Dietary protein requirement of fingerling sterlet sturgeon (Acipenser ruthenus)

Although sterlet sturgeon is a valuable candidate for aquaculture development, information regarding the nutrient requirements of this species is limited. Therefore, in this study, we aimed to determine the optimum protein level required by sterlet sturgeon fingerlings. Groups of fingerlings (mean initial body weight ± SD: 7.02 ± 0.02 g; N = 3) were assigned to one of the six dietary protein levels at 29.1%, 39.1%, 46.8%, 49.2%, 53.7%, and 57.6% on a dry matter basis. Isocaloric (18.3 ± 0.2 kJ GE/g diet) semi-purified diets were prepared using white fish meal and casein as the major sources of protein. After the 8-week feeding trial, the fingerlings fed the 49.2% protein diet exhibited the best results in terms of weight gain, feed efficiency, and whole-body nutrient composition (crude protein, crude lipid, gross energy, and amino acid contents), whereas those fed the 53.7% and 57.6% protein diets showed poor performance. Application of the two-slope straight broken-line and quadratic models on the dataset of weight gain revealed the optimum protein level to be 45.9% and 44.1%, respectively. Based on the model selection criteria, including the adjusted coefficient of correlation and corrected Akaike information criterion, the protein level (44.1%) estimated using the quadratic model was more accurate than that obtained using another model (45.9%). Taken together, 44.1% protein level (18.3 kJ GE/g diet on dry matter basis) resulted in the optimal growth of the sterlet sturgeon fingerlings (initial body weight: approximately 7 g).     

Effects of acute temperature change on the metabolism and swimming ability of juvenile sterlet sturgeon (Acipenser ruthenus,Linnaeus 1758)

The objective of this study was to provide information on changes in the metabolism and swimming ability of juvenile sterlet sturgeon, Acipenser ruthenus, caused by acutely low or high temperatures. Changes in critical swimming speed (U_crit), oxygen consumption rate (MO₂), tail beat frequency (TBF) and tail beat amplitude (TBA) were observed with a Steffensen‐type swimming respirometer, an oxygen electrode and a camera at different swimming speeds at three temperatures: 5°C, 15°C, and 25°C. Fish tested at 5°C and 25°C were maintained at 15°C (near optimal) for one week to simulate conditions below a dam. The U_crit value decreased significantly during acute temperature changes at 5°C and 25°C; U_crit was highest near the optimal temperature. Oxygen consumption rate (MO₂) increased with the swimming speed at 15°C; however, at 25°C and 5°C, the MO₂ decreased with the swimming speed. Both TBA and TBF decreased at 5°C and 25°C compared to values at 15°C. The slopes of the regression lines (TBF/U) at 5°C and 25°C seemed lower compared to 15°C.     

Juvenile growth of Acipenser ruthenus and 4 different sturgeon hybrids

Juvenile growth of A. ruthenus and four different sturgeon hybrids ( Huso huso x A. ruthenus, A. baerii x A. ruthenus, A. gueldenstaedtii x A. ruthenus, A. gueldenstaedtii x H. huso ) was studied for 71 days applying two feeding regimes, offering a fry diet (3.5 % daily rate) for 26 days for the hybrids and 41 days for A. ruthenus and a fattening diet (3 % daily rate) for 45 days and 30 days respectively. Significant growth differences occured at the end of experiment: The hybrid A. gueldenstaedtii x H. huso showed the highest, A. ruthenus the lowest growth.     

Effects of dietary vitamin C on tissue ascorbate and collagen status in sturgeon hybrids (Acipenser ruthenus L. x Acipenser baeri Brandt)

The aim of this study was to determine the vitamin C status of various tissues, and collagen concentration in cartilage, of a sterlet ( Acipenser ruthenus L.) x Siberian sturgeon ( Acipenser baerii Brandt) hybrid fed different dietary vitamin C rations. Growth, vitamin C status and collagen concentration were measured in groups fed diets supplemented with 150, 300, 600 mg.kg^-1 ascorbate-2-polyphosphate (APP); 100, 200, 400 mg.kg^-1 ascorbate-2-monophosphate (AMP), 1000 mg.kg^-1 L-ascorbic acid (AA) and with an almost ascorbate free diet (Total AA > 5 mg.kg^-1), as a control.
No significant differences in growth were observed, and no external symptoms of scurvy developed in the vitamin C-free group. No reduction in total vitamin C concentration was revealed in the tissues examined, as compared with initial concentrations. Ascorbate-2-phosphate esters were only found in kidney or hepatopancreas of fish fed with these vitamin C derivatives. Significantly (p<0.05) lower contents of collagen were observed in the control group after 8 weeks feeding. No significant differences were found among the groups after 16 weeks although the collagen concentration in the cartilage was slightly lower in the control group without vitamin C supplementation.     

