Determination of hormones inducing oocyte maturation in <Emphasis Type="Italic">Chalcalburnus tarichi</Emphasis> (Pallas, 1811)

Chalcalburnus tarichi is an endemic cyprinid species living in the Lake Van basin, in eastern Anatolia, Turkey. The present study was undertaken to determine which hormones induce oocyte maturation in C. tarichi. The levels of 17α,20β,21-trihydroxyprogesterone (20β-S), progesterone (P), 17α-hydroxyprogesterone (17α-HOP), 11-deoxycortisol (11-DOC), and 17α-hydroxy-20β-dihydroprogesterone (17,20β-P) were measured in fish caught from Lake Van and the Karasu River, and injected with human chorionic hormone (hCG) (1,000 and 1,500 IU/kg). Oocytes of fish caught from the lake were also incubated in vitro with different doses (50, 200, and 1,000 ng/ml) of 20β-S, 17α-HOP, 11-DOC, and 17,20β-P. 11-DOC was found to be the most effective hormone among those measured for inducing oocyte maturation in vivo and in vitro. 17,20β-P could not be determined in the plasma of any fish in vivo (P < 0.05). 1,000 IU/kg dose of hCG given by injection caused a statistically significant increase in all plasma hormone levels (P < 0.05). It was found that there was a significant decrease in the P level only at 1,500 IU/kg dose of hCG injected (P < 0.05), while the level of other hormones increased at this dose (P < 0.05). It was also determined that all the hormones were effective in germinal vesicle breakdown (GVBD) in in vitro oocyte culture (P < 0.05). However, 11-DOC was found to be the most effective hormone in GVBD at a dose of 200 ng/ml (70% GVBD). In conclusion, 11-DOC synthesized during final oocyte maturation in C. tarichi was found to be a potent inducer of GVBD, which shows that 11-DOC may be described as an oocyte maturation steroid in this species.     

Induction of diploid gynogenesis in turbot <Emphasis Type="Italic">Scophthalmus maximus</Emphasis> with left-eyed flounder <Emphasis Type="Italic">Paralichthys olivaceus</Emphasis> sperm

Turbot Scophthalmus maximus exhibits sexually dimorphic growth, with females growing faster and reaching larger adult sizes than males. Thus, development of techniques for preferentially producing females is necessary to optimize production of these species. In this paper, gynogenetic diploids of turbot were induced by activating egg development with ultraviolet (UV)-irradiated left-eyed flounder Paralichthys olivaceus sperm combined with cold shock to prevent extrusion of the second polar body. The results of UV irradiation experiments showed that survival, motility, and duration of activity of P. olivaceus sperm generally decreased with increase in UV dose. The typical Hertwig’s effect was observed after fertilized turbot eggs with UV-irradiated P. olivaceus sperm and the optimal UV dose for gynogenetic haploid production was 36,000 erg mm⁻². At 15°C, appropriate timing of cold shock for retention of the second polar body in turbot eggs was at 6 min after fertilization. Results of different combinations of two shock temperatures (1 or 3°C) and four shock durations (15, 25, 35 or 45 min) at 6 min after fertilization demonstrated that shock of 25 min at 1°C gave the highest production of diploid gynogens (39.58% relative to its diploid control). The results of this study reveal that the use of UV-irradiated P. olivaceus sperm for activation of turbot eggs and cold shock for polar body retention is an effective method to produce gynogenetic offspring.     

Masculinization of Convict Cichlid (<Emphasis Type="Italic">Cichlasoma nigrofasciatum</Emphasis>) by immersion in <Emphasis Type="Italic">Tribulus terrestris</Emphasis> extract

