We analyzed the polymerase chain reaction (PCR) and the associated random amplification of polymorphic DNA (RAPD) techniques in order to study the Miyazaki and Okinawa populations of the common silver-biddy Gerres oyena (Forsskål). Fourty-six bands were identified for the common silverbiddy using 13 random primers. Nine bands were observed in all of the common silver biddies in this study, and the Okinawa and Miyazaki populations can be distinguished by using the A29-260 marker. In the inter-stock, the mean genetic similarity index was 0.858±0.060 between individual specimens from the Miyazaki population, 0.822±0.085 for the Okinawa population and 0.634±0.086 between them, respectively. In addition, a dendrogram distributed the genotypes (the Okinawa and Miyazaki populations) into different clusters. These results showed that the Okinawa and Miyazaki populations of the common silver-biddy are genetically distinct.

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Genetic variability and differentiation in six breeds including three varieties of goldfish (Wakin, Ryukin, Comet, Oranda-shishigashira, Aka-, Kuro-, Sansyoku-demekin, and Ranchu) were examined by enzyme electrophoresis. Within a breed, average heterozygosity (H_e), estimated from allele frequencies at 26 loci, ranged from 0.046 to 0.100 with a mean of 0.072. Nei's genetic distance (D) between breeds was ranging from 0.0015 to 0.0239 with a mean of 0.0116, indicating that the level of differentiation among breeds falls into the range of local races. The dendrogram constructed from the genetic distance showed that the Wakin, Ryukin, and Comet were clustered together, the three varieties of Demekin and Oranda-shishigashira formed another cluster, and the Ranchu was only distantly related to the other breeds.

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First generation of hexaploid loach Misgurnus anguillicaudatus, which had been produced by inhibiting the second polar body extrusion after fertilization in a natural tetraploid pair, were raised until the adult size. The sex ratio of these hexaploids did not deviate from 1:1, as predicted from the almost equal number of females and males in tetraploids as well as the all-female gynogenetic tetraploids. These observations suggested an involvement of the male heterogametic system in sex determination. Hexaploid females laid significantly larger eggs than tetraploids and diploids. Mature eggs of four mature hexaploid females (three were eu-hexaploid, while one was a hexaploid/tetraploid mosaic) were individually fertilized with spermatozoa of hexaploid (6n), tetraploid (4n), and diploid (2n) males. The progeny were viable and the majority of surviving fish from 6n female×6n male, 6n×4n, and 6n×2n crosses were hexaploid, pentaploid, and tetraploid, respectively. Fertilization of eggs of diploid and tetraploid females with spermatozoa of hexaploids (2n×6n and 4n×6n) gave viable tetraploid and pentaploid progeny, respectively. Thus, hexaploid loach formed triploid gametes (eggs and spermatozoa) and produced viable second generation progeny.

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This paper reports a laboratory target strength (TS) measurement and theoretical modelling of walleye pollock and Pacific hake. The measurements are performed in a tank as a function of fish tilt angle, which is controlled in one-degree steps. A precision echo sounder is used and sphere calibrations are conducted before or after the measurement. A specialized soft X-ray imaging system is used to map the fish swimbladder. The X-ray images are digitized to obtain the fish morphological parameters. Based on these parameters, theoretical TS of the fish is calculated using the approximate deformed cylinder model and the general prolate spheroid model. An advantage of using these models lies in their simplicity and the fact that few parameters are required in the model computation. Calculations indicate a negligible contribution of the fish body to the total scattering in the wide range of tilt angle. The theoretical TS values are compared with the measured values. Agreement between measured and theoretical TS is reasonably good for near normal incidence, but deteriorates for larger tilt angles. The level of agreement is discussed. Obtained maximum and average TS values are nearly equal to or a little bit smaller than the published data. Using the deformed cylinder model, broad band TS characteristics are generated. Our results highlight the usefulness of the simple method to predict fish TS using the deformed cylinder model associated with X-ray imaging.

