Evidence of cylindrospermopsin uptake and clearance in fish (Oreochromis niloticus) under laboratory conditions

Cylindrospermopsin (CYN) is a cyanotoxin that has raised serious concerns about public health in many parts of the world. It can bioaccumulate and affect the health of aquatic organisms, but despite this, few studies have been conducted on CYN uptake and clearance in fish. In this paper, the authors evaluate the uptake and clearance of CYN in the muscle tissue and viscera of juvenile tilapia (Oreochromis niloticus) after exposure to aqueous extracts and whole cells of Cylindrospermopsis raciborskii (CYN-producer). CYN blended with commercial fish food, and three experiments were conducted. In the first trial, fish food, and aqueous extracts containing 0.31 μg CYN g⁻¹ of food per day, was administered to tilapia for 15 days. In the second trial, fish were provided food and intact cells (5.4 μg CYN g⁻¹ of food per day) for 15 days. In the last trial, they were provided fish food and aqueous extracts (0.8 μg CYN g⁻¹ of food per day) for 12 days, and for the next 10 days, the animals were fed food without toxic cell extracts (to simulate a clearance period). The concentration of CYN in muscle tissue and viscera was analysed using ELISA. In the case of juvenile tilapia, the presence of CYN was higher in viscera than in muscle tissue, and the toxin remained in the tissues even after 10 days without the addition of contaminated food. The results suggest that tilapia represents a potential source of CYN transfer through the food web, and this shows the need for a continuous monitoring of this compound in organisms that are used for human and animal consumption.     

Effect of exposure to cadmium on the tropical freshwater prawn Macrobrachium rosenbergii

The effect of cadmium (Cd) on mortality, resistance and bioaccumulation in the tropical freshwater prawn Macrobrachium rosenbergii from Egypt were studied. Survival of prawns exposed to Cd doses over 60 μg l^?1 were significantly lower than that of prawns exposed to lower doses. After 96 h prawns exposed to >40 μg l^?1 of Cd had a greater reduction in total haemocyte count and phagocytic activity than those exposed to lower concentrations. Bioaccumulation of Cd in the gills, hepatopancreas and muscles was variable. Cadmium accumulated significantly in the gills and hepatopancreas, but Cd accumulation in the muscles increased only marginally. Macrobrachium rosenbergii manifested histopathological alterations in the gills, hepatopancreas and muscles when exposed to various concentrations of cadmium.     

Measurements of digestibilities of cholesterol and fatty acids using 5α-cholestane as an inert marker in the tilapia, Oreochromis niloticus, and the freshwater prawn, Macrobrachium rosenbergii

An attempt was made to measure the apparent digestibility (AD) of cholesterol and fatty acids using 5aL-cholestane as a marker (cholestane-method) using tilapia. Oreochromis niloticus, and the freshwater prawn, Macrobrachium rosenbergii, as test animals. The results were compared with those obtained by the Cr₂O₃-method. Casein-based test diets containing 0.3% levels of cholestane and chromic oxide as markers were prepared for determination of AD of dietary cholesterol and fatty acids. After 2 weeks of acclimation, the test animals, tilapia (average 3.0 g wet weight) and freshwater prawn (average 0.5 g wet weight) were fed the test diets for 1 week, and faecal samples were collected over 6 subsequent days. Lipids in the diets and faeces were extracted with chloroform-methanol and saponified with 10% KOH in methanol. Gas-liquid chromatography (GLC) on 1.5% OV-17 of the unsaponifiable matters afforded the quantities of 5aL-cholestane and cholesterol, whereas that on 5% Shinchrom E-71 of the saponifiable matters provided the quantities of fatty acids. Chromic oxide was determined by a wet-digestion method. In tilapia, the AD of cholesterol determined by cholestane- and Cr₂O₃-methods were 78.1% and 73.5%, respectively. The AD of individual fatty acids differed with the types of fatty acids, but those of fatty acids determined by the two methods in this study were similar, except for a slightly higher value of several fatty acids such as 16:1 in the cholestane-method than in the Cr₂O₃-method. This indicates that 5α-cholestane as well as Cr₂O₃ can be conventionally used as a marker for determining AD of fatty acids and cholesterol. The cholestane-method is advantageous as it permits parallel analysis of cholesterol (or fatty acids) and 5α-cholestane by GLC using small amounts of faecal sample. The cholestane-method was also useful for analysing digestibilities of cholesterol and total lipids in freshwater prawn. However, slight differences between the cholestane- and Cr₂O₃-methods were observed in the AD of several fatty acids such as 14:0 and 16:1.     