We have examined the effects of Tribulus terrestris (TT), a non-toxic herb, on sex reversal in the Convict Cichlid Cichlasoma nigrofasciatum with the aim of introducing a new environmentally friendly method for masculinization in C. nigrofasciatum. TT is a natural plant product that elevates the testosterone levels in humans and animals. Different concentrations (0.0, 0.10, 0.20 and 0.30 g/l) of TT extract were tested for their effect on sex reversal in C. nigrofasciatum by immersing newly hatched offspring once weekly for 2 months in TT extract. Of the dosages used in the present study, 0.30 g/l TT was the most effective in terms of masculinization, resulting in a maximum male ratio of 87.23% (P < 0.001). Sex ratios of 79 and 85% at 0.10 and 0.20 g/l TT, respectively, were also significantly different from the expected 1:1 ratio (P < 0.001). Histological examinations revealed that the testes of fish treated with TT extract contained all stages of spermatogenesis, clearly demonstrating that the administration of TT extract to C. nigrofasciatum stimulated spermatogenesis. Total survival rates in all treatments and the control were uniformly high, ranging from 88.57 to 90% (P > 0.05). We concluded that TT had no negative effect on the survival rate of C. nigrofasciatum. In addition, all groups of TT-treated fish exhibited successful growth acceleration compared to the control group, but only the 0.30 g/l TT treatment significantly improved the growth rate of C. nigrofasciatum. (P < 0.01). Sex reversal in C. nigrofasciatum demonstrated that TT-treated 0-day-old larvae showed successful sex reversal, spermatogenesis and a better growth rate than untreated progenies.     

A non-lethal method for establishing sexual maturation in mussels (<Emphasis Type="Italic">Mytilus</Emphasis><Emphasis Type="Italic">galloprovincialis</Emphasis> (Lamarck, 1819)) during broodstock conditioning in hatcheries

The concentration of the sex steroids 17β-estradiol, testosterone and progesterone was determined by specific enzyme-linked immunosorbent assays in the hemolymph of the mussel Mytilus galloprovincialis and related to its sexual maturation. A non-lethal hemolymph sampling procedure was used in mussels matured in controlled conditions of a hatchery. To our knowledge, this is the first study on sexual steroid hormones of a bivalve species using the same specimens in consecutive samplings. Histological data showed that nearly 90 % of the mussels ripened during the experiment. Steroid analyses revealed that 17β-estradiol and testosterone were closely related to the onset of gamete development, while progesterone would have an important role at the end of gonadal maturity. This method can be used to assess sexual maturation in bivalves when conditioning in hatcheries, without killing them.     

Serum steroid concentrations and development of reproductive organs during puberty in male bonnethead sharks, Sphyrna tiburo

Puberty is a critical, hormone-mediated event during which an animal acquires the ability to breed and propagate. Despite the importance of this stage in animal reproduction, little is known regarding the physiological factors that regulate and/or accompany puberty in several vertebrate groups including elasmobranchs. To address the need for such information, the present study investigated morphological and hormonal changes that occur during puberty in male bonnethead sharks (Sphyrna tiburo). Serial changes in development of claspers, paired copulatory organs in male elasmobranchs, and serum steroid concentrations during puberty were evaluated in captive-held male S. tiburo. Captive-animal studies were supplemented by observations on gonadal development, gonaduct morphology, and serum steroid concentrations in feral, peripubertal male S. tiburo. Changes in size and histological architecture of testes and gonaducts of peripubertal sharks mirrored the seasonal progression of events that occur in these structures in mature males. Claspers grew in length continuously during puberty, but sharks did not reach functional maturity until a short period before mating activity commences in the mature population. Clasper growth appeared to be strictly regulated in S. tiburo, perhaps to ensure growth of these organs to sizes deemed critical for reproductive success. Serum concentrations of testosterone, dihydrotestosterone, progesterone, and 17β-estradiol increased in both captive and feral sharks during pubertal development, and may be associated with development of the gonads and gonaducts. Differences in hormone profiles of captive and feral sharks were observed at certain periods during puberty, but their origin remains unclear. This revised version was published online in July 2006 with corrections to the Cover Date.     

Comparison between α-LNA and DHA in early developmental stages of <Emphasis Type="Italic">Takifugu obscurus</Emphasis> and <Emphasis Type="Italic">Takifugu rubripes</Emphasis>

