Cellular elements of the test plates in the sea urchin Strongylocentrotus intermedius were examined using light and electron microscopes to promote an understanding of their roles in the growth of test plates. Seven types of cells were identified in the interstitial tissue of the test stereom: sclerocyte, spherule, rounded, phagocytic, vacuolated, strongly eosinophilic, and cuboidal undifferentiated. Small basophilic cells accumulated at the sutural areas, i.e., the growth sites of test plates, and were closely associated with the trabeculae. Ultrastructural aspects of the cells indicated apossibility that they participate in the trabecula formation. Therefore, the small basophilic cells should be sclerocytes. Morphological aspects of the spherule, rounded, and phagocytic cells of the test corresponded to those of the spherule, vibratile, and phagocytic coelomocytes, respectively. They are probably migrating coelomocytes. The vacuolated cells were distributed throughout the interstitial tissue and even in test epidermis. The strongly eosinophilic cells were located near the trabecule base and the suture, where they might have some association with collagen fibers. The cuboidal cells appeared to be undifferentiated, progenitor cell. The vacuolated, strongly eosinophilic, and cuboidal undifferentiated cells might originate in situ in the test tissue or may be forms of tansforming coelomocytes.

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Age, morphology and osteology of larval Japanese flounder settling at YanagihamaBeach, Nagasaki Prefecture in 1990 and 1991 were compared among birth months. Age and size at settlement (H stage larvae) decreased as the season advanced; from 25.2 days and 12.0mm SL to 20.4 days and 9.9mm SL in the February and April cohorts, respectively, in 1990. Then from 27.0 days and 11.3mm SL to 22.1 days and 9.5mm in the February and April cohorts, respectively, in 1991. Whereas no differences in morphometry and meristic counts of larvae were seen between monthly cohorts, the degree of vertebral ossification at settlement was greater in the February cohort than in the April cohort, suggesting inferior swimming ability in the latter.

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In order to obtain basic information on the population structure of Japanese seabass Lateolabrax japonicus, four sample lots were collected from coastal waters of Seto Inland Sea (Akashi, Murotsu, and Nandan) and from Japan Sea (Kasumi) in Hyogo Prefecture, Japan. Seventy-one to eighty specimens from each locality were examined for the screening of polymorphism among 12 loci in 10 enzymes. Being fitted for Hardy-Weinberg equilibrium, it was suggested that each lot was derived from simple Mendelian population. The proportion of polymorphic loci ranged between 0.55-0.67 and the expected heterozygosity (H_e) were between 0.172-0.199. Considering these much higher values compared with some other marine fishes, the high genetic variability should be retained in this species. The differentiations among localities were tested by chisquare values for heterogeneity. In MPI-1^* locus, highly significant difference (p<0.001) was detected in the case of Kasumi vs. each locality of Seto Inland Sea. The D values (0.0014-0.0082) were high, especially in the cases related with Kasumi (0.0054-0.0082). These results indicate the independence of the subpopulation ofKasumi from the others. Among the Inland lots, we could not get enough information about the existence of subpopulation except for the cases of significant difference in sIDDH^*.

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The present paper describes the karyotypes in the two species of abalones Nordotis discus and N. gigantea, which are found commonly in the coasts of warmer waters in Japan. Spawning was induced by ultraviolet irradiation stimulus. These larvae were obtained by artificial insemination in the laboratory. Chromosome preparations were made by the chopping method in trochophore larvae of these species. The diploid chromosome numbers were determined as 36 in these species. Thekaryotypes were composed of 10 pairs of metacentric chromosomes and 8 pairs of submetacentric chromosomes in N. discus and in N. gigantea. The fundamental number of both species was determinedas 72. Therefore, it was estimated that the chromosome complements of these species were remarkably similar. This suggests that these two species may be closely related to each other. These resultssupport successful production of the artificial hybrids between N. gigantea and N. discus in the laboratory.

