Primary structure and thermostability of bigeye tuna myoglobin in relation to those of other scombridae fish

ABSTRACT:   In the present study, the cDNA encoding myoglobin (Mb) of bigeye tuna Thunnus obesus was cloned and its amino acid sequence deduced in order to investigate the relationship between the primary structure and thermostability of scombridae fish Mb. An open reading frame of bigeye tuna Mb cDNA contained 444 nucleotides encoding 147 amino acids. The primary structure of bigeye tuna Mb was highly conserved when compared with those of bluefin tuna and yellowfin tuna Mb, the sequence identity being 95.2–100.0%. It also showed relatively high identity (82.3–89.1%) with the counterparts of scombridae fish. Myoglobin was then isolated from the dark muscle of four scombridae fish including bigeye tuna. Differential scanning calorimetry and circular dichroism measurements on these Mb revealed that the thermostability of bigeye tuna Mb was lowest and that of skipjack Katsuwonus pelamis Mb highest among the scombridae fish Mb examined. The α-helical contents of scombridae fish Mb at 10°C were in the range of 39.8–44.8%, clearly lower than that of horse Mb (55.3%), suggesting instability of fish Mb. The melting temperatures of these Mb fell in the range of 75.7–79.9°C, lower than that of horse Mb (84.2°C). These results strongly suggest the instability of fish Mb.     

Molecular characterization of southern bluefin tuna myoglobin (<Emphasis Type="Italic">Thunnus maccoyii</Emphasis>)

The primary structure of southern bluefin tuna Thunnus maccoyii Mb has been elucidated by molecular cloning techniques. The cDNA of this tuna encoding Mb contained 776 nucleotides, with an open reading frame of 444 nucleotides encoding 147 amino acids. The nucleotide sequence of the coding region was identical to those of other bluefin tunas (T. thynnus and T. orientalis), thus giving the same amino acid sequences. Based on the deduced amino acid sequence, bioinformatic analysis was performed including phylogenic tree, hydropathy plot and homology modeling. In order to investigate the autoxidation profiles, the isolation of Mb was performed from the dark muscle. The water soluble fraction was subjected to ammonium sulfate fractionation (60–90 % saturation) followed by preparative gel electrophoresis. Autoxidation profiles of Mb were delineated at pH 5.6, 6.5 and 7.4 at temperature 37 °C. The autoxidation rate of tuna Mb was slightly higher than that of horse Mb at all pH examined. These results revealed that tuna myoglobin was unstable than that of horse Mb mainly at acidic pH.     

Amino acid sequences of α-skeletal muscle actin isoforms in two species of rattail fish, Coryphaenoides acrolepis and Coryphaenoides cinereus

SUMMARY: The cDNA clones of α-skeletal actins were isolated from the skeletal muscle of two species of rattail fish, Coryphaenoides acrolepis and Coryphaenoides cinereus . The complete nucleotide sequences of the cDNA and their deduced amino acid sequences were determined. Each of the two species had two α-skeletal actin cDNA. The nucleotide sequences of the coding region of the two α-skeletal actin isoform genes within each species had 92.0 and 91.8% homology. From the cDNA sequences of the four α-skeletal actin isoforms in the two species, amino acid sequences of 377 amino acid residues were deduced. It was predicted that the two N-terminal amino acid residues of each protein are processed after translation. The amino acid sequences of α-skeletal actin 1 in the two Coryphaenoides species were identical, as were the amino acid sequences of α-skeletal actin 2 in the two species. The amino acid sequences of the two α-actin isoforms, α-skeletal actin 1 and α-skeletal actin 2, differed by only a single amino acid, Ala/Ser at the 155th position. Northern blot analysis showed that a similar amount of each of the two α-actin isoform mRNA was expressed in the skeletal muscle of the two Coryphaenoides species.     

Possibility for decreasing of mercury content in bluefin tuna <Emphasis Type="Italic">Thunnus orientalis</Emphasis> by fish culture

ABSTRACT:   The bluefin tuna tested were reared for 22 months from eggs before the beginning of the experiment, and sampling was performed every 3 months over the following year. The experimental results showed that the mercury concentration in the muscle ranged from 0.32 to 0.85 μg/g, which is lower than that found in wild bluefin tuna of a similar size. Increase in the mercury concentration corresponding to the increase in body weight was not shown, and it was quite different with wild bluefin tuna. Furthermore, no significant relationship was found between the lipid concentration and the mercury concentration in muscle. Among the internal organs of cultured bluefin tuna, the heart (0.32–0.66 μg/g), liver (0.43–0.99 μg/g) and spleen (0.59–1.0 μg/g) contained higher concentrations of mercury. It was estimated that the full-cycle cultured bluefin tuna had been fed small fish containing lower concentrations of mercury, and that the mercury concentration of tuna would be almost equal throughout the year because the effect of mercury accumulation would be weakened by body growth. Therefore, it was concluded that selecting diet fish species might decrease mercury contents in cultured bluefin tuna.     

