Analysis of motility parameters from paddlefish and shovelnose sturgeon spermatozoa

Ninety to 100% of paddlefish Polyodon spathula were motile just after transfer into distilled water, with a velocity of 175 μm s^-1, a flagellar beat frequency of 50 Hz and motility lasting 4–6 min. Similarly, 80–95% of shovelnose sturgeon Scaphirhynchus platorynchus spermatozoa were motile immediately when diluted in distilled water, with a velocity of 200 μm s^-1, a flagellar beat frequency of 48 Hz and a period of motility of 2–3 min. In both species, after sperm dilution in a swimming solution composed of 20 mM Tris–HCl (pH 8·2) and 20 mM NaCl, a majority of the samples showed 100% motility of spermatozoa with flagella beat frequency of 50 Hz within the 5 s following activation and a higher velocity than in distilled water. In such a swimming medium, the time of motility was prolonged up to 9 min for paddlefish and 5 min for sturgeon and a lower proportion of sperm cells had damage such as blebs of the flagellar membrane or curling of the flagellar tip, compared with those in distilled water. The shape of the flagellar waves changed during the motility phase, mostly through a restriction at the part of the flagellum most proximal to the head. A rotational movement of whole cells was observed for spermatozoa of both species. There were significant differences in velocity of spermatozoa between swimming media and distilled water and between paddlefish and shovelnose sturgeon.     

Phylogenetic relationships of Amur sturgeon Acipenser schrenckii Brandt, 1869 based on 18S rDNA sequencing

The phylogenetic relationships of Amur sturgeon A. schrenckii Brandt, 1869 with related species have been analyzed based on sequencing of the 18S rDNA small subunit. The complete sequence (1746 bp) of 18S rDNA has been estimated in seven individual A. schrenckii clones. The results show that the rDNA mutation profile of A. schrenckii 18S is very similar to that of A. fulvescens (Genbank data). Structural-functional and phylogenetic analyses allowed us to identify a presumably expressed variant, as well as taxon-specific mutation (adenine insertion after position 658 bp), for A. schrenckii 18S rDNA. Phylogenetic reconstructions performed with various approaches (NJ, MP, ML and Bayesian) support the monophyly of the genus Acipenser and point (1), based on which, in accordance with the classification based on ecological and morphological data (Artyukhin, 2006), the Amur sturgeon is closer to the sterlet than the Baltic sturgeon and (2) to substantial differentiation between North American (A. fulvescens) and Eurasian (A. schrenckii, A. ruthenus, and A. sturio) species of Acipenseridae.     

Movement of sturgeons (Acipenser oxyrinchus oxyrinchus and A. fulvescens) in the St. Lawrence Estuary: A 35-year tagging study

Movement of Atlantic sturgeon (Acipenser oxyrinchus oxyrinchus) and lake sturgeon (A. fulvescens) in the St. Lawrence Estuary (Québec, Canada) are not fully understood. To assess the movement extent of both species, a mark–recapture study was conducted in collaboration with commercial fishermen operating in the St. Lawrence Estuary. Between 1981 and 2015, 3,367 Atlantic sturgeon (fork length 21.8–199.5 cm) and 3,180 lake sturgeon (fork length 17.8–190.8 cm) were tagged and released. Of these, 673 Atlantic sturgeon and 42 lake sturgeon were recaptured. The maximum distances traveled between capture and recapture locations were 1,307 km for Atlantic sturgeon (8 years after initial capture) and 252 km for lake sturgeon (less than 1 year after initial capture). Statistical analyses identified differences in the dispersal distance of both species as revealed by a first component characterized by individuals with short dispersal distances (98% and <35 km for Atlantic sturgeon; 58% and <1 km for lake sturgeon) and a second component characterized by individuals with longer dispersal distances (2% and >600 km for Atlantic sturgeon; 42% and >190 km for lake sturgeon). We suggest that the short dispersal distances detected in the vast majority of Atlantic sturgeon recaptures likely reflect strong site fidelity, highlighting the importance of the St. Lawrence Estuary as a preferred habitat for juveniles and subadults. Although recaptures were low for lake sturgeon because this species is only marginally targeted by commercial fishermen in the St. Lawrence Estuary, our results also showed that this species uses estuarine habitats and that half of the population seems to exhibit strong site fidelity (67% of individuals were recaptured within 2 km).     

