Gonadal development, aromatase activity and P450 aromatase gene expression during sex inversion of protogynous red-spotted grouper Epinephelus akaara (Temminck and Schlegel) after implantation of the aromatase inhibitor, fadrozole

In the present study, we implanted 2‐year‐old female red‐spotted grouper, Epinephelus akaara, with a non‐steroidal aromatase inhibitor (AI), fadrozole, in the breeding season and examined changes in gonadal histology, serum sex steroids, aromatase activities and P450 aromatase (P450arom) gene expression in gonads after AI implantation. Aromatase inhibitor at doses from 0.1 to 10.0 mg kg^?1 BW induced a sex inversion and completion of spermatogenesis up to the functional male phase, but doses of 1.0 and 10.0 mg kg^?1 BW AI produced more males than 0.1 mg kg^?1 BW AI. Serum estradiol‐17β (E₂) levels decreased, but 11‐ketotestosterone (11‐KT) levels increased significantly in all the AI‐implanted groups, whereas testosterone (T) levels increased significantly only in the 1.0 mg kg^?1 BW AI‐implanted group. Aromatase activities and P450arom gene expression in gonads were inhibited significantly in the AI‐implanted groups, which was in accordance with the decrease in serum E₂ levels. These results suggested the optimal dose of AI to induce sex inversion to be 1.0 mg kg^?1 BW. Furthermore, the sex inversion induced by AI may be attributed to the inhibition of P450arom gene expression and aromatase activity and the resultant decrease in the biosynthesis of endogenous E₂. Meanwhile, the elevated 11‐KT levels were also associated closely with the occurrence of sex inversion in protogynous red‐spotted grouper.     

A non‐paradoxical dose response to 17α‐methyltestosterone by Nile tilapia Oreochromis niloticus (L.): effects on the sex ratio,growth and gonadal development

Commercial tilapia production is dependent on monosex culture, commonly obtained through the inclusion of an androgen in the diet for a brief period soon after hatch. To determine a minimum effective dose and identify the problems associated with over‐dosing, Nile tilapia Oreochromis niloticus fry were fed diets containing methyltestosterone (MT) at rates up to 1200 mg kg^?1 of diet for 28 days. The minimum effective dose for ≥95% males was 14 mg MT kg^?1 diet. Percent phenotypic males increased as the rate increased from 3.75 (80%) to 30 mg kg^?1 MT (99%). Methyltestosterone given at rates of 120 mg kg^?1 or more reduced efficacy but did not result in a reduced frequency of males relative to that of non‐treated fish. The term ‘paradoxical feminization’ does not adequately describe the observed sex ratios, where no fish were feminized but the efficacy of MT at high doses to masculinize females was reduced. At 1200 mg MT kg^?1, the frequency of females (48 ± 1%) was not different from that in the non‐treated population. The mechanism for the reduced efficacy is not clear and is not adequately explained as an aromatization of androgen to oestrogen response.     

Stocking density effects on growth and production of the threatened silurid catfish,Mystus cavasius (Hamilton) fingerlings in nursery ponds

The fingerling‐rearing experiment of the threatened catfish, Mystus cavasius was carried out at different stocking densities in earthen nursery ponds. Twelve‐day‐old fry were stocked at 200 000 ha^?1 in treatment‐1 (T₁), 250 000 ha^?1 in treatment‐2 (T₂) and 300 000 ha^?1 in treatment‐3 (T₃) respectively. The mean length and weight of fry at stocking was 1.24 ± 0.25 cm and 0.11 ± 0.04 g respectively. Fry in all the experimental ponds were supplemented with SABINCO nursery feed for the first 14 days and starter‐I feed for days 15–56. The physico‐chemical parameters and plankton population of pond water were within the suitable level for fish culture. Growth in terms of final weight, final length, weight gain, length gain and specific growth rate and survival of fingerlings were significantly higher in T₁ than those in T₂ and T₃. Feed conversion rate was significantly lower in T₁ followed by T₂ and T₃ in that order. Significantly higher number of fingerlings was produced in T₃ than that in T₂ and T₁. Even then, consistently higher net benefits were obtained from T₁ than those from T₃ and T₂. Among the treatments evaluated, 200 000 fry ha^?1 was the best stocking density considering the highest growth, production and net benefits of fingerlings of M. cavasius in nursery ponds.     

