family, is a Brazilian freshwater fish, which is important commercially, nutritionally and ecologically. It is encountered in the Rio da Prata Bay in Southern South America. Studies on the immune system of this fish are scarce, but the physiological mechanisms of the species are analogous to those of other vertebrates. Thus, this work discusses the present study, which correlates P. lineatus leukocytes and the generation of reactive oxygen species after modulatory stimuli. Leukocytes were characterized by light and electron transmission microscopy and investigated by the generation of H₂O₂ and O₂ ^?, using phenol red, flow-cytometry and electron transmission histochemistry. The study determined that monocytes and neutrophils are the main cells responsible for generating O₂ after stimulation with phorbol myristate acetate. Superoxide dismutase successfully inhibited the generation of reactive oxygen species in neutrophils and monocytes, but stimulated generation when in association with phorbol myristate acetate. Fish leukocyte samples from P. lineatus showed cross-reactivity with antibodies directed against human NADPH-oxidase antibody subunits (p47^phox and p67^phox). Thus, catalase enhanced the presence of p47^phox. Neutrophil mitochondria were shown to be generators of H₂O₂ (charged by cerium precipitate), being enlarged and changing their format. The present study contributes to a better understanding of the respiratory burst pathways in this species and suggests mitochondria as the organelle responsible for generation of reactive oxygen species.     

Zebrafish fin‐derived fibroblast cell line: A model for in vitro wound healing

The goal of this study was to develop and characterize a cell line from the caudal fin tissue of zebrafish and also its application as an in vitro model to study the effect of H₂O₂ in wound healing. Fibroblastic cell line was developed using explant culture method from caudal fin tissue of zebrafish and characterized. This cell line was named as DrF cell line. The DrF cells treated with 0–10 µM/ml H₂O₂ were tested for viability, proliferation and motility by MTT assay, trypan blue assay and chemotaxis assay, respectively. Among the different concentrations of H₂O₂, 4 µM was found to be nontoxic to study cell migration in in vitro scratch wound assay. Furthermore, the expression of proliferating cell nuclear antigen (PCNA) and chemokine receptor (CXCR4) genes was carried by qPCR. The cell survival, proliferation and migration were extremely enriched at 4 µM level of H₂O₂. We observed accelerated wound closure in DrF cells treated with H₂O_2. The qPCR results indicated that H₂O₂ markedly up‐regulated mRNA expression of PCNA and CXCR4. The findings from our study suggest that H₂O₂ at low levels promotes cell survival, proliferation, migration and wound healing in DrF cells.     

The effect of exogenous hormonal treatment on egg quality of Chilean flounder (Paralichthys adspersus)

In this study, the effects of hormonal treatments on egg mortality after fertilization in Chilean flounder were evaluated. In the control group, plasmatic estradiol-17β showed very low levels before spawning induction. However E₂, GnRHa and GnRHa/bGH treatments caused high plasma E₂ levels resulted in poor egg survival. Probably, the persistent high E₂ levels affected the timing of oocyte maturation.     

In vitro effects of estradiol-17β on myoid and fibroblastic cell proliferation in the immature rainbow trout testis

This study sought to determine whether estradiol-17β (E₂) could induce proliferation of interstitial cells (fibroblasts and myoid cells) in the immature rainbow trout (Oncorhynchus mykiss) testis. To investigate this, a floating organ culture system was utilized. The results showed that a dose of 18.3n M E₂ added to the medium significantly increased interstitial cell proliferation. Pretreatment of the tissue with the estrogen receptor antagonist ICI 182,780 nullifies this proliferative effect. These results show that E₂ can enhance interstitial cell proliferation in the immature rainbow trout testis before spermatogenesis begins.     

Identification of potential general markers of disease resistance in Exopalaemon carinicauda via three‐round challenge selection with an acute hepatopancreatic necrosis disease‐causing strain of Vibrio parahaemolyticus

Sixteen candidate disease‐resistant parameters were selected through which to evaluate the acute hepatopancreatic necrosis disease (AHPND)‐resistant capability of Exopalaemon carinicauda after three generations of selection for AHPND‐causing strain of Vibrio parahaemolyticus (VP_AHPND) resistance in our previous study. However, these parameters required further verification. In this study, another AHPND‐resistant E. carinicauda series was obtained through a short‐term selection procedure, consisting of three virulent challenge rounds of selection (about three‐week interval for each challenge) with VP_AHPND infection. After this selection, the survival rate at 144 hr post infection (hpi) increased from 23.33% to 37.78% and the observed 48‐hr LD₅₀ of VP_AHPND to shrimp increased from 10^5.5 cfu/ml to 10^6.5 cfu/ml. Then, the immune response of this AHPND‐resistant E. carinicauda was studied using the 16 candidate AHPND‐resistant parameters selected for in our previous study. The improved VP_AHPND clearance rate in hpi, increased total haemocyte counts, haemocyanin concentration, alkaline phosphatase activity and expressions of six immune‐related genes (Tollip and ALF in haemocytes and hepatopancreas; lysozyme, crustin and cathepsin B in hepatopancreas; and LGBP in haemocytes) at 24 hpi after the three‐round challenge selection suggest that these immune parameters may be reliable markers for the evaluation of the physiological status and potential AHPND‐resistant phenotypes in E. carinicauda.     

