In vitro assessment of the antiparasitic effect of caprylic acid against several fish parasites

The antiparasitic effect of caprylic acid against several fish parasites, i.e., the ciliate Cryptocaryon irritans (theronts), monogenean Benedenia seriolae (oncomiracidia and adults), copepod Pseudocaligus fugu (copepodids and adults) and myxosporean Kudoa shiomitsui (spores), was examined by in vitro trials.

Caprylic acid at a concentration of 1 mM had a parasiticidal effect against C. irritans theronts, B. seriolae oncomiracidia and K. shiomitsui spores and a contractile effect against B. seriolae adults, but had no clear effect against P. fugu copepodids and adults. These results suggest that caprylic acid may have an antiparasitic effect against various fish parasites, including parasites classified as monogenea, ciliates and myxosporea.     

Furunculosis in Atlantic salmon (Salmo salar L.) is not readily controllable by bacteriophage therapy

The potential of bacteriophage therapy to control bacterial disease in farmed fish was tested using, as an example, furunculosis of Atlantic salmon, caused by Aeromonas salmonicida subsp. salmonicida.

In vivo testing with Atlantic salmon and rainbow trout (Oncorhynchus mykiss Walbaum) showed no adverse effects, with bacteriophage generally cleared within 96 h of administration by either intraperitoneal (i.p.) injection or oral in-feed.

Juvenile Atlantic salmon were administered a combination of bacteriophage O, R and B (1.9 × 10⁸ pfu fish^− 1) by i.p. injection, after they had been challenged with A. salmonicida subsp. salmonicida 78027, also by i.p. injection. The fish that were injected with bacteriophage immediately after challenge died at a significantly slower rate then those that were either not treated with bacteriophage, or treated 24 h post-challenge. However, the end result (100% mortality) was not affected.

In further experiments the effects of oral (1.88 × 10⁵ pfu g^− 1 fish^− 1 daily for 30 days), bath (1.04 × 10⁵ ml^− 1 daily for 30 days) and i.p. (6.25 × 10⁷ pfu fish^− 1) phage treatment to control furunculosis in experimentally infected Atlantic salmon were compared with antibiotherapy (treatment with 10 mg kg^− 1 bw^− 1 day^− 1 oxolinic acid for 10 days), using an indirect cohabitation challenge. No protection was offered by any of the bacteriophage treatments, compared to the positive challenge group, although significant protection was offered by the oxolinic acid treatment. Analysis of samples taken from the trials demonstrated that bacteriophage were correctly administered to the fish and, on occasion, were isolated from fish that had succumbed to furunculosis. It was also shown that bacteriophage resistant A. salmonicida subsp. salmonicida isolates could be recovered from mortalities in all the treatment groups.

The results suggest that, although there were no safety problems associated with the approach, furunculosis in Atlantic salmon is not readily controllable by application of bacteriophage.     

Two microalgae Crypthecodinium cohnii and Phaeodactylum tricornutum as alternative source of essential fatty acids in starter feeds for seabream (Sparus aurata)

Despite oils extracted from algae and other microorganisms that may constitute excellent sources of HUFAs, few studies have determined the nutritional value of different microalgal species for young marine fish. Six thousand gilthead seabream (Sparus aurata) postlarvae (73 mg body weight) were fed for 57 days diets containing either fish oil as a single lipid source or 2 and 4% of Cryptecodinum cohnii or 5% Phaeodactylum tricornutum. Fish oil substitution by C. cohnii resulted in improved fish survival and a very good growth performance, in agreement with a higher proportion of DHA in diets and in total lipids of fish. Incorporation of DHA and other fatty acids was proportional to their contents in diet suggesting the good nutritional utilization of homogenized C. cohnii. Lower survival rates were found in fish fed P. tricornutum and could be related to an epithelial degeneration observed in the anterior intestine. This degeneration could be related to a higher lipid content in these fish or to the strong hornlike cornutate processes found in the valves of the diatom P. tricornutum.     