ABSTRACT:   Rearing experiments were conducted to investigate the essential fatty acid requirements in the early developmental stages of river puffer Takifugu obscurus and tiger puffer T. rubripes using two n-3 series unsaturated fatty acids, α-linolenic acid (18:3n-3, α-LNA) and docosahexaenoic acid (22:6n-3, DHA), under salinity of 30 and 18.5–20.3°C. River and tiger puffer larvae used in this study were 15 and 14 days old after hatching, and their average body weights were 30.1 and 20.8 mg, respectively. The results on fatty acid requirements of these two species were evaluated from fish growth, survival, fatty acid composition of the fish body and activity test results. The DHA groups of both river and tiger puffer exhibited better survival and weight gain. However, there was no difference in the mean final body weights of river puffer between two dietary groups. Also, the DHA group of tiger puffer showed better results in the recovery test from anesthetic condition than that obtained in the LNA group. In an examination of the fatty acid compositions of the whole body, the LNA group containing no dietary DHA resulted in 0.5% DHA in tiger puffer and 1.1% DHA in river puffer . These results suggest that α-LNA from Artemia converted to eicosapentaenoic acid (20:5n-3, EPA) and to DHA successively by their fatty acid metabolism. Symptoms following essential fatty acid deficiency were not observed in any experimental groups. As river puffer did not represent a significant difference in the dietary effects between α-LNA and DHA treatment groups, its essential fatty acid requirement was assumed to be somewhat closer to that of the freshwater fishes in comparison with that of marine fishes, including tiger puffer.     

The physiological tolerance of the grey carpet shark (<Emphasis Type="Italic">Chiloscyllium punctatum</Emphasis>) and the epaulette shark (<Emphasis Type="Italic">Hemiscyllium ocellatum</Emphasis>) to anoxic exposure at three seasonal temperatures

The epaulette shark (Hemiscyllium ocellatum) and the grey carpet shark (Chiloscyllium punctatum) are commonly found in periodically hypoxic environments. The ecophysiological time available for these animals to safely exploit these niches during different seasonal temperatures was examined. The time to loss of righting reflex (T _LRR) was examined in response to an open ended anoxic challenge at three seasonal temperatures (23, 25 and 27°C). Ventilation rates were measured in an open ended anoxic challenge at 23°C and during 1.5 h of anoxia followed by 2 h of re-oxygenation at 23 and 25°C. The mean T _LRR of epaulette and grey carpet sharks was inversely proportional to temperature. The T _LRR was similar between species at 23°C; however, grey carpet sharks had significantly reduced T _LRR at higher temperatures. During the standardised anoxic challenge, epaulette sharks entered into ventilatory depression significantly earlier at 25°C. During re-oxygenation, epaulette sharks exposed to anoxia at 23°C had no significant increase in ventilation rates. However, after anoxic challenge and re-oxygenation at 25°C, epaulette sharks showed a significant increase in ventilation rates during re-oxygenation. Grey carpet sharks displayed no evidence of ventilatory depression during anoxia. However, during re-oxygenation, grey carpet sharks had significantly elevated ventilation rates above pre-experimental levels and control animals. These data demonstrate that the anoxia tolerance times of both species were temperature dependent, with a significant reduction in the T _LRR occurring at higher temperatures. Epaulette sharks had a significantly greater T _LRR at higher temperatures than grey carpet sharks, which did not enter into a ventilatory depression.     

Substrate specificity of brain acetylcholinesterase and its sensitivity to carbamate insecticides in <Emphasis Type="Italic">Carassius auratus</Emphasis>

The substrate specificity of brain acetylcholinesterase (AChE) in adult Carassius auratus fish and its sensitivity to carbamate insecticides were investigated in vitro. The results showed that the order of four substrates hydrolyzed by brain AChE in C. auratus was acetylthiocholine iodide > β-methylthiocholine iodide > propionylthiocholine iodide > butyrylthiocholine iodide, and the maximum velocity (V _max) of AChE hydrolyzing acetylthiocholine iodide (ATCh) was the highest among the four substrates, and the V _max values were 0.067 and 0.082 mmol min⁻¹ mg⁻¹ for male and female fish respectively. But their Michaelis–Menten constants (K _m) were the lowest, only 0.071 and 0.072 mmol/l respectively. Compared with other carbamate insecticides, the sensitivity of brain AChE to carbofuran was the highest and the IC₅₀ values were 1.04 × 10⁻⁶ mol/l for females and 1.17 × 10⁻⁶ mol/l for males. The inhibitory tendencies of eserine, methomyl, and aldicarb to brain AChE were very similar, and the percentage inhibition increased with time at the concentration of 1 × 10⁻⁶ mol/l. The order of inhibition potential of the three inhibitors from the highest to the lowest was eserine, aldicarb, and methomyl.     

