Endocrine changes during the previtellogenic stage of the great sturgeon,Huso huso (Linnaeus, 1758)

The study aimed to investigate changes in serum concentrations of sex steroids, thyroid hormones, stress indices, cholesterol, triacylglycerol, calcium and phosphorus, as well as the growth in the 3‐year old previtellogenic stage of great sturgeon, Huso huso over a 6‐month period. Forty individually marked fish (mean weight 6913 g) were divided among three circular tanks under a natural temperature and photoperiod. Every 1.5 months fish were weighed and blood samples taken for serum analysis. Fish showed significant changes in 17β estradiol, testosterone, triiodothyronine (T₃), thyroxine (T₄), adrenocorticotropic hormone, cortisol, cholesterol, and phosphorus, but not in triacylglycerol and calcium concentrations. From the January onset of the experiment to the end of the trial in July, there was a decrease in sex steroids and cortisol concentrations as well as a decline in T₃ concentrations from March and in T₄ from February to July. The mean weight of fish increased significantly from the beginning to the end of the experiment. Although the oocyte diameter increased from 197 to 202 μm, the difference was not significant. The histological data on oocytes suggest that the great sturgeon, like the other sturgeons, has a heterogeneous asynchronous ovarian development in the early stage of reproduction. This data might be useful to the further understanding of the physiology of Huso huso and/or to improve its culture. Moreover, it leads to a general increase in the understanding of the basic reproductive biology of this valuable chondrostean species.     

Modifying the physiological responses to handling stress in beluga sturgeon Huso huso (Linnaeus, 1758); Interactive effects of feeding time and dietary fat

The present study investigated the interaction of feeding times with two dietary fat levels on physiological responses to handling stress in juvenile Beluga sturgeon. Fish were fed with two different diets (high energy; HE: 24% fat and low energy; LE: 12% fat) for 8 weeks at two feeding times; 09:00 and 16:00 (during the day) or 21:00 and 04:00 (during the night). At the end of the trial, blood samples were taken and the resting fish were held in a net out of water for 90 s as the handling stressor. Three hours after application of stress, post‐stress blood was taken. Cortisol, glucose, and lactate concentrations were considered as stress indicators. The mean values of cortisol and lactate levels did not reveal a significant difference between pre‐ and 3‐hr post‐stress samples, but the average concentration of glucose showed a significant difference. Cortisol, lactate and glucose concentrations were not influenced by the difference in the diets. Moreover, the cortisol and lactate concentrations were not affected by the different feeding times, while glucose levels were significantly affected by the feeding times with the lowest level in fish fed during the day. With respect to stress indicators, the results revealed that feeding times affected pre‐ and post‐stress secondary response to handling stress, but the rate and magnitude of metabolites (cortisol, glucose and lactate) were not affected by dietary fat levels. Therefore, it is necessary to examine the best feeding times with the interaction of feed ingredients in sturgeon farming.     

The effect of feeding rate on growth of pectoral fin spine of juvenile great sturgeon Huso huso (Linnaeus, 1758)

The effect of two feeding rates (0.5 and 1.5% of total body weight) was assessed on the growth of pectoral fin spines of captive juvenile great sturgeon, Huso huso, after second year of life. The fish received Oxytetracycline (OTC) twice in the first and second years of their lives under basic diet. During the 5‐month experimental period, juveniles (mean 1,187.4 g, 0.1 standard deviation [SD]), n = 50) were reared with two feeding rates under similar conditions in 10 fiberglass tanks (1.5 m³). The fish were fed manually with a commercial diet twice a day (35% Biomar, Nersac, France) throughout the experiment. The OTC marks were distinguished in all pectoral fin spine sections under ultraviolet light. The means of the first and second annular radii were 806.6 µm (27.2 SD) and 2,246.5 µm (50.2 SD), respectively. The marginal increment analysis beyond the second OTC mark revealed a significantly smaller marginal increment for low feeding rate treatment (143.9, 11.2 SD) as compared to the high feeding rate one (269.0, 14.6 SD). The results indicate the slower growth rate in the fish fed the low feeding treatments seen in the pectoral fin spine formation, which can be used as an indicator of recent feeding history in sturgeon juveniles. The best daily feeding rate for great sturgeon of 2,460 g was determined to be 1.5% body weight/day in this study.     