Effects of chloramphenicol, erythromycin, and furazolidone on growth of Isochrysis galbana and Chaetoceros gracilis

This study focused on determining the effects of antibiotics on microalgae used as food for scallop larvae. Six different dose levels of chloramphenicol, erythromycin, and furazolidone were added to cultures of Isochrysis galbana and Chaetoceros gracilis. An in vivo experiment was subsequently conducted to determine the effect of chloramphenicol and erythromycin on larval survival of the Pacific calico scallop Argopecten ventricosus in tanks and on the population of its associated bacteria. Results showed that growth of I. galbana was not significantly affected by chloramphenicol or erythromycin at the test doses of 0.5, 1.0, 3.0, 6.0, 9.0, and 12.0 mg/l. C. gracilis was significantly sensitive to erythromycin and chloramphenicol at doses higher than 0.5 and 3.0 mg/l, respectively. Furazolidone inhibited the growth of both I. galbana and C. gracilis at all test doses. Results showed that exposure of scallop larvae to a dose of 6 mg/l chloramphenicol or erythromycin did not significantly affect growth of I. galbana, significantly enhanced survival of the scallop larvae, and inhibited the growth of Vibrio spp. in tanks. This study demonstrated the adverse effect of chloramphenicol, erythromycin and furazolidone on I. galbana and C. gracilis microalgae but the positive effect on survival of the scallop larvae, decreasing associated bacterial population.     

Effects of dietary estradiol-17β in juvenile shortnose sturgeon, Acipenser brevirostrum, Lesueur

Shortnose sturgeon, Acipenser brevirostrum, were fed estradiol-17β (E₂) in commercial salmon feed in two separate experiments, with the aim of creating all-female populations for aquaculture. In the first experiment, 5-month-old fish were fed 0 (control), 10, 50 and 100 mg E₂/kg feed during a 9-month trial starting in September. This experiment was confounded by copper contamination of the heated water source that appeared to cause E₂-dose-dependent mortality. As a result, a second, parallel experiment was conducted on the same year class of fish starting at age 7 months in November, but with hormone dosage reduced to 0, 10, 25 and 50 mg E₂/kg feed in a 7-month trial. In both experiments, control and 10 mg E₂/kg groups remained active and fed well, but the other fish were less active and fed less as E₂ dosage increased. Histological observations indicated that anatomical differentiation of the ovaries had begun in 3 of 5 individuals examined at the start of Experiment 2. In January, sub-samples of 6 fish from each group showed that E₂-treated fish had mottled livers, swollen kidneys and reduced gonad size. The number of hepatocytes and their staining characteristics were similar in both the control and 10 mg E₂/kg groups. At 25 mg E₂/kg and higher, the number of hepatocytes and the presence and quantity of yolk protein increased with the E₂ dosage. At final sampling in May, all of the treatment groups showed some individuals with these pathologies evident macroscopically, and their frequency and severity increased with E₂ dosage such that all individuals of the 100 mg E₂/kg group were affected. In general, fish grew better and had lower mortality in Experiment 2 (8.5%, as compared to 15.2% for Experiment 1) during the period of E₂ treatment. Final sampling revealed that all treated fish were apparently feminized in both experiments, based on anatomical differentiation of the gonads. Control groups, on the other hand, were 65 and 58% female in Experiments 1 and 2, respectively.     

Substitution of Chaetoceros muelleri by Spirulina platensis meal in diets for Litopenaeus schmitti larvae