Protein and energy needs for maintenance and growth of Nile tilapia (Oreochromis niloticus)

A trial was undertaken with juvenile Nile tilapia fed graded levels of dietary protein (0–35% DP) over 7 weeks (28°C). Measurements of nitrogen and energy utilization were made using data on ADC, comparative carcass analyses, nitrogen excretion and oxygen uptake. Data indicate that the daily protein intake for maximum N gain was c. 12 g/kg/d and the maintenance protein requirements were about of 2g/kg/d. The optimal DP/DE ratio was found to be 18mg/kJ. Differences were observed in the data on endogenous nitrogen and energy utilization depending on the dietary treatments as well as on the criteria used.     

Scale regeneration of tilapia (Oreochromis niloticus) under various ambient and dietary calcium concentrations

• 1.1. Scale regeneration was examined in Oreochromis niloticus under normal, dietary calcium-deficient, and both dietary and ambient water calcium-deficient conditions.
• 2.2. Calcium contents of regenerating scales were dependent on the calcium status of the fish, but areal growth of the scales was independent.
• 3.3. Transient hypocalcemia accompanied with a loss of calcium from original scales was observed in descaled fish in dietary calcium deficiency, and intact and descaled fish in both dietary and ambient water calcium deficiency.
• 4.4. The results suggested that calcium mobilized from internal sources was insufficient and both water and dietary calcium were necessary for normal calcification of the regenerating scales.

     

Relationship of respiration to body weight in the tilapia Oreochromis niloticus under resting and normoxic conditions

• 1.1. The relationships of oxygen consumption (V_o2, ml/min/fish), ventilation volume (V_g, ml/min/fish), stroke volume (V_s, ml/stroke/fish), respiration frequency (RF, stroke/min) and oxygen utilization (U, %) to body weight (W, g) in the tilapia under resting and normoxic conditions were V_o2 = 0.002835W^0.754193, V_g = 0.688965W^0.722786, V_s = 0.002790W^1.017188, RF = 248.58210W^−0.296139, U = 0.000935W + 87.394288, respectively.

     

Behavior stereotypes of food acceptance and of the rejection of 'bitter' food in the freshwater prawn Macrobrachium rosenbergii

The sequence of food-intake and food-rejection behavior of thefreshwater prawn Macrobracium rosenbergü was investigated.Video recordings were carried out on 12 adult animals of bothsexes at intermolt phase. Plain and quinine-adultered (bitter)food pellets were presented, and the recorded sequences of behaviorwere analyzed by independent evaluators. Food acceptance andrejection were found to occur according to rigid motor stereotypes.An identical behavioral display typified the initial detectionof and search for both plain and bitter food pellets. Subsequentmaxilliped-handling behaviors displayed during ingestion ofplain pellets and in handling and rejection of the quinine-adulteratedpellets were clearly different. Both behaviors were quantifiedand compared.     

微囊藻毒素-LR在罗非鱼(Oreochromis niloticus)体内的动态分布

应用腹腔注射染毒法,研究不同作用时间下微囊藻毒素-LR(MC-LR)在罗非鱼各组织器官的动态分布。结果表明:除罗非鱼肌肉中MC-LR含量低于检出限(用ND表示未检出)外,罗非鱼肝脏、血清、胆囊、鳃、肠道都有MC-LR检出;且各组织器官MC-LR残留量的分布有明显的区别,其中以肝脏中残留量的平均值为最高,其变化区间为0.902～4.938mg·kg-1,血清、胆囊、鳃、肠道的MC-LR残留量变化区间分别为ND～0.390mg·L-1、ND～0.236mg·kg-1、0.0134～0.0369mg·kg-1、0.007～0.016mg·kg-1;染毒后84h,罗非鱼肝脏、鳃、血清、胆囊中的MC-LR残留量都显著降低,且血清和胆囊中的MC-LR含量低于检出限,表明罗非鱼对MC-LR具有很强的解毒机能;染毒后36h,胆囊和肝脏中的MC-LR残留量骤然下降,由此推测胆囊和肝脏可能是罗非鱼对MC-LR的主要解毒器官;试验期间,肌肉中MC-LR含量始终低于检出限,表明低剂量的MC摄入不会在罗非鱼肌肉中造成明显的残留。     

Quantification of lectin in freshwater prawn (Macrobrachium rosenbergii ) hemolymph by ELISA

