The sterol composition of marine invertebrates, crustaceans, molluscs, echinoderms, coelenterates, and sponges has been re-examined by using improved analytical techniques. As a result, the sterol composition of marine invertebrates has been shown to be more complex mixtures including many new types of sterols with unusual steroid nuclei or with non-conventional side chains. Some crustaceans and molluscs are of importance as seafood and aquaculture species. Crustaceans and some molluscs require dietary sources of sterol for growth and survival because of the absence of de novo sterol-synthesizing ability. The present paper gives a review of the requirement and nutritive value of sterols, biosynthesis and metabolism of sterols, and composition and structure of sterols in marine invertebrates.

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Reproductive biology of the big eye, Priacanthus macracanthus, is described based on 601 specimens caught by otter trawl in north-eastern Taiwan waters and landed at Tahsi fishing port from December 1997 to November 1998. Four methods, macroscopic appearance of ovary, gonadosomatic index, oocyte diameter frequency distribution, and histological examination suggested that the spawning season of the big eye in north-eastern Taiwan waters is from April to July and peaks in May and June. Five stages of oocyte development were determined based on histological examination and ovarian development can be divided as undeveloped, developing, matured, and spent stages. Mean fecundity (±SD) was estimated to be 131112±33379. Size at first maturity estimated from the Logistic model was 18.9 cm FL corresponding to a 3-year-old specimen.

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Five geographic information system (GIS) grid themes, water depth, bottom type, topographic slope, distance to coast and distance to fishing ports, were adapted to evaluate and select a site for artificial reef deployment in I-Lan Bay off north-eastern Taiwan. From AHP (analytic hierarchy process) analysis, the weights of suitability of the five GIS grid themes were 0.416, 0.141, 0.331, 0.074, and 0.038, respectively. The results show that water depth is the most important factor in selecting a site for an artificial reef. Using ArcView, a GIS software package, a suitability map was made based on the selected grid themes. The suitable sites for artificial reef deployment identified by the GIS method corresponded to the sites selected using the traditional non-GIS approach. Based on the individual GIS grid themes, the GIS software and AHP method can be integrated to select objectively the optimal sites for artificial reef deployment.

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Seasonal growth and maturation patterns of two Sargassum horneri populations were surveyed in Hiroshima Bay on the western Seto Inland Sea. Both S. horneri populations began to exhibit rapid growth in autumn, exceeding a daily increase in plant length of 10 mm/day. However, there are several differences in seasonal timing for maturation and reaching maximum plant size. Plants of the Ohno-seto population, inhabiting the innermost area of Hiroshima Bay, reached their maximum plant size in December and became senescent after January. Plants of the Matsugahana population, inhabiting the mouth of the bay, continued to grow during winter and reached their maximum plant size in April. Senescence occurred after May. The largest difference in the seasonal patterns of the two S. horneri populations is most likely to be in the timing of maturation. In the Ohno-seto population, receptacle formation of the plants begins in November, and the eggs are released from late November to January. The plants settled as embryos spend the winter in an early stage of development. In the Matsugahana population, on the contrary, the plants spend the winter as adults and gradually produce receptacles. Abundant egg release of the Matsugahana population occurred in April and May.

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The vertical movement of a chum salmon Oncorhynchus keta returning to Japan was examined using a depth-recording archival tag in the western North Pacific Ocean over a 23-day period. The chum salmon showed clear diel vertical movements, with an average swimming depth of 12.8 m during the day and 4.8 m at night. The chum salmon spent 48% of the day and 85% of the night in the upper 10 m, 30% of the day and 10% of the night in the 10-20 m layer, and 20% of the day and 5% of the night in the 20-60 m depth layer. The chum salmon remained in the top part of the water column and rarely went below 40 m and only went below 50 m on 3 days out of the 23 days. The diel vertical movement of the chum salmon may be related to maximization of foraging efficiency.

