Effectiveness of l‐ascorbyl‐2‐polyphosphate as an ascorbic acid source for sea cucumber,Apostichopus japonicus

A feeding trial was conducted to determine the adequate dietary ascorbic acid (AsA) levels and the effects on growth, meat quality and antioxidant status of sea cucumber (10.04 ± 0.06 g), Apostichopus japonicus. l ‐ascorbyl‐2‐polyphosphate (35% AsA equivalent) was supplemented separately to the basal diet to obtain five AsA levels, 0, 598, 1473, 4676 and 14340 mg kg^?1 diet respectively. After 60‐day feeding trial, the sea cucumbers fed diets containing 598 and 1473 mg AsA kg^?1 showed significantly higher (P < 0.05) body weight gain and specific growth rate values than the sea cucumbers fed control diets. The sea cucumbers fed diets containing 1473 and 4676 mg AsA kg^?1 showed significantly higher (P < 0.05) hydroxyproline contents than those of the sea cucumbers fed diets containing 0 and 598 mg AsA kg^?1. Antioxidant enzymes such as total antioxidant capacity, superoxide dismutase and glutathione peroxidase showed increasing trends with the increasing dietary AsA levels, but no significant differences (P > 0.05) were observed when the sea cucumbers fed diets with high dietary AsA levels. The content of malondialdehyde had the opposite trend of antioxidant enzymes. In conclusion, the adequate dietary AsA level focusing on growth performance of sea cucumber is between 598 and 1473 mg kg^?1 diet. Furthermore, high level of dietary AsA (between 598 and 4676 mg kg^?1 diet) improved meat quality and antioxidant status.     

Effects of dietary chitosan oligosaccharide complex with rare earth on growth performance and innate immune response of turbot,Scophthalmus maximus L.

An 8‐week‐feeding trial was conducted to investigate the effect of dietary chitosan oligosaccharide complex with rare earth (COS‐REE) on growth performance and innate immune response of turbot, Scophthalmus maximus L. (Initial average weight was (12.1 ± 0.1) g) as well as disease resistance against Edwardsiella tarda. Six practical diets (approximately 53.01% protein and 12.57% lipid) were formulated to contain graded levels (0, 75, 150, 300, 600 and 1200 mg kg^?1) of COS‐REE. Results of the present study showed that, compared to the control group (0 mg kg^?1), the specific growth rate (SGR) was significantly higher in fish fed the diet with 300 mg kg^?1 COS‐REE (P < 0.05), while the feed conversion ratio (FCR) significantly decreased (P < 0.05). The phagocytic index (PI) and the activity of super oxide dismutase (SOD) of serum in fish fed the diet with 300 mg kg^?1 COS‐REE was significantly higher than fish fed the control diet (P < 0.05), but no significant differences were observed in malondialdehyde (MDA) and hepatic metallothionein (MT) concentrations. After 8 weeks, fish were challenged by intraperitoneal injection with E. tarda, and COS‐REE‐treated fish demonstrated increased protection capability. These results suggested that COS‐REE could enhance growth, innate immunity and disease resistance in turbot, and the optimum dose was approximately 300 mg kg^?1.     

Effects of dietary heat‐killed Lactobacillus plantarum L‐137 (HK L‐137) on the growth performance,digestive enzymes and selected non‐specific immune responses in sea cucumber,Apostichopus japonicus Selenka

