In vivo evaluation of the effects of commercial Bacillus probiotics on survival and development of Litopenaeus vannamei larvae during the early hatchery period

Three in vivo experiments were conducted to measure the effectiveness of commercial Bacillus probiotics on survival and development of Litopenaeus vannamei larvae from nauplii 3–4 (N_3–4) to zoea 3 (Z₃) stages. Experiment I: Nine commercial Bacillus probiotics were individually added to larvae twice at N₆ and Z₁ at levels of 2, 20, and 100 mg L^?1. Only six of nine products at 20 mg L^?1 exhibited higher or significantly higher larval survival (P < 0.05) than the control. Experiment II: Two superior products from the first experiment were administered to larvae with six dose frequencies at 20 mg L^?1. For both products, three doses, once at each of N_3–4, N₆ and Z₁ stages, yielded the best larval survival and development rates (P < 0.05), and these were confirmed by enhanced activities of tryptase and amylase. Experiment III: The isolates of these two products, identified as Bacillus subtilis and Bacillus licheniformis, were delivered to larvae singly at concentrations of 1.0 × 10⁸ and 1.0 × 10⁹ CFU L^?1, or at the same concentrations by mixing the two equally. At 1.0 × 10⁹ CFU L^?1, B. subtilis exerted more beneficial effects on larvae than B. licheniformis or the mixture. Therefore, the optimal dosage and dose frequency of commercial Bacillus products should be evaluated prior to large‐scale application in shrimp hatcheries to avoid futility or even adverse effects, as spore counts and non‐bacterial ingredients are key factors influencing the efficacy of Bacillus probiotics.     

Efficacy of formalin,iodine and sodium chloride in improvement of egg hatching rate and fry survival prior to the onset of exogenous feeding in yellow perch

Egg disinfection is considered the most important routine work in hatcheries to avoid fungal and/or bacterial infection of fish eggs. The aim of this study was to determine the effectiveness of three disinfectants: formalin, iodine and sodium chloride on the hatching success of yellow perch eggs. The disinfectants were tested in triplicate at different concentrations for 15 and 30 min bath treatments. Two experiments were conducted; formalin at five concentrations (25, 50, 100, 150 and 200 mg L^?1) and 25 mg L^?1 iodine were tested in the first experiment. The second experiment involved formalin at three concentrations (250, 500 and 1000 mg L^?1), iodine at three concentrations (50, 100 and 250 mg L^?1) and sodium chloride at three concentrations (500, 1000 and 3000 mg L^?1) were used. Iodine and sodium chloride‐treated eggs hatched earlier than formalin‐treated eggs. The highest mean percentage of eyed stage, hatching rate and survival to first feeding fry was observed at 200 mg L^?1 formalin for 30 min, 50 mg L^?1 iodine for 15 min and 500 mg L^?1 sodium chloride for 30 min. High concentrations of formalin (1000 mg L^?1), iodine (250 mg L^?1) and sodium chloride (1000 and 3000 mg L^?1) showed toxicity to yellow perch eggs, resulting in low hatching rate and survival to first feeding fry. We recommended formalin at a concentration of 150–200 mg L^?1 for 30 min as an effective, easily available and low‐cost disinfectant for routine use to improve yellow perch hatchability.     

Morphological deformities in mud crab Scylla serrata juveniles exposed to antibiotics during the larval stage