The nutritional response of Litopenaeus schmitti larvae to substitution of Chaetoceros muelleri by Spirulina platensis meal (SPM) was evaluated. The substitution levels (S) were 0%, 25%, 50%, 75% and 100%, dry weight basis. Final larval length (FL) ranged from 1.98 to 3.16 mm for the different substitution levels. There was a significant relationship between S and FL, described by the following quadratic equation: FL = 2.853 + 0.01598S − 0.000233S². The substitution level (S) yielding maximum FL was 34.2%. Development index (DI) values ranged from 2.84 to 3.93 and were dependent on substitution level. The corresponding equation was DI = 3.799 + 0.00945S − 0.000189S² (P < 0.01). Maximum DI was obtained at 25.0% substitution. Survival was high (82–87%) and no significant differences were found between treatments. Protein digestibility of either microalgae was high, with 92% for SPM and 94% for C. muelleri, with no significant differences between them. The results in this study indicate that an adequate balance of nutrients in relation to the requirements of the species is critical. To simultaneously improve FL and DI, a 30% substitution of C. muelleri by SPM is suggested. This is equivalent to feeding 0.15 mg larvae^− 1 day^− 1 dry weight basis of a 70% C. muelleri/30% SPM diet, representing 0.078 mg protein larvae^− 1 day^− 1, 0.026 mg lipids larvae^− 1 day^− 1 and 2.732 J larvae^− 1 day^− 1.     

In vivo fish models for visualizing Aeromonas hydrophila invasion pathway using GFP as a biomarker

The invasion pathway of Aeromonas hydrophila in vivo was studied in Crucian carp (Carassius auratus gibelio) with a virulent strain A. hydrophila J-1 transformed with a plasmid encoding green fluorescent protein (pGFPuv) (A. h J-1^GFP), which had similar virulence characteristics (haemolysin, extracellular proteases production, toxicity for EPC cells, survival in fish serum and LD₅₀ value) as the parent strain. Fish were divided into four experimental groups: (1) normal fish; (2) bacteria bath challenged, unwounded fish; (3) skin artificially wounded by scalpel, bacterial bath challenged fish; and (4) skin mucus layer partially removed by paper towel, bacterial bath challenged fish. The number of bacteria from blood, gills, kidney, muscle, liver and intestine were detected at 2, 4, 8, 12, and 24 h post-challenge. High bacterial numbers were observed in the muscle of the artificial wound group, in the kidney of the mucus removed group and in the gills of all groups. In conclusion, the gills and damaged skin are likely to be the main routes of entry for A. hydrophila, and GFP can be used as a real-time biomarker to study intimate host–pathogen interaction in fish.     

Optimal conditions for egg storage, incubation and post-hatching growth for the freshwater turtle, Chelodina rugosa: Science in support of an indigenous enterprise

Incubation of northern snake-necked turtle (Chelodina rugosa) eggs and subsequent sale of hatchlings for the pet industry has the potential to provide culturally suitable employment for indigenous communities in northern Australia. Developmental arrest in response to egg inundation is unique to C. rugosa. Eggs can be stored under water for up to 10 weeks without appreciable impact on egg or embryo survival, allowing the transport and sale of eggs into niche markets without high levels of mortality, and permitting eggs to accumulate in diapause until there are sufficient numbers to incubate as batches. Eggs that are not inundated or inundated for short periods experience similar survival rates to eggs inundated for lengthy periods. Incubation temperature influences embryo survival and development period in C. rugosa. Embryonic survival is greatest at 26 °C, steadily declining as temperature increases to 32 °C. A similar increase in incubation temperature decreases incubation period by approximately 40 days, however almost half of this variation is attributed to the increase in incubation temperature from 26 to 28 °C. Hatchling growth in C. rugosa is characterized by two phases. There is an initial phase of relatively slow growth under the partial influence of initial egg size and incubation duration, followed by a second phase of relatively rapid growth under the partial influence of water temperature and mass at hatching. Post-hatching survival is negatively correlated with duration of egg inundation and water temperature. Evidence suggests that inundation of C. rugosa eggs for 6 weeks, incubation of embryos at 28 °C and raising hatchlings in 28 °C water will yield the best overall outcomes.     