A feeding trial was conducted to evaluate the efficacy of dietary heat‐killed Lactobacillus plantarum L‐137 (HK L‐137) on growth performance, digestive, non‐specific immune and phagocytosis of sea cucumber, Apostichopus japonicus. Sea cucumbers (initial body weight 1.35 ± 0.04 g) were fed diets supplemented with five levels of HK L‐137 (0, 0.005, 0.025, 0.05 and 0.25 g HK L‐137 kg^?1 diets) for 60 days. Results indicated sea cucumbers fed with diets containing 0.05 and 0.25 g HK L‐137 kg^?1 diets showed significantly (P < 0.05) higher body weight gain and specific growth rate than other groups. Sea cucumbers fed with diets containing 0.05 and 0.25 g HK L‐137 kg^?1 diets showed significantly (P < 0.05) higher protease activity than control group. Higher amylase, lysozyme and phagocytic activities were found in 0.25 g HK L‐137 kg^?1 diet group. Higher superoxide dismutase enzyme and alkaline phosphatase activity was found in 0.05 g HK L‐137 kg^?1. While no significant differences (P > 0.05) were found in acid phosphatase activity. These results suggested that dietary supplementation of 0.05 g HK L‐137 kg^?1 diets would have benefit on growth, digestive enzymes and several non‐specific immune parameters of sea cucumber.     

Total organic carbon budget of integrated aquaculture system of sea cucumber Apostichopus japonicus,jellyfish Rhopilema esculenta and shrimp Fenneropenaeus chinensis

The total organic carbon (TOC) budget of the integrated aquaculture system of sea cucumber Apostichopus japonicus, jellyfish Rhopilema esculenta and shrimp Fenneropenaeus chinensis was investigated in a cofferdam (1.202 km²) for 13‐month field study. The results showed that the total input of TOC to the system was 498 300 kg year^?1, of which photosynthesis accounted for 26.8% of input TOC. The TOC production of the sea cucumber was 3668 kg year^?1, and that of the jellyfish and the shrimp was 714 kg year^?1. The TOC utilization rate of the input TOC by sea cucumber production was 7.4‰, and that of the integrated system increased by 18.9% than that of sea cucumber monoculture model through integration with jellyfish and shrimp. The outflow TOC from the system was 336 600 kg year^?1, which account for 92.7% of the inflow TOC. The integrated aquaculture system not only produce aquatic products, but also eliminate 7.3% of the inflow TOC, therefore, this system was not only a production system of aquatic products, but also an organic matter purification system for the coastal environment. The TOC utilization rate of the system can be probably improved by means of increasing stocking density of the sea cucumber and/or the shrimp.     

Vitamin E requirement of sea cucumber (Apostichopus japonicus) and its' effects on nonspecific immune responses

This feeding trial was conducted to determine the vitamin E requirement of sea cucumber (Apostichopus japonicus) and to study the effects of vitamin E on nonspecific immune parameters. Commercial sea cucumber diets supplemented with 0, 50, 100, 200, 500 or 1000 mg kg^?1 vitamin E were fed to juvenile sea cucumber (7.96 ± 0.01 g) for 60 days. Specific growth rate and total coelomocyte counts were maximized in sea cucumber fed diets supplemented with 100 mg kg^?1 vitamin E. Reactive oxygen species production was maximized at 50 mg kg^?1 vitamin E, whereas superoxide dismutase activity was the highest in sea cucumber fed diets supplemented with 100 mg kg^?1 vitamin E. The vitamin E concentration in the body wall of the sea cucumber increased as the vitamin E level in the diet increased. Analysis of specific growth rate and total coelomocyte counts of the sea cucumber indicated that 88–92 mg kg^?1 vitamin E is required for maximal growth and nonspecific immune responses of A. japonicus and that 114.7 mg kg^?1 vitamin E is required to minimize lipid peroxidation levels.     