The effects of antibiotics on the external deformities, growth and survival of mud crab Scylla serrata larvae and juveniles were determined. Zoeae were exposed to oxytetracycline (OTC) (0, 3.0, 6.0, 9.0, 12 mg L^?1) and furazolidone (FZD) (0, 0.5, 1.0, 1.5, 2.0 mg L^?1) in the first and second experiments, respectively, until the late megalopa. The crab instars were grown in nursery tanks for 1 month. Larvae survived until megalopa only at 3.0 and 6.0 mg L^?1 OTC or 0.5 and 1.0 mg L^?1 FZD. These four concentrations were run simultaneously in another experiment. Morphological deformities in zoea 5 were bent dorsal, rostral and furcal spines. There was no significant difference (P > 0.05) on the deformities of zoea 5 in 3.0 and 6.0 mg L^?1 OTC and 0.5 and 1.0 mg L^?1 FZD. Significantly (P < 0.05) higher survival and faster growth were attained in 3.0 mg L^?1 OTC and 0.5 mg L^?1 FZD. Deformities observed in juveniles were fused frontal and lateral spines, asymmetrical and depressed tip of abdominal flap and gap between sternites. High percentage occurrence of deformities was observed in the 6.0 mg L^?1 OTC and 1.0 mg L^?1 FZD in the first and third experiments, respectively. There was no significant difference (P > 0.05) observed in the survival of juveniles in OTC and FZD treatments. However, growth was significantly (P < 0.05) faster in lower concentrations of the two antibiotics. The study shows the effects of OTC and FZD in the morphology of mud crab. Therefore, there is a need to eliminate the use of antibiotics and find alternatives.     

Efficacy of calcein as a chemical marker of green‐lipped mussel (Perna canaliculus) larvae and its potential use for tracking larval dispersal

An optimal chemical shell marking protocol was developed for the New Zealand green‐lipped mussel, Perna canaliculus with a view to its future use in larval tracking experiments. Larval P. canaliculus aged either 10, 15 or 19 days post fertilization were immersed in treatments of 50, 100 and 200 mg L^?1 of calcein for a period of 24 h before measurements of shell mark brightness were taken. There was 100% marking success in all calcein treatments for all age classes, with 19‐day larvae immersed in 200 mg L^?1 calcein producing the brightest mark. Growth was not affected by calcein immersion; however, 10‐day larvae exhibited significantly higher levels of mortality compared with 15‐ and 19‐day larvae suggesting a reduced resilience to the marking protocols in younger larvae. In a mass staining experiment, a solution of 100 mg L^?1 calcein was used to successfully stain15.6 million hatchery reared P. canaliculus larvae. Calcein, therefore, offers a low impact method with which to stain the sensitive early life stages of this species thus providing a rapid method for identifying individuals of interest, i.e. individuals released in the wild or specific family lines within a hatchery environment.     

Effects of incubation water hardness and salinity on egg hatch and fry survival of Nile tilapia Oreochromis niloticus (Linnaeus)

This study examined the effects of water hardness and salinity on yolk sac larvae and swim‐up fry survival of Nile tilapia, Oreochromis niloticus (Chitralada strain), eggs during artificial incubation. Four experiments were conducted to evaluate the effects of hardness, salinity and the sources of saline incubation water. High water hardness treatments (500–4200 mg L^?1 as CaCO₃) resulted in higher yolk sac larvae and swim‐up fry survival than low water hardness treatments (50.0 and 132 mg L^?1 as CaCO₃); although yolk sac larvae and swim‐up fry survival did not differ among the high or low hardness treatments. Salinity of 4.0 g L^?1 using seawater, and 4.0 and 8.0 g L^?1 using unprocessed common salt resulted in the higher survival rate of yolk sac larvae and swim‐up fry than other salinity treatments. Yolk sac larvae and swim‐up fry survival was found to decrease with the increase in salinity and increase with the increase in water hardness. The present study demonstrated the positive effects of increased water hardness level (>132 mg L^?1) on yolk sac larvae and swim‐up fry survival. The study also showed that seawater salinity of 4 g L^?1 was the most appropriate salinity level for incubating Nile tilapia eggs.     