An enzyme-linked immunoabsorbent assay was developed to quantify the lectin present in the hemolymph of the freshwater prawn Macrobrachium rosenbergii. This method involves the use of murine monoclonal IgG1 with kappa light chain (designated as 3G1) antibodies raised against the purified lectin, the assay that we developed recognized as little as 30 ng/ml of lectin, and was used to measure the lectin concentration in animals at different maturation stages. The highest concentration of lectin was identified in the hemolymph from post-larval prawns and the lowest in molt stage adult animals. The hemagglutination activity of the lectin was four-fold higher in adult than in juvenile specimens, although in all cases N-acetylated sugar residues, such as N-acetyl-D-glucosamine, N-acetyl-D-galactosamine, and N-acetyl-D-neuraminic acid were inhibitors of the lectin activity, suggesting that lectin plays a role in the transport of N-acetylated sugar in juvenile prawns. Our results indicate that lectin concentration and hemagglutinating activity could be influenced by developmental conditions of the freshwater prawn.     

Effect of the hydrostatic pressure on otolith growth of early juveniles of Nile tilapia Oreochromis niloticus

Nile tilapia Oreochromis niloticus early juveniles were maintained for 2 weeks in a pressurized system under a controlled photoperiod, at constant salinity and temperature. Groups of fish were exposed to one of three absolute hydrostatic pressure (HP) regimes: (1) a constant normal atmospheric pressure (100 kPa), (2) a constant 40 m pressure (500 kPa) or (3) a semi-diurnal cyclic vertical migration (100-500 kPa). No significant differences were detected in otolith size and incremental periodicity among the three HP treatments, suggesting that HP does not affect otolith growth of early juveniles O. niloticus.     

Effect of different stress factors on some physiological parameters of Nile tilapia (Oreochromis niloticus)

This study was conducted to determine the effect of different stress factors on some physiological measurements of Nile tilapia (Oreochromis niloticus).A total number of 160 Nile tilapia, the body weight ranging between 100 and 120 g, were exposed to three stress factors of hypoxia, overcrowding and starvation for different periods 24, 72 and 144 h. The results of cortisol level were 134.15, 144.27, 154.12 ng/ml and 140.18 ng/ml for control, hypoxia, overcrowding and starvation, respectively, while after 144 h did not show significant difference among treatments compared with control group. In contrast, the values of T₃ and T₄ observed reduction with significant difference that T₃ ranged between the highest value 122.12 ng/ml for control group to lowest value of starvation group 94.35, 93.81 and 88.46 ng/ml after 24, 72 and 144 h. Also, similar trend of results observed in T₄ and blood glucose among treatments. And the enzymatic activity of lactate dehydrogenise (LDH) increased in hypoxic group, while a significant reduction appeared in overcrowding and starved fish compared to control group. The pyruvate kinase (PK) activity decreased in hypoxic group but increased in other group.     

Muscle growth in Nile tilapia (Oreochromis niloticus): histochemical,ultrastructural and morphometric study

Muscle growth in Nile tilapia (Oreochromis niloticus) was studied focusing on histochemical, ultrastructural, and morphometric characteristics of muscle fibers. Based on body length (cm), we studied four groups: G1=1.36+/-0.09, G2=3.38+/-0.44, G3=8.90+/-1.47, and G4=28.30+/-3.29 (mean+/-S.D.). All groups showed intense reaction to NADH-TR in subdermal fibers and weak or no reaction in deep layer fibers. In G3 and G4, an intermediate layer was also observed with fibers presenting weak reaction; in G4, groups of fibers with intense reaction were observed in the subdermal region. The myosin ATPase (m-ATPase) activities were acid-stable and alkali-labile in subdermal fibers; most deep layer fibers were alkali-stable and acid-labile. Intermediate fibers were acid-labile and alkali-stable. Two fiber populations were observed near deep muscle layer: one large presenting weak acid- and alkali-stable and the other small alkali-stable.During growth, muscle fiber hypertrophy was more evident in intermediate and white fibers for G3 and G4. However, in these groups, the presence of fiber diameters < or =21 microm suggested that there is still substantial fiber recruitment, confirmed by ultrastructural study, but hypertrophy is the main mechanism contributing to increase in muscular mass.     