Optimum dietary protein level for growth and coelomic fluid non‐specific immune enzymes of sea cucumber Apostichopus japonicus juvenile

A feeding trial was conducted to determine the optimum dietary protein level of sea cucumber Apostichopus japonicus juvenile focusing on growth performance and non‐specific immune response. Diets with seven crude protein levels (42.0, 108.9, 155.2, 216.7, 258.0, 313.3 and 357.5 g kg^?1) were fed to sea cucumber juveniles (1.05 ±0.01 g) once a day for 100 days. More than 70% survival was observed, and there was no significant difference among all treatments. The sea cucumbers fed diets containing 108.9 g kg^?1 crude protein showed significantly (P < 0.05) higher body weight gain than those of the sea cucumbers fed diets containing 42.0, 216.7, 258.0, 313.3 and 357.5 g kg^?1 crude protein. No significantly differences (P > 0.05) were observed in moisture, crude protein, crude lipid, ash and carbohydrate content of the body wall among all treatments. The coelomic fluid catalase activity of the sea cucumbers generally increased with increasing dietary protein levels. Therefore, the acid phosphatase, superoxide dismutase and lysozyme activity increased with increasing dietary protein levels at first and decreased subsequently. The relationship between dietary protein levels and body weight gain was analysed by a second‐order polynomial regression analysis model. The result indicates that the optimum dietary protein level for sea cucumber juveniles is 135.4 g kg^?1.     

Effects of dietary folic acid on growth,antioxidant capacity,non‐specific immune response and disease resistance of juvenile Chinese mitten crab Eriocheir sinensis (Milne‐Edwards, 1853)

An 8‐week study was conducted to determine folic acid requirement and its effect on antioxidant capacity and immunity in juvenile Chinese mitten crab Eriocheir sinensis (Milne‐Edwards, 1853), followed by a challenge assay with the pathogen Aeromonas hydrophila for 2 weeks. Folic acid was added to a basal diet at seven levels (0, 0.5, 1.0, 2.0, 4.0, 8.0, 16.0 mg folic acid kg^?1 diet), and a diet free of folic acid and vitamin B₁₂ was also included as a control. Crabs were fed twice daily in 32 tanks with 7.76–8.17 mg oxygen L^?1, 25.0–31.0 °C and 7.5–8.3 pH. Growth and feed efficiency were significantly greater in crabs fed ≥2.0 mg folic acid kg^?1, but not significantly different between crabs fed diets >2.0 mg folic acid. The superoxide dismutase activity and glutathione S‐transferase activity were highest in crabs fed ≥2.0 mg folic acid kg^?1, followed by those fed 0.5 and 1.0 mg folic acid kg^?1, and the control diet. The malondialdehyde content was highest in crabs fed the control diet, followed by those fed 0 mg folic acid kg^?1, and the lowest value occurred in those fed ≥0.5 mg folic acid kg^?1. Phenoloxidase activity and total haemocytes were significantly higher in crabs fed ≥2.0 mg folic acid kg^?1 than other diets. Crabs fed 2.0 mg folic acid kg^?1 had the highest lysozyme, acid phosphatase and alkaline phosphatase activities but the lowest cumulative mortality. The optimum dietary folic acid requirement by E. sinensis was estimated at 2.29–2.90 mg kg^?1 diet.     

Survival and growth of the sea cucumber Holothuria leucospilota Brandt: a comparison between suspended and bottom cultures in a subtropical fish farm during summer

The feasibility of co‐culturing the sea cucumber Holothuria leucospilota Brandt in a subtropical fish farm was investigated in a field study. Sea cucumbers were cultured in the fish farm in cages suspended at 4 m deep (suspended culture) and directly on the seafloor (bottom culture). The survival and growth of the sea cucumbers were monitored twice during the 3‐month, summer experimental period (May 26–August 14, 2010). Results showed that the suspension‐cultured sea cucumbers exhibited excellent survival rate (100%) during the whole study period. There also occurred no mortality in the bottom‐cultured sea cucumbers during the first culture period (May 26–July 13); but all these died from anoxia caused by water column stratification during the second culture period (July 14–August 14). The specific growth rate of the bottom‐cultured sea cucumbers (1.05 ± 0.21 % day^?1) was nearly double that of the suspended culture animals (0.57 ± 0.21 % day^?1) during the first culture period, and the growth rates of the suspended culture sea cucumbers in the second culture periods (0.46 ± 0.24 % day^?1) was only a little lower than that of the first period. The sea cucumbers H. leucospilota could ingest and assimilate sediment with high organic matter content with an average assimilation efficiency of 14.9 ± 3.9%. This study indicated that fish farm detritus can be effectively used as a food source for the sea cucumber and that it can be turned into a valuable secondary crop in the form of the sea cucumber biomass.     