Ammonia tolerance of Litopenaeus vannamei (Boone) larvae

The tolerance of Litopenaeus vannamei larvae to increasing concentrations of total ammonia nitrogen (TAN) using a short‐term static renewal method at 26°C, 34 g L^?1 salinity and pH 8.5 was assessed. The median lethal concentration (24 h LC₅₀) for TAN in zoea (1‐2‐3), mysis (1‐2‐3) and postlarvae 1 were, respectively, 4.2‐9.9‐16.0; 19.0‐17.3‐17.5 and 13.2 mg L^?1TAN (0.6‐1.5‐2.4; 2.8‐2.5‐2.6 and 1.9 mg L^?1 NH₃‐N). The LC₅₀ values obtained in this study suggest that zoeal and post‐larval stages are more sensitive to 24 h ammonia exposure than the mysis stage of L. vannamei larvae. On the basis of the ammonia toxicity level (24 h LC₅₀) at zoea 1, we recommend that this level does not exceed 0.42 mg L^?1 TAN – equivalent to 0.06 mg L^?1 NH₃‐N – to reduce ammonia toxicity during the rearing of L. vannamei larvae.     

Effect of Sargassum wightii fucoidan on growth and disease resistance to Vibrio parahaemolyticus in Penaeus monodon post‐larvae

The polysaccharide – fucoidan was extracted from brown seaweed Sargassum wightii and its antibacterial activity was screened by agar well diffusion method. The maximum zone of inhibition observed was 15.66 mm in 20 mg mL^?1 concentration against Vibrio parahaemolyticus. The Minimum Inhibitory Concentration (MIC) of the fucoidan was 12 mg mL^?1 against V. parahaemolyticus. The fucoidan was then enriched with Artemia nauplii at four different concentrations such as 100, 200, 300 and 400 mg L^?1 for 12 h. The enriched Artemia nauplii were fed to Penaeus monodon post‐larvae for 20 days and the growth performance was assessed. The weight gain and SGR of the control group were 0.2432 g and 15.78%, respectively. But, in experimental groups fed with fucoidan enriched Artemia nauplii, the weight gain and SGR were increased and were respectively ranged from 0.2602 to 0.3161 g and from 16.11 to 17.05%. The P. monodon post‐larvae were challenged with V. parahaemolyticus for a period of 30 days showed a reduction in mortality percentage of experimental groups over the control group and it was ranged between 36.97 and 89.86%. During the challenge test, the V. parahaemolyticus load was also enumerated from the infected shrimp at every 10 day intervals. In the control group, the Vibrio load showed a linear increase in hepatopancreas and muscle tissues from 10th to 30th days of challenge test, whereas in the experimental groups, the Vibrio load established a declining trend with the advancement of challenged test.     

Survival and respiration of marbled rabbitfish (Siganus rivulatus) fingerlings at various oxygen tensions

Marbled rabbitfish, Siganus rivulatus, is an economically valuable herbivorous fish and a potential candidate for warmwater aquaculture. This study was carried out to: (1) assess the effect of various oxygen concentrations on survival and behaviour of S. rivulatus fingerlings and (2) investigate the response of S. rivulatus to hypoxia and determine its critical oxygen tension (P_crit). In the first experiment, groups of rabbitfish (15 fish per group) were maintained for 1 h in waters of various oxygen concentrations. They were then transferred to well‐aerated tanks and observed for 72 h. Survival was recorded, fish behaviour at low oxygen concentrations observed, and LC₅₀ after 1‐h hypoxia and 72‐h recovery evaluated. In the second experiment, a series of stop‐flow respirometry experiments were performed during which dissolved oxygen was allowed to drop to 0.5 mg L^?1 and respiration rate recorded at various oxygen concentrations. In the first experiment, all fish survived for 1 h at oxygen concentration of 1.44 mg L^?1 and greater, but started dying at oxygen concentrations below 0.65 mg L^?1 (16% survival). The LC₅₀ of S. rivulatus fingerlings was 0.6 mg L^?1. Results of the second experiment showed that S. rivulatus is an oxyregulator until P_crit (1.7 mg L^?1 O₂) is reached, becoming an oxyconformer below this concentration. Findings allow for a better understanding of environmental oxygen tolerances and minimum acceptable oxygen concentration in rabbitfish aquaculture.     

