We evaluated the effect of probiotics treatment on growth, digestibility and immunity against Vibrio bacteria in Kuruma shrimp, Marsupnaeus japonicus, which were reared in a closed recirculating system. The experimental shrimp were divided into four groups as follows: the control group was fed with the control diet and reared in the water without probiotics; the probiotics diet group was fed with the diet containing probiotics and reared in the water without probiotics; the water supply group was fed with the control diet and reared in the water with probiotics; the combination group was fed with the diet containing probiotics and reared in the water with probiotics. Growth in three groups treated with probiotics were significantly higher (P<0.05) than that in the control group. Protein, fat and phosphorus contents in the whole body of the shrimp in the probiotics treated groups were higher, while protein and fat contents in the feces were lower than those in the control group. The number of Vibrio bacteria in the shrimp bodies and in the rearing water in the probiotics treated groups, were significantly smaller (P<0.05) than that in the control group. As to the tests of air-exposed stress in sawdust and heat stress in water, the shrimp in the probiotics treated groups gained a higher tolerance of air-exposed and heat compared with the shrimp in the control group. Results indicated that probiotics supplied in the diet and/or in the rearing water, improved the growth of the shrimp and the digestibility of protein and fat, controlled the vibrio bacteria in shrimp bodies and in the rearing water, and therefore shrimp built up stronger resistance to stress conditions.

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Chemical characteristics of bottom sediment of mud flat and Zostera japonica meadow at intertidal zone, and sea bed were revealed. Total organic carbon (TOC), total nitrogen (TN) and acid volatile sulfide (AVS) of the intertidal zone were lower than those of sea bed. Furthermore C/N and N/P atomic ratios of intertidal zone were higher and lower than sea bed, respectively (p<0.05). From these results, it is considered that the environment of intertidal zone was aerobic and organic matters contents, especially nitrogen atoms, were smaller in comparison with the sea bed environment. On the other hand, in phosphorus composition, residual phosphorus compound (Re-P) was the most abundant phosphorus at the intertidal zone and there was a significant correlation between Re-P and TOC (p<0.001). This correlation indicates that the main component of Re-P was organic phosphorus compound, suggesting that phosphorus at the intertidal zone mainly exists as organic compound. In fact, chemical characteristic of bottom sediment of intertidal zone differ greatly from the sea bed, whose phosphorus compound is composed chiefly of calcium bound phosphorus (Ca-P). Relativity between phosphorus composition and TOC suggests that the source of phosphorus, which gives origin to Fe and Al bound phosphorus (Fe-P and Al-P), is from coastal land area, but the amount supplied is scarce.

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This study was aimed to investigate the physico-chemical property differences of the muscles in wild and cultured red sea bream obtained from Kagoshima region, southern Kyushu of Japan. Twenty live fish samples comprising 10 wild fish and 10 cultured fish (mean weight 1380±40.0 g) were obtained from the Kagoshima Bay fishery and the Azuma Marine Fish hatchery, Kagoshima. All specimens were slaughtered by hypothermia, divided into dorsal and ventral fillets and maintained under 0°C for 48 hours wrapped in aluminium foil. Monitoring of k-value, pH and muscle rheology was conducted by using the fillets at 6-hour intervals for 48 hours and sensory analyses were conducted by using samples prepared for Japanese sashimi. Approximately 200 g samples from dorsal and ventral fillets were then freeze-dried for proximate analysis. In the dorsal muscles, k-values at 24, 30 and 36 h increased to 5.6, 5.5 and 6.4% respectively in culture fish compared to 3.1, 3.5 and 3.8% respectively in wild fish. The k-value at 48 h was significantly higher (P<0.05) in cultured (17.5%) than that in wild (4.5%) ones. Ventral muscles showed slightly higher k-value at 0 hr in both cultured and wild fish, increasing significantly with time. Accordingly, deterioration rate of sashimi was faster in cultured than wild fish. On the contrary, at 36 h the pH values were significantly lower in cultured fish (6.0 in dorsal; 6.0 in ventral) compared to wild fish (6.5 in dorsal; 6.9 in ventral). Water and fat losses in ventral muscles in both wild and cultured fish were higher compared to those of dorsal muscles. The data of muscle rheology indicated textures of wild fish were harder than those of cultured fish. However, all samples deteriorated at 48h. Sensory scores for dorsal freshness, taste, odor and texture were generally similar and product acceptance of sashimi from both wild and cultured fish was not significantly different. Cultured fish showed significantly higher lipid in dorsal and ventral than in wild. Lipid contents were higher in dorsal muscles than in ventral muscles for both wild and cultured fish. Results demonstrated that physico-chemical properties differences between wild and cultured red sea bream were attributed to higher fat accumulation in cultured fish than in wild fish. Other parameters indicated no big differences between dorsal and ventral muscles of both wild and cultured fish within 48 h after slaughter. It can be concluded that wild and cultured red sea bream demonstrated similar quality characteristics within 48 hours after slaughter.