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The developmental processes of the olfactory organ, taste buds, lateral line system, and inner ear in the Japanese sardine Sardinops melanostictus were examined in reared and wild specimens. Both the ciliated and microvillous receptor cells in the olfactory organ were found shortly after hatching. Formation of the olfactory nostrils and lamellae began at about 20 mm standard length (SL). The calculated inflection points in the relationship between the number of olfactory lamellae and standard length were at 30.3 and 62.7 mm SL. Taste buds first appeared in a 16.1-day-old larva of 11.2 mm notochord length. Newly hatched larvae were equipped with 12 pairs of neuromasts on the head and trunk. The formation of the lateral line canal commenced at about 20 mm SL and the four canals had ossified by 32.5 mm SL. At hatching, the inner ear was an oval-shaped auditory vesicle with two otoliths. Three semicircular canals formed by the first-feeding stage, and larvae assumed an upright position. The pro-otic bulla was gas-filled and formation of the lagena pocket began at about 20 mm SL. The saccular sac was entirely formed by about 32 mm SL. From the present results, the basic structure of the sense organs was almost completed by 32 mm SL, and the adult condition might be attained at 60-70 mm SL, as suggested by the second inflection point in the relationship between the number of olfactory lamellae and SL.

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Historical catch and effort data collected from tuna longliners during 1962 and 1997 are used to investigate how the spatio-temporal distribution of the two most economically important species of tunas in the Tropical Pacific Ocean (TPO) (i.e. yellowfin Thunnus albacares and bigeye Thunnus obesus) varies with the ocean temperature fluctuations that occur during El Niño/Southern Oscillation (ENSO) events. Relatively high hook rates of both species were mostly associated with regions where sea surface temperature (SST) increased during El Niño or La Niña years. Conversely, relatively low hook rates were associated with regions where SST fell during La Niña years, especially in the eastern TPO. The hook rates of these regions varied concurrently or within 3 months after the onset of an ENSO, as determined by the Southern Oscillation Index (SOI). We speculate that the displacement of the preferable water column and the change in availability of fishing gear during ENSO are the two possible reasons causing the change of hook rates. During El Niño periods, the part of the water column preferred by bigeye tuna was expended in western regions of the eastern TPO and compressed in the east. This appeared to lead to a displacement of bigeye from east to west. During La Niña episodes, yellowfin tuna appeared to undergo a meridian displacement and expend their preferred range northwards.

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The common Japanese conger Conger myriaster is known to mature up to the mid-vitellogenic stage without hormonal treatment, in contrast to the Japanese eel Anguilla japonica, which does not mature spontaneously under captive conditions. In order to obtain basic information on the process of gonadal development in anguilliformes under natural conditions, we have characterized cDNA of two distinct gonadotropin β-subunits (GTH-Iβ and -IIβ) in the common Japanese conger, and examined changes in their mRNA levels during vitellogenesis in the absence of hormonal treatment. Northern blot analysis revealed that the mRNA levels of GTH-Iβ were low at the perinucleolus stage, increased at the oil droplet stage, and peaked at the primary yolk globule stage. However, the levels decreased significantly at the secondary yolk globule stage. On the other hand, GTH-IIβ mRNA was first detected at the oil droplet stage, and showed a drastic increase with the progression of ovarian development. These results suggest that the two GTHs have distinct functional roles in oogenesis in anguilliformes where GTH-I is involved in the early stages of oogenesis, and GTH-II in the late stages of oogenesis.