Acute toxicity of ammonia in Pacu fish (Piaractus mesopotamicus,Holmberg, 1887) at different temperatures levels

Piaractus mesopotamicus juveniles (total length 12 ± 0.5 mm) were exposed to different concentrations of ammonia‐N (un‐ionized plus ionized ammonia as nitrogen), using the static renewal method at different temperature levels (15, 20 and 25°C) at pH 7. The 24, 48, 72, 96 h LC₅₀ values of ammonia‐N in P. mesopotamicus juveniles were 5.32, 4.19, 3.79 and 2.85 mg L^?1 at 15°C; 4.81, 3.97, 3.25 and 2.50 mg L^?1 at 20°C; and 4.16, 3.79, 2.58 and 1.97 mg L^?1 at 25°C respectively. The 24, 48, 72, 96 h LC₅₀ values of NH₃‐N (un‐ionized ammonia as nitrogen) were 0.018, 0.014, 0.013, 0.009 mg L^?1 at 15°C temperature; 0.023, 0.019, 0.016 and 0.012 mg L^?1 at 20°C; 0.029, 0.026, 0.018 and 0.014 mg L^?1 at 25°C. The temperature increase from 15 to 25°C caused an increase of ammonia‐N susceptibility by 21.80%, 9.55%, 31.92% and 30.87%, after 24, 48, 72 and 96 h exposure respectively. Furthermore, we found that exposure of fish to ammonia‐N caused an elevation in total haemoglobin and blood glucose with an increase of 2 mg L^?1 concentration. Ammonia levels tolerated, especially in different temperatures levels, have important implications for the management of aquaculture.     

Immune responses of Litopenaeus vannamei to non‐ionic ammonia stress: a comparative study on shrimps in freshwater and seawater conditions

To investigate the effect of non‐ionic ammonia (NH₃‐N) stress (0.1 and 0.5 mg L^?1) on the immunity of Litopenaeus vannamei cultured in long‐term freshwater, the total haemocyte count (THC), the activity of phenoloxidase (PO), nitric oxide synthase (NOS), superoxidase dismutase (SOD) and the content of malondialdehyde (MDA) were determined and further compared with those of seawater shrimps. The results showed that NH₃‐N stress significantly reduced THC and the activity of PO and SOD (P < 0.05). Under 0.1 mg L^?1 NH₃‐N stress, NOS activity increased first and then decreased significantly, while it dropped dramatically under 0.5 mg L^?1 NH₃‐N stress (P < 0.05). During NH₃‐N stress, MDA content increased continuously, and the MDA content in hepatopancreas of freshwater shrimps was higher than that of seawater shrimps. It was concluded that NH₃‐N stress significantly influenced the non‐specific immunity and could also upset the balance of antioxidant system of L. vannamei in both freshwater and seawater shrimps. Compared with in seawater, the shrimps in freshwater were more vulnerable to NH₃‐N stress because of higher lipid peroxidation and lower immunity.     