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This study was aimed to investigate the effects of replacing fish oil with palm oil (P) blended with fish oil (F) as the lipid source on growth performance and lipid profiles of red sea bream. A 50-day feeding trial for juveniles (initial body weight 3.68±0.09 g) was conducted in triplicates using 4 isocaloric diets containing different ratio of fish oil and palm oil such as {10:0 (10F0P), 8:2 (8F2P), 6:4 (6F4P) and 4:6 (4F6P). Growth performance in terms of specific growth rate (SGR), body weight gain (BWG), feed intake (FI), feed conversion ratio (FCR), and hepatosomatic index (HSI) were taken. Protein, moisture, ash and lipid contents in both whole body and liver, fatty acid compositions in dorsal and ventral muscles, and liver were also compared among treatments. There were no significant differences in SGR of fish fed with 10F0P and 8F2P but the SGR in both treatments were significantly higher than that of 4F6P. Diet 6F4P and 4F6P groups demonstrated significant lower FI as compared to diet 10F0P and 8F2P groups. Whole body lipid deposition in fish fed diet 4F6P was significantly lower than those of other dietary groups. On the other hand, the dorsal lipid contents gradually decreased with increased palm oil level but there was no significant difference in ventral muscle. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) contents in both dorsal and ventral muscles gradually declined with increased dietary palm oil. A similar gradient degradation in n-3/n-6 ratio was also demonstrated in both dorsal and ventral muscles although no signs of EFA deficiency occurred throughout the experimental period. In conclusion, diets containing not exceeding 40% palm oil may be a suitable ratio in incorporating palm oil as a fish oil replacement.

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Gut microbes of red sea bream Pagrus major can ferment lactosucrose, an indigestible and fermentable oligosaccharide, in vitro. Dietary lactosucrose (LS) increases mechanical strength of intestinal tunica muscularis of this fish. Thus, the aims of this study were to evaluate hindgut fermentation of dietary LS by the intra-luminal existence of short-chain fatty acids (SCFA), and to confirm the effect of this sugar on weight of digestive organs in a marine teleost, red sea bream. Fish of approximately 70g body weight were randomly assigned to two groups and fed either a commercial diet (control group) or a commercial diet supplemented with 0.24 g/100 g LS (LS group) for two months. Fish fed LS had greater SCFA in intestinal contents than did control fish (P<0.001). The ratios of SCFA/total organic acids of hindgut contents exceeded 55% in fish fed LS and were significantly greater than in control fish (P<0.01). Contents of free (not bonded) water in hindgut contents of fish fed LS were lower than in control fish (P<0.05). Fish fed LS had heavier stomachs and intestines than in fish fed control diet (P<0.05). The above results suggest that hindgut microbes of red sea bream can ferment LS to produce SCFA. Fermentation of this sugar should affect intestinal water absorption by lowering free water content and affect digestive organs by SCFA fermentation metabolites.

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Pen shells Atrina pectinata (shell length 120-145 mm) were reared in the breeding seawater with the resuspended sediments collected from several sites in Ariake Bay, adjusting at 800 Nephelometric Turbidity Unit (NTU) for 15 days. The results showed that survival rate and glycogen content of the pen shells in the resuspended sediments were higher than those of the non-suspended and bentonite groups. In addition, the following thickness of sediments were covered over the pen shells (shell length 135-150 mm), 0 (control), 2, 5, 10, 20 and 35 mm. As a result, survival rate and glycogen content of these pen shells under surface accumulated soft sediments over 10 mm thickness declined in comparison with those of the soft sediments less than 5 mm thickness. These results suggest that the resuspended sediments could be one of the efficient food sources for pen shells. However, by the sediment over 10 mm thickness, glycogen consumptions and fatality risk increase for pen shells in order to avoid burial with thick sediments.