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The trackings of Lake Biwa catfish were executed to examine how migratory they were and what habitat they selected in Lake Biwa in 1993, 1995 and 1996 by ultrasonic biotelemetry. Eight individuals were tracked for a maximum of 118 days in the South Basin, and 11 individuals for a maximum of 319 days in the North Basin. In the South Basin experiments, 7 individuals quickly returned to their caught place, and 5 individuals didn't change their settled positions in the South Basin from July to October. In the Shiozuwann Bay experiments, 9 individuals settled around the shallow floor near their spawning ground in the Shiozuwann Bay throughout the year. It was shown that Lake Biwa catfish had settlement characteristics, less migratory, and mainly inhabited in the shallow area near the spawning ground all the year round.

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Motile enterococci were found in the range of 10³ to 10⁴ cfu/g in the muscle of fishery products on TCBS agar. Six strains of motile enterococci were isolated and classified on thebasis of morphological, biochemical, chemical, physiological, and genetic properties. One yellow-pigmented isolate was identified as Enterococcus casseliflavus and three strains of non-pigmented isolatesas E. gallinarum. Two non-pigmented strains were considered as a new species of the genus Enterococcus. The two strains divided the cells in two planes and had 43 mol% G+C of the DNA.

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Tolerability of the Kuruma prawn Penaeus japonicus exposed to air was investigated at two different relative humidities (R. H.). Under 14°C air conditions, 100% survival times, rates of water loss, respiratory activity, and muscular levels of lactate and ammonia were examined for the prawn previously acclimated to 18°C.
The 100% survival times were 23 and 51h at 73 and 93% R. H., respectively. Cumulative rates of water loss were influenced by R. H. and time; 17.28 and 4.71% at 73 and 93% R. H. after 28 and 70h air exposure, respectively. When stored at 93% R. H., the O₂ consumption of the prawndid not fall in 48h contrary to its fall at 73% R. H. in 16h. At 93% R. H., 60h aerial exposure produced lactate increase, contrary to its slight and not significant increase at 73% R. H. in 16h. For muscular ammonia, a slight but significant increase occurred in 16 and 24h air exposure at 73 and 93% R. H., respectively. Thus high R. H. prolongs the air tolerance of the prawn, proposing a new technical device to its live transportation.

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Five (GT./CA)_n microsatellite loci were isolated from a size-selected genomic library of red sea bream (Paqrus major). Primers for PCR amplification were constructed for the microsatellite loci and the loci characterized by screening polymorphisms in three wild populations of the red sea bream. All loci displayed a high degree of length polymorphism, as observed in the total number of alleles per locus (16-32), and a high degree of heterozygosity ranging from 0.675-0.907. Distinct differences were observed among three wild populations of red sea bream collected from coastal waters of Japan, both in the average number of alleles per locus and the frequency distributions of the alleles.
The primers developed for red sea bream were also tested for their ability to amplify homologous sequences from 5 closely related species of Sparidae.
These microsatellite loci show great potential as indicators for genetic variability and divergence among subpopulations of Pagrus major, and to a lesser degree with some of the related species tested.

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Nocturnal activity of hatchery-reared juvenile flounder was observed in the laboratory to understand their predation risk after release. The number of individuals which were “swimming”, “nonburying” and “solitary” was counted using CCD camera with infrared lamp, and were recorded by time-lapse video-recorder. The number of swimming individuals was higher at night than in the daytime. In a tank without sand, most fish aggregated along the edge of the bottom during the day, whereas at night fish dispersed all over the bottom. In a tank with sand, about 60% of fish emerged from the sand at night, while fish buried into the sand in the daytime. These results suggest that the activity of juvenile flounder might be nocturnal than the diurnal one ever understood, and that released flounder juveniles might suffer comparatively higher predation risk during the night because of their ap pearance on the sand and the off-bottom swimming behavior.

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The magnitude of intraspecific genetic differentiation of red sea bream Pagrus major collected from four areas of western Japan, was estimated using restriction fragment length polymorphism (RFLP) analysis of mitochondrial DNA (mtDNA) D-loop region. Sixty-one composite haplotypes, representing information from 6 restriction endonucleases, were generated for samples of 38-58 fish from each site in 1994, and 72-95 fish from each site in 1995 from Tanabe Bay, Tomogashima Suido, Bingo Nada, and the Japan Sea. Genetic variation was relatively high in all samples: the haplotypic diversity was 0.86-0.90 in 1994 and 0.88-0.92 in 1995, and the averages of the mean nucleotide sequence divergence between and within samples were 0.99 and 0.99%, respectively. On the contrary, the pure mean nucleotide sequence divergence between samples is low, 0.01% in average. Significant heterogeneity was not observed in the distribution of haplotypes among samples both in 1994 and 1995. Furthermore, significant differences were not observed in 23 pairs of 28 combinations of 8 samples. Consequently, differences among samples of red sea bream are not so large basically, although significant geographic differences were observed between Bingo Nada where the genetic structure may be unstable and Tanabe Bay where many hatchery released fishes were caught together with the natural fishes.