Suspension feeding and growth of ark shell <Emphasis Type="Italic">Anadara granosa</Emphasis>: comparison with ubiquitous species <Emphasis Type="Italic">Scapharca subcrenata</Emphasis>

The ark shell Anadara granosa is a species peculiar to the Ariake Sound. To determine why distribution of A. granosa in Japan is largely confined to this area, we examined feeding and growth of A. granosa as functions of environmental and biological variables. The results were compared with those of another ark shell Scapharca subcrenata, which is ubiquitous in Japan. Feeding experiments indicated that A. granosa is eurythermal and euryhaline, as is S. subcrenata, but is adapted to temperature slightly higher than S. subcrenata. Weight-specific clearance rate (CR) of A. granosa as a function of soft-body dry-weight (w) followed the power function of w (CR=2.7×w ^−0.37), with coefficient and exponent very close to those for S. subcrenata. Growth rate of A. granosa increased linearly with daily ration, similar to S. subcrenata. Thus, feeding and growth characteristics of A. granosa were comparable to those of S. subcrenata and no ‘peculiarities’ of the former were detected. Therefore, the factors that make A. granosa a species restricted to the sound are probably not directly related to feeding or growth characteristics.     

Purification and partial characterization of vitellogenin from shorthead redhorse (<Emphasis Type="Italic">Moxostoma macrolepidotum</Emphasis>) and copper redhorse (<Emphasis Type="Italic">Moxostoma hubbsi</Emphasis>) and detection in plasma and mucus with a heterologous antibody

Vitellogenin (VTG), the egg yolk precursor protein, was purified from plasma of estradiol-3-benzoate (E2B)-treated male shorthead redhorse (Moxostoma macrolepidotum) and immature copper redhorse (Moxostoma hubbsi) by a two-step chromatographic procedure without precipitation. Intact VTGs appeared as dimers with apparent molecular masses, determined by gel filtration, of ∼425 kDa (copper redhorse) and ∼450 kDa (shorthead redhorse). In native polyacrylamide gel electrophoresis (PAGE), dimeric redhorse VTGs appeared as a 520 kDa band. Both VTGs were reduced to a single monomer of ∼150 kDa in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) under reducing and nonreducing conditions, indicating that monomers are not linked by disulfide bonds in the dimer form. The purified proteins were characterized as phospholipoglycoproteins. Isoelectric focusing of both VTGs revealed components with isoelectric points ranging from 5.3 to 6.0, suggesting charge heterogeneity. The amino acid composition of both VTGs contains a high proportion of nonpolar amino acids and was similar to those of other teleosts. An antibody developed against carp (Cyprinus carpio) VTG showed cross-reactivity with VTG from both redhorse species. Using this antibody, VTG was detected in plasma and surface mucus of E2B-treated redhorse. This is the most extensive report on purification and characterization of vitellogenin from catostomidid species.     

Survey of genetic variation at three microsatellite loci in captive populations of endangered Japanese minnow <Emphasis Type="Italic">Aphyocypris chinensis</Emphasis> with implications for reduction of inbreeding

The Japanese population of the cyprinid minnow Aphyocypris chinensis is nearing extinction in the wild. The genetic diversity of three microsatellite loci in five captive populations was investigated, and an effective breeding strategy to reduce inbreeding from pairwise relatedness (R _xy ) between each captive line is discussed. The average number of alleles ranged 2.33–4.67 and the average heterozygosity ranged 0.283–0.602. The pairwise relatedness observed in most combinations showed a significant decrease between the populations. It is suggested that exchange of individuals between different breeding lines should effectively stop inbreeding. Studies show that the effective population size (N _e ) estimated from the number of parental individuals was 8.54 in one captive population, which is insufficient to maintain genetic diversity. It is recommended that more parental individuals should be used, and to exchange fish in a rotating mating mode between institutions participating in captive breeding of A. chinensis.     