Effects of Cu-exchanged montmorillonite on growth performance, microbial ecology and intestinal morphology of Nile tilapia (Oreochromis niloticus)

A total of 360 Nile tilapia at an average initial body weight of 3.9 g were randomly allocated to 4 treatments, each of which had three replicates of 30 fish/tank and used to investigate the effects of Cu²⁺-exchanged montmorillonite (Cu-MMT) on growth performance, microbial ecology of the skin, gill and intestine, and intestinal morphology. The dietary treatments were: 1) basal diet, 2) basal diet + 1.5 g/kg MMT, 3) basal diet + 30 mg/kg copper as CuSO₄ (equivalent to the copper in the Cu-MMT treatment group), or 4) basal diet + 1.5 g/kg Cu-MMT. The results showed that supplementation with Cu-MMT significantly improved growth performance as compared to control and fish fed with Cu-MMT had higher growth performance than those fed with MMT or CuSO₄. Supplementation with Cu-MMT reduced (P < 0.05) the total intestinal aerobic bacterial counts and affected the composition of intestinal microflora with a tendency of Aeromonas, Vibrio, Pseudomonas, Flavobacterium, Acinetobacter, Alcaligence, Enterobacteriaceae decreasing as compared with control. Measurements of villus and microvillus heights at different sites of the intestinal mucosa indicated that dietary addition of MMT or Cu-MMT improved intestinal mucosal morphology. However, the microbial ecology of the skin and gill was not affected by the addition of CuSO₄, MMT or Cu-MMT. Supplementation with CuSO₄ had no (P > 0.05) effect on the growth performance, microbial ecology and intestinal morphology. The results showed that Cu-MMT exhibited antibacterial activity in vivo and protected intestinal mucosa from invasion of pathogenic bacterium and toxins, resulting in a positive effect on the growth performance.     

Isolation and characterization of a regulator of G protein signaling (RGS16) gene homologue in yellow grouper Epinephelus awoara

Regulators of G protein signaling (RGS) are GTPase-activating proteins (GAP) which act as modulators of G protein-coupled receptors. We isolated a RGS16 homologue in yellow grouper (Epinephelus awoara) spleen using suppression subtractive hybridization and RACE-PCR. The nucleotide sequence of yellow grouper RGS16 full-length cDNA was 700 bp and contained an open reading frame of 537 bp, encoding a putative protein of 178 amino acids. The encoded protein shows 47–61% identities to other homologues. RT-PCR analysis demonstrated that RGS16 was expressed in yellow grouper spleen and up-regulated in kidney heart, liver, and anterior kidney by lipopolysaccharide. This study will help towards validating the specific function of RGS in marine fish.     

Streptococcus iniae infection and tissue distribution in hybrid striped bass (Morone chrysops×Morone saxatilis) following inoculation of the gills

This study was designed to test the possibility that Streptococcus iniae enters through the gills and causes infection in hybrid striped bass. To determine the dose response, four groups of fish were inoculated with S. iniae via the gills with a dose of 5.0×10⁵, 2.6×10⁶, 5.0×10⁶, or 1.0×10⁸ CFU/fish. One group of fish was inoculated with tryptic soy broth (TSB) via the gills to serve as controls. The cumulative percent mortality was 13%, 27%, 100% and 100% for 5.0×10⁵, 2.6×10⁶, 5.0×10⁶ and 1.0×10⁸ CFU/fish, respectively. We also examined the tissue dissemination of S. iniae at 0.5, 4, 8, 12, 24, 48 and 72 h after experimental gill inoculation. Fish were inoculated with 2.6×10⁶ or 5.0×10⁶ CFU/fish, which caused low and high mortality, respectively. Within 48 h, fish inoculated with the 2.6×10⁶ dose were culture positive on the gill surface, blood of the first and second gill arches, blood of the third and fourth gill arches and the nares. However, for the dose of 5.0×10⁶ CFU/fish, S. iniae was also isolated from the olfactory, optic and cerebellum regions of the brain, eye, head kidney, trunk kidney, spleen and liver at 48 h. For the 2.6×10⁶ dose, S. iniae was not isolated until 48 h post-inoculation, but was isolated at 12 h for the 5.0×10⁶ dose. The results of this study indicate that S. iniae can enter hybrid striped bass through the gills. However, mortality at similar S. iniae doses was lower than we previously observed by inoculation of the nares.     