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The larval stages of the crystal shrimp Penaeus (Farfantepenaeus) brevirostris were described based on the materials hatched and reared in the laboratory. The larval morphology of P. (F.) brevirostris was compared with four other Penaeus species that occurred off the eastern Pacific coast. The larval development of P. (F.) brevirostris consists of six naupliar, three protozoeal, and three mysis stages before the first postlarva. Differences in body size and setal formulae were recognized among the five species, and diagnostic keys were given for the fourth nauplius, the third protozoea, and all mysis stages. The chronology of nauplius and a non-molt growth of protozoea are discussed withthose of other Penaeus shrimps.

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Deep-sea bacteria were isolated from seawater samples collected from the Japan Trench by repeatedly culturing in liquid and solid media. A modified High-Pressure-Temperature Gradient Instrument (PTG apparatus) was constructed to observe growth responses of the isolated deep-sea bacteria under the combination of various pressure and temperature levels. Growth characteristics of two deep-sea bacterial strains isolated from different depths were studied with this PTG system. An obligate barophilic bacterium, strain JT 761, showed growth in the temperature range between 0 and 14°C and pressure between 30 and 70 MPa, and showed optimum growth, 0.054h^-1, at 60 MPa and 1.8°C. A facultative barophilic bacterium, strain JT 403, showed growth in the temperature range between 0 and 14°C and pressure between 0.1 and 60 MPa, and showed optimum growth, 0.047h^-1, at 20 MPaand 5.1°C.

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This paper examines the hypothesis that spectral cone types exhibit different retinomotor movements in response to monochromatic radiation and can thus be identified histologically in the retina. Red sea bream Pagrus major, 79-105mm in total length, were dark adapted in a glass aquarium, then exposed overhead to various intensities of white light and monochromatic stimuliwith peaks at 337, 368, 466, 551 and 609 nm. The retinae from fish from each stimulus were examinedhistologically and scored for thickness of the cone layer and the pigment layer relative to the total retinal thickness. The red sea bream tested showed retinomotor response to 368 nm but not to 337 nm, and therefore were considered sensitive to ultraviolet stimulus. As additional single cones were absent in the cone mosaic, UV perception by central single cones or twin cones was suggested. White light and three other monochromatic stimuli elicited the retinomotor movement of all single and twin cones. These results and similar data from the literature show that the light-adaptive retinomotor movement of different spectral cone types is triggered by light perception in rods and thus do not vary by wavelength of visual stimulus between 368 and 609 nm.

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This study examined whether the efficiency of resting egg production of the marine rotifer Brachionus plicatilis is improved by employing the semi-continuous culture method. Rotifers were cultured in 0.5m³ polycarbonate tanks for 17-23 days by feeding frozen and concentrated Nannochloropsis oculata. An average harvest of 9.0×10⁴ resting eggs per a gram (dry weight) N. oculata was achieved, which was 2.5 times more efficient than that from batch cultures. Although the water quality was comparatively improved (less concentrations of NH₄-N, NH₃-N and DON) in semicontinuous rotifer cultures, two out of four cultures collapsed on 15-20 th days, probably due to the bacterial effect. The semi-continuous cultures were stabilized by placement of a nylon filter to remove particulate organic materials. With a filter, 1.3 times more resting eggs were harvested on average.