Effects of continuous and alternate administration of β‐glucan and mannan‐oligosaccharide on the growth,immunity and resistance against Vibrio splendidus of sea cucumber Apostichopus japonicus (Selenka)

A 4‐week growth trial was conducted to compare the effects of different feeding strategies of dietary immunostimulants on the growth, immunity and resistance against Vibrio splendidus of sea cucumbers Apostichopus japonicus (Selenka). Six feeding strategies were set, including feeding immunostimulants‐free diet continuously (control), feeding dietary β‐glucan (1.25 g kg^?1 diet) continuously, feeding dietary mannan‐oligosaccharides (MOS; 2.00 g kg^?1 diet) continuously, feeding β‐glucan 2 days followed by MOS 5 days alternately, feeding β‐glucan 5 days followed by MOS 2 days alternately and feeding β‐glucan 7 days followed by MOS 7 days alternately. The sea cucumbers fed immunostimulants showed higher specific growth rate (SGR) and lower cumulative mortality than control (P < 0.05). When sea cucumbers were fed with β‐glucan continuously, total coelomocytes counts and superoxide anion were significantly higher than control on the 4th day (P < 0.05). However, these two immune parameters were not significantly higher than those in control after the 18th day (P > 0.05). While sea cucumbers continuously fed MOS, these two immune parameters were not significantly higher than control until the 15th day. All immune parameters of the sea cucumbers fed with β‐glucan and MOS alternately were significantly higher than those in control during the experiment (P < 0.05). The sea cucumbers fed with β‐glucan 7 days followed MOS 7 days alternately showed the highest SGR and second lowest cumulative mortality. It was suggested that this feeding strategy is most suitable for sea cucumbers.     

Acute and chronic effects of aqueous ammonia on marbled spinefoot rabbitfish,Siganus rivulatus (Forsskål 1775)

Ammonia is a metabolite of aquatic organisms which might reach deleterious levels in intensive fish farms. The aim of the present study was to determine median lethal concentrations (96‐h LC₅₀) of total ammonia nitrogen (TA‐N) on marbled spinefoot rabbitfish (Siganus rivulatus) and chronic effects of TA‐N on survival, growth and behaviour of juvenile rabbitfish over a 50 day period. In the first experiment, fish were exposed to 0, 2, 4, 6, 8, 10, 12, 14, 16, 18 and 20 mg L^?1 TA‐N for 96 h and survival evaluated. In the second experiment, 12 fish were stocked per 50‐L tank and treated with one of 0, 2, 4, 6, 8, 10 and 12 mg L^?1 TA‐N with three replicate tanks per treatment. Survival and growth were determined and histopathological alterations of gills due to chronic ammonia exposure were studied by light and electron microscopy. The 96‐h LC₅₀ values were 16–18 mg L^?1 TA‐N. In the chronic exposure experiment, fish reared in water with 0 mg L^?1 TA‐N had 100% survival and had 50% weight increase in 50 days. Fish at 2 and 4 mg L^?1 TA‐N all died whilst fish in 6, 8, 10 and 12 mg L^?1 TA‐N survived and grew albeit less than in treatment 0 mg L^?1. Gills from ammonia treated fish displayed severe histological and ultrastructural alterations including hyperplasia, hypertrophy and fusion of secondary lamellae, aneurysms and presence of pleomorphic altered cells. Chronic exposure to ammonia is deleterious to marbled spinefoot rabbitfish and low concentrations of ammonia appear to kill the fish in <50 days whilst fish can survive for more than 50 days at concentrations between 6 and 12 mg L^?1 TA‐N.     

Effect of different total suspended solids levels on a Litopenaeus vannamei (Boone, 1931) BFT culture system during biofloc formation