Induction of meiotic gynogenesis in ship sturgeon Acipenser nudiventris using UV-irradiated heterologous sperm

Diploid gynogenesis was induced in ship sturgeon Acipenser nudiventris using UV-irradiated sperm from Siberian sturgeon Acipenser baerii. The optimal condition for the retention of the second polar body in ship sturgeon was determined to be 10 min after activation/fertilization in experiments. The temperature of cold shock and its duration were 2.5 °C and 30 min, respectively. A total of 30 gynogens of known parentage from experimental treatments were screened using microsatellite DNA analysis, and uniparental transmission in meiogens was confirmed. The results show that heterologous Siberian sturgeon sperm is applicable as UV-irradiated sperm for the induction of gynogenesis in ship sturgeon. This technique may recover the critically endangered sturgeon species that are becoming extinct.     

从线粒体DNA控制区核苷酸序列论达式鲟、亚洲和北美洲中吻鲟的分类地位

尽管古老的鲟形目鱼类的分类及系统演化一直是中外学者感兴趣的研究课题,但迄今仍有诸多谜团未解。其中,关于亚洲远东地区和北美地区的中吻鲟(Acipenser medirostris)的分类地位争论已久。长江水系的达式鲟(A.dabryanus)和中华鲟(A.sinensis)及其它鲟属(Acipenser)鱼类之间的亲缘关系近年来也有异议。为了给上述争议提供更多的科学依据,作者测定了线粒体DNA(mtDNA)控制区 (D-loop) 的核苷酸序列,并进行了分子系统学和遗传差异分析。研究结果表明：⑴UPGMA,NJ和MP分子系统学分析方法以及遗传差异分析都支持了亚洲远东地区中吻鲟和北美中吻鲟为一个有效种的观点;⑵同样研究方法并结合相关群体遗传资料表明,长江达式鲟与中华鲟关系最近,提出了达式鲟为中华鲟的一陆封类群的假说。最后,作者认为本文提出的观点和假说还需要更多的研究来证实。     

Motility of spermatozoa from shovelnose sturgeon and paddlefish

The spermatozoa in the seminal plasma from shovelnose sturgeon Scaphirhynchus platorynchus and paddlefish Polyodon spathula were immotile with only a few spontaneously motile spermatozoa for 5-10 and 10-20 s, respectively. Spermatozoa of shovelnose sturgeon were observed to be 100% motile immediately after sperm dilution in 10 m m NaCl and 20 m m Tris-HCl, pH 8.5. The duration of mass progressive movement was 2-3 min; and 1 to 5% of spermatozoa remain active after 360 s (P<0.01). Spermatozoa of paddlefish demonstrated the best motility 10 s after dilution in 10 m m NaCl with 20 m m Tris-HCl, pH 8.5. The duration of mass progressive movement was 2-3 min and 1 to 5% of spermatozoa remained active after 370 s ( p <0.01). The spermatozoa of shovelnose sturgeon and paddlefish were motile in a range of osmotic pressure from 0 to 100 mosmol kg⁻¹ and 0 to 120 mosmol kg⁻¹, respectively. The best results with short-term storage of sperm from shovelnose sturgeon and paddlefish were observed in 100 m m glucose + 20 m m Tris-HCl, pH 8.5 and 150 m m glucose + 20 m m Tris-HCl, pH 8.5.     

从线粒体DNA控制区核苷酸序列论达式鲟、亚洲和北美洲中吻鲟的分类地位

尽管古老的鲟形目鱼类的分类及系统演化一直是中外学者感兴趣的研究课题 ,但迄今仍有诸多谜团未解。其中 ,关于亚洲远东地区和北美地区的中吻鲟 (Acipensermedirostris)的分类地位争论已久。长江水系的达式鲟 (A .dabryanus)和中华鲟 (A .sinensis)及其它鲟属 (Acipenser)鱼类之间的亲缘关系近年来也有异议。为了给上述争议提供更多的科学依据 ,作者测定了线粒体DNA (mtDNA)控制区(D loop) 的核苷酸序列 ,并进行了分子系统学和遗传差异分析。研究结果表明 :⑴UPGMA ,NJ和MP分子系统学分析方法以及遗传差异分析都支持了亚洲远东地区中吻鲟和北美中吻鲟为一个有效种的观点 ;⑵同样研究方法并结合相关群体遗传资料表明 ,长江达式鲟与中华鲟关系最近 ,提出了达式鲟为中华鲟的一陆封类群的假说。最后 ,作者认为本文提出的观点和假说还需要更多的研究来证实。     