Hormonal sex reversal of tilapias: implications of hormone treatment application in closed water systems

The phenomenon of incidental sex reversal was observed in experiments originally designed to determine the importance of feeding duration and feed quality in the optimization of masculinization and feminization treatments of Oreochromis niloticus (L.) fry by oral application of hormones. Four separate experiments were conducted: (1) masculinization and (2) feminization using three feeding durations and (3) masculinization and (4) feminization using three alternative feed mixtures. Masculinization experiments incorporating 17 α-methyltestosterone (MT) into the feed at 40 mg kg^?1 were applied to putative monosex male progeny whilst feminization treatments were applied to putative all female fry using diethylstilboestrol (DES) at 1000 mg kg^?1. Treatments were applied outdoors in small cages suspended in 900-l circular concrete tanks with a flow through rate of 1-l min^?1. Replicate cages for all treatments and the controls in each experiment were reared together in the same tank. Only the first experiment produced significant differences in sex ratio, with the control treatment producing a slightly lower proportion of males than the three MT treatments at different durations. The most significant finding was that in all experiments the sex ratio of the control differed significantly from expectations and in all but experiment 1, did not differ from those of the treated fish in which sex reversal had clearly been effected. It appears likely that incidental sex reversal occurred in the control fish as a result of the accumulation of active metabolites from the treated fish and/or leachates from uneaten food given to fish in the same water body. The results are discussed in the context of improving the efficiency and safety of hormonal sex reversal treatments.     

Dietary supplementation with the microalgae Parietochloris incisa increases survival and stress resistance in guppy (Poecilia reticulata) fry

Dietary enrichments with the arachidonic acid (ARA)‐rich microalga, Parietochloris incisa, on the survival of guppy (Poecilia reticulata) fry were examined. Diets were applied via Artemia enrichment to fish from two commercial farms for 34 and 36 days of experimental period (trials 1 and 2, respectively). In trial 1, Artemia nauplii were enriched with dry biomass of whole algal cells at 0 (control), 0.1, 0.2 and 0.4 mg mL^?1. Fry fed with Artemia enriched with 0.4 mg mL^?1 demonstrated the lowest mortality rates (24% and 1% in farms 1 and 2, respectively) compared with controls (36% and 13% in farms 1 and 2, respectively). In trial 2, fry were fed with Artemia, enriched with whole algal cells (0.4 mg ml^?1), algal hexane extract (HE; containing primarily ARA‐rich triacylglycerols and β‐carotene; 0.19 mg ml^?1) or the extraction residue (0.28 mg ml^?1). Acute stress (5 min air exposure) was applied after 18 days. The lowest mortality was recorded in the whole alga‐fed group (av. 26% and 2.6% in farms 1 and 2, respectively), with a slightly, but not significantly higher mortality in the HE‐fed group (av. 29% and 6.2% in farms 1 and 2, respectively). Elevated lysozyme was associated with the reduced mortality. Overall, the use of P. incisa as a dietary supplement for guppy fry during their first month of life enhanced their survival and stress resistance.     

Effects of oestradiol-17β or 17α-methyltestosterone administration on gonadal differentiation of largemouth bass Micropterus salmoides (Lacepède)

Monosex populations can be a valuable management tool in culture of larger size largemouth bass (>400 g). In this study, we investigated the effective mode and duration of oestrogen and androgen administrations to produce monosex largemouth bass populations. The experiment consisted of nine treatments. In oral administration groups, we fed 40-day-old fry either 200 mg of an oestradiol-17β (E₂) kg⁻¹ diet or 60 mg of a 17α-methyltestosterone (MT) kg⁻¹ diet for 30, 45 or 60 days. In bath treatments, we immersed fry in a 1 mg MT L⁻¹ solution for 5 h a day on three or six occasions. For control treatment, we fed fry an ethanol-treated diet for 45 days. The frequency of females in the control group was 53.1%. Oral administration of E₂ at all durations resulted in slight increases in the frequency of females (59.8–70.5%). Both modes of androgen administration at all durations were ineffective in altering phenotypic sex. The experimental results of our study indicated that male differentiation passed the point of being completely and functionally influenced by exogenous oestrogens, while female differentiation had already taken place and was no longer responsive to exogenous androgens in 40-day-old (33.5 mm) largemouth bass fry.     