Characterization of xenobiotic metabolizing enzymes in sturgeon (Acipenser baeri)

1. Cytochrome P-450, NADPH-cytochrome c reductase, benzo(a)-pyrene hydroxylase (AHH), 7-ethoxycoumarin-O-deethylase (7-ECOD), epoxide hydrolase (EH), UDP-glucuronyltransferase (UDPGT) and glutathione S-transferase (GSHST) activities in sturgeon (Acipenser baeri) have been measured and partially characterized. 2. Cytochrome P-450-dependent monoxygenase (MO), EH, and conjugation reactions were detected in liver and to a lesser extent in kidney and gills. 3. Hepatic enzyme activities in the sturgeon were equally high or higher than in rainbow trout liver, with the exception of UDPGT whose activity was 14% of that in trout liver. 4. The MO and EH activities displayed the expected pH maxima of 7.5, whereas transferases were relatively independent of the pH in the 6.5-7.5 range. 5. The temperature optima for MO and EH were close to those reported in other fish species, whereas for conjugation reactions the temperature optima were 45 and 60 degrees C for GSHST and UDPGT respectively.     

史氏鲟外周血细胞的显微及超微结构

采用血细胞计数、光学显微及电子显微技术对二龄史氏鲟外周血细胞的数目、形态及结构进行了研究。二龄史氏鲟红细胞的数目为47.75×104个/mm3,白细胞数目为2.9万个/mm3,其中淋巴细胞所占比率最高。史氏鲟的外周血中除正常红细胞外,还有处于分裂状态及未成熟的红细胞。史氏鲟外周血中的白细胞有四种类型,分别为淋巴细胞、粒细胞、血栓细胞和单核细胞。其中粒细胞有两种,即嗜中性粒细胞和嗜酸性粒细胞。嗜中性粒细胞含有多种形状的核,其中分叶的核数目较多,粒细胞及淋巴细胞均类似于哺乳动物。对史氏鲟外周血细胞细微结构的观察显示:红细胞中具有少量的细胞器;淋巴细胞结构典型;单核细胞较粒细胞稍小且具有较多线粒体;血栓细胞具有梭形和圆形两种,胞质较少,其中梭形的血栓细胞胞质几乎透明;对粒细胞的颗粒按照形状和电子密度进行了分类。     

Site heteroplasmy in the mitochondrial cytochrome b gene of the sterlet sturgeon Acipenser ruthenus

Sturgeons are fish species with a complex biology. They are also characterized by complex aspects including polyploidization and easiness of hybridization. As with most of the Ponto-Caspian sturgeons, the populations of Acipenser ruthenus from the Danube have declined drastically during the last decades. This is the first report on mitochondrial point heteroplasmy in the cytochrome b gene of this species. The 1141 bp sequence of the cytb gene in wild sterlet sturgeon individuals from the Lower Danube was determined, and site heteroplasmy evidenced in three of the 30 specimens collected. Two nucleotide sequences were identified in these heteroplasmic individuals. The majority of the heteroplasmic sites are synonymous and do not modify the sequence of amino acids in cytochrome B protein. To date, several cases of point heteroplasmy have been reported in animals, mostly due to paternal leakage of mtDNA. The presence of specific point heteroplasmic sites might be interesting for a possible correlation with genetically distinct groups in the Danube River.     

Morphology, chemical contents and physiology of chondrostean fish sperm: a comparative study between Siberian sturgeon (Acipenser baerii) and sterlet (Acipenser ruthenus)