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Different life stages of ayu Plecoglossus allivelis, including downstream migrating yolk-sac larvae, larvae and juveniles distributed in the estuary or coastal waters, and upstream migrating juveniles, were collected in the Shimanto River and adjacent coastal waters from October 1996 to June 1997. The objective of this study was to examine mortality process of larval and juvenile ayu by comparing the seasonal change in number of newly hatched larvae with the birth date distributions of the later life stages. For yolk-sac larvae during the downstream migration, two-peaks in abundance were found in mid-Novem-ber and between late December and mid-January. In the later life stages, November hatched larvae and juveniles were few, while larvae and juveniles hatched between late December and early January were more abundant. This indicates a high mortality of the larvae hatched in mid-November 1996. From October to November 1996, water temperatures in the Shimanto Estuary and adjacent coastal waters were over 20°C, in which the survival rate of larval ayu was considerably low. We propose that the unusually high water temperature of the sea in autumn 1996 caused the high mortality of larval ayu hatched in November.

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Two length frequency data sets of the big eye, Priacanthus macracanthus, from the adjacent waters of Guei-Shan Island during the periods of 1981-1982 and 1997 were analyzed with MULTIFAN to estimate parameters of the von Bertalanffy growth equation. Age structures and growth parameters of the two periods were compared. Parameters of the von Bertalanffy growth equation for 1981-1982 data set were estimated as: asymptotic fork length L∞=42.5cm, growth coefficient K=0.151/yr and age at zero length t₀=-0.790yr for females and L∞=43.8cm, K=0.122/yr and t₀=-1.458yr for males. Five age classes were separated and dominant age classes were 3⁺, 4⁺, 2⁺ for both sexes. Parameters of the von Bertalanffy growth equation for 1997 data set were estimated as: L∞=46.8cm, K=0.118/yr and t₀=-1.255yr. Six age classes were separated and dominant age classes were 3⁺, 4⁺. Significant difference on growth for the big eye was found between the two periods.

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Population status of Sphyrna lewini in the northwestern Pacific was estimated by demographic analyses using the best available life history parameters. Demographic tables were constructed using estimates of natural mortality (M) of 0.558/year for age 0 and 0.279/year for ages 1-15 with maximum age of 15. A fixed fecundity was set as 12.9 female embryos per litter and an age-specific fecundity was from N=(-26.105+0.179L)/2. Females mature at age 5 and reproduce every two years. The population increase rate was estimated to be 22.8%/year, and the generation time was 7.57 years without exploitation. The net reproductive value, generation time and intrinsic rate of natural increase decreased with the increase of fishing mortality. At F=0.3 and fishing started at age 5, the population increase rate was 12%/year, the generation time was 6.26 years and population doubled every 6.1 years. Sphyrna lewini population had strong resilience as long as fishing started at an age older than age at maturity but would decline when intensive fishing started at an age younger than age at maturity.

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Histological characteristics and the distribution pattern of the visual cells, single cones, twin cones, and rods in the retina of the eye of wild adult Japanese sardine Sardinops melanostictus were investigated. The developmental process of the visual cells was also examined in the reared and wild larvae. In the specialized part (area temporalis) of the adult retina, slender twin cones were regularly distributed in a square mosaic pattern with central single cones. In the unspecialized part, twin cones with stout ellipsoids were regularly distributed and many rods were recognizable. A newly-hatched larva had developing lens and undifferentiated retinal cells. The retinal differentiation proceeded rapidly, with the pigment epithelial cells and visual cells already formed in a 31.5 hour-old larva. In a 79.5 hour-old larva at the first-feeding stage, the visual cell layer consisted of only single cones. A so-called pure-cone retina was still recognized in a larva of 18.6mm standard length (SL). In a 20.9mm SL larva, rod-like cells with thin ellipsoids and outer segments appeared and twin cones also observed. In larvae greater than about 20mm SL, the rods and twin cones rapidly increased in number. The retina of a 35.6mm SL juvenile basically did not differ from the adult one.