The GABA a agonist muscimol enhances locomotor activity, but does not alter the behavioural effects of CRH in juvenile spring chinook salmon (Oncorhynchus tshawytscha)†

The present study investigated the effects of manipulating the GABAergic system on locomotor activity in juvenile spring chinook salmon, Oncorhynchus tshawytscha. In addition, we evaluated whether the GABAergic system is important for mediating the behavioural effects of corticotropin-releasing hormone (CRH). An intracerebroventricular (ICV) injection of the GABA_a agonist muscimol caused an acute and dose dependent increase in locomotor activity in juvenile spring chinook salmon. ICV injections of the GABA_a antagonist bicuculline prevented the increase in activity when administered concurrently with muscimol. The GABA_b agonist baclofen had no effect on locomotor activity in this study. Furthermore, we found no evidence that the locomotor response to exogenous CRH was altered by the concurrent administration of muscimol or bicuculline. These results provide evidence to support the hypothesis that endogenous GABA within the central nervous system is involved in the control of locomotor activity in fish. The data also suggest that there is no interaction between the GABAergic system and CRH with regards to the control of locomotor activity in this species.     

The effects of commercial bacterial products on macrobenthos community in shrimp culture ponds

This study was conducted to assess the effects of commercial bacterialproducts on the composition and abundance of macrobenthos in commercial tigerprawn Penaeus monodon culture ponds. Nine brackishwaterponds, with an area of 0.5 ha each were used for this study. Twocommercial bacterial products (T₁ and T₂) were applied inthree ponds each. Another three ponds served as the control. Sediment samplesfrom all ponds were analyzed for the nitrogen, phosphorus and organic mattercontents, and macrobenthos. Total nitrogen and total carbon concentrations inthe sediments of T₁ were significantly lower (p< 0.05) thanT₂. In addition, total nitrogen in T₁ pond sediment wassignificantly lower (p < 0.05) than the control towardsthe end of the culture period. There were no significant differences in totalphosphorus amongst treatments. The study revealed the presence of four majorgroups of macrobenthos in all treatments; gastropods, bivalve, polychaetes andinsect larvae. The total density of macrobenthos in all ponds, exceptgastropods, showed a decreasing trend with the progress in culture indicatingeither predation by shrimp or deterioration of pond bottom. T₁ hadsignificantly (p < 0.05) higher density of polychaetescompared to the T₂ and the control.     

The effects of commercial bacterial products on macrobenthos community in shrimp culture ponds

This study was conducted to assess the effects of commercial bacterialproducts on the composition and abundance of macrobenthos in commercial tigerprawn Penaeus monodon culture ponds. Nine brackishwaterponds, with an area of 0.5 ha each were used for this study. Twocommercial bacterial products (T₁ and T₂) were applied inthree ponds each. Another three ponds served as the control. Sediment samplesfrom all ponds were analyzed for the nitrogen, phosphorus and organic mattercontents, and macrobenthos. Total nitrogen and total carbon concentrations inthe sediments of T₁ were significantly lower (p< 0.05) thanT₂. In addition, total nitrogen in T₁ pond sediment wassignificantly lower (p < 0.05) than the control towardsthe end of the culture period. There were no significant differences in totalphosphorus amongst treatments. The study revealed the presence of four majorgroups of macrobenthos in all treatments; gastropods, bivalve, polychaetes andinsect larvae. The total density of macrobenthos in all ponds, exceptgastropods, showed a decreasing trend with the progress in culture indicatingeither predation by shrimp or deterioration of pond bottom. T₁ hadsignificantly (p < 0.05) higher density of polychaetescompared to the T₂ and the control.     