The present study was carried out with two sturgeon species, Siberian sturgeon (Acipenser baerii) and sterlet (A. ruthenus) to compare spermatological parameters to better understand inter‐species differences. Significant differences between morphometric features were observed such as acrosome length, acrosome width, head length, midpiece width and flagellar length, while midpiece length did not reveal such differences. The sterlet has a shorter spermatozoon than the Siberian sturgeon. Ultrastructural parameters vary significantly in terms of length of the nucleus, diameter of the endonuclear canals (EC), size of posterolateral projections (PP) and diameter of flagellum. Mean values for density of spermatozoa in the semen, seminal plasma pH, osmolality (mOsmol kg^?1), along with Ca²⁺, Na⁺, K⁺, Cl^? ions concentrations (mm ) were determined to be 0.61 ± 0.37 × 10⁹, 8.16 ± 0.18, 77.20 ± 52.28, 0.24 ± 0.06, 31.39 ± 10.21, 3.51 ± 1.10, 14.00 ± 4.30 in A. baerii and 0.41 ± 0.32 × 10⁹, 8.13 ± 0.19, 50.74 ± 6.27, 0.16 ± 0.11, 20.11 ± 3.78, 1.26 ± 0.54, 6.11 ± 0.60 in A. ruthenus, respectively. Significant differences were observed in Na⁺, K⁺ and Cl^? concentrations in the seminal plasma as well as in sperm velocity. The percentage of motile spermatozoa did not show any significant difference between the two species. Comparing the results of this study with published literature data on sturgeon spermatozoa reveals that morphological and ultrastructural parameters of spermatozoa together with some parameters of the seminal fluid and spermatozoa velocity can be used in comparative spermatology to better understand inter‐species differences. The observed biochemical and physiological differences should be also considered for the development of methods for controlled reproduction and for sperm cryopreservation techniques.     

Fertility of females of sturgeon hybrids obtained from species with different levels of ploidy (Acipenser ruthenus and A. dauricus) and their cloning

Main purpose of this work is the identification of females of artificial sturgeon hybrids capable to produce unreduced oocytes. The importance of this task is due to the ability to receive clonal all-female lines. Experiments were performed on the previously obtained reciprocal hybrids of sterlet, Acipenser ruthenus (S) with ~120 chromosomes and kaluga, Acipenser dauricus (K) with ~260 chromosomes. Karyotypes of backcross hybrids of (S × K) female (obtained by crossing sterlet female with kaluga male) and sterlet male included 180 – 190 chromosomes. This means that (S × K) female produced eggs with ~125 chromosomes and its karyotype consisted of ~250 chromosomes. This number was confirmed by a comparative analysis of erythrocyte size in this female and species with different ploidy. Karyotype with ~250 chromosomes can occur in (S x K) female only as a result of fertilization of a diploid sterlet egg (120 chromosomes) with kaluga haploid sperm (~130 chromosomes). Eggs of hybrid fertile (S × K) female, inseminated with inactivated sperm of Amur sturgeon and sterlet, developed into viable gynogenetic offspring, confirmed by the analysis of five microsatellite loci in this progeny, (S x K) female, and males used for UV-inactivated sperm. These data allow us to propose a method for obtaining fertile females of sturgeon hybrids from species with different ploidy. For this, experimentally obtained diploidized eggs from diploid 120-chromosome species must be fertilized by 250–270-chromosome male. Karyotypes of backcross hybrids of (K × S) female (obtained by crossing kaluga female with sterlet male) and sterlet male included ~250 chromosomes and hybrids of this female with kaluga male had ~320 chromosomes. These results proved an ability of hybrid (K × S) female to produce unreduced eggs, resulting in triploid backcrosses. The absence of reduction during egg development is well known in clonal forms (species) of vertebrates, which are of hybrid origin, and in artificially created fish hybrids. However, this has not been reported previously for sturgeons. Insemination of eggs of (K × S) female with UV-inactivated sperm of sterlet and Amur sturgeon led to offspring generation for which the genetic identity to their mother was proved using microsatellite analysis. That is, clonal inheritance was observed. These results suggest the possibility of developing a technology to produce all-female offspring. Artificial production of clonal lines in hybrid vertebrates can be also considered as experimental reproduction of the first stages of reticular speciation in nature.     