Efficacy of bithionol as an oral treatment for amoebic gill disease in Atlantic salmon Salmo salar (L.)

This study examined the efficacy of bithionol as an oral treatment for Atlantic salmon Salmo salar affected by amoebic gill disease (AGD). The current commercial management strategy of AGD is a costly 3 h freshwater bath. It is labour intensive and the number of baths needed appears to be increasing; hence, there is an effort to identify alternative treatments. Efficacy was examined by feeding AGD-affected Atlantic salmon twice daily to satiation with bithionol, an antiprotozoal, at 25 mg kg^− 1 feed. Three seawater (35‰, 17 °C) re-circulation systems were used each consisting of three tanks containing 32 Atlantic salmon smolts with an average (± SEM) mass of 90.4 g (± 5.2). Three feeds were examined in the trial including bithionol, plain commercial control and oil coated commercial control. Feeding commenced 2 weeks prior to exposure to Neoparamoeba spp. at 300 cells L^− 1 and continued for 28 days post-exposure. Efficacy was determined by examining gross gill score and identifying percent lesioned gill filaments twice weekly for 4 weeks post-exposure. Bithionol when fed as a two-week prophylactic treatment at 25 mg kg^− 1 feed delayed the onset of AGD pathology and reduced the percent lesioned gill filaments by 53% and halved the gill score from 2 to 1 when compared with both the plain and oil controls during an experimental challenge. There were no palatability problems observed with mean feed intake of bithionol over the trial duration with fish consuming higher levels of the bithionol diet compared to both the oil and plain controls. This study demonstrated that bithionol at 25 mg kg^− 1 feed, when fed as a two-week prophylactic treatment for Neoparamoeba spp. exposure, delayed and reduced the intensity of AGD pathology and warrants further investigation as an alternative to the current freshwater bath treatment for AGD-affected Atlantic salmon.     

Effect of docosahexaenoic acid enrichment in Artemia on growth of Pacific bluefin tuna Thunnus orientalis larvae

Feeding enriched Artemia induces growth failure in Pacific bluefin tuna (PBT) Thunnus orientalis larvae; however, feeding of yolk-sac larvae of marine fish promotes larval growth. It is considered that this growth failure partly results from dietary docosahexaenoic acid (22:6n-3, DHA) deficiency. Therefore, we examined the effect of DHA contents in enriched Artemia on the growth of PBT larvae. Artemia nauplii were enriched with graded levels of DHA ethyl ester, and fed to PBT larvae for 9 days. Yolk-sac larvae of Japanese parrotfish Oplegnathus fasciatus were used as a reference diet. The DHA contents in Artemia increased with the enrichment from 0 mg g^− 1 dry weight basis (DW) to 25 mg g^− 1 DW, while the content in the reference diet was 21 mg g^− 1 DW. Feeding of enriched Artemia significantly improved the growth of PBT larvae. However, this improvement was negligible when compared with the excellent growth of the larvae that were fed the reference diet. PBT larvae that were fed the reference diet accumulated approximately twice or much higher levels of DHA in the neutral and polar lipids in the body when compared with the larvae that were fed enriched Artemia. These results show that PBT larval growth cannot be promoted by feeding enriched Artemia even if the DHA contents in Artemia are elevated to the same levels as those of yolk-sac larvae. The incorporation of dietary DHA into phospholipids in the fish body may be desirable for the normal growth of PBT larvae.     