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Growth of the bivalve-feeder Halla okudai (Polychaeta: Lysaretidae) was evaluated in relation to species and abundance of prey under wild and rearing conditions. The field survey was conducted from May to July 1995 at the M. L. W. S. in 11 intertidal sandy flats of Hiroshima Bay, the Seto Inland Sea. H. okudai habitats were classified into four types by major prey organisms: clam, clam & oyster, oyster, and mussel. The condition factor was highest in the clam & oyster-type habitat with the highest density of Ruditapes philippinarum. The condition factor also increased with increasing the density of R. philippinarum, whereas this was not seen in Crassostrea gigas and Mytilus galloprovinciallis. In laboratory experiment, the growth efficiency was significantly higher in the R. philippinarum-group than in the M. galloprovinciallis- and the C. gigas-groups. Thus, H. okudai is suggested to select prey to maximize the growth.

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Three microsatellite loci, Cca-17^*, Cca-21^*, and Cca-30^*, were isolated from a partial genome library of common carp Cyprinus carpio. Primers for PCR amplification of template DNA were designed based on the unique sequences flanking each motif. The total number of alleles detected from nishikigoi and wild type common carp population were five, six and nine at loci Cca-17^*, Cca-21^*, and Cca-30^*, respectively. Inheritance was shown to be Mendelian by analysis of genotype ratio in F1 offspring of six families of nishikigoi. Allele ^*69 at locus Cca-21^* and allele ^*266 at locus Cca-30^* were detected only in wild type common carp with frequencies of 0.304 and 0.458, respectively, while allele ^*288 was observed only in the nishikigoi with the frequencies ranged from 0.600-0.610.

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The protective value of antibody of maternal origin was examined in larvae of red sea bream. Eggs and larvae from the mother fish, immunized with Vibrio vaccine, were incubated for about 1 month, and the immunoglobulin (IgM) amount and antibody titre were monitored. IgM and antibody against Vibrio were recognized in the eggs. However the levels acutely diminished within a few days after hatching. On day 20 to 25, the IgM concentrations but not antibody titres increased again, indicating the beginning of their synthesis. These results imply the maternal antibody has no protective function in the larvae.

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An ultrasonographic technique for non-invasive sexing of barfin flounder Verasper moseri was established based on sexual dimorphism of gonadal appearence. We used here a high frequency transducer (10MHz) attaching to ultrasonography to improve echoic images. Although gonads of 23-month-old males were impossible to discern, those of females were visible as bright oval contours in the ultrasonographic images. Accurate identification of sex was accomplished in 63 fish ranged from 204 to 274mm in total length of 14-month-old fish, 288 to 401mm of 19-month-old fish and 252 to 370mm of 23-month-old fish, except for three false diagnosis. On the other hand, it was difficult to determine sex in 11-month-old fish, 98-135mm in total length, by this technique. The present results indicate the practical usefulness of ultrasonography for a rapid, non-invasive and realtime sexing of immature flounders.

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Effects of two harmful dinoflagellate blooms, Gymnodinium mikimotoi and Heterocapsa circularisquama on the clearance rate of the blue mussel Mytilus galloprovincialis and the pacific oyster Crassostrea gigas were studied during both red tide periods occurred in Hiroshima Bay, Seto Inland Sea, Japan, 1995. The mean clearance rate of mussel and oyster decreased to 19.8% and 14.4% of control in the G. mikimotoi red tide, respectively. In the oyster, red tide filtrate of G. mikimotoi slightly inhibited the filtration activity. The mean clearance rate of the mussel and the oyster decreased to the 8.8% and 1.5% of the control seawater in the H. circularisquama red tide, respectively. However, filtration rates of both mussels and oysters were not inhibited in the filtrate of H. circularisquama red tides. Both bivalves exposed to red tide exhibited various negative responses such as retraction of their mantles and gradual valve closure. The present study demonstrated that red tides of G. mikimotoi and H. circularisquama strongly inhibited the filtration rate of bivalves.