Effect of dietary alginic acid on juvenile tilapia (Oreochromis niloticus) intestinal microbial balance, intestinal histology and growth performance

The aim of the present study was to assess the effect of a commercial alginic acid source (Ergosan) on tilapia Oreochromis niloticus intestinal microbial balance, intestinal morphology, and growth parameters. Fish were fed a basal control diet or the basal diet plus a source of alginic acid (5?g?kg(-1) Ergosan; Schering-Plough Aquaculture, UK) for 9?weeks. At the end of the trial, light and electron microscopy demonstrated that the morphology of the intestinal tract at the gross and ultra-structural level was not affected by dietary alginic acid inclusion. Both groups of fish displayed healthy, normal morphology with no signs of disease, cell or tissue damage. Intestinal epithelial leucocyte infiltration was not affected by dietary alginic acid. Molecular bacterial profiles derived from PCR-DGGE illustrated highly similar microbial communities (both within the lumen and associated with the intestinal mucosa) in the respective treatment groups. Microbial ecological parameters (e.g. species diversity and richness) also remained unaffected. Although not significant, trends towards elevated survival and body protein content were observed in the alginic acid-fed fish. These results are suggestive that alginic acid does not adversely impact the indigenous gastrointestinal microbial balance and subsequently does not impact upon the epithelial brush border integrity. Validation of non-detrimental impacts of immunostimulatory products on gastric microbiota and epithelial integrity should be pursued in future studies as maintaining microbial balance and epithelial integrity is essential for proper gut functionality.     

Microsatellite loci in the eastern form of the giant freshwater prawn (Macrobrachium rosenbergii)

Six microsatellite loci were identified and characterized in the eastern form of the widespread and commercially important giant freshwater prawn (Macrobrachium rosenbergii). The loci were detected by randomly screening for dinucleotide and trinucleotide repeat units within a partial genomic library developed for the species. In a sample of 29 prawns, number of alleles and heterozygosity per locus ranged from 12 to 18 and from 0.66 to 0.90, respectively. These markers provide powerful tools for the conservation and management of wild stocks, the improvement of cultured stocks of M. rosenbergii, and for investigating evolutionary processes underlying genetic divergence among populations.     

Transfer of foreign gene to giant freshwater prawn (Macrobrachium rosenbergii) by spermatophore-microinjection

We developed a spermatophore-microinjection (SMI) technique that allows exogenous DNA fragments to be transferred easily into the giant freshwater prawn (Macrobrachium rosenbergii), an important aquacultural shellfish and aquatic invertebrate model. From 28 to 1, 000 ng of the circular plasmid pGL, in a total volume of 1 microl, were directly microinjected into spermatophores. Fertilization and hatching of prawns created with SMI were completed in vivo. Fertilization and hatching rates in the SMI treatments did not differ from those of the untreated control group. The genomes of free swimming, SMI-created larvae (21 days after fertilization) were analyzed using PCR and Southern blot analyses. A product with a molecular mass of 680 bp was amplified. It corresponded to amplifications of pGL, and Southern blot analysis revealed that the amplified band was positive. The gene transfer rate was primarily dependent on the concentration of DNA during SMI. The higher the concentration of pGL, the higher the rate of gene transfer. PCR and Southern blot analyses detected the existence of foreign DNA in 16 of 23 samples (70%) of genomic DNA isolated from hatched larvae in the 750 ng pGL SMI treatment. SMI, described here for the first time, is the simplest and most efficient method for mass producing transgenic giant freshwater prawns.     

Effect of feeding pineapple waste on growth performance,texture quality and flesh colour of nile tilapia (Oreochromis niloticus) fingerlings

The study aims to evaluate the effects of pineapples waste on the growth, texture quality and flesh colour of Nile tilapia (Oreochromis niloticus) fingerlings. Fingerlings were fed with four different levels of pineapple waste diets throughout 56 days, which contain a control group (Diet 1) and experimental diets that formulated with 10% (Diet 2), 20% (Diet 3) and 30% (Diet 4) of pineapple waste. The experimental diet was formulated with rice bran, fish meal, soybean meal, vitamin and mineral premix, vegetable oil and binder to attain 32% dietary protein. The results revealed that the formulated fish diet with pineapple waste given the optimum weight gain, weight gain percentage, specific growth rate than the control group, where Diet 4 has shown the highest value (p < 0.05). There were no effects of the pineapple waste diet on the texture quality of the fillet, while only red chromaticity (a*) showed a significant difference (p < 0.05). In conclusion, the addition of pineapple waste can improve the growth rate of Nile tilapia, and the supplementation level of the pineapple waste in the diet was 30% of the total feed formulation.     