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We examined the stomach contents of seven pelagic fish species collected by longline in the eastern tropical Pacific Ocean: yellowfin tuna Thunnus albacares, bigeye tuna T. obesus, swordfish Xiphias gladius, striped marlin Tetrapturus audax, dolphinfish Coryphaena hippurus, lancetfish Alepisaurus ferox, and pelagic thresher Alopias pelagicus. Fifty fish species from 31 families were identified in 222 stomachs examined. The choice of prey fish was very similar between bigeye tuna and swordfish, yellowfin tuna and swordfish, yellowfin tuna and dolphinfish, and big eye tuna and pelagic thresher. In contrast, similarity was low between striped marlin and lancetfish, dolphinfish and pelagic thresher, and lancetfish and pelagic thresher. According to the habitat depth range of prey fish, the seven predator species were divided into three groups: (i) wide range feeders from surface to midwater (yellowfin tuna and striped marlin); (ii) epipelagic feeders (dolphinfish); and (iii) midwater (mesopelagic) feeders (bigeye tuna, swordfish, lancetfish, and pelagic thresher). The most important prey fish belonged to the following families: Sternoptychidae, Phosichthyidae, Paralepididae, Omosudidae, Myctophidae, Exocoetidae, Hemiramphidae, Bramidae, Gempylidae, and Scombridae. Although pelagic fish predators extensively use these prey families, different dominant families and feeding depths of each predator are considered to reduce trophic competition among pelagic fish in the eastern equatorial Pacific.

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Two experiments were conducted with flounder to examine phytase pretreatment or dietary phytase supplement for improving phosphorus (P) utilization of soybean meal (SBM) or soy protein concentrate (SPC). In experiment I, three SBM based-diets, SBM (basal), phytase-supplemented SBM (supplement), and phytase-pretreated SBM (pretreatment), were fed to flounder, and digestibilities of P, gross energy (GE) and crude protein (CP) were measured by an indirect method. In experiment II, three SPC based-diets including P-supplemented mineral mixture (control), P-free mineral mixture (unsupplemented), or P-free mineral mixture supplemented with phytase (supplement) were fed to flounder for 50 days, and growth, whole-body P composition, and plasma mineral concentration were determined. In experiment I, a significantly higher absorption of P was found in fish fed the pretreatment or supplement diet compared to the fish fed the basal diet. In experiment II, the growth of fish fed the supplement diet was comparable to that of the control and better than that of fish fed the unsupplemented diet. Whole-body P retention and plasma P concentrations of supplement-fed fish were significantly higher than those of fish fed the unsupplemented diet. These results suggest that phytase supplementation to the SBM- or SPC-based diets improves phytase-bound P availability for Japanese flounder.

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Although essential for fisheries resources management, little is known about the population structure of the Japanese conger Conger myriaster. To search for useful molecular markers for the study of population structures, mitochondrial (mt) and nuclear DNA diversities in the Japanese conger were examined using DNA sequencing and amplified fragment length polymorphism (AFLP) analysis, respectively. A region spanning from part of the 3' end of the cytochrome b gene to the central part of the control region (585 base pairs/individual) was sequenced for 73 individuals from five different localities around Japan. A total of 99 variable positions were found, and a total of 67 haplotypes were identified in the 73 individuals. The average sequence differences within samples (1.8-2.4%) was comparable to those between samples (1.9-2.3%). The results of a hierarchical analysis of the sequence data did not suggest clear genetic subdivision among the local samples. Although a total of 499 bands were detected in the AFLP analysis from 58 individuals with three primer sets, no genetic structuring could be suggested. Nonetheless, to elucidate the subtle structure of populations, if any, many more individuals from more localities should be examined in detail. The genetic variations detected in the present study would serve as useful molecular markers for such studies.

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The supplemental effect of amino acids (AAs) to a soy protein concentrate (SPC) diet for red sea bream Pagrus major was evaluated for different-sized fish. The fish were fed artificial diets based on 52% SPC for 60 days in juvenile (mean weight 11.7 g) and for 153 days in yearling (mean weight 178 g). To these diets L-lysine hydrochloric acid (LS), DL-methionine (MS) and both L-lysine and DL-methionine (LMS) were supplemented. A control diet without AA supplementation (NOS) and a reference diet based on brown fish meal (BFM) were also tested. In the juvenile fish, the growth performance of the MS group was significantly higher than the NOS group. The growth performance was further improved by the dietary supplementation of both Lys and Met (LMS group). Lenticular opacity was not observed in these two dietary groups. On the other hand, the growth performance of the NOS and LS groups was lowest, and the incidence of lenticular opacity of the LS group was higher rather than the NOS group. In the yearling fish, feed conversion of the LMS group was slightly higher than the NOS group, but the growth performance of the LMS group was similar to the NOS group. These parameters were similar or slightly lower in the LS or MS groups compared with the NOS group. Lenticular opacity was not observed in any dietary groups for yearling fish. In both juvenile and yearling fish, the performances of the BFM groups were superior to the other dietary groups. The results for juvenile fish indicate that the quality of the SPC diet is improved by the Met supplementation, and further improved by both Lys and Met supplementation. Furthermore, the results in the present study suggest that the AA requirement of red sea bream changes with growth and/or age.

