We attempted to improve the breed of red sea bream Pagrus major by means of selective breeding in order to establish red sea bream seedling for aquaculture capable of growing more quickly than natural red sea bream. In about 1964, natural young fish were reared into broodstock and, among young fish obtained from these broodstock, those growing quickly were selected and reared into broodstock. This selective breeding has been repeatedly carried out for more than 25 years. The following results were obtained. (1) As the selective breeding was repeated, the body weight of broodstock aged 4 years showed an increase. (2) The selected red sea bream seedling grew at an obviously elevated rate as the selective breeding was repeated. Namely, the time (days) required for attaining a commercially available fish size (about 1kg) was shortened. (3) The average realized heritability, which was determined by the average body weight of 4-year-old broodstock and body weight of 4-year-old fish in the growth curve of each generation, was 0.33±0.28.

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The growth of the Hatatatenumeri-dragonet Repomucenus valenciennei was studied based on 13, 287 specimens collected by three kinds of trawl survey in Tokyo Bay during the period from June 1990 to September 1993. Growth and the life span were estimated by analyzing length frequency data. Year classes of 1990 and 1991 were used to estimate the parameters of the growthcurves. The growth of the dragonet showed seasonal oscillation and was best expressed by modified logistic rmulae, in which the growth rate changes with a cycle in a year. The life span was estimated to be two years for both sexes, reaching 94mm for females and 110mm for males in mean standard length.

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Morphological changes of unfertilized and fertilized eggs of the Japanese sardine Sardinops melanostictus were observed continuously. Unfertilized sardine eggs could be divided into three types on the basis of their morphology: a distorted type, a narrow perivitelline-spacetype, and a normal perivitelline-space type. The first type of eggs disintegrated and sank to the bottom of the containers in a short time. The latter two types of eggs formed not only a perivitelline space but also a blastodisc. They disintegrated gradually within 12 hours after stripping. Common to eggs of these three types, the egg membrane finally broke and the egg's contents were mainly lost. The examination of the field-collected eggs showed that some wild eggs were very similar in their morphological characteristics to unfertilized and disintegrated sardine eggs mentioned above. These facts suggest that unfertilized sardine eggs may be commonly present in natural spawning.

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The distribution of pelagic elasmobranchs was analyzed using bycatch data from the high seas salmon surveys by research gillnets from 1981 to 1991 in the North Pacific Ocean and the Bering Sea. Five species of sharks and one stingray were reported. Blue and salmon sharks and spiny dogfish were dominant and constituted 98% of the total shark catch. It is considered that the research area includes the southern part of the distribution for spiny dogfish and salmon sharks and the northern part of that for blue, shortfin mako, cookiecutter sharks and pelagic stingrays. These elasmobranchs showed spatial segregation by species in distribution according to oceanographic conditions. Intraspecific sexual and growth dependent segregations were also confirmed. Length frequency distributions of blue andsalmon sharks suggest that their nursery grounds exist around the transitionaldomain of the subarctic boundary.

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On the basis of the clear distinction between fish schools from the two subpopulations of skipjack tuna Katsuwonus pelamis in the western Pacific Ocean reported already, an attempt was made to clarify the ecological characteristics of fish schools of the central-west Pacific subpopulation. Comparisons of body length frequency distribution as well as the relationship between body length and weight between fish schools of the western Pacific subpopulation and those of the central-west Pacific subpopulation, which both migrated to the east coast region of themainland of Japan, showed the following: 1) marked differences were observed between the two subpopulations on the modal sizes of fish schools sampled in the latter half of the fishing season, August through November, 2) analyses on the seasonal progression of body length frequency distribution strongly suggested the probable occurrence of three spawning seasons a year in the western Pacific subpopulation, but it is necessary to examine where the anticipated fish schools of the central-west Pacific subpopulation originating from one of the three spawning seasons migrate, and 3) fish of the central-west Pacific subpopulation were slightly fatter than those of the western Pacific subpopulation on average.