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Aeropyrum pernix, a strictly aerobic marine hyperthermophilic archaeon, has been studied as a potential biotechnological source. In order to clarify the phylogenetic position of A. pernix, the nearly complete nucleotide sequence of the 23S rRNA was determined and compared with the other hitherto known archaeal counterparts. To avoid the artifact caused by disparities of G+C content, transversion evolutionary distance analysis was adopted. The result confirmed that A. pernix is closely allied to the Pyrodictium-Desulfurococcus-Staphylothermus group (the order “Igneococcales”) within the kingdom Crenarchaeota. However, it also revealed that the genus Aeropyrum diverges sharply from the other genera within the family Desulfurococcaceae. Thus, we concluded that the decision to transfer the genus Aeropyrum from the family Desulfurococcaceae is reasonable in light of our present data, as well as the physiological characteristics of this archaeon reported previously.

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To find the adequate feeding condition in ayu Plecoglossus altivelis culture, fish averaging 5.1g in body weight were reared on different feeding regimes. Group I and III were satiated two and four times daily, respectively. Group II was fed a diet equivalent to 60% of the diet fed to group III with four daily feedings. The effects were assessed by growth, anatomical measurements, and biochemical parameters.
Growth and feed conversion efficiency were not significantly different among the three groups. While stomach volume was not significantly different, the intestine was longer in the fish fed four times daily with restricted diets (group II). The serum glucose level increased with feeding frequency. Hepatic enzymes, glucose-6-phosphatase and alanine aminoacyl transaminase, which are associated with glycol-ysis and amino acid turnover, were higher in group II.
Following the feeding experiment, the fish were kept without feeding for 23 days. Mortality and body weight loss were lower in the group reared on four daily feeding with restricted diets. Feeding four times daily with restricted rations resulted in high growth performance and normal physiological conditions.

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Identification of the causative bacteria Vibrio spp., which were isolated from diseased sea urchin Strongylocentrotus intermedius and their rearing cages at low seawater temperatures at Date, Shiriuchi and Shikabe Fisheries Breeding Centers in Hokkaido was undertaken based on their biochemical properties, DNA-DNA homology and serological analysis. All fifteen strains, five from each Breeding Center were almost identical in biochemical properties except indole production. However, the strains isolated at Date differed from the strains at Shiriuchi and Shikabe in DNA-DNA homology and analysis of thermostable and thermolabile antigenic compositions. These results indicated that the strains from Shiriuchi and Shikabe belonged to the same species but they were different from those of Date. Moreover, the causative Vibrio spp. differed in biochemical, genotypical and serological properties from any strains of Vibrio spp. isolated from the intestine of healthy sea urchin and any of type strains of the genus Vibrio.

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Prooxidant activities of fish skin were assayed for 31 marine and fresh water species. Prooxidant activity was measured by increase in absorption at 234 nm due to the conjugated diene produced during incubation with linoleic acid or eicosapentaenoic acid. Prooxidant activity for linoleic acid was present in all of the fish skin extracts tested, and ranged from 1.24 units/mg extractable protein in the John dory Zeus japonicus to 9.16 units/ mg extractable protein in the yellow tail Seliora quinqueradiata. The activities of extracts from fishes of the Order Clupeiformes tended to be high and labile to heating. Since specific lipoxygenase inhibitors, oleic acid and eicosatetraynoic acid, as well as hemoprotein inhibitors, potassium cyanide and o-phenanthroline, inhibited the prooxidant activity of some skin extracts, it is likely that the hemoprotein and lipoxygenase-like enzyme may contribute to the prooxidant activity in fish skins. Catechin mixtures prepared from tea also effectively prevented the prooxidant activity in fish skin extracts as well as catechin and epigallocatechin. The use of catechins is advantageous from a viewpoint of preventing both enzymatic and non-enzymatic lipid peroxidation in fish skins.