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The growth, maturation and feeding habits of Acanthogobius hasta were studied in Ariake Sound. In June, a new cohort ranging from 16 to 64mm S L appeared. They were thought tohave hatched in the spring of the same year. The cohorts grew slowly throughout summer and autumn. Their growth rate became faster thereafter, with most of them attaining over 200mm SL between January and February. Males grew faster than females and attained sizes as large as 400mm SL. After April, they declined rapidly in number, and disappeared by July. A. hasta is an annual fish. Its bodyform changed with growth and differed between males and females at SL≥ca. 100mm. GSI started toincrease in December, reached a peak in mid-February, and then decreased. Females having postovulatory follicles were first observed in mid-February and accounted for all the catch in mid-April. Females that had spawned did not seem able to spawn again. They spawn only once in life from mid-Februaryto early April. Stomach contents revealed that feeding habits changed with growth and differed between virgins and primiparas.

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A microbiological survey was carried out to clarify the causative organism of spotting disease of sea urchin Strogylocentrotus intermedius, which occurred at Shakotan-Cho Fisheries Breeding Center, Hokkaido at the end of August in 1993. A filiform bacterium which formed brightly yellowish colonies on solid media was isolated from coelomic fluid of diseased animals. Gliding movement on seawater agar plate and 40.5 of DNA GC mol% confirmed that the isolate belongs to the family Cytophagaceae.
Experimental infection of sea urchin with the isolate was perfomed by the immersion method using 10⁶, 10⁵ and 10⁴ CFU/m/ at 20°C, 23°C and 25°C. All the animals challenged with 10⁶ CFU/ml at 23°C and 10⁴ CFU/ml at 25°C died, some of which showing spotting lesions, at one and three days after the challenge, respectively. No mortality was observed at 20°C. The bacterium was reisolated from coelomic fluids and surfaces of dead sea urchins. These results suggested that the bacterium was the pathogen of spotting disease of Strongylocentrotus intermedius.

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The distribution of Vibrio anguillarum serotype J-O-1, a causative agent of vibriosis of ayu Plecoglossus altivelis in Lake Biwa, was investigated by both fluorescence antibody method using specific monoclonal antibody against V. anguillarum J-O-1 (FA-DAPI method) and aconventional cultural method. There were 3.2×10⁴ cells/cm³ of V. anguillarum J-O-1 deposited on the bottom sediment surface (0-0.1cm) under the net cages of an ayu culturing farm in Lake Biwa. In ayu culturing ponds, which are concrete tanks on the lakeshore, we also detected this pathogen(1.6×10⁵ cells/cm³) in the bottom sediments collected from the drains for outflow of pond water intoLake Biwa. In non-culturing areas of the lake, V. anguillarum J-O-1 cells were not found in any water, bottom sediment nor plankton samples. In the same samples from which V. anguillarum J-O-1 cells weredetected by the FA-DAPI method, no cells were recognized by the conventional cultural method. This is the first report for direct detection of V. anguillarum J-O-1 from freshwater environments.

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As a part of the on-going efforts to increase the use of alternative proteins inpractical fish feeds, this study was conducted to investigate whether or not fingerling rainbow trout can grow normally on the same non-fish meal diets that resulted in the green-liver for yellowtail.
Two feeding experiments were carried out with fingerling rainbow trout for 20 weeks. The first experiment was conducted to examine the effect of supplemental crystalline amino acids, with or without coating, in the non-fish meal diets on the growth performance of rainbow trout. In the second, the effect of supplemental levels of crystalline amino acids in the non-fish meal diets was investigated.
Feeding of non-fish meal diets for 20 weeks produced excellent growth and feed performances for rainbow trout without any abnormalities. Supplementation of essential amino acids (EAA) in the non-fish meal diets to balance amino acid profiles comparable to the control fish meal diets resulted in no further enhancement of growth over the control fish meal diet group. The experimental fish could grow well on the non-fish meal diets without EAA supplement. This has shown that the content of EAA in the non-fish meal diets used in this study was sufficient to satisfy the EAA requirement of rainbow trout.