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Flow-cytometric assays were made for DNA contents of erythrocytic and other somatic cells in the progeny of natural tetraploids (4 n) of loach Misgurnus anguillicaudatus. When relative DNA contents of normal diploid loach are expressed as 2 C, the progeny produced from various fertilizations of gametes from diploids and tetraploids as well as chromosome manipulations such as duplication of maternalchromosomes and gynogenesis showed DNA contents which were theoretically expected.
Absolute DNA contents measured on the basis of chicken erythrocytic DNA for diploid, triploid, tetraploid, pentaploid and hexaploid loach were 2.53, 3.82, 5.10, 6.37, and 7.63 pg/nucleus, respectively. Therefore, there is a proportional relationship between the DNA contents and the ploidy status in the loach.
The relative DNA contents of the progeny of triploid loach confirmed that the triploid females laid both large triploid eggs and small haploid eggs. Those of the progeny of pentaploid loach demonstrated that pentaploid female laid only diploid eggs.
Active formation of 1 C (haploid), 2 C (diploid), and 3 C (triploid) spermatozoa was detected in the testes of diploid, tetraploid, and hexaploid males, respectively. In contrast, no evidence of maturation division of spermatogenesis was flow-cytometrically obtained in the triploid males examined. Pentaploid males gave a small peak at 2.3 C DNA contents, suggesting the formation of aneuploid spermatozoa.

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A bloom of a colony-forming diatom of the genus Thalassiosira suddenly occurred in almost all areas of the eastern Seto Inland Sea during the period from late fall to winter of 1994-1995. In order to obtain basic information on the bloom and for species identification, we examined the morphological characteristics of individual cells and colonies by light and electron microscopes. We also investigated its distributions and the relationship between the bloom and environmental factors. Since the observations by microscope revealed that the morphological characteristics of the species agreed with previous descriptions, we identified the species as Thalassiosira diporocyclus. In the eastern Seto Inland Sea, this bloom was first found around the southern Harima-nada in late October of 1994, andit spread widely throughout the eastern Seto Inland Sea during November and December 1994. The density of colonies began to decline in early January of 1995, and was hardly observed in any of the areas in early February. The high abundanceof this species caused low concentrations of nutrients and nori production was economically damaged. This is the first record on the bloom of T. diporocyclus inthe eastern Seto Inland Sea. Careful monitoring of this species is necessary toprevent recurrent damage to fisheries in this area.

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The larval development of Penaeus occidentalis was described and illustrated based on laboratoryreared samples. Morphology of the larvae was also compared withthat of other Penaeus species in the eastern Pacific coast. The larvae of P. occidentalis passed 6 naupliar, 3 protozoeal, and 3 mysis stages before the first postlarva. The comparison reveals some mophological differences in segmentation, setal arrangement, body size, and body formation. The chronological pattern in the naupliar stage of P. occidentalis was the same as that of the other species.

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We examined the possibility that growth increments in otolith of southern bluefin tuna Thunnus maccoyii are deposited daily and investigated the growth in the juvenile stage of this species. Among samples collected from off the west and southwest coast of Australia between 1990 and 1993, otoliths of 122 fish from 247 to 820mm FL were observed under SEM. The number of increments of these otolithsranged from 78 to 877. The following findings support the daily deposition of increments: 1) The features of increments in the nucleus region were the same as those of larvae which were verified as daily increments, 2) The back-calculated date of first increment deposition corresponded well with the spawning period, and 3) Growth of tag-recapture fish agreed with that from otolith examination assuming daily deposition. Growth of this species in the juvenile stage was linear and 0.761mm/day. Estimated sizes were 508mm FL at age 1 and 786mm FL at age 2, which was much larger than previously estimated.

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Age and growth of the pleuronectid fish Kareius bicoloratus in Tokyo Bay were estimated from left sagittal otoliths of 717 fish. Examination of the outer margins of the otoliths showed that the translucent zone was formed once a year, can be used as an annulus and its formation completed at the beginning of their spawning season (December). Growth of the fish was expressed by von Bertalanffy's equation as L_t=197.0{1-exp[-1.890(t-0.305)]} for males and L_t=307.8{1-exp[-1.075(t-0.325)]} for females, where L_t is standard length in mm and t is age in years. It was found that the growth of females and males differs, with the females showing a higher growth than the males at each estimated age. Growth in body weight was also examined. Most of the fish examined were found tobe of ages 1 and 2, although the oldest fish were 3 years old for males and 5 years old for females.Their growth and age composition in Tokyo Bay could be subjected to area-specific effects.