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The location and timing of spawning of the Japanese eel Anguilla japonica was studied during two research cruises of the R/V Hakuho Maru and R/V Suruga Maru in the North Equatorial Current of the western North Pacific Ocean during June-September 1998. There were 38 A. japonica leptocephali (10.0-43.2 mm in total length (TL)) collected in three areas: 24 specimens around the Arakane and Pathfinder Seamounts in June (approx. 16°N and 143°E) and five specimens at the southernmost station (13°N) and nine specimens at the northernmost station (17°N) of a transect along 137°E in September. The average total lengths of the leptocephali were significantly different among the three areas, with those around the seamount being smallest, those at the northern station being largest, and none being collected along the easternmost 144°E transect. This and the currents in the region suggested that spawning of A. japonica occurred near some of the seamounts in the West Mariana Ridge. Back calculated spawning dates indicated that most leptocephali were born during the new moon, supporting the hypothesis that A. japonica spawns around the new moon. Analysis of otolith daily rings found a strong correlation between total length and age (r=0.97), and the average daily growth rate was about 0.5 mm/day.

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The process of gonadal sex differentiation in the F₂ hybrid sturgeon, the bester (Huso huso female × Acipenser ruthenus male), was investigated histologically. In fish more than 6 months old, two types of gonads were distinguishable. In the putative ovary, the germinal epithelium had invaginations in the stroma of the gonad, and a few germ cells were localized immediately below the epithelium. In putative testis, invaginations were lacking and germ cells were observed deep in the stroma. However, active mitotic and meiotic germ cells were not observable even by 16 months of age in ovaries, and only some of the testes showed initiation of spermatogenesis by 21 months of age. The invaginations in the stroma of putative ovaries developed into ovarian lamella-like structures, whereas the gonadal surface of putative testes remained smooth. These results suggest that the formation of invaginations in the stroma of gonads is the morphological index of ovarian differentiation and that it is necessary to start hormone treatment for functional sex reversal before 6 months of age in these fish.

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To determine the thermal limit for normal spawning of Japanese whiting and to evaluate thermal effects on spawning in the normal temperature range, two experiments were conducted. In Experiment 1, the temperature of three experimental groups (Groups A, B and C) was increased from 25 to 28, 31 or 33°C, respectively. Subsequently, temperatures in all groups were decreased to 28°C. During the experimental period, the number of eggs, egg size, and hatching rate were monitored. According to the results, the upper thermal limit of normal spawning was considered to be 28-29°C, because the number of eggs and the hatching rate decreased at temperatures over 28°C. However, egg size tended to be reduced at 28°C, and was found to be strongly dependent on temperature. In Experiment 2, the number of eggs, egg size and time of spawning were compared between fish held at a constant temperature of 22°C (Group CT) and those held at increasing temperatures from 22 to 28°C (Group IT). Group IT spawned more in number and had smaller-sized eggs than did Group CT, while spawning was delayed in group IT. To identify the thermal effects on spawning of Japanese whiting, the mechanisms governing egg size must be determined.

