Physiological response,blood chemistry profile and mucus secretion of red sea bream (<Emphasis Type="Italic">Pagrus major</Emphasis>) fed diets supplemented with <Emphasis Type="Italic">Lactobacillus rhamnosus</Emphasis> under low salinity stress

Environmental stressors caused by inadequate aquaculture management strategies suppress the immune response of fish and make them more susceptible to diseases. Therefore, efforts have been made to relieve stress in fish by using various functional feed additives in the diet, including probiotics. The present work evaluates the effects of Lactobacillus rhamnosus (LR) on physiological stress response, blood chemistry and mucus secretion of red sea bream (Pagrus major) under low salinity stress. Fish were fed four diets supplemented with LR at [0 (LR0), 1 × 10² (LR1), 1 × 10⁴ (LR2) and 1 × 10⁶ (LR3) cells g^?1] for 56 days. Before stress, blood cortisol, urea nitrogen (BUN) and total bilirubin (T-BIL) showed no significant difference (P > 0.05), whereas plasma glucose and triglyceride (TG) of fish-fed LR2 and LR3 diets were significantly lower (P < 0.05) than those of the other groups. Plasma total cholesterol (T-CHO) of fish-fed LR3 diet was significantly (P < 0.05) lower than that of the other groups. Furthermore, total plasma protein, mucus myeloperoxidase activity and the amount of mucus secretion were significantly enhanced in LR-supplemented groups when compared with the control group (P < 0.05). After the application of the low salinity stress test, plasma cortisol, glucose, T-CHO and TG contents in all groups showed an increased trend significantly (P < 0.01) compared to the fish before the stress challenge. However, plasma total protein and the amount of secreted mucus showed a decreased trend in all groups. On the other hand, BUN, T-BIL and mucus myeloperoxidase activity showed no significant difference after exposure to the low salinity stress (P > 0.05). In addition, the fish that received LR-supplemented diets showed significantly higher tolerance against low salinity stress than the fish-fed LR-free diet (P < 0.05). The physiological status and the detected immune responses, including total plasma protein and mucus myeloperoxidase activity in red sea bream, will provide a more comprehensive outlook of the effects of probiotics to relieve stress in fish.     

Reproductive traits of the Ryukyu long-furred rat (Diplothrix legata) on Okinawa-jima Island

The Ryukyu long-furred rat, Diplothrix legata, is a large rodent distributed only on Amami-ohshima Island, Tokuno-shima Island and Okinawa-jima Island, Japan. This animal is endangered as a result of deforestation, predation by introduced carnivores and mortality caused by vehicles. We performed theriogenological examinations of 32 male and 25 female Ryukyu long-furred rats carcasses collected from wild populations on northern Okinawa-jima Island from December 2005 to September 2013. Adult males had remarkably large preputial glands. Seminiferous diameter of adult was significantly small (136 ± 28 µm, n=8) from April to August. Numerous spermatozoa were observed from September through February, and seminiferous diameter was significantly large (216 ± 27 µm, n=12) during this time in adults; testes length changed in a similar pattern. These findings indicate that the mating season may occur from September through February. Size (body length) at sexual maturity was estimated to be >560 mm in both sexes. From observation of corpora lutea and placental scars, litter size was estimated to range from 2 to 12 (average=6, n=4). These results provide fundamental knowledge that will be beneficial for in situ and ex situ conservation of this rare species.     

Growth,nutrient utilization,oxidative condition,and element composition of juvenile red sea bream <Emphasis Type="Italic">Pagrus major</Emphasis> fed with fermented soybean meal and scallop by-product blend as fishmeal replacement

A feeding trial was conducted to evaluate the effect of fermented soybean meal and scallop by-product blend (3:2) (FP) on the performance of juvenile red sea bream. Five isocaloric diets were prepared by replacing 0% (FP0), 15% (FP15), 30% (FP30), 45% (FP45), and 60% (FP60) fishmeal (FM) protein with FP, respectively. Triplicate groups of fish (initial mean weight 2.83 g) were fed the test diets for 45 days in a flowthrough seawater system. The results demonstrated that growth rates of fish fed FP0, FP15, and FP30 were similar, and significantly higher (P < 0.05) than those of FP45 and FP60. Nutrient utilization was significantly lower in FP60, with no differences found among the other groups. Dietary heavy-metal contents were affected by inclusion of FP, impacting on whole-body heavy-metal contents. In terms of oxidative stress, fish fed the FP30 diet were in the best condition, since this fish group showed the least oxidative-stressed condition as well as the highest tolerance against oxidation. In conclusion, the approach of utilizing this fermented mixture is promising and it could replace at least 30% FM protein in red sea bream diet without negative effects on performance, body composition or health of fish.     