Age and growth of threeline grunt <Emphasis Type="Italic">Parapristipoma trilineatum</Emphasis> along the south-western coast of Kii Peninsula,Japan

Age and growth were determined for threeline grunt Parapristipoma trilineatum based on otolith readings of 1043 specimens collected along the south-western coast of Kii Peninsula, Japan. Observations of the otolith margin, together with comparisons of ring mark counts with age in artificially reared specimens, verified that such marks were produced once a year. The formation period corresponded approximately to the main spawning season (May–June). Both surface and cross-section methods were used for age determination; the surface method often resulted in underestimations of age for fish older than 2 years. Discrepancies between the two methods became greater with fish growth. The von Bertalanffy growth curve, based on ages inferred from the cross-section method, was FL _t=331×{1−exp[−0.283×(t+1.45)]}. No substantial difference was detected between male and female growth parameters. The oldest maximum ages inferred from the cross-section method were 15 and 21 years for females and males, respectively. It seems likely that P. trilineatum grows more slowly and lives longer than previously thought, and the difference is attributed primarily to differences in aging methods used in the present and previous studies.     

Physiological responses of over-wintering common carp (<Emphasis Type="Italic">Cyprinus carpio</Emphasis>) to disturbance by Eurasian otter (<Emphasis Type="Italic">Lutra lutra</Emphasis>)

Using a tame animal, the impact of otter (Lutra lutra) disturbance on over-wintering carp (Cyprinus carpio) was monitored in two experiments, 133 and 140 days, respectively, over two consecutive winters (November–April). The level of stress in over-wintering carp exposed to various intensities of disturbance by otters was quantified using biological indicators of stress (cortisol, cortisone, indices of nitrogen, carbohydrate, lipid and mineral metabolism and activity of basic blood plasma enzymes) taken from blood plasma of stocked carp at the end of the winter seasons (when the photoperiod was 12 light:12 dark, respectively, 13L:10D). Moreover, condition (Fulton’s coefficient of condition and fat content in muscles) and mortality rate of that carp were measured after over-wintering and also after the subsequent vegetation period. The analysis of blood and tissue samples of experimental fish showed changes in nitrogen, carbohydrate and mineral metabolism as well as levels of hormones and fat reserves. Higher response to stress in metabolism of carp with lower intensity of disturbance by otter suggests that high level of disturbance can lead to metabolic adaptation of carp to stress. The effect of stress on the mortality rate of carp during the over-wintering is not clear. Nevertheless, the negative effect of stress on survival, condition and growth rate of carp in the subsequent vegetation period was not observed.     

Effects of fish CYP inducers on difloxacin <Emphasis Type="Italic">N</Emphasis>-demethylation in kidney cell of Chinese idle (<Emphasis Type="Italic">Ctenopharyngodon idellus</Emphasis>)

A drug–drug interaction occurs when the effect of one drug is altered by the presence of another drug which is generally associated with the induction of cytochrome P450s (CYPs) activity. Thus, unexpected treatment failures often happen resulting from inappropriate coadministration in fisheries. However, little information is available about CYP induction in fish. The reaction of difloxacin (DIF) biotransformation to sarafloxacin (SAR) belongs to N-demethylation catalyzed mainly by CYP(s). In order to supply useful information on CYP induction, the present study assessed the effects of fish-specific CYP inducers on DIF N-demethylation and enzyme kinetics in kidney cell of Chinese idle (CIK; grass carp (Ctenopharyngodon idellus)) by RP-HPLC. Results demonstrated that the amounts of SAR formation and enzymatic parameters Clint and Vmax were significantly increased due to β-naphthoflavone (BNF) pretreatment. Therefore, we suggest that CYP1A may be involved in DIF N-demethylation in CIK. This study provides instructive information to ensure treatment success via avoiding CYP induction in fisheries.     

Tench (<Emphasis Type="Italic">Tinca tinca</Emphasis> L.) larvae rearing under controlled conditions: density and basic supply of <Emphasis Type="Italic">Artemia</Emphasis> nauplii as the sole food

After artificial reproduction of tench, larvae must be maintained indoors, and studies on rearing conditions are needed, focussing on the reduction of labour and costs. Three experiments on larvae (5th day post-hatch) were conducted for 25 days using Artemia nauplii as the sole food in order to determine basic feeding and density conditions during the first rearing period. Tench were maintained in 25 l fibreglass tanks, supplied with an artesian water flow throughout of 0.2 l min⁻¹. Water temperature was 22.5 ± 1°C, and the photoperiod was natural. Larvae fed on a restricted amount of nauplii reached high survival rates, even with the minimum of 50 nauplii larva⁻¹ day⁻¹. This amount of food may be sufficient at least for the first 25 days of exogenous feeding if fast growth is not the priority, and high densities can be maintained with good survival rates (over 90% up to 160 larvae l⁻¹ and 77% with 320 larvae l⁻¹). When food was supplied in excess once a day, high survival rates were achieved (91–97%), without differences among the densities tested. Animals at a density of 100 l⁻¹ reached the highest length (15.57 mm) and individual weight (46.8 mg). This growth is greater than those reported in studies feeding several times a day. It could be deduced that, while live food remains available for tench, it is not necessary to feed so frequently. Considering the relationship among the initial number of animals, final survival and growth and ration supplied, the new data reported here are useful to establish suitable stocking densities under both culture and experimental conditions.     

