The use of microsatellite loci for identification of sturgeon species (Acipenseridae) and hybrid forms

The genetic polymorphism of ten sturgeon species that inhabit the territory of the Russian Federation (Russian sturgeon, Siberian sturgeon, Amur sturgeon, Sakhalin sturgeon, Persian sturgeon, ship sturgeon, sterlet, stellate sturgeon, beluga, and kaluga) was examined at five microsatellite loci (Afug41, Afug51, An20, AoxD161, AoxD165) (in total, 3821 individuals). The examined loci were successfully amplified with the same primer set in all species tested and demonstrated a high level of variation. Alleles specific to different species have been identified, which allows them to be used to identify species of sturgeon and their food products. In addition, the possibility of identifying hybrid forms was demonstrated. The assignment test performed in the STRUCTURE software program showed a high probability of correctly assigning each individual to its species based on genotyping with five microsatellite loci examined (96–98%, on average). However, for Russian and Persian sturgeon, the rate of proper species assignment was considerably lower (75 and 84%, respectively).

     

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 state of natural reproduction of stellate sturgeon Acipenser stellatus in the Lower Volga

Data obtained during 1979–2005 on the natural reproduction of stellate sturgeon Acipenser stellatus and water level in the Volga River were analyzed. The relationship between the number of stellate sturgeon larvae migrating down the river, flow quantity during the summer period of low water and the number of spawners entering spawning grounds was shown. Fish productivity of spawning grounds located in the river channel in different zones of the Lower Volga was estimated. Recommendations are presented concerning the increase in the level of stellate sturgeon natural reproduction in the lower pool of the Volgograd hydrosystem.     

Migrations and resources of sterlet <Emphasis Type="Italic">Acipenser ruthenus</Emphasis> (Acipenseridae) from the lower reaches of the Volga River

Based on long-term materials, spawning, postspawning, and downstream migrations of the Lower Volga sterlet Acipenser ruthenus were studied. The dates, extent, pattern (two-step and one-step) and main parameters of the spawning and postspawning migrations of the previously separated three populations of the Lower Volga starlet—Volgograd and eastern and western delta ones were established. The main differences of the downstream migration of this potanadromous species from that of anadromous species of acipenserids were revealed. It was found that, after downstream migration, juveniles of sterlet form a continuous chain of aggregations from the dam of the Volgograd Hydraulic Power System to the Caspian Sea. Each link of this chain acts as a center of formation of local groups of sterlet that are components of the distinguished populations. It is shown that the observed distribution of sterlet in the lower reaches of the Volga is determined by the location of the spawning grounds of individual populations and the extent of downstream migrations of prolarvae and larvae. The total extent of migrations of sterlet is small (it does not exceed 250 km), which is a cause of the relative isolation of the previously distinguished populations. The stocks of Volgograd sterlet are exhausted at the present time; the recovery of its stock is possible, first of all, at the expense of a decrease in illegal fishing. The resources of the eastern and western delta populations of sterlet are in a satisfactory condition.     

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.     

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.     

High‐throughput SNP‐genotyping analysis of the relationships among Ponto‐Caspian sturgeon species

Legally certified sturgeon fisheries require population protection and conservation methods, including DNA tests to identify the source of valuable sturgeon roe. However, the available genetic data are insufficient to distinguish between different sturgeon populations, and are even unable to distinguish between some species. We performed high‐throughput single‐nucleotide polymorphism (SNP)‐genotyping analysis on different populations of Russian (Acipenser gueldenstaedtii), Persian (A. persicus), and Siberian (A. baerii) sturgeon species from the Caspian Sea region (Volga and Ural Rivers), the Azov Sea, and two Siberian rivers. We found that Russian sturgeons from the Volga and Ural Rivers were essentially indistinguishable, but they differed from Russian sturgeons in the Azov Sea, and from Persian and Siberian sturgeons. We identified eight SNPs that were sufficient to distinguish these sturgeon populations with 80% confidence, and allowed the development of markers to distinguish sturgeon species. Finally, on the basis of our SNP data, we propose that the A. baerii‐like mitochondrial DNA found in some Russian sturgeons from the Caspian Sea arose via an introgression event during the Pleistocene glaciation.     