Effect of dietary L‐malic acid supplementation on growth,feed utilization and digestive function of juvenile GIFT tilapia Oreochromis niloticus (Linnaeus, 1758)

Two feeding trials (FTs) were conducted in 2013 and 2014, respectively, to determine the optimal L‐malic acid (LMA) level for juvenile GIFT (Genetically Improved Farmed Tilapia) Oreochromis niloticus. Except for the LMA level, the FT1 and FT2 had a similar diet formulation. In FT1, LMA was included at 0 (basal diet), 1, 4, 8, 16 and 32 g kg^?1, respectively. After 20 weeks, daily weight gain and feed conversion ratio were improved but not differentiated with 1–8 g kg^?1 LMA. Further increasing the LMA supply initially decreased the feed intake (16 g kg^?1), and then decreased both feed intake and feed utilization (32 g kg^?1), thus impairing the fish growth. FT2 was subsequently conducted with a smaller LMA range (0, 0.5, 1, 2, 4 and 8 g kg^?1, respectively) but was unfortunately terminated at the end of 8 weeks because 20% of the fish were badly injured during weighing. Unexpectedly, growth and feed utilization were still improved but not differentiated with 0.5–8 g kg^?1 LMA. In FT2, beneficial effects of LMA inclusion on the digestive function (pepsin, foregut amylase and foregut lipase), the activities of serum lysozyme and hepatic superoxide dismutase, and liver lipid peroxidation (malondialdehyde concentration) were found. Taking the results of FT1 and FT2 together, it could be concluded that dietary LMA supplementation at low concentrations (0.5–8 g kg^?1) could improve growth and feed utilization, but excess LMA (≥16 g kg^?1) might compromise feed intake and/or feed utilization, thus impairing fish growth. To reduce feed costs in commercial practice, 0.5 g kg^?1 LMA is recommended in the feed of juvenile GIFT tilapia based on the results of this study.     

Electrochemical measurement of hydrogen peroxide in plasma: evaluation of freshwater prawn (Macrobrachium rosenbergii) and crucian carp (Cyprinus carpio) phagocytes under natural conditions

An electrochemical technique for the real-time detection of hydrogen peroxide (H2O2) was employed to describe respiratory burst activity (RBA) of phagocytes in plasma which can be used to evaluate the ability of immune system and disease resistance. The method is based upon the electric current changes, by redox reaction on platinum electrode of extracellular hydrogen peroxide (H2O2) released from phagocytes stimulated by the zymosan at 680 mV direct current (d.c.). Compared with the control, activation of respiratory burst by zymosan particles results in a high amperometric response, and a current peak was obtained during the whole monitoring process. The peak current was proved by addition of Cu2+ and other controls, to be the result of intense release of H2O2 from phagocytes. The peak area was calculated and used to evaluate the quantity of effective H2O2, which represents the quantity of H2O2 beyond the clearance of related enzymes in plasma. According to Faraday's law, the phagocytes' ability of prawns to generate effective H2O2 was evaluated from 1.253 x 10(-14) mol/cell to 6.146 x 10(-14) mol/cell, and carp from 1.689 x 10(-15) mol/cell to 7.873 x 10(-15) mol/cell. This method is an acute and quick detection of extracellular effective H2O2 in plasma and reflects the capacity of phagocytes under natural conditions, which could be applied for selecting species and parents with high immunity for breeding in aquaculture.     

Clock-controlled endogenous melatonin rhythms in Nile tilapia (Oreochromis niloticus niloticus) and African catfish (Clarias gariepinus)

The purpose of this work was to investigate the circadian melatonin system in two tropical teleost species characterized by different behavioral habits, Nile tilapia (diurnal) and African catfish (nocturnal). To do so, fish were subjected to either a control photoperiod (12L:12D), continuous light (LL) or darkness (DD), or a 6L:6D photoperiod. Under 12L:12D, plasma melatonin levels were typically low during the photophase and high during the scotophase in both species. Interestingly, in both species, melatonin levels significantly decreased prior to the onset of light, which in catfish reached similar basal levels to those during the day, demonstrating that melatonin production can anticipate photic changes probably through circadian clocks. Further evidence for the existence of such pacemaker activity was obtained when fish were exposed to DD, as a strong circadian melatonin rhythm was maintained. Such an endogenous rhythm was sustained for at least 18 days in Nile tilapia. A similar rhythm was shown in catfish, although DD was only tested for four days. Under LL, the results confirmed the inhibitory effect of light on melatonin synthesis already reported in other species. Finally, when acclimatized to a short photo-cycle (6L:6D), no endogenous melatonin rhythm was observed in tilapia under DD, with melatonin levels remaining high. This could suggest that the circadian clocks cannot entrain to such a short photocycle. Additional research is clearly needed to further characterize the circadian axis in teleost species, identify and localize the circadian clocks, and better understand the environmental entrainment of fish physiology.