Comparison of early life history stages of the bay scallop, Argopecten irradians: Effects of microalgal diets on growth and biochemical composition

The culture of bay scallops, Argopecten irradians, is limited by a reliable and affordable supply of spat and the ability to ensure that animals attain market size within a single growing season. The main goals of our study were thus: (1) to develop growth-optimizing algal diets for implementation in hatcheries, and (2) to identify and compare bay scallop postlarval and juvenile dietary requirements, especially of lipids and fatty acids, which if met may enhance production. Nutritional needs of postlarval bay scallops (present study) are compared with those of sea scallops, Placopecten magellanicus, offered the same diets in a previous companion study. To this end, postlarval (initial shell height, SH = 240 μm) and juvenile (initial SH = 10 mm) bay scallops were offered 6–7 microalgal diet combinations at 20 °C, for 3 weeks. A similar growth ranking among diets was observed between the two developmental stages. A combination diet of Pavlova sp. (CCMP 459) and Chaetoceros muelleri was far superior to any other diet tested, yielding growth rates of 58 and 357 μm day^− 1 which were 65% and 25% higher than the next highest performing diet of Tetraselmis striata/C. muelleri in postlarvae and juveniles, respectively. The T. striata/C. muelleri diet, which is limited in the n-3 fatty acid docosahexaenoic acid (DHA), yielded very poor growth of sea scallop postlarvae in a prior study, indicating that bay scallops may have less stringent requirements for DHA than sea scallops. The Pav 459/C. muelleri diet, which also supported the highest growth of sea scallop postlarvae, is characterized by elevated levels of the n-6 fatty acids, arachidonic (AA) in C. muelleri and 4,7,10,13,16-docosapentaenoic (DPA) in Pav 459. The two diets deficient in AA and n-6 DPA, Pavlova lutheri/Thalassiosira weissflogii and P. lutheri/Fragilaria familica, yielded the lowest growth rates in both bay scallop postlarvae and juveniles. Tissue enrichment of these two fatty acids relative to the diet, as well as overall enrichment in ∑n-6 fatty acids was observed across developmental stages and dietary treatments. A similar pattern has previously been observed in sea scallop postlarvae, suggesting a dietary requirement for n-6 fatty acids in pectinids that has often been overlooked in the past.     

四氢异喹啉衍生物的合成及体外抗多子小瓜虫活性的研究

合成四氢异喹啉衍生物并进行杀多子小瓜虫(Ichthyophthirius multifiliis)药效评价。以1-甲基-1,2,3,4-四氢异喹啉为起始原料,在2位的胺基上引入与环己甲酰氯、苯甲酰氯、噻吩甲酰氯、乙酰氯以及氯乙酰氯等不同的酰基进而合成5种四氢异喹啉衍生物(化合物1~化合物5),研究其对小瓜虫掠食体和包囊的杀虫活性,并对杀虫活性物质进行安全性评价。结果显示,5种化合物均具有一定的杀虫活性,其中,化合物1 [(1-甲基-3,4-二氢异喹啉-2(1H)-基)-苯基-甲酮]的杀虫活性最强,其对多子小瓜虫掠食体4 h的100%杀灭浓度为24.0 mg/L,对包囊6 h的100%杀灭浓度为60.0 mg/L,对掠食体的半数致死浓度(LD₅₀)为16.4mg/L。急性毒性实验结果显示,化合物1对翘嘴红鲌(Erythroculter ilishaeformi)的48 h LD₅₀为234.3 mg/L,其安全浓度为64.1 mg/L。研究表明,化合物1[(1-甲基-3,4-二氢异喹啉-2(1H)-基)-苯基-甲酮]是一种具有较好开发前景的杀小瓜虫药物。     

Diurnal variation of tryptic activity in larval stage and development of proteolytic enzyme activities of malabar grouper (Epinephelus malabaricus) after hatching

Diurnal variation in tryptic activity and developmental changes in proteolytic enzyme activities of malabar grouper larvae (Epinephelus malabaricus) were examined. Five different groups were prepared for the experiment of diurnal variation of tryptic activity in larvae: larvae were fed Thai-type rotifers Brachionus rotundiformis from the time of mouth opening, fed rotifers from 6 h after mouth opening, 12 h, 24 h and not fed rotifers (starved control). The experimental tanks were placed in temperature-controlled baths at 28 °C under 24 h light. Developmental changes in proteolytic activity of trypsin and pepsin-like enzyme were measured from hatching to 57 days after hatching (DAH).