Anaesthetic efficacy of clove oil,benzocaine, 2‐phenoxyethanol and tricaine methanesulfonate in juvenile marbled spinefoot (Siganus rivulatus)

Anaesthetics are used in aquaculture and fisheries to facilitate routine procedures, such as capture, handling, transportation, tagging, grading and measurements that can often cause injury or induce physiological stress. Two experiments were performed to assess the efficacies of four anaesthetic agents, clove oil, benzocaine, 2‐phenoxyethanol and MS‐222 on juvenile marbled spinefoot rabbitfish (Siganus rivulatus). In the first experiment we tested the lowest effective doses that produced induction and recovery times in 3 min or less and 5 min or less respectively. Dosages were 70 mg L^?1 for clove oil, 60–70 mg L^?1 for benzocaine, 400 μL L^?1 for 2‐phenoxyethanol and 100–125 mg L^?1 for MS‐222. In the second experiment, we determined optimal concentrations of the four anaesthetics if they were to be used to transport rabbitfish fry. Anaesthetic concentrations suitable for handling and transport were: 10–15 mg L^?1 of MS‐222, 5–10 mg L^?1 of benzocaine, 5 mg L^?1 of clove oil and 50–100 μL L^?1 of 2‐phenoxyethanol. All anaesthetic agents are acceptable for use on S. rivulatus, however, 2‐phenoxyethanol, MS‐222 and clove oil appear to be more suitable than benzocaine. Further studies need to be conducted on effects of high and low doses of anaesthetic agents on physiology of marbled spinefoot.     

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.     

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.     

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.     

A screening of multiple classes of pharmaceutical compounds for effect on preadult salmon lice Lepeophtheirus salmonis

The salmon louse, Lepeophtheirus salmonis Krøyer, is the major obstacle facing a sustainable future for farmers of salmonids in the North Atlantic Ocean. Medicinal compounds have been the most utilized tool to prevent salmon lice infestation; however, the active compounds have become less effective or considered environmentally unfriendly in the past years. Novel medicinal compounds are thus highly desired. In two experiment series, 26 medicinal compounds were screened for their efficacy against salmon lice, in a 30‐min exposure and 24‐h exposure, respectively. Pyriprole, imidacloprid, cartap and spinetoram were effective at 50 mg L^?1 in the short‐time exposure. In the 24‐h exposure, pyriprole, propoxur, cartap, imidacloprid, fenoxycarb, pyriproxyfen, nitenpyram, spinetoram, spiromesifen and diflubenzuron induced a high level of immobilization at 5 mg L^?1. The EC₅₀ values of the effective compounds were calculated in further titration studies for both exposure periods. Several physiological and biochemical pathways were discovered as possible targets for medicinal intervention against the salmon louse.     

Acute ammonia toxicity and gill morphological changes of Japanese flounder Paralichthys olivaceus in normal versus supersaturated oxygen

Ammonia toxicity and morphological changes in gills of juvenile Japanese flounder Paralichthys olivaceus (5.76 ± 0.12 g) were investigated when fish were separately exposed to normal dissolved oxygen (DO) at 6.5 ± 0.5 mg L^?1 and supersaturated oxygen at 16.0 ± 2.0 mg L^?1 at different ammonia concentrations. Under normal oxygen, ammonia concentrations were tested from 0.04 (control) to 93.3 mg L^?1 total ammonia nitrogen (TAN), whereas under oxygen supersaturation, ammonia concentrations ranged from 0.04 (control) to 226.7 mg L^?1 TAN in the trial. After exposure to ammonia for 96 h, the ammonia LC₅₀ for fish was 62.48 mg L^?1 TAN (0.50 mg L^?1 NH₃–N) at normal oxygen and 160.71 mg L^?1 TAN (0.65 mg L^?1 NH₃–N) at oxygen supersaturation. Light microscopic observations confirmed that gill damage in normal oxygen was more profound than in oxygen supersaturation when fish were exposed to the same level of TAN (93.3 mg L^?1). Furthermore, electron microscopic scanning also showed more crimple, retraction and fibrosis on the secondary lamella surface in fish exposed to normal oxygen than those in fish exposed to supersaturated oxygen at the same TAN (93.3 mg L^?1). This study suggests that supersaturated oxygen can increase ammonia tolerance in Japanese flounder through reducing gill damage by ammonia, which partially explains the merit of using pure oxygen injection in intensive fish farming.     