Annual variation of potential predation impacts on larval and juvenile marbled sole <Emphasis Type="Italic">Pseudopleuronectes yokohamae</Emphasis> by sand shrimp <Emphasis Type="Italic">Crangon uritai</Emphasis> in Hakodate Bay,Hokkaido

An investigation was conducted to evaluate the annual variation in potential predation impact (PPI) on larval and juvenile Pseudopleuronectes yokohamae by Crangon uritai in Hakodate Bay using predator-prey size relationships. Laboratory experiments were designed to estimate the favorable prey size of C. uritai through back-calculation of body length (BL) of P. yokohamae from sagittal otolith diameter observed in the stomachs of C. uritai. The most favorable prey-predator size ratio (BL of P. yokohamae-total length (TL) of C. uritai) class was 0.15–0.19, and ranged from 0.12–0.31. There was a significant positive correlation between the BL increase of P. yokohamae and the bottom water temperature in the field, such that BLs stagnated during the cold year of 1999 from April to June, and increased during the warm year of 2002. In contrast, no significant correlation was found between the TL increase of C. uritai and the bottom water temperature. Moreover, there were no significant differences in the mean TL of C. uritai between months (April–June) or years (1998–2002). Variation of PPI depended on the relationships between the growth rates of P. yokohamae and bottom water temperature. Therefore, the warm year of 2002 resulted in higher potential predation impact (PPI≥20), and it was at least 20 days shorter than that of the cold year of 1999. These results suggest that increased bottom water temperature in the nursery area was one of the most important factors for cumulative predation loss.     

Effects of triploidy on the Caspian salmon <Emphasis Type="Italic">Salmo trutta caspius</Emphasis> haematology

The aim of this study was a comparison of key haematological features of diploid (2n) and triploid (3n) Caspian salmon (Salmo trutta caspius). Morphometric indices of erythrocytes were determined on blood smears by light microscopy. Triploidy significantly (P < 0.001) increased all morphometric indices measured in the erythrocytes including cell size, cell surface area, and cell volume. The increase in cell size was larger for the major (27%) axis than for the minor (22%) axis, thus making erythrocytes of 3n Caspian salmon more ellipsoidal. The estimated increase in erythrocyte nuclear volume (87%) was bigger than the theoretical expected 50% increase. Haematological indices were measured manually by hemocytometry. Triploids had lower numbers of red blood cells (RBC: 1,120,000 cells/mL in 2n vs. 700,000 cells/mL in 3n; P < 0.001) but they were larger in size (mean erythrocytic volume [MEV]: 363.1 nm³ in 2n vs. 483.3 nm³ in 3n; P < 0.001). The decrease in RBC number was not compensated by the increase in MEV and, thus, triploidy affected the haematocrit (Hct: 38.8% in 2n vs. 33.06% in 3n; P < 0.05). Total blood hemoglobin concentration was lower in triploid fish (Hb: 9.9 g/dL in 2n vs. 8.9 g/dL in 3n; P < 0.05). In contrast, mean erythrocytic hemoglobin (MEH: 95 μg in 2n vs. 133.2 μg in 3n; P < 0.001) was higher for 3n Caspian salmon as a result of their larger erythrocytes, although MEH concentration (MEHC: 0.26 g/dL in 2n vs. 0.27 g/dL in 3n) did not significantly differ (P > 0.05). White blood cell (WBC) counts (lymphocytes and neutrophiles) were measured and WBC/RBC ratios were calculated. There were no significant differences in WBC (15,710 cells/mL in 2n vs. 12,683 cells/mL in 3n; P > 0.05), lymphocytes, and neutrophils as %WBC as well as WBC/RBC ratios between two ploidy levels (P > 0.05). Triploid Caspian salmon showed higher erythrocyte abnormalities such as ‘twisted’, ‘tailed’, and ‘anucleated’ cells as well as high portions of immature RBC in blood smears in comparison with diploids (P < 0.001).     