Effect of water temperature on the feeding activity and the resultant mercury levels in the muscle of cultured bluefin tuna Thunnus orientalis (Temminck and Schlegel)

Tuna muscle often contains high levels of mercury, and fish samples with mercury concentrations ten times higher than the specified safety standards have been reported. Here, we report on the relationship between water temperature and the concentration of mercury in the tail muscle tissue of cultured bluefin tuna Thunnus orientalis. The fish used in this study were cultured at Fisheries Laboratory of Kinki University (Amami Experimental Station, Kagoshima, Japan). One hundred fish weighing 26.2–89.4 kg were selected for analysis between February 2007 and January 2008. Water temperature during rearing ranged from 21 to 29 °C. The total mercury levels were measured using the reduction vaporizing atomic absorption method after acid digestion. Body weight increased approximately 1.5 times that observed in a previous study, despite feeding activity either being the same or less than that observed previously. The average mercury concentration in white muscle was 0.353 mg kg^?1, remaining almost constant and independent of body growth. Unlike previous studies, seasonality was not observed in this study. Based on these findings, water temperatures within a certain range were considered to stabilize feeding activity and increase feeding efficiency. Consequently, water temperature is considered to have a moderating effect on seasonal fluctuations in muscle mercury concentrations in cultured bluefin tuna.     

Replacement of squid and krill meal by snail meal (Buccinum striatissimum) in practical diets for juvenile of kuruma shrimp (Marsupenaeus japonicus)

A 60‐day feeding trial was conducted to investigate the effects of substituting squid and krill meal with marine snails (Buccinum striatissimum) into the diets of juvenile kuruma shrimps (Marsupenaeus japonicus). Five experimental diets were formulated to contain varying levels of snail meal at 0%, 25%, 50%, 75% and 100% (D1, D2, D3, D4 and D5, respectively) and fed to juvenile kuruma shrimps (initial mean weight 0.27 ± 0.02 g). The results showed that weight gain, feed intake and specific growth rate were improved significantly in D4 and D5 groups when compared with D1 group (p < 0.05). Significant differences were not detected in survival rate among all shrimps fed diets containing several levels of snail meal (p > 0.05). Crude protein content of shrimps fed the control diet was significantly lower than other treatments (p < 0.05). Lipid content in shrimps fed 50% snail meal were significantly higher than the control while cholesterol content in shrimps fed 100% snail meal were significantly decreased and polyunsaturated fatty acid contents were significantly increased in shrimps fed 75%–100% snail meal (p < 0.05). These results suggest that supplementing snail meal for complete replacement of squid and krill meal can be done to improve juvenile kuruma shrimps’ growth and reducing their cholesterol levels.     

Can fermented soybean meal and squid by‐product blend be used as fishmeal replacements for Japanese flounder (Paralichthys olivaceus)?

An experiment was carried out to evaluate fermented soybean meal and squid by‐product blend (1:1) (FP) as replacement of fishmeal (FM) for Japanese flounder, Paralichthys olivaceus. Five isocaloric (19 kJ g^?1) diets were prepared by replacing 0 (FP0), 12 (FP12), 24 (FP24), 36 (FP36) and 48% (FP48) FM protein with FP. Triplicate groups of juveniles (mean weight of 3.9 g) were delivered the test diets for 8 weeks in a flow‐through sea water system. The results showed that there were no significant differences (P > 0.05) among the growth rates of fish fed FP0, FP12, FP24 and FP36 diets. Growth and nutrient utilization parameters were significantly reduced in fish fed FP48 diet. Although, whole body proximate composition of fish were not significantly affected by the dietary treatments compared to the control; methionine and phenylalanine contents were significantly decreased in FP48 group. Protein retention was also significantly decreased in the similar group of fish. Dietary treatments did not alter most of the plasma metabolites, while some of the health parameters were improved in the replacement groups. Results suggested that FP is a potential candidate for alternative protein ingredient in aquafeed and can replace 36% FM protein in the diet of Japanese flounder.     