Production of recombinant great sturgeon (Huso huso,Linnaeus, 1758) growth hormone (GH) by Pichia pastoris (Guillierm, 1956)

In this research, the encoding cDNA of growth hormone (GH) was cloned from the pituitary gland of great sturgeon Huso huso (three adults: two females and one male, 7–9 years old, 70–90 kg, reared in concrete ponds). In order to obtain the great sturgeon recombinant GH expression in Pichia pastoris, the mature encoding cDNA was first cloned in TA vector PTZ57R and then sequenced. After confirmation of the correct GH sequence, the GH coding sequence was subcloned into pHILS1 expression vector. The yeast Pichia pastoris GS115 strain was transformed with the expression plasmid. Results obtained from this study showed that great sturgeon GH recombinants were expressed upon induction with methanol and exported into the medium. The level of expression was examined using RNA analysis, SDS‐PAGE, and western blot analysis. RNA analysis of the recombinant strains showed a sharp, specific band in 800 bp. The specific band in transformants indicated the presence of GH RNA in the yeast. SDS‐PAGE and western blot analysis showed a specific 21 kDa band for the growth hormone. Culture conditions were optimized for pH = 6 and incubation time (after 24 h induction, peaking at 72 h) for maximal protein production. The results provide useful information for the future production of recombinant growth hormones in other sturgeon species.     

Effects of dietary l‐carnitine supplements on growth and body composition in beluga sturgeon (Huso huso) juveniles

The effects of dietary l ‐carnitine on growth performance, whole body composition and feed utilization were studied in beluga, Huso huso. Fish were randomly allocated in 15 tanks (30 fish per tank) and triplicate groups were fed to satiety during 84 days one of five isonitrogenous (41% CP) and isoenergetic (20 MJ kg^?1) diets, each differing in l ‐carnitine content [0 (control), 300, 600, 900 and 1200 mg kg^?1 diet]. At the end of the trial, fish grew from 19‐ to 23‐fold in weight, from 8.4 g to a maximum of 191 g. Fish fed 300–600 mg l ‐carnitine had the highest specific growth rate (SGR, 3.69 and 3.72% day^?1) and protein efficiency ratio (PER, 0.95 and 0.99), and the lowest feed conversion ratio (FCR, 1.4 and 1.3) than the other groups (P < 0.0001). SGR, PER and FCR were the poorest for fish fed 1200 mg l ‐carnitine, while fish fed the unsupplemented and 900 mg l ‐carnitine supplemented diet showed intermediate performance. Body lipid concentration decreased significantly from 5.8 to 5.1% (P < 0.0001) with dietary l ‐carnitine supplementation increasing from 0 to 300 mg. Energy content was significantly lower in fish fed the 900 and 1200 mg l ‐carnitine diet (5.8 MJ kg^?1), when compared with the other treatment groups (6.4–6.6 MJ kg^?1). The results indicated that feeding sturgeon on diets supplemented with 300 mg l ‐carnitine kg^?1 diet improved growth performance, and stimulated protein‐sparing effects from lipids.     

Effects of dietary prebiotic GroBiotic®‐A on growth performance,plasma thyroid hormones and mucosal immunity of great sturgeon,Huso huso (Linnaeus, 1758)

The present study was conducted to evaluate the effects of Grobiotic^®‐A, a commercial prebiotics, when administered in feed on the growth performance, plasma thyroid hormones and mucosal immunity of great sturgeon (Huso huso). The commercial prebiotic mixture was supplemented in the diets at four different levels (i.e. 0.0% as control, 0.5%, 1% and 2%, in three replicates, 20 fish per replicate) and fed to the fish for an 8‐week period wherein 240 fish were cultured in 1,800‐L fiberglass tanks that formed part of a flow‐through system. Water temperature was maintained at 20.4 ± 1.5°C. Significant changes in growth performance parameters were observed, but only in those groups fed with 1% and 2% prebiotics. Specifically, marked improvements relative to the control group were observed in percentage weight gain, body weight gain, feed conversion ratio and specific growth rate in prebiotic‐fed fish. The levels of plasma thyroid hormones, specifically thyroxine and thyroid stimulating hormones were significantly elevated in the group receiving 2% prebiotics. Activities of lysozyme and alkaline phosphatase in skin mucus were significantly enhanced in prebiotics‐fed groups, particularly at an inclusion level of 1% and higher (2% group compared to the control). Inhibitory activity of the skin mucus against pathogens, particularly Streptococcus iniae and Yersinia ruckeri, was significantly improved following prebiotic feeding. Taken together, dietary inclusion of GroBiotic^®‐A promoted growth, modulated thyroid hormones, and enhanced mucosal immunity of H. huso. This prebiotic mixture has the potential for use in improving the growth performance and health status of farmed great sturgeon.     