Contemporary status of Caspian sturgeon (Acipenseridae) stock and its conservation

The paper describes the contemporary status of sturgeon stocks in the Caspian Sea. The effect of natural reproduction and hatchery production on sturgeon population restocking is estimated. Variations in the status of spawning populations of sturgeons migrating into the Volga River were analysed. Recommendations on appropriate measures for conservation of the Caspian sturgeon stocks are presented.     

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.     

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.     

Population viability analysis of the Danube sturgeon populations in a Vortex simulation model

Populations of six sturgeon species in the Danube River (beluga, Russian sturgeon, stellate sturgeon, sterlet, ship sturgeon and Atlantic sturgeon) have experienced severe decline during the last several decades, mostly due to the unsustainable fishery, river fragmentation and water pollution. Present lack of knowledge on basic sturgeon demography, life history and relative effects of different negative factors is further hindering implementation of efficient policy and management measures. In the present study, population viability analysis in a Vortex simulation model has been conducted in order to assess the state of the six Danube sturgeon species, their future risk of extinction and to determine the most suitable conservation and management measures. Population viability analysis has revealed a large sensitivity of the Danube sturgeon populations to changes in the natural mortality, fecundity, age at maturity and spawning frequency. It was also confirmed that the sturgeons are highly susceptible to even moderate levels of commercial fishery, and that their recovery is a multi-decadal affair. Stocking with adult individuals was shown to produce considerably greater effect on population persistence than stocking with juveniles, but the latter approach is probably still preferable since it avoids many inherent problems of aquaculture cultivation. This study represents the first population viability analysis of the Danube sturgeons.     

Outmigration pathways of stocked juvenile European sturgeon (Acipenser sturio L., 1758) in the Lower Rhine River,as revealed by telemetry

Working towards a future Rhine Sturgeon Action Plan the outmigration pathways of stocked juvenile European sturgeon (Acipenser sturio L., 1758) were studied in the River Rhine in 2012 and 2015 using the NEDAP Trail system. A total of 87 sturgeon of 3 to 5 years old (n = 43 in 2012, n = 44 in 2015) were implanted with transponders and released in May and June in the river Rhine at the Dutch‐German border, approximately 160 km from the sea. In total three sturgeons (3%) were found dead on river banks within seven days after the release. Based upon their wounds these sturgeons were likely hit by ship‐propellers. Tracking results were obtained from 57 (66%) of the sturgeons, of which 39 (45%) indicated movement into the Port of Rotterdam. Here the sturgeons remained for an average of two weeks, which suggests they spent time to acclimatize to higher salinities before entering the North Sea. Of the 45 (52%) sturgeons that were confirmed to have entered the North Sea, ten (22%) were recaptured (mainly by shrimpers and gill‐nets) close to the Dutch coastline; nine were alive and were released. From the results we obtained the preferred outmigration pathways, movement speeds and an indication of impacting factors (i.e. ship propellers and bycatch). Bycatches provided also localisations information in the coastal area. A next step to complete this work would be to assess habitat selection in freshwater and downstream migration of young of the year (YOY sturgeons) in the Lower Rhine.     

Mitochondrial DNA variation, effective female population size and population history of the endangered Chinese sturgeon, Acipenser sinensis

The anadromous Chinese sturgeon (Acipenser sinensis), mainly endemic to the Yangtze River in China, is an endangered fish species. The natural population has declined since the Gezhouba Dam blocked its migratory route to the spawning grounds in 1981. In the near future, the completion of the Three Gorges Dam, the world's largest hydroelectric project, may further impact this species by altering the water flow of the Yangtze River. Little is currently known about the population genetic structure of the Chinese sturgeon. In this study, DNA sequence data were determined from the control region (D-loop) of the mitochondrial genome of adult sturgeons (n = 106) that were collected between 1995–2000. The molecular data were used to investigate genetic variation, effective female population size and population history of the Chinese sturgeon in the Yangtze River. Our results indicate that the reduction in abundance did not change genetic variation of the Chinese sturgeon, and that the population underwent an expansion in the past. AMOVA analysis indicated that 98.7% of the genetic variability occurred within each year's spawning populations, the year of collection had little influence on the diversity of annual temporary samples. The relative large effective female population size (N _ef) indicates that good potential exists for the recovery of this species in the future. Strikingly, the ratio of N _ef to the census female population size (N _f) is unusually high (0.77–0.93). This may be the result of a current bottleneck in the population of the Chinese sturgeon that is likely caused by human intervention. This revised version was published online in July 2006 with corrections to the Cover Date.     