Synergistic effects of dietary vitamin E and selenomethionine on growth performance and tissue methylmercury accumulation on mercury‐induced toxicity in juvenile olive flounder,Paralichthys olivaceus (Temminck et Schlegel)

An 8‐week feeding trial was conducted to evaluate the synergistic effects of dietary vitamin E and selenomethionine (SeMet) on induced methylmercury (MeHg) toxicity in juvenile olive flounder Paralichthys olivaceus. Nine semi‐purified diets were formulated to contain three different vitamin E levels as DL‐α‐tocopheryl acetate (0, 100 and 200 mg TAkg^?1 diet) and three different selenium (Se) levels (0, 2 and 4 SeMet mg kg^?1 diet) on the constant mercury toxicity level (20 mg MeHgkg^?1 diet). Nine experimental diets, in a 3² factorial design (E₀Se₀, E₀Se₂, E₀Se₄, E₁₀₀Se₀, E₁₀₀Se₂, E₁₀₀Se₄, E₂₀₀Se₀, E₂₀₀Se₂ and E₂₀₀Se₄), were fed to triplicate groups of fish averaging 2.3 ± 0.04 g (mean ± SD) in the semi‐recirculation system. After 8 weeks of feeding trial, vitamin E and Se showed significant effects on weight gain (WG) of fish (P < 0.05). We found that there was a clear trend of increasing WG with elevating vitamin E and Se levels in the diets. Feed efficiency (FE), specific growth rate (SGR), protein efficiency ratio (PER) and survivability exhibited a similar trend with WG. Both antioxidants had significant interaction effects on FE and PER (P < 0.05). Methylmercury concentrations in fish muscle, liver and kidney decreases in a dose‐dependent manner as dietary vitamin E and Se levels increase. Interestingly, the most significant interactive effects of vitamin E and Se were found in liver tissue for depleting Hg concentrations (P < 0.05). These findings suggest that dietary vitamin E more than 100 mg TA kg^?1 diet with 2 or 4 mg SeMet kg^?1‐supplemented diets could have synergistic effects on growth and liver mercury bioaccumulation on MeHg‐induced toxicity in juvenile olive flounder.     

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

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

Effects of ascorbic acid enrichment by immersion of rainbow trout (Oncorhynchus mykiss, Walbaum 1792) eggs and embryos

This study was conducted to examine the effects of different forms and concentrations of ascorbic acid (vitamin C), and different enrichment times (24 and 48 h post ovulation) on egg, embryo and alevin ascorbate concentrations and survival of rainbow trout (enrichment was at the ova stage). In experiments 1 and 2, fertilized eggs were immersed in water containing ascorbate at 0 (control), 100, 1000 mg L^?1 l ‐ascorbic acid (AA) and 2000 mg L^?1 l ‐ascorbyl monophosphate (AP). In experiment 3, 0 (control), 500 and 1000 mg L^?1 AA neutralized (N) with NaOH, 1000 mg L^?1 AA non‐neutralized (NN), 1000 and 2000 mg L^?1 AP immersions were used. The mean total ascorbic acid (TAA) and dehydroascorbic acid (DHA) concentrations were measured before fertilization, at 3 and 24 h after fertilization, at the eyed stage, and in hatched alevins. We observed significant differences in TAA concentration at different immersion levels at 3 and 24 h after fertilization. Survival decreased significantly depending on the level of vitamin C, pH of the solutions and immersion time. We suggest that when broodstock rainbow trout do not have enough vitamin C in their ovaries, immersion of eggs in 1000 mg L^?1 of neutralized AA may be useful.     

Relative efficacy of dietary administration of 3,5,3'-triiodothyronine (T3) to different stages of an Indian major carp, Cirrhina mrigala (Hamilton): growth and economics

Different stages of an Indian major carp, Cirrhina mrigala (Hamilton), i.e. hatchling, fry and fingerling, were fed diets supplemented with 3,5,3′-triiodothyronine (T₃) at doses of 0, 1, 5 and 10 mg kg^?1 diet for 60 days, and then reared on a hormone-free diet for 120 days to study the growth performance of fish during the treatment period (TP) and post-treatment period (PTP). Oral administration of T₃ at all the stages resulted in a significantly higher specific growth rate (SGR) and better food conversion ratio (FCR) in comparison to the control group (P<0.05). However, the growth response of C. mrigala towards T₃ administration appeared to be stage-specific. At the end of the experimental period of 180days (60days TP+120days PTP), the optimum dosage of T₃ appeared to be 5.0 mg kg^?1 diet for both hatchlings and fry, and 1.0 mg kg^?1 diet for fingerlings, resulting in 62.08%, 30.22% and 13.33% higher weights than in the control group, respectively. The cost of hormone for production of 1 kg of fish was lowest in the case of hatchlings (Rs 0.54), followed by fry (Rs 7.00) and fingerlings (Rs 16.07). Hence, based on the degree of growth response of the fish towards T₃ administration in addition to its economics, only incorporation of T₃ in the diet of hatchlings appeared to be economical.