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Lipases (triacylglycerol acylhydrolase, EC 3. 1. 1. 3) were purified from Aspergillus repens and Eurotium herbariorum NU-2 strain by using a DEAE-Sephadex A-50 column and preparative electrophoresis. The purified enzymes from A. repens and NU-2 had molecular weights estimated by SDS-PAGE to be 38, 000 and 65, 000, respectively. Lipase from A. repens had a pH optimum of 5.3 and a temperature optimum of 27°C, while for the NU-2 strain corresponding values were pH 5.2 and 37°C. The specific activity of NU-2 was about twice that of A. repens. Substrate specificity toward olive oil or triolein and positional specificity for hydrolyzing the 1 (3)-position ester bonds of triacylglycerol are discussed for both enzymes.

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As a series of studies to clarify the relationship between serum lipoprotein profile and nutritional physiological metabolism of fat in fish body, the difference of serum lipoprotein profile between cultured and wild red sea bream Pagrus major was examined by disc electrophoresis and density gradient ultracentrifugation. The low density lipoprotein (LDL) level was higher in cultured red sea bream than in wild red sea bream. In the high density lipoprotein (HDL), though the HDL₃ level was similar between both groups, the HDL₂ level was remarkably lower in cultured fish than in wild fish. Both the levels of thickness coefficient and fat in internals within the limits of macroscopic observation were significantly higher in cultured fish than in wild fish, and the levels of triglyceride and cholesterol contents in serum also exhibited a higher tendency in cultured fish. From these findings, it was suggested that in cultured fish, the nutritional physiological condition of fat in fish body might be unbalanced toward fat accumulation. Furthermore, although the function of HDL₂ in relation to fat carrier in serum of fish could not be clarified, it was considered that the HDL₂ might be not specially concerned with fat accumulation.

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The ATP/IMP ratio of stressed fish muscle in summer (from June to August) was lower at the initial stage than the ratio in winter (from January to March), and reached to the ultimate value 3h earlier. Though stressed fish muscle in summer showed larger contraction than that in winter, almost the same tension values were observed in both seasons. This result was ascribed to the structure weakening due to large contraction in summer.
The ATP /IMP ratio of unstressed fish muscle in summer was lower at the initial stage than the ratio in winter, and reached to the ultimate value 4h earlier. The initial ATP /IMP ratios of spinal cord destroyed fish muscles at two seasons were both high values. The ratio in summer was slightly lower at the initial stage than the ratio in winter, and reached to the ultimate value 10h earlier. Unstressed and spinal cord destroyed fish muscles in summer contracted more than those in winter, and showed larger tension values. This result means that the structure weakening was not remarkably induced in the muscles obtained in summer. In spinal cord destroyed fish muscles the slow but continuous tension development induced the structure weakening. Therefore the tension increment in summer was not as large as in unstressed fish muscle.

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As part of study on the relationship between muscle fiber types and rigor mortis progress in dorsal ordinary muscle of fish, the histochemical discrimination of muscle fiber types in dorsal ordinary muscle was examined by the acid and alkaline stabilities of actomyosin ATPase activity, and a comparison was carried out among various fishes and parts. The optimum combination of pH and time for acid preincubation was remarkably different within the ranges from pH 4.48 to pH 4.54 and from 0.5min to 4.0min among ten sample fishes, while that at alkaline preincubation was almost similar at pH 10.40 although the time slightly differed. In any case, the muscle fiber types in dorsal ordinary muscle of various fishes were discriminated by the acid and alkaline stabilities of actomyosin ATPase activity. The four kinds of muscle type were identified in ten sample fishes and these muscle types were only the IIa like subtype of white muscle in one fish, the IIa and IIb like subtypes of white muscle in two fishes, the pink muscle and IIa or IIb like subtype of white muscle in two fishes, and the pink muscle and IIa and IIb like subtypes of white muscle in six other fishes, respectively. Furthermore, the muscle fiber types in dorsal ordinary muscle were remarkably different between wild and cultured fish, among specimens of the same fish, and among parts along the body length and at varying depths.