Intraperitoneal injection urocortin-3 reduces the food intake of Siberian sturgeon (Acipenser baerii)

Urocortin-3 (UCN3), one of the corticotropin releasing factor (CRF) family peptides, which was discovered in 2001, has a variety of biological functions. However, the researches of UCN3 in fish were scarce. In order to understand whether UCN3 play a role in regulating food intake in fish, we first cloned the ucn3 cDNAs sequence of Siberian sturgeon (Acipenser baerii Brandt), and investigated the ucn3 mRNA levels in 11 tissues. The Siberian sturgeon ucn3 cDNA sequence was 1044 bp, including an open reading frame (ORF) of 447 bp that encoded 148 amino acids with a mature peptide of 40 amino acids, a 5ʹ-terminal untranslated region (5ʹ-UTR) of 162 bp and a 3ʹ-terminal untranslated region (3ʹ-UTR) of 435 bp. The result of tissue distribution showed that ucn3 widely distributed in 11 tissues with highest expression in brain. We also assessed the effects of periprandial (pre- and post-feeding), fasting and re-feeding on ucn3 mRNAs abundance in brain. The results showed the expression of ucn3 mRNA in brain was significantly elevated after feeding, decreased after fasting 17 days and increased after re-feeding. To further investigate the food intake role of UCN3 in Siberian sturgeon, we performed intraperitoneal (i.p.) injection of Siberian sturgeon UCN3 (SsUCN3) with three doses (60, 120 or 240 ng/g) and recorded the food intake. Acute and chronic i.p. injection SsUCN3 reduced the food intake in a dose-dependent pattern. In conclusion, this study indicates that SsUCN3 acts as a satiety factor to inhibit the food intake of Siberian sturgeon.     

Motility and fertility of cryopreserved semen in Persian sturgeon,Acipenser persicus,stored for 30–60 min after thawing

We investigated the effect of storage times of frozen–thawed Persian sturgeon (Acipenser persicus) semen on the duration of sperm motility, percentage of motile sperm, and fertilization and hatching rates of fresh sperm and sperm stored for 0, 30, and 60 min at 4 °C post-thawing. Frozen thawed semen analyzed immediately after thawing had similar quality characteristics as fresh semen. For cryopreserved semen stored for 30 min after thawing the characteristics did not differ to fresh semen and cryopreserved semen. For cryopreserved semen stored for 60 min a significant decline in the parameters was observed. Fertilization and hatching rates were not affected by storage times of maximally 30 min of storage.     

In vitro sperm maturation in sterlet,Acipenser ruthenus

The aim of the study was to examine sperm maturation in sturgeon and to establish the localization of the maturation. We demonstrated that sperm maturation occurs in sturgeon outside the testes via dilution of sperm by urine. The process involves the participation of high molecular weight (>10 kDa) substances and calcium ions.     

Spatiotemporal changes in juvenile lake sturgeon abundance in a large river

Monitoring trends in a long-lived species such as lake sturgeon (Acipenser fulvescens) has it challenges. Targeting the correct life stage to monitor can provide different and contrasting results. Early life stages experience high variable mortality and are often not a good predictor of population trend. Juvenile lake sturgeon (i.e., 5–10 years) experience high survival and could be a good life stage to monitor for assessing population trends. A structured, randomized gillnetting program, Broad-scale Monitoring, which was selective to juvenile lake sturgeon, was conducted over three cycles (i.e., 5 + years apart) in three contiguous Ottawa River reaches supporting lake sturgeon. In this study, we assessed spatiotemporal changes in juvenile lake sturgeon abundance among cycles, among and within the three river reaches. Density rasters based on catch records of lake sturgeon spatially were created using multilevel B-splines. Spatial differences in abundance among cycles was determined by obtaining the difference between the rasters. Three cycles of Broad-scale Monitoring sampled 334 lake sturgeon at a mean CPUE of 0.90 (0.12 SE) sturgeon • gang⁻¹ and a mean total length 673 mm (115 SD) from all three reaches. The relative abundance of juvenile lake sturgeon did not significantly vary among sample cycles, however, CPUE varied within reaches among cycles. There was a spatiotemporal change in density with a decrease in the lowest reach whereas juvenile lake sturgeon density increased the most in the upper reach. The approach used in this study has the potential to monitor spatiotemporal changes in juvenile lake sturgeon abundance populations and may prove effective when assessing rehabilitation or fisheries management efforts.     