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Diet and food selectivity of Calliurichthys japonicus and Repomucenus huguenini were investigated from March 1992 to February 1993 in Tosa Bay, Kochi Prefecture. The diet of C. japonicus was composed of polychaetes, amphipods, gastropods, ophiuroids, bivalves, cumaceans, and other small benthic animals. Amphipods were abundant in number both in spring and winter months, while polychaetes and ophiuroids were the most dominant in autumn and summer, respectively. Calliurichthys japonicus actively selected gastropods and showed a more randomized feeding character than R. huguenini. In R. huguenini, polychaetes and amphipods were the most dominant prey items throughout the year. Polychaetes were most abundant in number from spring to autumn, while the amphipods dominated in winter. Although R. huguenini had a similar diet to that of C. japonicus, R. huguenini had a more selective and specialized diet than the C. japonicus, and actively selected decapods, isopods, ostracods, and cumaceans. The difference in feeding habits between the two species may be attributed to their difference in adult body-size and their ability to visually differentiate prey at their different depth distributions.

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To determine the distribution of neuropeptide Y (NPY)-like immunoreactivity in the brain (cephalic ganglion) of the land crab Chiromantes haematocheir, an immunohistochemical study was carried out using the streptavidin-biotin technique, with special regard to a possible relationship with the molluscan cardioexcitatory tetra-peptide (FMRFamide), oxytocin, and urotensins I and II. NPY-like immunoreactivity was found to occur widely in the crab brain, including several cell groups in the protocerebrum and deutocerebrum, while NPY-positive fibers were seen in many portions of the neuropiles. In most of the NPY-positive cells, FMRFamide-like immunoreactivity was demonstrated, suggesting colocalization of both peptides. Although an immunopositive oxytocin-like substance was detected sporadically in the crab brain, no urotensins were detected. These results suggested that NPY-like peptide, in conjunction with FMRFamide-like peptide in the brain of C. haematocheir, functions as a neurotransmitter or neuromodulator during the process of integration.

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The nutritional contribution of Pseudomonas sp., a predominant bacteria in Artemia-rice bran culture, in the diet of Artemia was determined. Artemia nauplii hatched out aseptically were fed with autoclaved rice bran, heat-dried (HD) Pseudomonas sp. or its combination for 9 days under axenic laboratory conditions. Amino acids analysis by the HPLC method was performed forboth diets. A higher essential amino acid content was obtained in HD Pseudomonas compared to rice bran. Artemia fed with rice bran and HD Pseudomonas resulted in better survival and significant growth than the Artemia fed with rice bran only. Some resident bacteria, in particular Pseudomonas, growing in Artemia culture seem to constitute a major source of protein and amino acids needed for thegrowth and survival of Artemia.

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Age and growth of the starspotted dogfish Mustelus manazo in Tokyo Bay were examined from centrum analysis using soft x-radiography. Based upon seasonal centrum edge analysis, a dark ring was formed annually, mostly in June. The von Bertalanffy growth equations were described as follows, male; L_t=1241[1-exp{-0.120(t+2.59)}], female; L_t=1341[1-exp{-0.113(t+2.55)}], where L_t is total length in mm and t is age in years. The maximum age was 8 years old for males and 10 years old for females. The growth until 2 years was similar in both sexes, but over 2 years females tended to grow larger than males. Sex ratios were approximately 1:1 until age class 4; after age class 5 the proportion of males was reduced.

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Five antioxidative substances in the brown alga Eisenia bicyclis were isolated by silicic acid column chromatography and thin-layer chromatography. They were identified to be phloroglucinol and its polymers, namely eckol, phlorofucofuroeckol A, dieckol, and 8, 8'-bieckol on the basis of spectroscopic evidence (IR, FABMS, and ¹H and ¹³C NMR). Their potent antioxidant effect extended the induction time of autoxidation of methyl α-linolenate.

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Myoglobins from yellowfin tuna, bonito and yellowtail exhibited two distinct endothermic peaks reflecting multiple states of structural unfolding, whereas those of sheep and sperm whale myoglobins followed a two-state process. This indicated that the thermal unfolding of fish myoglobins is predominantly a multi-state process differing from that of the mammalian ones. Changes in dependence on temperature of an α-helix content and a tryptophan fluorescence intensity were found to be decreased more for fish myoglobins compared with those of the mammalian myoglobins. Consequently, fish myoglobins showed a lower recovery of the α-helix and tertiary structure than that of the mammalian proteins. These results are thermally attributed to low conformational stability of the fish proteins. Therefore, fish myoglobins exhibit a rather labile structural folding, suggesting a greater susceptibility to heat denaturation than that of the mammalian myoglobins.