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In order to evaluate seed quality for release projects, body constituents of artificially-produced black sea bream Acanthopagrus schlegeli were monitored after release. The individually marked fish (42mm in total length) were released into a Daiô Bay of Nino-shima Island, Hiroshima, in August 1993. The hatchery-produced fish which had high lipid reserves lost them abruptly within one month after release. Thereafter, the lipid level became lower than that of wild fish at the 100 th day. Intestinal length of hatchery-produced fish was shorter than that of wild fish, but extended to the same length to wild fish by living in the sea. Activity of the hepatic enzyme glucose-6-phosphate dehydrogenase (EC 1. 1. 1. 49) involved in lipogenesis was higher in hatchery-produced fish but reached the same level as that in wild fish at one month after release. Activity of hepatic 6-phosphogluconate dehydrogenase (EC 1. 1. 1. 44) remained constant during the experimental period. The changes in the body constituents could be helpful in evaluating fish quality in releasing projects.

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Common mackerel Scomber japonicus was used for the experiments as an easy-to-disintegrate species. The occurrence of himodori (disintegration) due to the difference of heating rate was investigated in the fish pastes heated at 60°C in a water bath. It was judged from the modified himodori-index that himodori in the center of heat-induced gel occurred more easily than that of thesurface, because of the difference in their heating rates. Also, chemical changes due to himodori were proved by reduction of myosin heavy chain in the heat-induced gel. Thus, it was suggested that the occurrence of himodori could be reduced by rising the heating rate more than 1.0°C/min at least in thecenter of fish pastes as far as the range of temperatures from 50 to 60°C.

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In order to investigate the gel-forming ability of tilapia muscle with freshness-lowering of its raw fish, the heat-induced gels from salt-ground meat of tilapia (i.e., tilapia paste: TP) were evaluated by the K value, jelly strength test, and the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).
When TP prepared from the fresh raw fish (K value=16%) was heated at 80°C for 20 and 120min, its heat-induced gels showed a desirable texture with high elasticity corresponding to that of surimibased products (i.e., kamaboko) on the market. The TP heated at the setting (suwari) temperature (40°C) for short time (20min) could produce a soft elasticity at high fresh conditions of raw fish below 17% of K value. The disintegration or degradation (himodori) could be controlled under the high fresh conditions of raw fish.
Accordingly, it was suggested that the tilapia muscle with the high freshness below 17% of K value could produce an available gel by setting and reducing of himodori.

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To clear the molecular-weight distribution of 6 kinds of agar molecules and the stiffness of the molecule in hot solution with hydrodynamic model, sedimentation velocity measurement was performed.
Distribution of the sedimentation constant of agar molecules showed broad and asymmetrical curves in all samples. According to the theories of Yamakawa et al. for a cylindrical wormlike chain model, the translational friction coefficient and the intrinsic viscosity were approximated. The excluded volume also was estimated by use of the second virial coefficient. From the combination of the persistence length with the excluded volume, agar molecular chain showed the behavior of a typical flexible chain; the chain of higher molecular weight was flexible, and that the lower one was relatively rigid in the hot aqueous solution.
It was predicted that the properties of the commercial agar-gel did not simply depend on the average molecular weight and that the flexibility of the chain had an effect on the gel strength.

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Heating at 40°C: By washing the meats of lizardfish, the gel forming ability wasdepressed, that is, suwari of maeso Saurida undosquamis and wanieso S. wanieso was weakened and modori of tokageeso S. elongata was strengthened. Along with the depression of gel formation, the amount of substances (MHC-A) between myosin heavy chain (MHC) and actin on SDS-PAGE patterns of the gels increased, and that of the larger substances (P) than MHC decreased, accompanied by the increase in the amount of 5% trichroloacetic acid soluble peptides.
Heating at 60°C: By washing the meats, the gel forming ability was improved, that is, modori of three species, especially of tokageeso, was weakened in contrast to the case of heating at 40°C. The degradation of MHC was reduced along with the supression of producing MHC-A and little change in P.
From the above results, it was concluded that the lowering of gel forming ability at 40°C by the washing was related to both the depression of polymerization and the enhancement of degradation of MHC, while the improvement of it at 60°C was related mainly to the depression ofdegradation of MHC.