Molecular cloning and functional analysis of Chinese sturgeon (Acipenser sinensis) growth hormone receptor

A full length cDNA encoding the growth hormone receptor (GHR) of Chinese sturgeon was cloned in order to investigate the mechanism of growth hormone in regulating the growth of Chinese sturgeon. The open reading frame of the cloned Chinese sturgeon growth hormone receptor (csGHR) cDNA encodes a trans-membrane protein of 611 amino acids containing all the characteristic motifs of GHR. By sequence alignment, substitutions of amino acid residues highly conserved in other species were identified. Using the CHO cell culture system, the function of csGHR and the biological significance of the amino acid substitution in csGHR were examined. The promoter of serine protease inhibitor 2.1 (Spi2.1) was trans-activated upon stimulation of seabream GH (sbGH) in the csGHR-expressing CHO cells. Furthermore, CHO cells stably expressing csGHR were stimulated to proliferate by sbGH. In agreement with our previous report, Chinese sturgeon growth hormone-binding protein (csGHBP) was detected in the culture medium of CHO cells stably expressing csGHR. Mutation of Asp residue in the ligand binding motif in csGHR to Glu significantly enhanced csGHR’s biological function, whereas mutation of Asp residue to Ala decreased its biological function. The results demonstrated that the cloned csGHR was of full biological function and the csGHBP could be generated through proteolysis of csGHR. These findings might provide new insights into thoroughly understanding the regulatory mechanism of Chinese sturgeon growth.     

Verification of meiotic gynogenesis in Siberian sturgeon (Acipenser baeri Brandt) using microsatellite DNA and cytogenetical markers

Diploid gynogenesis was induced in Siberian sturgeon Acipenser baeri using ultra violet (UV)-irradiated bester ( Huso huso × Acipenser ruthenus ) sperm. The higher survival rate of meiotic diploids was noted after 1350 ergs mm⁻² UV irradiation. A total of 80 meiotic diploids of known parentage from two different experimental treatments were screened using microsatellite DNA and cytogenetical analysis, and uniparental transmission in meiotic diploids was confirmed.     

Reproductive Structure of the Adult Green Sturgeon, Acipenser medirostris, Population in the Rogue River, Oregon

The primary objective of this study was to determine the reproductive structure of the adult green sturgeon population in the Rogue River. Green sturgeon were captured by gillnet in the lower 11.6–68.4 river kilometers in April to July 2000–2003 and September and October 2002–2003. Gonadal tissue, collected by biopsy, was processed histologically, blood was collected from the caudal vasculature, and fork length (FL) and total length (TL) (±0.5 cm) were measured for each individual. Sex steroids, testosterone (T), 11-ketotestosterone (11-KT), and estradiol-17β (E2), were measured by radioimmunoassay. Biological samples were collected from a total of 88 green sturgeon of which 37 females and 41 males were confirmed by histological analysis. Four gravid females, captured in the spring, were visually identified, and oocyte polarization index and ovarian follicle diameter indicated that these females were in spawning condition. Gonadal samples collected from six individuals did not contain gonial cells, hence the sex and stage of maturity in these individuals remains unknown. Of the 20 females captured in the spring, 1 was vitellogenic, 4 were post-vitellogenic, and 15 were post-ovulatory. Twenty-one females were captured in the fall of which 6 were pre-vitellogenic, 7 vitellogenic, and 8 post-ovulatory. Of the 16 males captured in the spring, 2 were pre-meiotic, 8 were ripe or actively spermiating, and 6 were post-spermiation. Twenty-five males were captured in the fall: 11 pre-meiotic males and 14 post-spermiation. The majority of green sturgeon captured in the Rogue River were reproductively active or had recently spawned indicating the importance of this river for the preservation of green sturgeon.     