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The gametophytes of derived from sporophytes of Ecklonia cava and E. kurome collected at six sites in Wakayama Prefecture were cultivated for 30 days under 14, 18, 20, 22, 25 and 28°C water temperature conditions. The growth of all gametophytes of both species increased with increasing temperature between 14 to 25°C, but they grew in abnormal at 28°C. The optimal water temperatures for maturation of the gametophytes and for early development of sporophytes were different among individuals from different sites. In addition, the shape of the juvenile sporophytes showed considerable differences among individuals cultured under different water temperatures. They took on round shape under 14°C culture, whereas did slender under above 18°C culture. These results suggested that response of gametophytes and sporophytes to water temperature didn’t reflect the difference between E. cava and E. kurome.

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In order to find out a effect of low water temperature treatment in reducing the reddening adductor disease caused mortality of the Japanese pearl oyster Pinctada fucata martensii in Nagasaki Prefecture, mortality and physiological traits were compared each month for one year between the different groups of oysters kept in various pearl farms where temperatures were below 13°C during winter, whereas control shells were kept in a warmer area. As an indicator of low temperature burden, the low-water-temperature index [LTI₁₃=Σ(13-Tn), Tn<13] was calculated from the water temperature measured each day at 10 o’clock (am) for each group of oyster. As compared to control, the 100 degree-day group showed lower mortality, lower a-value (value obtained from the spectrophotometer and used as a color indicator for the reddening adductor), slower gonad maturation, higher glycogen content in the adductor muscle and higher phyto-pigment content in the digestive diverticula after wintering particularly from spring to autumn when the reddening adductor disease progressed. The high survival rate in the 100 degree-day group at the end of experiment may be attributed to the delayed onset of the reddening adductor disease which was a result of higher glycogen content, delayed maturation and increased food intake of the oysters.

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We examined seasonal changes in the gonadosomatic index of wild fish and detailed the first spawning period of each year of cultured fish over several years to determine the spawning season and lunar-related spawning of camouflage grouper Epinephelus polyphekadion in a subtropical area of southern Japan. Additionally, we observed captive females’ and males’ spawning behavior and elucidated the mating system of this species. The main spawning season of wild fish was April-May of the solar calendar (Gregorian calendar), when gonadosomatic index (GSI) values of females and males peaked. The first spawning periods varied in the solar calendar. However, they were observed from the 19th to 23rd day in the third month of the lunar calendar (lunisolar calendar) during the full moon and the last quarter moon, excluding the years when spawning occurred one month later, because of low water temperatures before the spawning season in those years. This result indicates that camouflage grouper shows lunar-related spawning. The basic mating system of camouflage grouper is pair spawning between the largest dominant male and an ovulated female. However, some non-dominant males undertook sneaker spawning.

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From December 2004 to February 2006, growth, maturation, standing crops of Sargassum nipponicum and oceanographic conditions were surveyed at depths of 1-3 m on the coast of Hachijo-jima Island, central Japan. S. nipponicum germinated in October. Their growth rate was slow until December and rapid from January to April 2005. Their thallus length was the greatest during May to July, reaching to main 55 cm. Their receptacles were apparently observed from June to July 2005. Their maximum and minimum of standing crops were 2479.6 g, dry/m² in June 2005 and 12.2 g, dry/m² in November 2005, respectively. The oceanographic conditions at the study site were higher water temperature (average monthly water temperature 18.7-26.5°C) and lower nutrient concentration (DIN: 0.4-2.2μM, DIP: 0.01-0.22μM), characteristic of the Kuroshio Water. It is noteworthy that S. nipponicum showed very high values in standing crop at the study site despite of higher water temperature and lower nutrient concentration there.

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We studied the specific immunostimulant effect of heat-killed Enterococcus faecalis FK-23 (FK-23) when the artificial diet including FK-23 was fed on Japanese flounder and ocellate puffer. The diets with 5 different concentrations of FK-23 was prepared (0-1.25%). Fishes were injected formalin-killed cells of Vibrio anguillarum and the injection was performed two times. The second injection was done 3 weeks after the first injection. The agglutination titer of antibody for V. anguillarum cells in their serum was evaluated every week after the first injection and the evaluation was performed six times. The agglutination titer in the serum of ocellate puffer fed the diet including highest concentration (1.25%) of FK-23 increased more intensively with the progress of rearing than other concentrations. However those puffers fed the diet with 1.25% of FK-23 did not grow during the experimental rearing period. Although the agglutination titer in the serum of flounder did not increase, this is possibly because the non-specific immunity worked more intensively than specific one. Consequently FK-23 is possible to increase the vaccination effect of puffer and maintain its effect, and it also make the non-specific immunity of flounder intensify.

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Growth rings were clearly visible in rotula of red sea urchin, Pseudocentrotus depressus, by heating at 350°C for 10-180 min. After confirming a single formation of annulus in a year from December to January, we confirmed that growth rings in rotula were useful for age determination of red sea urchin.