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Apomyoglobins from both fish (yellowfin tuna, bonito, and yellowtail) and mammals (sheep and sperm whale) exhibited a single endothermic peak, reflecting a two-state processof thermal unfolding. A native structure of fish apomyoglobins had smaller α-helix content and showedless thermal stability of the α-helix structure than that of the mammalian ones. The conformational stability of a tertiary fold of the fish apoproteins observed from a tryptophan fluorescence intensity and the fluorescence intensity of ANS-apomyoglobin complex was also found to be lower than that of the mammalian apoproteins. These results suggest that even though the thermal unfolding process is predominantly similar, the fish apoproteins particularly show structural conformity with less compact and less stable globin than that of the mammalian apoproteins. The results were also compared with those of their holomyoglobins.

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Fresh samples of milkfish were stored in ice for up to two weeks or frozen at -20°C for up to two months. At due time intervals, the dorsal part of ordinary muscle excised was subjected to washing by various media (water, 40mM phosphate buffers of pH 5.6-8.0, alkaline wash), andthe effects of washing media on the extracting efficiency of pigment (myoglobin) and the tristimulus color values (L, a and b) of the extract and the meat paste (washed mince) were investigated. The results obtained showed that the pigment extracting efficiency decreased during ice or frozen storage. On theother hand, as the pH values of washing media were raised, the L values of both the extract and washedmince decreased, while a and b values became higher for the extract and tended to be lower for the washed mince.

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Juvenile yellowtail Seriola quinqueradiata were fed five isonitrogenous and isoenergetic diets which contained increasing levels of carbohydrate (3-24%) and decreasing levels of lipid (16-8%) and a high protein control diet for 30 days. Growth, body composition, and hepatic enzyme activities were determined to clarify the metabolic response to dietary carbohydrate and lipid. Growth, feed conversion, and protein-sparing effect were lowest in fish fed the highest-carbohydrate lowest-lipid diet, and these parameters tended to increase with decreasing dietary carbohydrate to lipid ratios. In fish fed high lipid diets, the activities of hepatic glucose-6-phosphate dehydrogenase, phosphogluconate dehydrogenase, malic enzyme, glucose-6-phosphatase, and alanine aminotransferase together with glycogen content were markedly lower, while hepatic fat and plasma free fatty acid concentrationswere higher than in those fed the control diet. These findings suggest that dietary lipid depressed hepatic lipogenesis, gluconeogenesis, and amino acid degradation while it was effectively utilized asan energy source. These metabolic responses seem to explain the yellowtail's high capability to utilize dietary lipid and its high protein-sparing effect.

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The enzyme sensor for ATPase activity consisted of an immobilized membrane of two enzymes, purine nucleoside phosphorylase and xanthine oxidase, and an oxygen electrode.
The optimum conditions for using the sensor were as follows; temperature, 32°C; pH, 7.0; flow rate of buffer, 0.4ml/ min; flow rate of substrate, 0.6ml/min; inosine concentration, 0.2mg/ml; ATP concentration, 0.04mg/ml; and sample volume; 40 μl: ATPase activity in fish serum measured by the proposed sensor system was in good agreement with activity determined by a conventional colorimetric method. Correlation coefficient was 0.986. One assay could be completed within 3min. Continuous determination was possible for 80min (16 assays). The mean current output was 0.28 μA and standard deviation was ±0.25%.

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Fouling macroalgae cause serious problems byfi settling on ships' hulls, power plant cooling systems, aquaculture systems, and other marine infrastructure. In order to solve this problem, it is necessary to find antifouling substances without toxicity or with lower toxicity than that of toxic agents such as TBTO [bis-(n-tributyltin)-oxide]. For this purpose, an assay for antifouling substances is important. First, the effects of environmental factors on attachment and germination of spores of the fouling macroalga Ulva conglobata were examined. The optimum conditions for attachment and germination of spores were: temperature, 20_??_25°C; salinity, 33; light intensity, 3000 lx underlong-day regimen. Under such conditions, a new and efficient assay method for antifouling substances against the attachment of fouling macroalgae was developed. Antifouling activity was statistically examined.

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Transglutaminase (TGase, EC 2. 3. 2. 13) from walleye pollack Theragra Chalcogramma liver was purified to electrophoretical homogeneity by Q-Sepharose and S-Sepharose chromatographies. The purified enzyme of 0.34mg was obtained from 15g of liver tissue and 591-fold purification was achieved from the liver extract. The molecular weight was estimated to be 77 kDa by SDS-polyacrylamide gel electrophoresis. The optimum pH and temperature for monodansyl cadaverine incorporation to N, N'-dimethylated casein were 9.0 and 50°C, respectively. The purifiedenzyme required Ca²⁺ above 3mM for the maximum activity, and Sr²⁺ also fully activated the enzyme. The activity was inhibited by sulfhydryl reagent, suggesting this enzyme was a thiol enzyme, the same as mammalian TGases. By this purified TGase, the gelation of myosin B solution was catalyzed, possibly through the polymerization of myosin heavy chains.