The tryptic activity of all fed groups showed the same pattern, and the diurnal variation of tryptic activity was clearly observed from 3 to 6 DAH. The highest tryptic activities were found at 19:00, and the activities were lowest from 01:00 to 07:00. In contrast, that of non-fed larvae was low compared to the fed groups, however the diurnal variation of tryptic activity was shown same tendency to the fed groups. Interestingly, both groups (fed and non-fed) were exhibited a circadian rhythm under the 24 h light conditions and delaying of first-feeding. Tryptic activity of larvae notably increased from 40 to 45 DAH and markedly decreased at 52 DAH. In contrast to the tryptic activity, that of pepsin-like enzyme clearly increased from 47 to 51 DAH. The results suggest that a functional change of protein digestion occurs from 40 to 50 DAH related with metamorphosis in malabar grouper. These results could contribute to determining appropriate feeding schedules, such as feeding time, frequency and optimal time to change food items, in mass-scale production of the present species.     

Production of a high percentage of male offspring in growth-enhanced transgenic tilapia using Oreochromis aureus ZZ selected pseudofemales

Gene transfer has offered a new tool for the development of improved fish strains for aquaculture. Monosex fish populations could minimize the impact of genetically modified organisms in the environment. In Oreochromis aureus, the use of pseudofemale spawners (sex-reversed male with a female phenotype) is an alternative technique for producing genetically male tilapia offspring. O. aureus fry were treated with 17β-estradiol at 100 mg/kg of food for 45 days. We obtained 77.1% females and 45.9% in the control group. Females randomly taken from the treated group were crossed with normal males. Fry from pseudofemales producing more than 90% male progenies were submitted to 17 β-estradiol treatment to obtain F₂ pseudofemales. The results of the sex-reversal were low and variable ranging between 66.0 and 84.3% females. F₂ pseudofemales were crossed with transgenic males from the F70 line (O. aureus × O. urolepis hornorum) and non-transgenic (O. aureus) males. The sex ratio of progeny of F₂ pseudofemale deviated significantly (P < 0.01) in favor of males in the crosses with transgenic (90.2%) and non-transgenic (89.3%) males compared to the results observed with normal females (51.0 and 52.3%, respectively). The mean fry production with pseudofemales (per m²/day) was similar to the normal females in the crosses with transgenic and non-transgenic males. To our knowledge this is the first report on the production of a near monosex population in genetically modified fish.     

The effect of spectral composition and light intensity on melatonin, stress and retinal damage in post-smolt Atlantic salmon, Salmo salar

Metal halide lights are currently used as standard in commercial Atlantic salmon sea cages as a means of enhancing productivity through grilse inhibition. However, such systems create bright point light sources that are neither environment specific nor species specific and could potentially compromise fish welfare. Light emitting diodes (LEDs) are a new form of lighting technology currently being developed for the fish farming industry that can be tuned to environment and species sensitivities through narrow bandwidth outputs. However, prior to implementing these new high energy alternatives, any potential adverse effects must be determined in fish. The objectives of this study were thus (1) to determine the effect of increasing intensities of blue LED light (0.199–2.7 W m^− 2, at 0.1 m from the light source) on light perception and stress response, and (2) to examine potential retinal damage under these conditions in post-smolt Atlantic salmon, Salmo salar. A white LED light was also tested, as well as a very high intensity metal halide positive control. Results demonstrated firstly that salmon perceived blue LED light (basal melatonin levels maintained) irrespective of intensity. Secondly, fish exposed to high intensity blue LED light showed an increase in plasma cortisol and glucose levels within 3 h, returning to a basal state 24 h post-light onset. This typical acute stress response was not observed in fish exposed to the white LED light and lower blue light intensities which could indicate differential sensitivities to spectral content of the light. No effects on the non-specific immune system (lysozyme activity) were observed. Finally, extensive histological examination of the retina from fish exposed to these various light treatments revealed no signs of damage. This demonstrates the efficiency of the adaptive mechanisms to light developed in fish.     