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In 1999, spotted halibut Verasper variegatus were experimentally reared at 9, 12, 15, 18, and 21°C from eggs through early juveniles at the Miyako Station of Japan Sea-Farming Association in Iwate Prefecture. Tissue concentrations of thyroxine (T₄) increased to maximum levels during metamorphic climax (H-stage) at all the temperatures. The peak T₄ levels were higher at lower temperatures, whereas no relationship was observed between tissue 3, 5, 3'-triiodothyronine levels and rearing temperatures. However, epithelial cell height of thyroid follicles and the number of vacuoles in colloid were not different among rearing temperatures. Although the peak T₄ levels were different, thyroid activity to produce T₄ might not be so different at all temperatures. Therefore, much T₄ observed at low temperatures was possibly due to the accumulation of the unused hormone in blood and tissue.

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In the present study, the effects of bilateral eyestalk ablation on molt interval, ovarian development, and hemolymph levels of ecdysteroid and vitellogenin were investigated in the giant freshwater prawn, Macrobrachium rosenbergii in order to examine the roles of the eyestalk hormones. In destalked males and females, ecdysteroid levels increased rapidly, and molt intervals were significantly shortened in comparison with control non-destalked prawns. All destalked females exhibited a reproductive molt cycle accompanied by ovarian development and oviposition even under continuous hemolymph sampling. In contrast, in control females, the percentages of animals undergoing a reproductive molt cycle were 69 and 51% with and without sampling, respectively. The duration of ovarian development was shortened in the destalked females. These results indicate the inhibitory roles of the eyestalk hormones on molting and vitellogenesis. On the other hand, after eyestalk ablation, male prawns molted more than twice, and destalked females showed a correlation of vitellogenin level to molt stage similar to control reproductive females. These results indicate that the molt cycle and the correlation between vitellogenesis and molting are maintained in the absence of eyestalk hormone regulation.

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The sea catfish Plotosus lineatus has four pairs of barbels of almost the same length as major taste and tactile organs. The patterns of innervation in the barbels of the sea catfish were studied to reveal peripheral neural organization for taste and mechanosensory systems. Taste buds are located in the epidermis of each barbel throughout the length of barbel. The density of taste buds is low at the proximal and increases distally and at the transverse levels, highest at the rostral, fairly high at the caudal and very low in the intermediate sides throughout the longitudinal axis of the barbel. Observations of cross-sections of the barbels show that the taste buds are usually located as pairs innervated by two different nerve strands originating from the same nerve bundle. The carbocyanine dye, Dil, was applied to the nerve stump in dissected fixed barbel specimens to trace the peripheral course of nerve fibers. The barbel nerves enter the caudal region at the base of the barbel as a trunk. The trunk sends various sizes of bundles to the rostral side coursing toward the tip. These bundles move in turn to the rostral and caudal epidermis and ramify repeatedly to make networks under the epidermis, but few networks were found in the intermediate sides. The networks usually make hexagons in shape. Their size is smaller in the rostral than in the caudal side and becomes smaller toward the apical region in both sides. The maximum and minimum diameters for the hexagons range 240-400 μm and 100-250 μm, respectively. Nerve strands appear in pairs from each network to innervate taste buds. The number of strands originating from one network ranged 20-50. Each strand divides into two substrands, each of which enters the same bud laterobasally opposite one another. Perigemmal and extragemmal fibers, which end around taste buds and in the epidermis, respectively, were also frequently found. These results show the unique organization of nerve fibers in the barbel taste and tactile systems of the sea catfish.

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The effects of muscle types and their cooling conditions on browning in the packing media solution of canned skipjack were investigated. Cooling conditions of dorsal muscle influenced browning of water fraction in packing media solution mainly due to amino-carbonyl reaction associated with non-protein nitrogen and sugars. For ventral muscle, brown color formation of oil as well as water fractions occurred due to degradation of amino nitrogen and non-protein nitrogen, and lipid oxidation. The separation of whole meat into dorsal and ventral muscles after precooking, cooling at different appropriate temperatures (≤10°C for 16 h and -15°C for 3 h for dorsal and ventral muscles, respectively), and packing with lower ventral muscle content (≤20%) retarded browning of packing media solution in canned skipjack.