Optimal stocking density for beluga,Huso huso,and ship sturgeon,Acipenser nudiventris during the grow‐out phase

The influence of three different initial stocking densities (SD) in flowthrough systems was evaluated on growth performance and feed utilizaition in beluga, Huso huso, and ship sturgeon, Acipenser nudiventris, juveniles in three different grow‐out phases for 228 days. In each grow‐out phase, fish were randomly distributed in 18 concrete square tanks (2.0 × 1.0 × 1.2 m³; 2 m³ of volume) according to the experimental SD. At the first phase, both fish species were subjected to three SD including 1.5, 3 and 6 kg/m² for 60 days. In the second phase, fish were submitted to three SD including 3, 6 and 9 kg/m², whereas in the third phase, beluga were stocked at 6, 9 and 12 kg/m² and ship sturgeon were stocked at 6, 8 and 10 kg/m² and each phase lasted for 12 weeks. In both species, the growth performance and feed intake (FI) significantly decreased with increasing SD at the end of the first phase (p < 0.05). In the second phase, growth performance and FI in beluga decreased with increasing SD, whereas in ship sturgeon neither growth nor FI were affected by SD. Interestingly, the growth performance and feed utilization in both species were not decreased with increasing SD in the third phase indicating adaptability of these species to high stocking density during grow‐out period. According to the result of this study, the appropriate initial SD for beluga within the range of 100–500, 500–2,000 and 1,500–3,000 g were at 1.5, 6 and 12 kg/m², respectively in an open flow‐throw system. Regarding to ship sturgeon, initial SD at 1.5 and 10 kg/m²recommended for fish within the range of 100–300 and 300–1,200 g, respectively in an open flow‐throw system.     

Early sex identification of 18‐month cultured beluga sturgeon (Huso huso) using ultrasonography,small surgery and plasma steroid hormones

Since sturgeons do not show clear sexual dimorphism particularly when are small in size, attempts were made to determine the best methods to identify early sex in farmed beluga sturgeon (Huso huso). The present study describes the ultrasonography, small surgery and plasma steroid hormone methods to determine gender at 18‐month fish, which no research has been conducted yet into the fish at such small ages. Twenty one cultured beluga sturgeon's gonad were imaged using an ultrasonograph unit with a 9–13 MHz linear transducer. Overall accuracy of sex determination using ultrasonography was 80.95%. Plasma testosterone (T) levels were significantly higher in males than in females whereas 17α,20βOH‐P levels were significantly higher in females than in males. Testosterone (T) and 17α,20βOH‐P were not correlated with morphometric parameters (TL, SL, W, CF) in 18‐month beluga sturgeon. Results of this study indicated that sex could be identified by each of ultrasonography, small surgery and analysis of blood plasma in such a small size (18‐month). Although direct observation was more efficient than the other methods, ultrasonography was the simplest and cost‐effective tool in sturgeon's sex determination compared to other methods, and the least invasive.     

A comparative study of feeding methods: Effect on the growth,behaviour and biochemical performance of juvenile Beluga sturgeon (Huso huso Linnaeus, 1758)

This study compared the influence of feeding methods on growth parameters of young‐of‐year Beluga sturgeon Huso huso in a 6‐week trial. Fish with an average weight 150.3 ± 0.8 g (±SE) were stocked into nine circular concrete tanks (30 fish per tank) in an open circular system with water temperature of 18.9°C. All fish were fed by three different feeding methods: (a) hand‐fed (HF), (b) continuously available (automated feeder; AF), (c) half of daily feed provided by hand, and another half by automated feeder (combined feeding). For the hand‐feeding method, fish were fed at 09:00, 14:00, 19:00, and 24:00. The entire automatic feeding groups were fed with the same amount of feed. The mean final body weight was the highest in fish fed by AF compared to fish fed by HF. Body weight increase, condition factor, specific growth rate, and feed conversion ratio did not differ among the feeding groups. Fish fed by AF revealed higher swimming activity than the HF group. No significant changes were found in hematocrit, glucose and total protein concentrations among treatments. The results showed less dependence of growth and physiology of Beluga sturgeon on feeding method, but automated‐feeding was shown to be suitable for sturgeon rearing because of further low labour costs in rearing systems.     

Behavioural responses to various chemical incentives in hybrid beluga x Russian sturgeon (Huso huso x Acipenser gueldenstaedtii) fry

Behavioural contact feeding responses to various chemical incentives were studied in hybrid beluga x Russian sturgeon ( Huso huso x Acipenser gueldenstaedtii ) fry. Solutions of 29 amino-acids (10–2 M); aqueous extracts of various vegetable, animal and microbiological components of commercial fish diets (30 min infusion of 100 mg in 100 ml), and various artificial and natural substances were tested. The most attractive amino-acids were lysine, methionine and alanine. Most of the dietary extracts tested did not stimulate feeding behaviour, indicating that widespread commercial sturgeon diets (such as ST-07 or ST-04Az) consist of components with little attractiveness for sturgeon fry. In most cases extruded diet components stimulated more bites than granulated ones. Among the diet components, only fish protein concentrate (FPC), dry milk and krill meal stimulated feeding behaviour. Flavour enhancers such as concentrated low molecular weight nitrogenous substancs (CLMNS), sodium glutamate, and "Finnstim" all stimulated feeding behaviour of fry. Some of these substances are recommended as flavour enhancers in sturgeon diets.     