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Marine algae were collected from March 2004 to February 2005 at Sabusawa Island, Hanabuchihama, and Funairi Island in Matsushima Bay (38°18´N, 141°5´E). A total of 134 species were collected, of which 54 species newly listed as marine plants in Matsushima Bay. The I/H value, the ratio of the number of species with isomorphic alternation of generations or no alternation (generally found in warm current areas; I) to that of species with heteromorphic alternation of generations (generally found in cold current areas; H) in Matsushima Bay was 1.7, clearly higher than the values of 1.3-1.4 found in localities to the southern coast of the Oshika Peninsula (38°24´N, 140°22´E). This shows that Matsushima Bay belongs to the temperate floral region phytogeographically. The I/H value were higher than those found in the past. Cole’s coefficient index of similarity between past and present marine algal flora in Matsushima Bay showed no significant similarity. The marine algal flora in Matsushima Bay has changed to a warm current type in synchrony with the 1.5°C increase in the average sea surface temperature from 1926-1935 to 1995-2004.

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Lactobacillus species can be used as supplementary probiotics, although the mechanism of protection of host animals from pathogenic bacteria remains unknown. The protective abilities of four candidate probiotics (L. brevis, L. plantarum, L. salivarius and L. rhamnosus) were examined in in vitro and in vivo systems. Zone of growth inhibition on the assay plates revealed that the extracellular products (ECP) and bacterial cells (BC) of some Lactobacillus species have a bacteriostatic effect on E. tarda. Serum obtained from fish fed with L. salivarius and L. rhamnosus in the 7th and 14th days post infection of E. tarda significantly inhibited the growth of E. tarda as compared to serum from fish fed with L. brevis and L. plantarum. Fish fed of L. plantarum, L. rhamnosus and L. salivarius showed low mortalities, while fish fed of L. brevis and control fish did high mortalities. Furthermore, histological examination revealed hyperplastic appearance of macrophage with cytoplasmic enlargement in the early period of E. tarda infection of fish fed fish of L. salivarius and L. rhamnosus and eventually the formation of pyogranuloma. These results implied that four Lactobacillus species have different effects to protect fish from E. tarda infection with increasing macrophage response and lower mortality of fish.

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The Japanese silver-biddy Gerres equulus (Temminck and Schlegel) is one of the most common shore fishes in the costal waters of southern Japan and commercially important for the local fishery. The batch fecundity of G. equulus was examined from the specimens collected in the Yatsushiro Sea, western Kyushu, Japan in June-September, 2001 and 2002. Batch fecundity (BF) ranged from 15,083 to 92,356. The regression correlation between BF and standard length (SL) for the early and the late spawning period was significantly different and these regressions were expressed as BF=1.00×10^-4 SL^3.79 (early spawning period) and BF=1.30×10^-4 SL^3.33 (late spawning period), respectively. The BF of females at age 2-10 years was higher in the early spawning period than in the late spawning period.

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The expanding populations of the great cormorant Phalacrocorax carbo require a management program to reduce damage on freshwater fisheries. The piscivorous predator has recently invaded into the Kofu Basin located at more than 70km inland from the river mouth. Visual censuses identified birds foraging along 80km-watercourses of the Kamanashi and Fuefuki rivers. We tentatively regarded areas where diving and feeding of birds occurred as a foraging zone with the neighboring areas left unexploited as a control zone. Riparian forest on the upper courses concealed foraging birds, and shoreline areas on the lower courses were broad enough to help conceal birds remote from the bank. They were capable of catching fish in a variety of stream conditions including a shallow riffle area with 20cm depth or an area of rapids with 160cm/s flow. Collections by casting net revealed that fishes, especially ayu Plecoglossus altivelis, were more abundant in the foraging zones than in the related control zones. The birds appear to forage at a site within a temporal habitat of ayu, and thereby their diet is biased towards ayu. Preventing parental birds from foraging for ayu is deemed to be effective to make the invasive population decline.

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The marble goby, Oxyeleotris marmoratus is the highest priced freshwater fish in Southeast Asia. However, the stable seed production technique has not yet been established due to the high mortality occurring in the larval stage. To solve the problem, 2 rearing experiments were conducted with Brachionus sp. feeding in different salinity levels (SaL). In Experiment 1, the effects of different SaLs on the survival and growth were investigated in the larval stage, during the period from 1 to 40 days after hatching (d AH). In Experiment 2, the larval stage was divided into 4 periods (1-10, 11-20, 21-30, and 31-40 d AH), and the optimum SaL for each period was determined. The optimum SaL was 10 ppt through out the whole larval stage. It was understood that the most important period of the salinity adjustment was in 1-10 d AH for the larval rearing. Although 10 ppt is the optimum SaL in the later rearing periods of 11-20, 21-30, and 31-40 d AH, larval rearing was possible in both freshwater and seawater. The successful utilization of Brachionus sp. as feed for freshwater fish O. marmoratus was a substantial progress for stable mass seed production in future.