Pigmentation, carotenoids, lipid peroxides and lipid composition of skin of red porgy (Pagrus pagrus) fed diets supplemented with different astaxanthin sources

A feeding experiment was carried out to determine the efficiency of different commercial sources, chemical forms and levels, of dietary astaxanthin, to appropriately pigment the red porgy (Pagrus pagrus) skin. According to this, total carotenoid content, profiles and chemical forms present in the skin were determined. In order to establish the potential for antioxidant protecting role of astaxanthin supplemented diets, peroxide levels and lipid composition of skin were also determined.

Red porgy alevins were fed six dietary treatments in triplicate; a basal diet (B) without carotenoids; two diets (N25 and N50) formulated to supply either 25 or 50 mg kg^− 1 of an esterified source of astaxanthin (Haematococcus pluvialis, NatuRose™); two diets (CP25 and CP50) with either 25 or 50 mg kg^− 1 of unesterified astaxanthin (Carophyll® Pink); and a positive control diet (B + S) proved as a successful pigmenting-diet in previous experiences (B + S, 88% basal diet:12% frozen shrimp) [Cejas, J., Almansa, E., Tejera, N., Jerez, S., Bolaños, A., Lorenzo, A., 2003. Effect of dietary supplementation with shrimp on skin pigmentation and lipid composition of red porgy (P. pagrus) alevins. Aquaculture 218, 457–469].

All fish fed carotenoid supplemented diets displayed a pink-coloured skin after 4 months of feeding in contrast to the greyish appearance displayed by fish fed the basal diet not supplemented with carotenoids (B). Furthermore, astaxanthin diesters were the major carotenoid in the skin of pink fish. A second carotenoid, tentatively identified as tunaxanthin diester, was also detected. The best results in terms of skin natural reddish hue, total carotenoid and astaxanthin contents were found by using the esterified forms of dietary astaxanthin (N25, N50 and B + S). Interestingly, the lowest levels of lipid peroxides were found in the fish fed these three treatments. However, no effect of treatment on lipid composition was found. In conclusion, red porgy alevins are able to efficiently utilise dietary natural or synthetic astaxanthin, and deposit this pigment in its esterified form to acquire an acceptable pink-coloured skin compared to that of the wild fish.     

A simple test to estimate the salinity resistance of fish with specific application to O. niloticus and S. melanotheron

In order to develop a simple and accurate index of the salinity resistance of tilapia, batches of 10 juveniles (5 to 20 g) of two different species Oreochromis niloticus and Sarotherodon melanotheron reared in freshwater were subjected to gradual increases in salinity until 100% mortality. Seven daily increments of salinity were tested with 4 replicates: 2, 4, 6, 8, 10, 12 and 14 g l⁻¹ day⁻¹, while control batches were kept in fresh water. The temperature was maintained at 27 °C. The concentration of oxygen, ammonia and the pH were not limiting factors. The mortality, monitored on a daily basis, appeared after 2–51 days and was spread out over 1–20 days, depending on the increment of salinity. The higher the daily rate in salinity increase, then the shorter the time lapse before total mortality occurred. The cumulative mortality as a function of salinity fit well with simple linear regressions. The criterion of the resistance to salinity was the index MLS (median lethal salinity) defined at each daily rate as the salinity at which 50% of fish died. For S. melanotheron, the mean MLS was 123.7±3.5 g l⁻¹ whatever the daily rate in salinity. For O. niloticus, the MLS was 46.3±3.4 g l⁻¹ for daily increases in salinity ranging from 2 to 8 g l⁻¹ day⁻¹ and decreased significantly (P<0.05) above this level. The MLS-8 g l⁻¹ day⁻¹ ,which takes into account the full capacity of the fish to adapt to the increasing salinity, appeared to be a simple, optimized and efficient criterion for assessing the resistance to salinity for O. niloticus and S. melanotheron. This criterion can be a useful tool for ranking the different parental strains and hybrids of different genus and species of tilapia used in programmes of genetic selection for growth and salinity tolerance.