Effects of culture conditions on feeding response of larval Pacific red snapper ( Lutjanus peru, Nichols & Murphy) at first feeding

Feeding incidence or number of larvae with preys (FIC) and intensity or number of prey per larvae (FIT) at first feeding of Pacific red snapper ( Lutjanus peru) larvae was investigated under different conditions: prey type (rotifer and copepod nauplius) and density, nauplii size, light intensity, water temperature, salinity and microalgae concentration. Rotifers were not consumed at any prey density and FIC increased significantly when a high nauplii density (10 > 1, 0.1 mL^?1) and light intensity (2000 > 1000, 500, 0 lx) were supplied. In a multifactorial experiment where light intensity (2000, 2500, 3000 lx), tank colour (grey and black) and prey type (nauplii and a mixed diet: rotifers and nauplii) were tested, a significant difference was found only for light intensity and prey type with a significant interaction between these factors. FIC was significantly higher with nauplii stage I–III than IV–VI and also at 25 °C than at 28 °C. Green water (0, 0.3 × 10⁶ or 1 × 10⁶ cells mL^?1) and salinity (25, 30, 35 gL^?1) did not affect FIC. FIT was not affected by any variables tested except in the density experiment where it was significantly higher at 10 nauplii mL^?1.     

In vitro and in vivo efficacy of drugs against the protozoan parasite Azumiobodo hoyamushi that causes soft tunic syndrome in the edible ascidian Halocynthia roretzi (Drasche)

It was discovered recently that infection by a protozoan parasite, Azumiobodo hoyamushi, is the most probable cause for soft tunic syndrome in an edible ascidian, Halocynthia roretzi (Drasche). In an attempt to develop measures to eradicate the causative parasite, various drugs were tested for efficacy in vitro and in vivo. Of the 20 antiprotozoal drugs having different action mechanisms, five were found potent (24‐h EC₅₀ < 10 mg L^?1) in their parasite‐killing effects: formalin, H₂O₂, bithionol, ClO₂ and bronopol. Moderately potent drugs (10 < 24‐h EC₅₀ < 100 mg L^?1) were quinine, fumagillin, amphotericin B, ketoconazole, povidone‐iodine, chloramine‐T and benzalkonium chloride. Seven compounds, metronidazole, albendazole, paromomycin, nalidixic acid, sulfamonomethoxine, KMnO₄, potassium monopersulphate and citric acid, exhibited EC₅₀ > 100 mg L^?1. When ascidians were artificially infected with A. hoyamushi, treated using 40 mg L^?1 formalin, bronopol, ClO₂, or H₂O₂ for 1 h and then monitored for 24 h, very low mortality was observed. However, the number of surviving parasite cells in the ascidian tunic tissues was significantly reduced by treating with 40 mg L^?1 formalin or ClO₂ for 1 h. The data suggest that we might be able to develop a disinfection measure using a treatment regimen involving commonly available drugs.     

Replacement of live food with a microbound diet in feeding Litopenaeus setiferus (Burkenroad) larvae

Microbound feeds have been well accepted by shrimps and farmers in many penaeid shrimp hatcheries. The present study focused on an adequate level of replacement of Artemia nauplii and microalgae by a microbound diet for rearing Litopenaeus setiferus (Burkenroad) larvae. A microbound diet (MBD) consisting of fishmeal, squid meal, shrimp meal, yeast meal and soybean meal was used. The first experiment was designed to obtain the optimum level of MBD to complete the live feeding schedule, from Protozoea (PZ_III) to Mysis (M_III). The experimental levels of the microbound diet tested were 2, 4, 6 and 8 mg MBD L^?1 day^?1. The next step was to determine the Artemia nauplii replacement level from PZ_I to M_III by MBD. These experiments were carried out either in the presence (Experiment 2) or in the absence of algae (Experiment 3). Four replacement levels were tested: 0% (4 mg MBD L^?1 day^?1: 1 Artemia nauplii mL^?1), 40% (5.5 mg MBD L^?1 day^?1: 0.6 Artemia nauplii), 60% (6.5 mg MBD L^?1 day^?1: 0.4 Artemia nauplii) and 100% (8 mg MBD L^?1 day^?1: 0 Artemia nauplii). In all experiments growth, survival, development, quality index (QI) and performance index (PI), were used to determine the optimum concentration of microbound diet. Results showed that 6 mg MBD L^?1 day^?1 can be recommended as a complement to live food for L. setiferus larvae from PZ_III to M_III. In the presence of algae, maximum growth and survival may be obtained in 40–60% (5.5–6.5 mg MBD L^?1 day^?1) of Artemia nauplii replacement levels. In the absence of algae, the Artemia nauplii replacement resulted in slower development, less salinity resistance, lower growth and lower survival than was obtained in larvae fed with algae.     