Experimental assessment of predation risk for juvenile green sturgeon,Acipenser medirostris,by two predatory fishes

Predation is a common cause of early life stage mortality in fishes, with reduced risk as individuals grow and become too large to be consumed by gape-limited predatory fishes. Large-bodied species, such as sturgeon, may reach this size-refuge within the first year. However, there is limited understanding of what this size threshold is despite the value of this information for conservation management. We conducted laboratory-based predation experiments on juvenile green sturgeon, Acipenser medirostris, to estimate vulnerability to predation during outmigration from their natal reaches in California to the Pacific Ocean. Two highly abundant and non-native predatory fish species (largemouth bass, Micropterus salmoides, and striped bass, Morone saxatilis) were captured in the wild to be tested with developing juvenile green sturgeon from the UC Davis Green Sturgeon Broodstock Program. Experimental tanks, each containing five predators, received thirty prey for 24-hr exposures. Between sturgeon prey trials, predators were exposed to alternative prey species to confirm predators were exhibiting normal feeding behaviors. In addition to green sturgeon mortality data, trials were video recorded and predatory behaviors were quantified. Overall, these predator species displayed much lower rates of predation on juvenile green sturgeon than alternate prey. Predation decreased with green sturgeon size, and predation risk diminished to zero once sturgeon reached a length threshold of roughly 20–22 cm total length, or between 38% and 58% of predator total length. Behavioral analyses showed low motivation to feed on green sturgeon, with both predators attempting predation less frequently as sturgeon grew. Results of this study imply that optimizing growth rates for larval and juvenile sturgeon would shorten the time in which they are vulnerable to predation. Future experiments should assess predation risk of juvenile green sturgeon by additional predator species common to the Sacramento-San Joaquin watershed.     

Osmo- and ionoregulatory responses of green sturgeon (<Emphasis Type="Italic">Acipenser medirostris</Emphasis>) to salinity acclimation

The green sturgeon is a long-lived, highly migratory species with populations that are currently listed as threatened. Their anadromous life history requires that they make osmo- and ionoregulatory adjustments in order to maintain a consistent internal milieu as they move between fresh-, brackish-, and seawater. We acclimated juvenile green sturgeon (121 ± 10.0 g) to 0 (freshwater; FW), 15 (estuarine; EST), and 24 g/l (SF Bay water; BAY) at 18°C for 2 weeks and measured the physiological and biochemical responses with respect to osmo- and ionoregulatory mechanisms. Plasma osmolality in EST- and BAY-acclimated sturgeon was elevated relative to FW-acclimated sturgeon (P < 0.01), but there was no difference in muscle water content or abundance of stress proteins. Branchial Na⁺, K⁺-ATPase (NKA) activity was also unchanged, but abundance within mitochondrion-rich cells (MRC) was greater in BAY-acclimated sturgeon (P < 0.01). FW-acclimated sturgeon had the greatest NKA abundance when assessed at the level of the entire tissue (P < 0.01), but there were no differences in v-type H⁺ATPase (VHA) activity or abundance between salinities. The Na⁺, K⁺, 2Cl⁻ co-transporter (NKCC) was present in FW-acclimated sturgeon gills, but the overall abundance was lower relative to sturgeon in EST or BAY water (P < 0.01) where this enzyme is crucial to hypoosmoregulation. Branchial caspase 3/7 activity was significantly affected by acclimation salinity (P < 0.05) where the overall trend was for activity to increase with salinity as has been commonly observed in teleosts. Sturgeon of this age/size class were able to survive and acclimate following a salinity transfer with minimal signs of osmotic stress. The presence of the NKCC in FW-acclimated sturgeon may indicate the development of SW-readiness at this age/size.     