Use of behavioral and physiological indicators to evaluate Scaphirhynchus sturgeon spawning success

Thirty gravid, female shovelnose sturgeon (Scaphirhynchus platorynchus) were captured in the Lower Missouri River in March 2004 to evaluate the effectiveness of physiology, telemetry and remote sensor technology coupled with change point analysis in identifying when and where Scaphirhynchus sturgeon spawn. Captured sturgeons were instrumented with ultrasonic transmitters and with archival data storage tags (DST) that recorded temperature and pressure. Female sturgeon were tracked through the suspected spawning period. Thereafter, attempts were made to recapture fish to evaluate spawning success. At the time of transmitter implantation, blood and an ovarian biopsy were taken. Reproductive hormones and cortisol were measured in blood. Polarization indices and germinal vesicle breakdown were assessed on the biopsied oocytes to determine readiness to spawn. Behavioral data collected using telemetry and DST sensors were used to determine the direction and magnitude of possible spawning‐related movements and to identify the timing of potential spawning events. Upon recapture observations of the ovaries and blood chemistry provided measures of spawning success and comparative indicators to explain differences in observed behavior. Behavioral and physiological indicators of spawning interpreted along with environmental measures may assist in the determination of variables that may cue sturgeon reproduction and the conditions under which sturgeon successfully spawn.     

Spawning Periodicity, Spawning Migration, and Size at Maturity of Green Sturgeon, Acipenser medirostris, in the Rogue River, Oregon

The Rogue River, Oregon represents one of three important spawning systems for green sturgeon, Acipenser medirostris, in North America. In this paper we describe the spawning migration, spawning periodicity, and size at maturity for green sturgeon caught in the Rogue River during 2000–2004. Green sturgeon were caught by gill net or angling; 103 individuals were tagged with radio or sonic transmitters (externally or internally). Green sturgeon caught by gill net and angling ranged from 145 cm to 225 cm total length. Histological and visual examinations of gonad tissues indicated that most green sturgeon were spawning or post-spawning adults that entered the Rogue River to spawn. Ripe individuals were caught when water temperature was 10–18°C. Specimens carrying transmitters migrated 17–105 km up river; reaches consisting of likely spawning sites were identified based on sturgeon migratory behavior. Most green sturgeon remained in the Rogue River until late fall or early winter when flows increased, after which they returned to the ocean. Eight green sturgeon (males and females) returned to the Rogue River 2–4 years after leaving, entering the river during March, April, and May when water temperatures ranged from 9°C to 16°C. None of the 103-tagged individuals entered the Rogue River during successive years. There appear to be few known natural threats to adult green sturgeon in the Rogue River. However, our data suggest that a high percentage of adults that spawn in the Rogue River (particularly males) were susceptible to harvest by commercial, Tribal, and sport fisheries after leaving the system because they were not adequately protected by maximum size limits during the period of this study. The implications of maximum size limits (or lack of size limits) to green sturgeon are discussed, and recent actions taken by Oregon and Washington Fish and Wildlife Commissions to manage green sturgeon more conservatively are presented.     

Using adaptive resolution imaging sonar to investigate Chinese sturgeon (Acipenser sinensis Gray, 1835) behaviour on its only spawning ground in the Yangtze River

The present study investigated the presence of Chinese sturgeon Acipenser sinensis on its only remaining spawning ground (below the Gezhauba Dam), and monitored the behaviour under different environmental conditions from 24 December 2015 to 23 January 2016. A fixed ARIS (Adaptive Resolution Imaging Sonar) system was used and a total of 72 Chinese sturgeon were detected during nine observations. Detections initially recorded a few A. sinensis in the early days of late‐December 2015, with an increase in recordings, leading to a peak in early‐January 2016 and declining thereafter. Water temperature slowly decreased during the study period from 18.1 to 15.7°C. During the middle of this temperature decline the sturgeon observations peaked, suggesting that Chinese sturgeon could have an optimal temperature range. The sturgeon Detections Per Unit Effort (DPUE) was higher in the night hours, peaking before dawn, suggesting a circadian behaviour rhythm. Sturgeon spawning was not observed during the investigation period. A delay in the decrease in water temperature caused by the Three Gorges Reservoir and the few numbers of reproductively mature individuals are suspected to have contributed to the failure in natural breeding.     