In a Biofloc Technology System (BFT), there is constant biofloc formation and suspended solids accumulation, leading to effects on water quality parameters that may affect the growth performance of cultured shrimp. This study aimed to analyse during biofloc formation the effect of different total suspended solids (TSS) levels on water quality and the growth performance of Litopenaeus vannamei shrimp in a BFT system. A 42‐day trial was conducted with treatments of three ranges of TSS: 100–300 mg L^?1 as low (TL), 300–600 as medium (TM) and 600–1000 as high (TH). The initial concentrations of 100 (TL), 300 (TM) and 600 mg L^?1 (TH) were achieved by fertilization before starting the experiment. Litopenaeus  vannamei juveniles with an average weight of 4.54 ± 1.19 g were stocked at a density of 372 shrimp m^?3. Physical and chemical water parameters and shrimp growth performance were analysed. After 6 weeks, TSS mean concentrations were 306.37, 532.43 and 745.2 mg L^?1 for, respectively, TL, TM and TH treatments. Significant differences (P < 0.05) were observed in TSS, settleable solids, pH, alkalinity and nitrite, especially between the TL and TH treatments. Similarly, differences (P < 0.05) were observed in the growth performance parameters, specifically final weight, survival, feed conversion and productivity. The water quality parameters at lower range of total suspended solids concentration (TL) treatment resulted in a better performance of L. vannamei in the BFT system. The maintenance at range of 100–300 mg L^?1 TSS is thus important to the success of shrimp culture.     

Effects of dietary lipid level on growth performance,body composition and digestive enzymes activity of juvenile sea cucumber,Apostichopus japonicus

A feeding trial aimed to determine the effects of dietary lipid level on growth performance, body composition and digestive enzymes activity of juvenile sea cucumber, Apostichopus japonicus. Diets with six crude lipid levels (1.9, 13.8, 29.1, 43.6, 59.6 and 71.6 g kg^?1) were fed to sea cucumbers (initial weights 0.65 ± 0.01 g) at a density of 30 juveniles, once a day. After 60 days, body weight gain (BWG), specific growth rate (SGR), feed intake (FI) and protein efficiency ratio (PER) decreased with increasing dietary lipid levels. The sea cucumbers fed 1.9 g kg^?1 crude lipid showed significantly higher (P < 0.05) BWG than those of the sea cucumbers fed 59.6 and 71.6 g kg^?1 crude lipid. Intestinal protease and lipase activities generally increased with increasing dietary lipid levels. Eicosapentaenoic acid (EPA) content of body walls generally increased with increasing dietary lipid levels. Docosahexaenoic acid (DHA) content of body walls reached the maximum value at a dietary lipid level of 13.8 g kg^?1. N‐3 highly unsaturated fatty acid content followed the same pattern of DHA. According to the growth performance and body composition of sea cucumbers, it can be indicated that the optimum dietary lipid level for juvenile sea cucumbers is between 1.9 and 13.8 g kg^?1.     

Effectiveness of copper sulphate,potassium permanganate and peracetic acid to reduce mortality and infestation of Ichthyobodo necator in channel catfish Ictalurus punctatus (Rafinesque 1818)

Ichthyobodo necator is a single‐celled biflagellate parasite, which in high density can cause significant mortality in young fish. Copper sulphate (CuSO₄), potassium permanganate (KMnO₄) and peracetic acid (PAA) were evaluated for effectiveness against ichthyobodosis. Treatments were: untreated control, 2.1 mg L^?1CuSO₄, 3.0 mg L^?1 KMnO₄, 1.5 mg L^?1 PAA and 3.0 mg L^?1 PAA, and were applied to flow‐through tanks on three consecutive days. The study was designed to simulate the flow‐through systems utilized in the commercial rearing of juvenile channel catfish (Ictalurus punctatus). Mortality was monitored daily to compare survival rate among treatments. Parasite intensity was assessed pre chemical exposure and 20–24 h after the third application to determine effectiveness of the treatment. An assessment was also done 7 days post application to investigate possible reoccurrence. Copper sulphate, KMnO₄ and PAA (3.0 mg L^?1) significantly reduced the infestation rate of I. necator. Copper sulphate significantly improved the survival of I. necator infested channel catfish after three flow‐through applications compared with the untreated control. The 3.0 mg L^?1 PAA resulted in significantly lower survival than the untreated control, the 1.5 mg L^?1 PAA and the KMnO₄ were not statistically different from the untreated control.     