Polyphyly of mtDNA lineages in the Russian sturgeon, Acipenser gueldenstaedtii: forensic and evolutionary implications

Molecular species identification methods are an important component of CITES monitoring programs for trade in sturgeon and caviar. To date, obtaining molecular evidence for distinguishing caviar from four closely related Eurasian sturgeon species Acipenser baerii (Siberian sturgeon), A. gueldenstaedtii (osetra), A. persicus (Persian sturgeon), A. naccarii (Italian sturgeon) remains problematic. Using approximately 2.3 kb of mtDNA sequence data (cytochrome b, NADH5, control region), we find this to be attributable to the polyphyletic nature of these mitochondrial DNA markers in the Russian sturgeon, A. gueldenstaedtii. Two mitochondrial lineages are present within this species: one is phylogenetically affiliated with A. persicus and A. naccarii, while the other clusters with A. baerii. These findings have a direct impact on molecular testing of commercial caviar and demonstrate the necessity of using large sample sizes when constructing forensic databases. Furthermore, the results affect current taxonomic designations for these species as well as hypotheses concerning their evolutionary origins.     

Comparative stable isotope analyses of green sturgeon (Acipenser medirostris) and white sturgeon (A. transmontanus) in the San Francisco estuary

Green sturgeon (Acipenser medirostris) and white sturgeon (A. transmontanus) are closely related, sympatric species that inhabit the San Francisco estuary. Green sturgeon have a more marine life history but both species spawn in the Sacramento River and reside for some duration in San Francisco Bay. These sturgeons are of conservation concern, yet little is known about their dietary competition when they overlap in space and time. To examine evidence of dietary differentiation, we collected whole blood and blood plasma from 26 green sturgeon and 35 white sturgeon in San Francisco Bay. Using carbon and nitrogen stable isotope analyses, we compared their relative trophic levels and foraging locations along the freshwater to marine gradient. Sampling blood plasma and whole blood allowed comparison of dietary integration over shorter and longer time scales, respectively. Plasma and whole blood δ¹³C values confirmed green sturgeon had more marine dietary sources than white sturgeon. Plasma δ¹⁵N values revealed white sturgeon fed at lower trophic levels than green sturgeon recently, however, whole blood δ¹⁵N values demonstrated the two species fed at the same trophic level over longer time scales. Larger individuals of both species had higher δ¹³C values than smaller individuals, reflecting more marine food sources in adulthood. Length did not affect δ¹⁵N values of either species. Isotope analyses supported the more marine life history of green than white sturgeon and potentially highlight a temporary trophic differentiation of diet between species during and preceding the overlapping life stage in San Francisco Bay.     

Cell culture and karyotype of Sakhalin sturgeon Acipenser mikadoi

A cell culture of rare and threatened species of Sakhalin sturgeon Acipenser mikadoi was established from a fragment of the pectoral fin and neighboring tissues. Initially, the culture consisted of different cell types including typical fibroblasts as well as cells of epithelial origin, myofibroblasts, etc. After approximately five passages, the culture largely consisted of cells with fibroblast morphology. Under normal culture conditions, these cells grew for more than one year at a constant rate and passed about 80 population doublings. In the absence of serum, cells entered the state of proliferative quiescence (G₀-state). When cultured without medium replacement for a long period of time, cells fused to form myofibers of about 1 cm in length. These myofibers could branch and acquired cross striation with time. About forty days after myofibers emerged, they degenerated, lost their shape, detached from the substrate, and finally died. The induction of adipogenic differentiation arrested cell proliferation and introduced lipophilic inclusions formed in a minor fraction of cells. The number of these inclusions was low, and cells with inclusions demonstrated various morphology distinct from typical adipocytes. The induction of osteogenic differentiation gave rise to cells that produce mineralized extracellular matrix and bone nodules. Chromosome analysis revealed a set of chromosomes typical for “high chromosome” sturgeon species. The variation in the chromosome number was very high (mean, 247 ± 33; modal value, 248). The analysis involving AT- and GC-specific fluorochromes has demonstrated that the telomeric and centromeric regions of all chromosomes are enriched in GC content. The distribution of AT- and GC-rich sequences along the chromosomes was heterogeneous. Long chromosomes were preferentially stained by the AT-specific dye, whereas small chromosomes demonstrated brighter fluorescence after 7-amino-actinomycin D staining; in particular, several small chromosomes fluoresced extremely brightly. This work is the first report of cell culture and karyotype analysis of Sakhalin sturgeon.