Effect of Combined Non‐ionized Ammonia and Dissolved Oxygen Levels on the Survival of Juvenile Dourado,Salminus brasiliensis (Cuvier)

The aim of this study was to assess the mean lethal concentration (LC₅₀) of dissolved oxygen in high ammonia concentration and also the LC₅₀ of ammonia under hypoxic conditions for juveniles of dourado, Salminus brasiliensis. In the first experiment, the non‐ionized ammonia (NH₃) concentrations were: 0.026, 0.447, 0.612, 0.909, and 1.334 mg/L, and the dissolved oxygen concentration was maintained at approximately 1.65 mg/L. In the second experiment, the dissolved oxygen concentrations were: 1.64, 1.99, 3.33, 5.10, and 7.77 mg/L, and the non‐ionized ammonia concentration was kept at approximately 0.927 mg/L. The mean lethal concentrations of non‐ionized ammonia varied from 0.584 to 0.577 mg/L, indicating that LC₅₀ values were almost unaffected by exposure time. The estimated LC₅₀ of dissolved oxygen varied from 4.02 to 5.02 mg/L, indicating a slight increase in the mean lethal concentrations as the exposure time increased. Results from this study indicate that interaction between these two parameters increases mortality and also suggest that dourado is susceptible to the combination of high ammonia with low dissolved oxygen concentrations.     

Response of Rainbow trout (Oncorhynchus mykiss) to unrefined peanut oil diets: Effect on growth performance,fish health and fillet fatty acid composition

A 60‐day feeding study with rainbow trout, Oncorhynchus mykiss, was conducted to determine the effects of replacement of fish oil (FO) by unrefined peanut oil (PO) on growth performance, feed utilization, body composition, fatty acid composition and serum biochemical and haematological parameters. Rainbow trouts (51.60 ± 0.75 g) were fed five experimental diets formulated by replacing dietary FO with PO at levels of level 0 (PO₀), 1/4 (PO₂₅), 1/2 (PO₅₀), 3/4 (PO₇₅) and 4/4 (PO₁₀₀), respectively. As a result, the best growth performance was observed in fish fed with PO₀ and PO₅₀ diet. No significant differences were detected among the groups in terms of body compositions. Fatty acid profiles of the fish fillets reflected the fatty acid profiles of the feeds that the fishes were fed with. In this study, the haematological parameters detected that there were no significant differences compared to the control group, whereas the serum biochemical parameters generally worsened as the ratio of peanut oil in the ration exceeded half of fish oil. As a conclusion, the results of the study suggested that the unrefined peanut oil could be used as a replacer of fish oil in diets for rainbow trout.     

Microsatellite multiplex panels for population genetic analysis of red sea bream <Emphasis Type="Italic">Pagrus major</Emphasis>

In this study, multiplex PCR panels were developed to amplify 20 microsatellite markers for red sea bream, Pagrus major, with the aim of reducing labor and material costs associated with genetic analysis of this species. The usefulness of these panels was validated in 200 fish collected at five sampling locations. The occurrence of null alleles was suggested at five markers, which were dropped from further analysis. The remaining 15 microsatellite loci showed high levels of heterozygosity (H _E range 0.34–0.97, H _O range 0.32–1) and a wide range in the number of alleles per locus (A; range 5–46). Increasing the number of microsatellite loci from three to ten and 15 improved the performance of the panels for population differentiation (F _ST) and estimation of Nei’s (D _S) genetic distance. The results of this study confirm the usefulness of genotyping a large number of microsatellite DNA markers to expand our knowledge of red sea bream population genetics.     

Feminization and growth of juvenile fat snook Centropomus parallelus fed diets with different concentrations of estradiol-17β

In this study, we evaluated the effect of diets with various concentrations of estradiol-17β (E₂) on the feminization, growth and survival of juvenile fat snook. Over a 45-day period, the juveniles were fed with five diets containing 0, 25, 50, 75 and 100 mg E₂/kg feed. Three replicates were run for each diet. After this hormonal treatment, the fishes were transferred to 1-m³ net cages, where they remained for 12 months. During this period, they were fed a commercial diet without added hormones. During the feminization, the control fish grew more than those in the other treatments, and the survival rates did not differ significantly among the treatments. At the end of the experiment growth did not differ among the treatments. In the control treatment, 80.85 % of the fish were male, and the females were significantly larger than the males. All of the treatments with E₂ produced 100 % females. No intersex fish were found in this study. This research showed that it is possible to obtain 100 % fat snook females using feeds with 25 mg E₂/kg for 45 days without impairing the growth or survival of the fish.     