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The marble goby, Oxyeleotris marmoratus (Eletoridae) is the largest freshwater gobioid fish in the world and is the most expensive freshwater fish in Southeast Asian countries. Rapid development for the culturing techniques of this fish has increased the demand of the juvenile. However, the stable seed rearing technique has not yet been established due to high mortality occurred during the early juvenile stage or after 40 days after hatch (d AH). To develop a seed rearing technique, an experiment on different salinity levels (SaLs) was conducted to determine the optimum SaL on growth and survival at the early juvenile stage (40 to 100 d AH). The optimum SaL was 10 ppt throughout the whole period in this experiment. The mean body weight (BW) and survival rate at 100 d AH in 10 ppt SaL were 0.31 g and 85.6% respectively, those being significantly higher (P<0.05) than those in the other SaLs. The most important period of the salinity adjustment was in 40 to 70 d AH for the juvenile rearing of O. marmoratus. After 70 d AH it is possible to rear the fish either in freshwater or brackishwater.

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To establish a seed production technique for a new hybrid orange-spotted grouper Epinephelus coioides × tiger grouper E. fuscoguttatus the egg and larval development were observed under artificial conditions. Newly ovulated eggs from a female E. coioides of 7.5 kg in body weight were measured at 0.81±0.02 mm (mean±SD) in diameter and weighed 3,505 eggs/g. After fertilization with sperm obtained from a male E. fuscoguttatus, its diameter measured 0.83±0.02 mm. The eggs hatched from 17 h 30 min (17:30 h) to 19:00 h after fertilization at 29.0°C at 30.0 ppt salinity. The fertilization and hatching rates were 93.9% and 50.3%, respectively. Newly hatched larvae were 1.52±0.01 mm in total length (TL) and floated motionless at water surface. Larvae commenced feeding at 3 days after hatched (d AH) when mouth and digestive tract were formed and eyes became deeply pigmented. Larvae showed typical early Epinephelus type pigmentation and differentiation of second dorsal-fin and pelvic-fin spines was observed prior to 10 d AH and thereafter elongated. Larvae started shifting habitat from pelagic to benthic at 40 d AH. Seven hundred and fifty tails 50 d AH juveniles with mean TL of 22.8±3.6 mm were produced from 21,500 newly hatched larvae.

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Serum protein content of pearl oyster Pinctada fucata martensii, an indicator of the selection of male and female parent-shells, was measured in 2002 and 2003, in order to develop a method of producing disease-resistant pearl oysters with high survival rates against the reddening adductor disease. The survival rate of 1-year-old oysters produced from parent-shells of high serum protein was compared with those obtained from the low content group. The serum protein of parent-shells ranged from 0.53 to 1.62 mg/ml in 2002 and from 0.13 to 0.86 mg/ml in 2003. The higher and lower content groups were established from high 10% and low 17-35% oysters of the frequency histogram, respectively. The oysters obtained from high content group showed high serum protein and total body weight from autumn thereafter as compared to lower group. At the end of the experiment, higher group was 5.0-20.4% higher in survival rates than lower group. Above results suggest that selection of parent-shells with high serum protein can be effective in improving survival rate during seed production.

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The histopathology of Nile tilapia Oreochromis niloticus eggs with fungal infection was described. Numerous hyphae were shown to spread over the outer layer of the egg envelope, infiltrating deeper into the yolk granules and accumulating in the cytoplasm. Invasive pores of hyphae were seen within some areas of the egg envelope and zoospore germination was also observed outside the egg envelope.

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Spheroplasts (PS) were prepared by degradation of the cell wall of a low quality laver Porphyra yezoensis with enzymes, and were used for a new artificial diet for short-neck clams Ruditapes philippinarum. Short-neck clams in aquarium were given the PS diet or condensed natural diatom Chaetoceros gracilis for 45 days. The PS diet induced a superior nutritional effect on the growth performance and carcass composition of short-neck clams as good as the diatom. As results, PS was found to be a useful ingredient of new artificial diet for short-neck clam that can be substituted for live microalgae.

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