Acute toxicity of nitrite on white shrimp Litopenaeus vannamei (Boone) juveniles in low‐salinity water

The nitrite toxicity was estimated in juveniles of L. vannamei. The 24, 48, 72 and 96 h LC₅₀ of nitrite‐N on juveniles were 8.1, 7.9, 6.8 and 5.7 mg L^?1 at 0.6 g L^?1; 14.4, 9.6 8.3 and 7.0 mg L^?1 at 1.0 g L^?1; 19.4, 15.4, 13.4 and 12.4 mg L^?1 at 2.0 g L^?1 of salinity respectively. The tolerance of juveniles to nitrite decreased at 96 h of exposure by 18.6% and 54.0%, when salinity declined from 1.0 to 0.6 g L^?1 and from 2.0 to 0.6 g L^?1 respectively. The safe concentrations at salinities of 0.6, 1.0 and 2.0 g L^?1 were 0.28, 0.35 and 0.62 mg L^?1 nitrite‐N respectively. The relationship between LC₅₀ (mg L^?1), salinity (S) (g L^?1) and exposure time (T) (h) was LC₅₀ = 8.4688 + 5.6764S – 0.0762T for salinities from 0.6 to 2.0 g L^?1 and for exposure times from 24 to 96 h; the relationship between survival (%) and nitrite‐N concentration (C) for salinity of 0.6–2.0 g L^?1, nitrite‐N concentrations of 0–40 mg L^?1 and exposure times from 0 to 96 h was as follows: survival (%) = 0.8442 + 0.1909S – 0.0038T – 0.0277C + 0.0008ST + 0.0001CT–0.0029SC, and the tentative equation for predicting the 96‐h LC₅₀ to salinities from 0.6 to 35 g L^?1 in L. vannamei juveniles (3.9–4.4 g) was 96‐h LC₅₀ = 0.2127 S² + 1.558S + 5.9868. For nitrite toxicity, it is shown that a small change in salinity of waters from 2.0 to 0.6 g L^?1 is more critical for L. vannamei than when wider differences in salinity occur in brackish and marine waters (15–35 g L^?1).     

Bioassay‐guided isolation and identification of active compounds from Macleaya microcarpa (Maxim) Fedde against fish pathogenic bacteria

Four alkaloids (Sanguinarine, 6‐Methoxyl‐dihydro‐chelerythrine, Cryptopine and β‐Allocryptopine) were isolated from aerial parts of Macleaya microcarpa (Maxim) Fedde using bioassay‐guided isolation method, and the inhibitory activity of ethanolic extract, various fractions and these four alkaloids against four fish pathogenic bacteria (Aeromonas hydrophila, Aeromonas salmonicida, Vibrio anguillarum and Vibrio harveyi) was assessed in vitro using the agar dilution method and the microdilution assay method respectively. A. hydrophila was the most sensitive strain to all the tested compounds. Minimum inhibitory concentration (MIC) values were lower for sanguinarine against all tested Gram‐negative strains than other three alkaloids, with MIC values of 12.5 mg L^?1 for A. hydrophila and 50 mg L^?1 to other pathogenic bacteria. Followed by 6‐methoxyl‐dihydro‐chelerythrine, which showed considerable antibacterial activity with MIC values of 80 mg L^?1 for A. hydrophila, 100 mg L^?1 for V. harveyi, and 125 mg L^?1 for both V. anguillarum and A. salmonicida. Cryptopine and β‐allocryptopine revealed similar inhibitory activity with MIC values of 100 mg L^?1 for A. hydrophila and 200 mg L^?1 for other three bacterial species. These finding provided evidence that extract, as well as isolated compounds from M. microcarpa might be potential sources novel antibacterial agents for the treatment of fish infectious diseases.     