Comparative coelomic fluid composition of sterlet sturgeon Acipenser ruthenus Linnaeus, 1758, Siberian sturgeon Acipenser baerii Brandt, 1869, and Russian sturgeon Acipenser gueldenstaedtii Brandt & Ratzeburg, 1833

The objectives of this study were to compare the physicochemical properties of coelomic fluid (CF) in three sturgeon species, sterlet sturgeon Acipenser ruthenus (age 5–8 years), Siberian sturgeon Acipenser baerii (age 15–20 years), and Russian sturgeon Acipenser gueldenstaedtii (age 13–18 years). For the study, CF was collected by plastic pipette from the eggs of mature female sterlet sturgeon (N = 10), Siberian sturgeon (N = 7) and Russian sturgeon (N = 4); osmolality, pH, ionic composition (Na⁺, K⁺, Ca²⁺, Mg²⁺, Cl^?), glucose, cholesterol and total protein of the CF were then analyzed. Mean concentration of K⁺ (4.4–6.11 mmol/L), Cl^? (89.8–98.0 mmol/L), Ca²⁺ (0.87–0.96 mmol/L), Mg²⁺ (0.57–0.67 mmol/L), cholesterol (0.13–0.16 mmol/L), total protein (2.41–3.57 g/L), pH (7.92–7.98), and osmolality (190–213 mOsm/kg) of CF were not significantly different among these three species. However, the mean concentration of Na⁺ was significantly lower in sterlet sturgeon (104.6 ± 7.7 mmol/L; p < .05) than in the Siberian (126.4 ± 6.2) and Russian (123.0 ± 5.9) sturgeon. For these three species, Na⁺, K⁺, Cl^?, and Mg²⁺ were the dominating ions; several significant correlations were observed among these ions and other physiochemical properties of CF. This is the first report on the ionic and biochemical composition of the CF of sterlet, Siberian and Russian sturgeon, which can be used as a reference point for further development of artificial media for the short‐term storage of unfertilized sturgeon eggs as well as for the standardization of the fertilization protocol for these species in controlled reproduction.     

First evidence of autotriploidization in sterlet (Acipenser ruthenus)

Polyploidization has played an important role in vertebrate evolution. Acipenseridae bring clear examples of polyploidy ancestry and, also, polyploidization seems to be an ongoing process in these fishes. In the present study, the genetic origin of six triploid specimens morphologically determined as Acipenser ruthenus from commercial aquaculture was analyzed using a combination of mitochondrial and nuclear markers. A further five successive statistical analyses including median joining of mitochondrial DNA control region sequences, principal coordinate analysis (PCA), factorial correspondence analysis (FCA), STRUCTURE assignation, and NewHybrids status determination for microsatellite data were applied for the clarification of the origin of one extra chromosome set added in these triploids genomes. Although interspecific hybridization had been suggested as a source of these triploids, the statistical analyses showed that the investigated triploids originate from autotriploidization rather than from interspecific hybridization. Therefore, we conclude that a combination of molecular markers with suitable statistical analyses should be used to verify the origin of unusual ploidy level. Evidently, such an approach is critically essential in aquaculture, where interspecific hybridization is very common and usually detected by changes in ploidy levels only.     

Cryopreservation of sterlet (Acipenser ruthenus) spermatozoa using different cryoprotectants

The present study examined the effectiveness of different cryoprotectan uses for cryopreservation of sterlet (Acipenser ruthenus) sperm. Four different cryoprotectans [dimethyl‐sulfoxide (DMSO), dimethylacetamide (DMA), ethyleneglycol (EG) and methanol (MET)] in concentrations of 5 and 10% in the extender media (30 mm sucrose, 1 mm KCl, 25 mm Tris–HCl pH 8.5) were used for the cryopreservation of sperm from five sterlet males. Percentages of motility, velocity, fertilization and hatching rate were measured. The highest post‐thawing motility was observed in sperm samples cryopreserved with 10% MET (46 ± 19%), 10% DMA (47 ± 18%) and 10% DMSO (45 ± 7%). Fertilization rate in the control (fresh sperm) was 69 ± 9% and the hatching rate 61 ± 8%. A higher hatching rate after cryopreservation was obtained with 10% MET (32 ± 17%) and 5% DMA (23 ± 3%) when compared to data obtained with 10% DMA (9 ± 5%), 5% MET and 5 and 10% DMSO, wherein the hatching rate was around zero. Our results demonstrate that DMA can be used for cryopreservation of sturgeon spermatozoa.     