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Approximately 8% of the total collagen in the mantle muscle of the squid Todarodes pacificus was solubilized by extraction with 50mM Tris-HCl, pH 7.5, containing 4m guanidine hydrochloride (G/HCl). The G/HCl-soluble collagen (GSC) was partially characterized by biochemical and immunochemical techniques for the purpose of obtaining information on the structure of intact collagen in the squid muscle. The GSC showed four a chain-sized components, termed chainse, e, f, g, and h, in SDS-PAGE. The SDS-PAGE pattern was changed by pepsin digestion to show quite a similar pattern to that of Type SQ-II collagen (quantitatively minor collagen) from pepsin-solubilized collagen. The pepsin-digested GSC and the pepsin-solubilized Type SQ-II collagen displayed quite similar peptide maps with lysyl endopeptidase and V-8 protease to each other. These results indicated that the GSC consisted predominantly of intact Type SQ-II collagen. In addition, the chains e and g were identified as intact forms of α1 (SQ-II) and α2 (SQ-II) components, respectively, by peptide mapping and immunoblot analysis of the GSC and its pepsin digests.

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The available phosphorus (P) content in various types of phosphates supplementedin semipurified diets at different levels was determined by the indirect method in common carp and rainbow trout, which have different digestive systems, and was compared with the amount of P extracted by deionized water (Fr. I), 80% acetic acid (Fr. II), and 0.25M hydrochloric acid (Fr. III) by fractionation, in order to simplify the estimation of available P content in fish feed.
Available P content in the experimental carp diets supplemented with various types of phosphates was almost equivalent to the amount of P extracted by deionized water (Fr. I) regardless to forms and levels of phosphates in the diets. The available P content in the rainbow trout diets supplemented with mono-or di-basic phosphate at the level of the P requirement was almost equal tothe amount of P extracted by deionized water alone. The available P content in the diet supplemented with tri-basic calcium phosphate was intermediate between the sum of Frs. I+II and Frs. I+II+III.

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In order to investigate the effects of feed restriction and starvation on the fatty acid synthesis and the oxidation of glucose and alanine to CO₂ in the hepatopancreas of carp Cyprinus carpio, the fish were fed a commercial diet at feeding rates of 100%, 50%, and 0% (starvation) ofsatiation for 30 days. The rate of fatty acid synthesis from D-[U-14C] glucose and L-[U-14C] alanine, theactivities of lipogenic enzymes, and the levels of serum triglyceride and body lipid were decreased by feed restriction and starvation. Moreover, the rate of glucose oxidation together with the glycolytic enzyme activities was significantly low in starved fish. However, the rate of alanine oxidation was scarcely changed, and alanine was more actively oxidized to CO₂ than glucose regardless of feeding rate. Theseresults indicate that the fatty acid synthesis and glycolysis are depressed by feed restriction including starvation and suggest that amino acids are the major energy sources for the hepatopancreas regardless of feeding rate.

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Eel hepatocytes cultured in the presence of 10^-8M thyroxine stimulated phospholipid, cholesterol, and triacylgrycerol syntheses from ¹⁴C-acetate, and the increase in phospholipidand cholesterol contents was induced in the thyroxine treated cells. The lipid efflux from the cells to the culture medium was stimulated by 143% in the thyroxine treated cells. Triacylglycerol and phospholipid contents increased significantly in the medium in which cells were cultured with thyroxine. Furthermore, in the thyroxine treated cells the incorporation of ³H-leucine into the apoproteins of the secreted lipoprotein was specifi cally stimulated by 2-fold compared to that in control cells, but that into other proteins except the secreted lipoprotein was not different between the thyroxine treated and controlcells. The incorpora tion of ¹⁴C-acetate into the secreted lipoprotein was also stimulated in the thyroxine treated cells. We conclude that the primary function of thyroxine is to stimulate lipogenesis, andthe increase in lipids, particularly phospholipid and cholesterol, in thyroxine treated cells seems to promote the lipoprotein synthesis and secretion.

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An unusual salt comprised of three 2-aminoimidazolium cations and halistanol sulfate anion was isolated from a marine sponge Topsentia sp. along with known halistanol trisodium salt. Their structures were determined by spectroscopic methods. The number of 2-aminoimidazoliumions in the molecule was estimated from the signal intensity in the ¹H NMR spectrum, and that of sodium ions was from the atomic absorption spectrometry. This is the first report of a unique salt consisted of 2-aminoimidazolium cations in natural source. Both halistanol sulfate salts showed antimicrobial activities against Flavobacterium marinotypicum and Mortierella ramaniana at 10 μg/disk.

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