Occurrence of toxic <Emphasis Type="Italic">Dinophysis acuminata</Emphasis> (Dinophyceae) in Uranouchi Inlet,Japan

In the Uranouchi Inlet, which has a commercial fishery of littleneck clam Ruditapes philippinarum, potentially toxigenic species of Dinophysis acuminata showed an annual occurrence except for the summer of 2000–2002. Relatively high abundances of D. acuminata were observed in April 2000 and November 2002. Some samples of D. acuminata cells, which were collected from the inlet in autumn 2002, contained pectenotoxin-2 (0.9–2.8 pg/cell) and okadaic acid (1.9–2.8 pg/cell). Cell length and hypotheca width of D. acuminata found in the Uranouchi Inlet were 33.7–45.0 and 21.0–31.6 μm, respectively. The length-width ratios ranged 1.21–1.71. Sequences of the 5.8S ribosomal RNA gene and flanking internal transcribed spacers, 1 and 2 of cells of D. acuminata from the inlet fell into a D. acuminata complex clade that included, D. acuminata-D. sacculus-D. acuta reported by others. In the inlet, occurrences of D. acuminata were detected within a broad water temperature range between 13.0 and 26.9°C, and those showing more than 25 cells/L were within 16.4–17.3°C for temperature and 27.6 and 34.1 for salinity. This is the first report that cells of Dinophysis acuminata from southern Japanese coastal waters contained okadaic acid and pectenotoxin-2.     

Parentage assignment of stocked black sea bream <Emphasis Type="Italic">Acanthopagrus schlegelii</Emphasis> in Hiroshima Bay using microsatellite DNA markers

The genetic contribution of 51 broodstock, comprising 29 females and 22 males, reared at Hiroshima City Marine Products Promotion Center for the production of stocked black sea bream was monitored during two consecutive years using seven microsatellite DNA loci. The high discrimination ability of these markers was reflected in the polymorphic identification content (PIC=0.831), the exclusion probability (Q≈1), and the low probability of identity index (I=3.635⁻¹⁰). The total number of breeders contributing to the mating process was estimated at 32 (62.7%) in 2000 and 30 (58.8%) in 2001. On pedigree reconstruction, 69.3% of the offspring were successfully assigned to a single broodstock pair. Loss of alleles accounted for 16.9% during seed production; nevertheless, 90.9% of males and 69.0% of females participated in the mating process. Based on microsatellite genetic tagging, 58.9% of the fish sampled during the two months after release were identified as hatchery stock, presenting no significant differences from wild conspecifics in either fork length or body weight.     

Economic analysis of tiger grouper <Emphasis Type="Italic">Epinephelus fuscoguttatus</Emphasis> and humpback grouper <Emphasis Type="Italic">Cromileptes altivelis</Emphasis> commercial cage culture in Indonesia

This study presents an economic analysis of tiger and humpback grouper at different production scales in Indonesia. The results highlight the non-viability of small-scale tiger grouper farming, with a 5-year projected negative cumulative cash flow of −IDR 18,102,650.00 and a negative net present value (NPV) of −IDR 22,059,576.28. An increased production scale of tiger grouper highlights a marginal viability for medium-scale farms (with a 5-year projected cumulative cash flow of IDR 198,320,673.00, a positive NPV of IDR 105,578,440.42; a benefit cost ratio of 1.25; an internal rate of return (IRR) of 88% and a payback period of 0.99 years), and an economically viable large-scale cage culture (with a 5-year projected cumulative cash of IDR 707,746,923.00; a NPV of IDR 406,801,749.07; a benefit cost ratio of 1.33; an internal rate of return of 157%; and a payback period of 0.57 years). The economic analysis of humpback grouper at different production scales highlighted a positive cumulative cash and NPV, a benefit cost ratio over 2, an internal rate of return over 300% and a payback period <1 year. A sensitivity analysis revealed that increased survival rate up to 80% would increase cumulative cash and NPV of small-scale tiger grouper cage culture. Additionally, improved profitability performance was associated with decreasing major production costs, increasing production and price of the product.