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The effect of pepsin digestion on each type of collagen from the mantle muscle, mantle skin, and arm muscle of the squid Todarodes pacificus was examined by biochemical and immunochemical techniques. An α chain-sized component, termed chain c, in the SDS-PAGE pattern of the residue after alkali extraction (RS-AL) was reactive for the antiseum against the α 1 (SQ-I) component which was one of the subunits of the major collagen, Type SQ-I. The band pattern of chain c in immunoblot analysis was not changed by pepsin digestion. These results indicate that chain c is an intact form of the α1 (SQI) component with relatively small telopeptides. On the other hand, another α chain-sized component, named chain a, in the RS-AL was reactive for the antiserum against the α 1 (SQ-II) component which was one of the subunits of the minor collagen, Type SQ-II. Chain a had a slower mobility on SDS-PAGE than the α1 (SQ-II) component. Only the α1 (SQ-II) component was reactive for the anti-α 1 (SQ-II) component serum in the pepsin-solubilized collagen, indicating that the α 1 (SQ-II) component was produced from chain a by pepsin digestion. Additionally, chain a and the α 1 (SQ-II) component showed quite similar peptide maps of V-8 protease digests to each other. These results indicated that chain a was an intact form of the α 1 (SQ-II) component with relatively large telopeptides.

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The water-soluble proteins (WSP) of otoliths were electrophoretically and immunochemically studied in the tilapia Oreochromis niloticus. WSP were separated into three or more undefined bands by gradient native PAGE and into 14-16 bands with a main band at 52 kDa by gradient SDS-PAGE. The main band was PAS-positive and had a moderate calcium-binding capacity. Proteinsextracted from the sacculus showed an intense calcium-binding capacity in a single band in the low molecular region. Immunoelectrophoresis and immunoblotting revealed that anti-WSP rabbit serum waspositive to the WSP, serum, and extracts from various tissues, including the sacculus, semicircular canal, heart, liver, gill lamella, and brain. These immunoreactions, except those of the WSP and the brain, disappeared after absorption with tilapia serum. Two-dimensional electrophoresis revealed thatat least two serum proteins at 20 kDa and 21 kDa immunoreacted with the antiserum. The amino acid composition of the 20-kDa protein was dominated by Glx but was not necessarily similar to that reported for tilapia otoliths.

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In stressed and unstressed fish muscles, the I-band disappeared and the M-and Z-lines became dense at 3 and 10h postmortem, respectively. In spinal cord destroyed fish muscle, the I-band disappeared at 24h, but the M-and Z-lines remained a light color. This state remained unchanged till 48h. In stressed fish muscle, the interfibrillar space was detected immediately afterdeath, and the space increased as the postmortem time elapsed. In unstressed fish muscle, the space began to increase at 10h. In spinal cord destroyed fish muscle, the slight space appeared in only some cases at 24h, and in all cases at 48h.
The distinct space detected in the muscle of stressed fish immediately after death was due to the generation of tension. The increased space in stressed fish muscle at 3h and in unstressed fish muscle at 10h was due to both the peak tension and the low muscle pH. The spinal cord destroyed fish muscle which showed the slight space at 48 h was not weakened by the presently receiving tension nor the experienced peak tension. The lack of proteolytic degradation with higher muscle pH was related to the less structure weakening in this muscle.

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Upon heating of carp myosin, actin-activated Mg-ATPase activity decreased much faster than its Ca-ATPase activity. Sepharose CL 4 B gel filtration revealed that myosin formed oligomer, which retained its Ca-ATPase activity; oligomer at void volume (Vo) on Sepharose CL 4 B exhibited ATPase activity. However, the Mg-ATPase/Ca-ATPase activity ratio for the Vo fraction was very low. Myosin filament formed by the oligomeric myosin at Vo was irregular. It was concluded that the low actin activated Mg-ATPase activity is responsible for the filament formed by oligomeric myosin. Oligomers were easily salted-out at 40% saturated ammonium sulfate, leaving monomers in the soluble fraction.

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