Exotic sturgeons in the Vistula Lagoon in 2011, their occurrence,diet and parasites,with notes on the fishery background

The Siberian sturgeon Acipenser baerii and the sterlet Acipenser ruthenus were recorded for the first time in the Vistula Lagoon in 2011. Among 66 sturgeons collected between April and December 2011, the Siberian sturgeon was the most numerous species (77%); however, a significant seasonal variability was observed, with sterlet dominating in the catches in late autumn of the same year. The stomach contents of the two species differed widely: Siberian sturgeon (14.9–42.2 cm standard length, SL) fed on crustaceans (Cercopagis pengoi, Oithona sp., Neomysis integer) (10.7% IRI), larvae and pupas of insects (Chaoborus sp., Chironomus sp., Polypedilum sp., Procladius sp., Culex sp.) (88.9% IRI) and fishes (Neogobius melanostomus, Osmerus eperlanus) (0.5% IRI), whereas sterlet (24.0–34.4 cm SL) consumed crustaceans (N. integer) (64.3% IRI), larvae of insects (Chironomus sp., Polypedilum sp.) (20.3% IRI) and fish (N. melanostomus) (15.4% IRI). Single Siberian sturgeon (4.3%) were found to harbour the parasitic nematode (Raphidascaris acus).     

Environmental cues for natural reproduction of the Chinese sturgeon,Acipenser sinensis Gray, 1835, in the Yangtze River,China

For effective conservation, it is important to explore the environmental cues initiating the spawning activities of a fish species. Based on monitoring data gathered between 1998 and 2011, the relationships between spawning activities of the Chinese sturgeon, Acipenser sinensis, and several environmental cues were analyzed using the rare events logistic regression ‘Relogit’ method, which indicated that water temperature, 1‐day ?‐discharge, and atmospheric pressure were among the key spawning cues for A. sinensis (P < 0.05). It is suggested that Chinese sturgeon might have an optimal environment window of 17–20°C water temperature, high day‐to‐day discharge increase, and low atmospheric pressure for spawning. In support of Chinese sturgeon reproduction, suggested modifications to the operational procedures for the Three Gorges Reservoir (TGR) to trigger spawning are: lowering the downstream water temperature to below 20°C before mid‐October and expanding the period with water temperatures of between 17 and 20°C; to create a day‐to‐day intermittent increase in the discharge to an optimal spawning water temperature; and to regulate flow at nights with a low atmospheric pressure.     

On the populational structure of sterlet <Emphasis Type="Italic">Acipenser ruthenus</Emphasis> (Acipenseridae) from the Volga Lower reaches

Trawl surveys made in the Lower Volga and the Caspian Sea revealed that the full life cycle of the Lower Volga sterlet is confined to fresh water and only single specimens living in the Volga-Caspian canal can live in the canal and swim to weakly saline and stagnant waters. Therefore, the sterlet living in the Volga-Caspian canal with its expressed current should be attributed to potamodromous forms. In consideration of the data on distribution, the variation of the complex of plastic characteristics, growth rate, and infestation with the parasite Amphilina foliacea it is concluded that within the Lower Volga population of sterlet three populations of the lower rank may be discerned: the Volgograd population living from the Volgograd Hydroelectric Complex dam to the Kamennyi Yar settlement; the eastern population living in the top of the Volga Delta and Buzan channel; and the western population in the Volga-Caspian canal. These populations differ in some plastic characteristics, linear growth rate, the part of fish infested with the parasite A. foliacea, and in the occurrence of anomalies in the structure of mature ovicells.