Morphometric recognition of two morphs in sterlet (Acipenser ruthenus) population induced by different reproductive behaviour

Four species of sturgeons appear in the Yugoslav part of the Danube River. Among them three are anadromous, beluga ( Huso huso ), Russian sturgeon ( Acipenser gueldenstaedtii ) and stellate sturgeon ( Acipenser stellatus ) and migrate to Danube from the Black Sea to spawn. Only sterlet ( Acipenser ruthenus ) is exclusively freshwater. There are obvious differences between vernal and winter races of anadromous sturgeons, due to their different migratory patterns. Russian ichthyologist Berg (1934) claimed that there are two races (morphs) in the starlet population as well. This stimulated long discussion in Russian and Yugoslav literature, and it was the starting point for our research on sterlet in the Yugoslav part of the Danube River.
Morphometric analysis was preformed on 159 individuals caught in Danube River among 1123 and 1173 km, on four localities during September and October 2002. Analysis included 24 morphological and 3 meristic characters. K‐means cluster analysis indicated existence of two different clusters (groups), one with 95 (59·75%) and other with 64 (40·25%) samples. Significant differences were found for preorbital, preoral and prebarbel length. Mean values (in % of head length) for first group were 50·13 ± 1·71, 57·61 ± 1·87, 39·74 ± 2·20 and for second group 53·50 ± 2·46, 62·07 ± 2·20 and 43·30 ± 2·08 respectively. Findings were confirmed by Principal Components & Classification Analysis. These results speak in favour of existence of two morphs in sterlet population, one with long and pointed rostrum (vernal) and other with short rostrum (winter). According to the literature that is probably influenced by different reproductive behaviour (time separation in spawning).     

Primary projections of the trigeminal nerve in two species of sturgeon: Acipenser oxyrhynchus and Scaphirhynchus platorynchus

Horseradish peroxidase histochemical studies of afferent and efferent projections of the trigeminal nerve in two species of chondrostean fishes revealed medial, descending and ascending projections. Entering fibers of the trigeminal sensory root project medially to terminate in the medial trigeminal nucleus, located along the medial wall of the rostral medulla. Other entering sensory fibers turn caudally within the medulla, forming the trigeminal spinal tract, and terminate within the descending trigeminal nucleus. The descending trigeminal nucleus consists of dorsal (DTNd) and ventral (DTNv) components. Fibers of the trigeminal spinal tract descend through the lateral alar medulla and into the dorsolateral cervical spinal cord. Fibers exit the spinal tract throughout its length, projecting to the ventral descending trigeminal nucleus (DTNv) in the medulla and to the funicular nucleus at the obex. Retrograde transport of HRP through sensory root fibers also revealed an ascending bundle of fibers that constitutes the neurites of the mesencephalic trigeminal nucleus, cell bodies of which are located in the rostral optic tectum. Retrograde transport of HRP through motor root fibers labeled ipsilateral cells of the trigeminal motor nucleus, located in the rostral branchiomeric motor column.     

First evidence of hybridization between endangered sterlets (Acipenser ruthenus) and exotic Siberian sturgeons (Acipenser baerii) in the Danube River

Most natural populations of Eurasian sturgeons have declined dramatically during recent decades, reaching historic low levels today. During the same period, sturgeon has become very popular in European aquaculture. Because many hatcheries are located near rivers, their unintentional escape is often documented, especially during floods. Until now, no cases of successful hybridization of these escaped fish with wild stocks have been reported. In this study, the genetic structure of a highly threatened population of sterlets (Acipenser ruthenus) from the Upper Danube was analysed as a requirement for their conservation. Surprisingly, we observed genotypes and morphotypes of Siberian sturgeon (Acipenser baerii), as well as hybrids between this species and native sterlets. This hybridization poses a serious threat for the survival of this isolated sterlet population in the upper part of the Danube. For the first time, natural reproduction is documented for Siberian sturgeon outside their natural range in Europe. This finding demonstrates the risk of extinction by hybridisation of endangered populations. We would like to stress that taking into consideration the risks for native sturgeon populations, farmed sturgeon should not be released into the wild, and all measures should be taken to prevent their accidental escape.     

俄罗斯鲟仔鱼初次摄食时间对生长及存活的影响

以人工繁育的俄罗斯鲟仔鱼为对象,采用实验生态学方法,研究了初次摄食时间对生长及存活的影响。结果表明：在水温17.4 ℃下,仔鱼在9～10日龄初次摄食,初次摄食前投喂对俄罗斯鲟仔鱼的生长和存活无显著性影响,延迟2 d以上初次摄食对生长影响明显,存活率在18日龄初次摄食时显著下降,24日龄时饥饿仔鱼全部死亡;23～24日龄时仔鱼如不能建立外源营养即达到饥饿不可逆点（PNR）。人工养殖条件下,俄罗斯鲟仔鱼必须在初次摄食后14 d内建立起主动摄食能力才能保证其仔鱼的正常发育、生长和存活。