Early metaphase II oocytes treated with dibutyryl cyclic adenosine monophosphate provide suitable recipient cytoplasm for the production of miniature pig somatic cell nuclear transfer embryos

We investigated the effects of in vitro maturation duration and treatment with dibutyryl cyclic adenosine monophosphate (dbcAMP) on the blind enucleation efficiency and developmental competence of miniature pig somatic cell nuclear transfer (SCNT) embryos. Oocytes were cultured for 22 h in NCSU-23 medium with or without 1 mM dbcAMP and then additionally cultured in dbcAMP-free NCSU-23 for 14, 18, or 22 h. Regardless of dbcAMP treatment, the rate of nuclear maturation reached a plateau at 36 and 40 h. However, mitochondrial distribution, a marker for cytoplasmic maturation, differed between the dbcAMP-untreated oocytes at 36 h and dbcAMP-treated oocytes at 40 h. The metaphase II chromosomes were adjacent to the first polar body in 68.8% and 63.5% of the dbcAMP-untreated oocytes at 36 h and dbcAMP-treated oocytes at 40 h, respectively. Furthermore, the blind enucleation efficiency by removing a small volume of cytoplasm was significantly higher in the dbcAMP-untreated oocytes at 36 h (82.9%) and dbcAMP-treated oocytes at 40 h (89.9%) than other groups. The rate of blastocyst formation was highest in the dbcAMP-treated oocytes at 40 h. Hence, this study demonstrated that dbcAMP-treated early metaphase II oocytes are suitable for the production of miniature pig SCNT embryos.     

Effect of simulated acid fog on membrane-bound calcium (mCa) in fir (Abies firma) and cedar (Cryptomeria japonica) mesophyll cells

The decline of virgin fir (Abies firma) forest at Mt. Oyama has been reported. Related field observations suggest that high acidity fog is linked with its decline. However, cedar (Cryptomeria japonica) in the same area shows no symptoms of decline. For assessing effects of acid fog on membrane-bound calcium (mCa) of the leaf mesophyll cells, 9-year-old seedlings of fir (Abies firma) and 8-year-old seedlings of cedar (Cryptomeria japonica) were exposed twice a week to simulated acid fog (SAF at pH 3 with pH 5 as control) for 2 h per day in a chamber during May–December 2007 (except August). Current and 1-year-old needles were collected from seedlings and analyzed at 1-month intervals. For current-year needles of fir, mCa levels in cells exposed to SAF at pH 3 were significantly lower than in cells exposed to pH 5, especially during September 2007–March 2008. In contrast, it is noteworthy that mCa levels of cedar were maintained as virtually constant irrespective of SAF acidity, indicating that fir is more sensitive to acid fog than is cedar. Based on these results, mCa loss by acid fog might also be caused in the declining virgin fir forest at Mt. Oyama.     

First report of blueberry red ringspot disease caused by <Emphasis Type="Italic">Blueberry</Emphasis><Emphasis Type="Italic">red</Emphasis><Emphasis Type="Italic">ringspot</Emphasis><Emphasis Type="Italic">virus</Emphasis> in Japan

Virus-like symptoms—red ringspots on stems and leaves, circular blotches or pale spots on fruit—were found on commercial highbush blueberry (Vaccinium corymbosum) cultivars Blueray, Weymouth, Duke and Sierra in Japan. In PCR testing, single DNA fragments were amplified from total nucleic acid samples of the diseased blueberry bushes using primers specific to Blueberry red ringspot virus (BRRV). Sequencing analysis of the amplified products revealed 95.7–97.7% nucleotide sequence identity with the BRRV genome. This paper is the first report of blueberry red ringspot disease caused by BRRV in Japan. The nucleotide sequence data reported in this paper are available in the GenBank/EMBL/DDBJ database as accessions AB469884 to AB469893 for BRRV isolates from Japan.     

Dormancy Characteristics of Malus ‘Snowdrift’ and Effects of Spraying 6-benzyladenine on Its Germination and Flowering in Long-day Treatment

[Objective] The dormancy characteristics of Malus ‘Snowdrift’ in long-day treatment were studied, and 6-BA was used to break the dormancy, with the aim to achieve the purpose of flowering in autumn. [Method] The new shoots of ‘Snowdrift’ in long-day treatment were conducted with hydroponics to investigate their dormancy time. And cytokinin 6-benzyladenine(6-BA)was used to treat the dormant shoots, to investigate the budding, flowering and flower bud differentiation. [Result] The shoots in long-day treatment entered endodormancy after August 7~(th) and the shoots in the natural daylight entered endodormancy before July 18~(th). In long-day treatment, 116 buds, 198 flowers were observed after 6-BA spraying. [Conclusion] The optimum concentration of 6-BA was 300 mg/L. And the flower bud differentiation of ‘Snowdrift’ in long-day treatment was faster than those in natural daylight after 6-BA spraying.