Factors influencing the natural reproduction decline in the beluga (Huso huso,Linnaeus, 1758), Russian sturgeon (Acipenser gueldenstaedtii,Brandt & Ratzeburg, 1833), and stellate sturgeon (A. stellatus,Pallas, 1771) of the Volga–Caspian basin: A review

The review describes the changes in natural reproduction of three important sturgeon species in the Volga–Caspian basin: (a) the beluga (Huso huso Linneaus, 1758), (b) the Russian sturgeon (Acipenser gueldenstaedtii Brandt &Ratzeburg, 1833), and (c) the stellate sturgeon (Acipenser stellatus Pallas, 1771). Since the past 60 years, these species responded to severe influences of natural and anthropogenic factors. On the basis of original and published data, an analysis has been made of (a) the numbers of larvae migrating from spawning sites (according to plankton net survey), (b) fecundity and histological anomalies in gonad development, (c) the numbers of adult sturgeons in the Caspian Sea and of spawners migrating to the Volga River (according to trawl and beach seine survey), and (d) foraging resources for the sturgeons. The results show that their natural reproduction in the Volga–Caspian basin has declined drastically during the past decades under the impact of (a) fluctuations of the Caspian Sea level and flow discharge from the Volga River, (b) blockage of sturgeon migration routes and loss of spawning sites because of dam construction, (c) water pollution in the lower reaches of the Volga River and in the Caspian Sea, and (d) intensive and selective illegal and unreported fishing. The relative significance of these factors has been changing during the study period.     

The RAPD technique failed to identify sex-specific sequences in beluga (Huso huso)

Because the sex of sturgeon cannot be determined externally until the pre‐spawning phase, they are currently sexed by means of minor surgery. Although the survival rate is nearly 100%, this surgery is invasive. One effective solution is to use DNA markers to diagnose the sex. In this study we employed bulked segregant analysis methodology to screen for genetic markers associated with the sex of the beluga (Huso huso). DNA bulks (male and female) were created by combining equal amounts of genomic DNA from 10 fish of both sexes. A total of 310 randomly amplified polymorphic DNA primers was used to screen the bulks, resulting in 4146 bands that were present in both sexes. Our results suggest that either the sex‐specific DNA in sturgeon may be comprised of sequences not complementary to 310 random decamer primers or that sex chromosomes are weakly differentiated in the sturgeon genome.     

Proteome modifications of juvenile beluga (Huso huso) brain as an effect of dietary methylmercury

Methylmercury (MeHg) is the most toxic form of mercury which is bioaccumulated in the aquatic food chain. It has been shown that one of the main targets of MeHg toxicity is the brain, but there is little knowledge of the molecular mechanisms of its toxic effects. In this work we used a proteomics analysis to determine the changes in the brain proteome of juvenile beluga (Huso huso) exposed to dietary MeHg. The juvenile beluga were fed the diet containing 0.8 ppm MeHg for 70 days. Proteins of the brain tissue were analyzed using two-dimensional electrophoresis and MALDI-TOF/TOF mass spectrometry. We found eight proteins with significant altered expression level in the fish brain exposed to MeHg. These proteins are involved in different cell functions including cell metabolism, protein folding, cell division, and signal transduction. Our results support the idea that MeHg exerts its toxicity through oxidative stress induction and apoptotic effects. They also suggest that chronic MeHg exposure would induce an important metabolic deficiency in the brain. These findings provide basic information to understand possible mechanisms of MeHg toxicity in aquatic ecosystems.     

Effect of prebiotic mannan oligosaccharide on hematological and blood serum biochemical parameters of cultured juvenile great sturgeon (Huso huso Linnaeus, 1754)

The nutritional effects of prebiotic mannan oligosaccharide were evaluated using hematological and blood serum biochemical parameters in cultured juvenile great sturgeon (Huso huso). Fish were offered formulated diets containing two levels of prebiotic mannan oligosaccharide (2 and 4 g kg^?1); a basal diet with no prebiotics was used as control. The experiment lasted for 46 days. Blood samples were collected from the caudal veins of 18 apparently healthy fish (average weight 217.77 ± 29.8 g) at the end of the trial. No significant differences were found in the serum enzyme activity levels between treatments (P > 0.05). However, adding mannan oligosaccharide as a supplement to the basal diet resulted in significant differences in lymphocytes and eosinophils between the control and the 2 g kg^?1 treatment (P < 0.05) as well as a significant difference in the creatinine factor in the 2 g kg^?1 mannan oligosaccharide treatment (P < 0.05). The results show that it would be advantageous to add 2 g kg^?1 mannan oligosaccharide to the diets of juvenile great beluga sturgeon (Huso huso).