Efficacy of clove oil and ethanol against Saprolegnia sp. and usability as antifungal agents during incubation of rainbow trout Oncorhynchus mykiss (Walbaum) eggs

Inhibitory concentrations of clove oil and ethanol against growth of Saprolegnia sp. hyphae were screened by a modification of the hemp (Cannabis sativa L.) seed MicroPlate (HeMP) method and their usability as antifungal agents during incubation of rainbow trout Oncorhynchus mykiss eggs was tested. In vitro experiment showed that in continuous static exposure, clove oil at 100 mg L^?1 significantly inhibited the growth of Saprolegnia, whereas in bath exposures, clove oil at 500 mg L^?1 had no significant effect at any exposure time tested (15, 60 and 240 min), but clove oil at 10 000 mg L^?1 significantly inhibited growth at all exposure times. Clove oil and ethanol treatments had no visible effects on the onset or spread of the fungus during incubation of rainbow trout eggs. Clove oil at 1000 mg L^?1 resulted in 95–100% mortality before the eyed stage was reached. Sublethal concentrations of clove oil and ethanol had no effects on the development rate of the embryo or growth and yolk utilization efficiency after hatching. This study suggests that clove oil and ethanol may not be options in controlling aquatic fungi infestations during incubation of rainbow trout eggs.     

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).     

Effects of dietary α‐ketoglutarate supplementation on the antioxidant defense system and HSP 70 and HSP 90 gene expression of hybrid sturgeon Acipenser schrenckii ♀ × A. baerii ♂ exposed to ammonia‐N stress

This study was conducted to determine the effects of dietary α‐ketoglutarate (AKG) supplementation on the antioxidant defense system and gene expression of heat shock protein (HSP) 70 and HSP 90 in hybrid sturgeons Acipenser schrenckii ♀ × A. baerii ♂ exposed to ammonia‐N stress. A 2 × 3 factorial experiment was arranged, in which each diet (0%, 1% AKG) was randomly assigned to 0.25 (control) 5 and 10 mg L^?1 ammonia‐N groups with three replicate aquaria for each 72 h. The 10 mg L^?1 ammonia‐N significantly increased serum ammonia concentrations and intestinal Gln concentrations and GS activity compared with the 0.25 or 5 mg L^?1 ammonia‐N groups. The intestinal Gln concentration and GS activity increased, and the serum ammonia concentration decreased, in fish given dietary supplementation of 1.0% AKG compared with fish given diets without AKG. Superoxide dismutase (SOD) and glutathione peroxidase (GPx) activity in serum, gills and intestines decreased when fish were exposed to 5 or 10 mg L^?1 ammonia‐N, and their activity increased in fish given diets with 1% AKG. Catalase in the serum and gills decreased when fish were exposed to 5 or 10 mg L^?1 ammonia‐N and increased in fish given diets with 1% AKG. The 10 mg L^?1 ammonia‐N or 1% AKG supplementation increased HSP 70 and HSP 90 gene expression in the liver. The increased activity of antioxidant enzymes, and increased HSP 70 and HSP 90 gene expression in fish fed diets containing 1% AKG suggested higher tolerance to ammonia‐N stress.