Acute toxicity of ammonia in Meagre (Argyrosomus regius Asso, 1801) at different temperatures

Argyrosomus regius (3.0 ± 0.9 g) were exposed to different concentrations of ammonia in a series of acute toxicity tests by the static renewal method at three temperature levels (18, 22 and 26°C) at a pH of 8.2. Low temperature clearly increased the tolerance of the fish to total ammonia nitrogen (TAN) and unionized ammonia (NH₃) (P < 0.05). While the 96‐h LC₅₀ values of TAN were 19.79, 10.39 and 5.06 mg L^?1, the 96‐h LC₅₀ of NH₃ were 1.00, 0.70 and 0.44 mg L^?1 at 18, 22 and 26°C respectively. The safe levels of NH₃ for A. regius was estimated to be 0.10, 0.07 and 0.04 mg L^?1 at 18, 22 and 26°C respectively (P < 0.05). This study clearly indicates that A. regius is more sensitive to ammonia than other marine fish species cultured on the Mediterranean and Eastern Atlantic coasts.     

The efficacy of MS‐222 as anaesthetic agent in four freshwater aquarium fish species

The efficacy of anaesthetic tricaine methanesulfonate (MS‐222) was evaluated in four freshwater aquarium fish species, Zebrafish (Danio rerio), Guppy (Poecilia reticulata), Discu (Symphysodon discus) and Green swordtail (Xiphophorus helleri). The correct dose of anaesthetic should induce the plane 4 of anaesthesia in less than 180 s, recovery in less than 300 s and must survive when exposed during 30 min to anaesthetic. Fishes were exposed to six concentrations of anaesthetic (75, 100, 125, 150, 200 and 250 mg L^?1) and the time of fish reaching plane 4 of anaesthesia, post exposure recovery, and the percentage of survival when fish were subject to 30 min in the anaesthetic were recorded. The optimal doses varied according to the species: D. rerio – 75, 100 and 125 mg L^?1, P. reticulata – 125, 150 and 200 mg L^?1, S. discus – 75 and 100 mg L^?1 and X. helleri – 125 and 150 mg L^?1. The induction time generally decreased significantly with increasing concentration of MS‐222 for all of the species evaluated. The recovery time had a tendency to increase with the increase of the MS‐222 concentration for D. rerio, P. reticulata and S. discus. On the other hand, X. helleri recovery time decreased with the increase of MS‐222 concentration. MS‐222 proved to be effective in anaesthesia for all the freshwater ornamental species studied. The main results clearly show that the optimal dose to anesthetize is fish species dependent and it is completely wrong to extrapolate optimal anaesthetic concentrations between different species.     

Preliminary success using hydrogen peroxide to treat Atlantic salmon,Salmo salar L., affected with experimentally induced amoebic gill disease (AGD)

Currently, the only effective and commercially used treatment for amoebic gill disease (AGD) in farmed Tasmanian Atlantic salmon is freshwater bathing. Hydrogen peroxide (H₂O₂), commonly used throughout the aquaculture industry for a range of topical skin and gill infections, was trialled in vitro and in vivo to ascertain its potential as an alternative treatment against AGD. Under in vitro conditions, trophozoites of Neoparamoeba perurans were exposed to three concentrations of H₂O₂ in sea water (500, 1000 and 1500 mg L^?1) over four durations (10, 20, 30 and 60 min) each at two temperatures (12 and 18 °C). Trophozoite viability was assessed immediately post‐exposure and after 24 h. A concentration/duration combination of 1000 mg L^?1 for >10 min demonstrated potent amoebicidal activity. Subsequently, Atlantic salmon mildly affected with experimentally induced AGD were treated with H₂O₂ at 12 and 18 °C for 15 min at 1250 mg L^?1 and their re‐infection rate was compared to freshwater‐treated fish over 21 days. Significant differences in the percentage of filaments affected with hyperplastic lesions (in association with amoebae) and plasma osmolality were noted between treatment groups immediately post‐bath. However, the results were largely equivocal in terms of disease resolution over a 3‐week period following treatment. These data suggest that H₂O₂ treatment in sea water successfully ameliorated a clinically light case of AGD under laboratory conditions.