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Tyrosinase inhibitor-producing microorganisms were isolated from the marine environment and characterized. Out of about 500 isolates from various sea areas, only three strains that were isolated from sediment samples of Off-Izu Islands and Sagami Bay were found to produce the inhibitors. Among the three, strain H1-7 that was isolated from Off-Izu Islands had significantly and constantly the strongest inhibitory activity (1000-2500 U/mL) and was selected for further studies on the identification of the strain and production of the inhibitor. The strain showed abundant to moderate growth covered with aerial mycelium on most media except for Miura's medium and cornmeal agar, forming a white sparse mycelial mat at first and later a conidial area shaded yellowish white to greenish yellowish gray. The diameter of the aerial mycelium without seawater was much larger than the medium with seawater on all the media tested. It had antimicrobial activity against most microbes tested. Based on the taxonomical characteristics, strain H1-7 belonged to the genus Trichoderma and was named Trichoderma sp. H1-7. The optimal medium for the production of the inhibitor was investigated by using shake flasks and an improved basal medium. The maximum production of the inhibitor was observed in a medium consisting of 2% soluble starch, 1% glucose, 0.5% Bacto-peptone (Difco) and 0.5% Bacto-yeast extract (Difco) at pH 7.5 in 50% (v/v) seawater for 2 days with shaking.

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The survey on scallop toxicity in association with causative dinoflagellates was conducted when the cultured scallops Patinopecten yessoensis were found to accumulate paralytic shellfish toxins in late November of 1997 at Otsuchi Bay, Iwate Prefecture, Japan. Alexandrium catenella was observed in high density at the beginning of the survey. The cell density of A. catenella decreased and completely disappeared in early December. The scallop toxicity also decreased with decrease of the cell density of A. catenella. The toxin profile of the scallop was similar to that of natural and cultured cells of A. catenella isolated from the Bay, showing that scallop toxicity is due to A. catenella. This is the first case in which paralytic shellfish toxins were confirmed in scallops during a bloom of A. catenella in Sanriku coast of Iwate Prefecture.

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A clean-up procedure using a solid phase extraction technique and a high-pressure liquid chromatography (HPLC) method for the determination of histamine in fish sauce from Thailand is introduced. The method was tested on fish sauce samples by adding various concentrations of histamine standards: 50-1000 mg/L of fish sauce, five replicates for each concentration. The samples were mixed with methanol and shaken for 1 min, applied to Bond Elut strong cation exchanger, then eluted with a mixed buffer solution of 0.5 M KH₂PO₄ (pH 6.5): methanol (1 : 1, v/v). Analysis was carried out by using HPLC with a post-column reaction with o-phthalaldehyde to form a fluorescent derivative. The recovery from fish sauce ranged from 90.5 to 95.5% with the limit of detection at 5 ng of histamine. The analysis of 549 commercial fish sauce samples showed that the samples contained histamine at various levels with the majority in the range of 200-600 mg/L.

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Soluble and insoluble dietary fibers are recognized to have different physiological responses. Therefore, we worked out a new method for binding of bile salts to soluble and insoluble dietary fibers in seaweeds. Dried and pulverized Porphyra yezoensis, Undaria pinnatifida, Laminaria japonica, and Hizikia fusiformis were suspended in cholate, chenodeoxycholate, or deoxycholate solution, and incubated. After centrifugation and then precipitation with ethanol according to the analytical method of soluble and insoluble dietary fibers, the concentration of cholates not binding to both dietary fibers was measured by enzymatic colorimetry. The percentage of bile salts bound to insoluble dietary fiber was higher than that to soluble dietary fiber, because the amount of insoluble dietary fiber was higher in seaweeds. The binding ability of soluble dietary fiber seemed to be greater than that of insoluble dietary fiber. Adsorption of cholate to seaweeds was lower in comparison with chenodeoxycholate and deoxycholate. Thus, seaweed dietary fibers have a different capacity to adsorb cholates.

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