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Inhibitory potencies against α-glucosidase activities were compared among bromophenols obtained from extracts of three Rhodomelaceae red algae, Symphyocladia latiuscula, Odonthalia corymbifera, and Polysiphonia morrowii. The bromophenols from these species are characterized by a number of Br atoms per benzene ring: S. latiuscula, three Br atoms (1 and 2); O. corymbifera, two Br atoms (3-7); and P. morrowii, one Br atom (8 and 9). The bromophenols exhibited mixed inhibition against yeast α-glucosidase reaction. In particular, symmetric dibenzyl ethers, bis(2, 3, 6-tribromo-4, 5-dihydrox-ybenzyl) ether (2) and bis(2, 3-dibromo-4, 5-dihydroxybenzyl) ether (5), exhibited about 10³ fold smaller K_i values than the other bromophenols. Their inhibition modes are disclosed to be mixed inhibition closed to competitive inhibition. Inhibitory potencies of the bromophenols increased with the increasing degree of bromo-substitution per benzene ring and the decreasing degree of methyl-substitution. In contrast to the strong inhibitory activity against yeast α-glucosidase activity, the bromophenols moderately inhibited rat-intestinal sucrase and maltase activity. The inhibitory potencies were all comparative among the bromophenols examined.

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To make clear the polymerization behavior of myosin heavy chain (HC) through SS bonding upon heating, SDS-PAGE and the measurement of total SH groups (TSH), reactive SH groups (RSH) and surface hydrophobicity were performed on those actomyosin (AM, 0.5% protein) from carp, flying fish and rabbit which were heated for 1h at temperatures ranging from 10 to 80°C.
Results were similar in common among three kinds of AM.
In the presence of EDTA (1mM), surface hydrophobicity and RSH content enormously increased during the heating at the temperature between 30 and 40°C followed by the gradual increase with the rise in temperature up to 80°C without the decrease in TSH.
In the presence of Cu²⁺ (6μM), TSH content decreased obviously with the rise in temperature above 30°C and RSH decreased above 40-50°C, although surface hydrophobicity behaved as in the presence of EDTA. HC dimer through SS bonding was observed up to 30°C, though its amount was unchanged without decrease in TSH upon heating. Above 30°C, the higher molecules than HC dimer such as trimer, tetramer and so on were found and the polymerization of HC was promoted with the rise in temperature. Actin was also polymerized above 30°C.
These results suggest that up to 30°C the dimer of HC was formed through the oxidation of SH groups without the unfolding of the protein and above 30°C HC polymer was formed along with the oxidation of SH groups which were emerged with the unfolding. In addition, it was suggested that actin is oxidized to high polymer above 30°C as well as HC.

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To elucidate the role of S-1 and rod in the polymerization of myosin heavy chain through SS bonding upon heating of actomyosin, each myofibril of carp (with 100μg/ml leupeptin), flying fish and rabbit was digested by ∝-chymotrypsin to S-1 and rod followed by heating for 1h at temperatures ranging from 10 to 80°C with added Cu²⁺ 12μM for carp and 100μM for other samples to enhance the oxidation of SH groups. SDS-PAGE and the measurement of SH content and surface hydrophobicity were performed. Behaviors of S-1 and rod from 3 species in common were as follows.
With the rising of temperature above 30°C, polymer at the top of 3% gel through SS bonding gradually increased with the concomitant decrease in S-1 and actin with Cu²⁺, while less polymer was found without Cu²⁺. This polymer was confirmed to be composed of S-1 and less amount of actin by two-step SDS-PAGE. Rod dimer through SS bonding was found almost at any heating temperatures except at 50°C where the degradation of rod occurred strongly.
Surface hydrophobicity increased clearly from 30°C, followed by gradual increase above 50°C. This behavior coincided very well with the increasing behavior of RSH upon heating without Cu²⁺. Oxidation of TSH was promoted with added Cu²⁺ slightly up to 30°C and greatly above 30°C, while RSH did so only a little below 30°C, and then greatly above 40°C in flying fish and 50°C in rabbit.
It was, therefore, concluded that the polymer formation of HC upon heating of actomyosin was mainly due to the oxidation of SH groups in S-1, while the dimer formation of HC was due to the oxidation of SH groups in rod.

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