Waste excretion of marble goby (Oxyeleotris marmorata Bleeker) fed with different diets

Marble goby (Oxyeleotris marmorata Bleeker), with its high demand and price, has a great potential as a profitable commercial aquaculture candidate in Malaysia and Southeast Asia region. Efforts are being made to produce this species in a better controlled culture environment like recirculating aquaculture system (RAS) due to poor growth performance and disease problems shown by conventional cage and outdoor pond culture systems. Quantification of waste excreted by fish is critical to RAS design. This study was conducted to characterize the waste excretion rates of marble goby fed with different diets (live food and minced fish). Ammonia-N (TAN), urea-N, nitrite-N (NO₂-N), nitrate-N (NO₃-N), total-N (TN), organic-N (ON), feces-N, 5-day biochemical oxygen demand (BOD₅) and total suspended solid (TSS) produced from marble goby were determined over a 72-h excretion period. Under given experimental conditions, the results showed that feed type had significant influence on the waste excretion rates, with marble goby fed live tilapia (Oreochromis niloticus) exhibiting significantly (P < 0.05) the lowest amount of waste excretion comparable to that of fish fed live common carp (Cyprinus carpio) and minced scads (Decapterus russellii). This indicates that feeding marble goby with tilapia poses less adverse effects on water quality and is thus a suitable diet for this species. The waste excreted by the fish is composed of nitrogenous excretion (TAN, Urea-N, ON, Feces-N), and productions of dissolved biodegradable organic substances (BOD₅) and TSS (TSS_feces + TSS_water). About 58-71% of the nitrogen consumed in food was excreted and its rate depended mainly on the feed type. TAN was the chief end-product of protein metabolism; about 74-84% of the daily total nitrogenous excretion was TAN. Urea-N accounted for 13-21% of the daily total nitrogenous excretion indicating that urea-N is an important nitrogenous excretory end-product in marble goby. The waste excretion data presented in this study can be served as a pre-requisite for designing a RAS for this species. The overall BOD₅ and TSS production found in this study also point to the need for including bio-filtration unit and suspended solids removal mechanism in the RAS design.     

Effects of thyroxine administration on the growth and survival of pike silverside (Chirostoma estor) juveniles

The pike silverside has high aquaculture potential despite its slow growth. In this study, thyroxine (T₄) concentrations (0.13, 1.3 and 13 nM), a control with no hormonal supplement and a negative control containing 4.5 mM methimazole (MMI) were tested to evaluate the growth of this species. Juveniles (0.2 g) were exposed by immersion to these treatments for 8 h every second day for 120 days, and growth evaluations were performed monthly over the entire trial period. In addition, tissue samples from each treatment group were assayed for triiodothyronine (T₃) and deiodinase type 2 (D2) activity . The survival rates in all T₄ groups were high, and a significant increase in growth was observed (average of 58%). The MMI treatment caused an increase in mortality and a reduction in the final body weight compared with the control. T₄ administration did not affect the tissue levels of T₃, and it decreased muscular D2 activity only after 30 days of exposure. These results demonstrated that low concentrations of T₄ in the culture environment could improve the growth of this species without affecting tissue hormone levels. The technique may have useful applications for early‐stage aquaculture of this and other economically important species.     

Effects of CO2 levels and light intensities on growth and amino acid contents in red seaweed Gracilaria lemaneiformis

The seaweed Gracilaria lemaneiformis is largely maricultivated in China, for use as food and as a material in the agar industry. This alga experiences ocean acidification caused by rising atmospheric CO₂ levels, and experiences changing light levels caused by self‐shading during the later period of mariculture. In this study, growth and amino acid (AA) content responses of G. lemaneiformis to different CO₂ levels (the present and the predicted increased levels) and varying light levels at 28 (±1)°C temperature conditions were investigated. The results showed that a higher light level enhanced algal growth and decreased water loss, but reduced AA accumulation. Decreased pH levels (as a result of CO₂ elevation) also enhanced algal growth and reduced AA contents, but the decreases in the AA score at the lower pH levels were not significant under the two light level treatments. In this study, the light treatments had greater influences on growth and AA contents than CO₂ levels. The results suggest that G. lemaneiformis quality will be negatively affected during the later mariculture production period as levels of CO₂ rise and global temperatures increase.     

Economic evaluation of Piaractus mesopotamicus juvenile production in different rearing systems

In this study, the costs and gross income related to the production of pacu Piaractus mesopotamicus juveniles were evaluated. This evaluation took into consideration a semi-intensive rearing, with direct stocking of the larvae into fertilized ponds (IL₀), or an initial intensive larviculture system, in which the larvae were fed in the laboratory for 3 (IL₃), 6 (IL₆), or 9 days (IL₉) before being transferred to the ponds. After 45 days of rearing, a gradual increase in production costs was observed as intensive larviculture time increased. Gross income also increased due to better survival rates (11.0, 25.3, 45.4, and 54.0% for IL₀, IL₃, IL₆, and IL₉, respectively). Therefore, increased profits were obtained under intensive larviculture (US$ 0.27, US$ 6.07, US$ 11.99, and US$ 13.16 per one thousand larvae in treatments IL₀, IL₃, IL₆, and IL₉, respectively).In a larger scale production simulation, the results obtained with initial intensive larviculture also showed evident economic advantages, confirming the feasibility of this system in comparison with the direct stocking of larvae in ponds for the production of pacu juveniles.