In vitro amino acid absorption using hydrolysed sardine muscle or soybean meal at different intestinal regions of the Pacific bluefin tuna (Thunnus orientalis)

The amino acid (AA) absorption along the intestinal tract of the Pacific bluefin tuna (Thunnus orientalis) was evaluated using two hydrolysed protein sources (fresh sardine muscle and soybean meal) with the everted intestine technique. Pork pepsin and pancreatic enzyme extract from the bluefin tuna were used to hydrolyse the protein from fresh sardine (FSH) and soybean meal (SMH) under optimal bluefin tuna fish physiological conditions. Both of the hydrolysate solutions were tested within three intestinal sections from the bluefin tuna. The everted intestinal fractions immersed in the hydrolysate solutions were sampled at different times to analyse for AA and absorption rate calculations. Fresh sardine and SMH contained greater amounts of essential amino acids (EAA) than those of non‐essential amino acids (NEAA); however, the profiles of AA absorbed showed higher absorption of NEAA in both cases. Using a similar concentration solution, the absorption rates within the intestinal fractions showed a preferential absorption in the proximal and distal regions for Arg and His when FSH was used. However, the absorption rates for Lys resulted in a decreasing proximal‐to‐distal gradient between the different intestinal regions for FSH and SMH. The possibility of a catabolic role of certain AAs in the enterocytes being able to explain the differences in absorption is discussed.     

Purification and characterization of glutathione peroxidase 1 in the red muscle of Pacific bluefin tuna Thunnus orientalis

Glutathione peroxidase GPx1 was purified from the red muscle of Pacific bluefin tuna Thunnus orientalis and its enzymatic properties were characterized. The enzyme was optimally active at pH 7.4 with a specific activity for hydrogen peroxide and a K _m of 6.7 μM. cDNA was also isolated and it contained a predicted open reading frame (ORF) encoding a 188 amino acid protein. The phylogenetic tree shows that fish including Pacific bluefin tuna, pufferfish, and zebrafish, but not mammals, possess two genetically distinct types of GPx1, i.e., GPx1a and GPx1b. The purified enzyme was classified as a fish GPx-1b enzyme on the basis of the phylogenetic tree of the GPx1 family.     

美济礁附近海域3种金枪鱼肌肉成分检测与营养评价

为分析中国南海美济礁附近海域大目金枪鱼(Thunnus obesus)、蓝鳍金枪鱼(T.thynnus)、黄鳍金枪鱼(T.albacares)的营养成分与营养价值,该研究采用国标法检测了3种金枪鱼的水分、蛋白质、灰分、脂肪酸和氨基酸成分及其结构,并进行营养评价。结果显示,3种金枪鱼中黄鳍金枪鱼蛋白质含量最高,蓝鳍金枪鱼脂肪和灰分含量最高;以氨基酸评分(AAS)和必需氨基酸指数(EAAI)为评分标准,3种金枪鱼氨基酸评分均大于等于1;3种金枪鱼各种必需氨基酸构成均优于联合国粮食及农业组织(FAO)/世界卫生组织(WHO)模式;蓝鳍金枪鱼的油酸含量远高于其他2种金枪鱼,且差异显著(P<0.05);蓝鳍金枪鱼的单不饱和脂肪酸(MUFA)和多不饱和脂肪酸(PUFA)种类最多且含量最高。结果表明,3种金枪鱼均具有较丰富的营养成分、较完美的营养比例和较高的营养价值;3种金枪鱼相比,黄鳍金枪鱼可提供更多的蛋白质,蓝鳍金枪鱼提供更多的不饱和脂肪酸。美济礁附近海域3种金枪鱼具有很高的开发价值,成分检测对研究设计这3种金枪鱼专用驯化养殖饲料有重要的参考价值。     

Occurrence of two distinct molecular species of cathepsin B in carp Cyprinus carpio

ABSTRACT:   We purified cathepsins B₁ and B₂ from the ordinary muscle of carp Cyprinus carpio . The N-terminal amino acid sequences (12 residues) of 29 kDa bands of cathepsins B₁ and B₂ are the same and showed high homology of 75% and 83%, respectively, with the heavy chain of rat and human cathepsins B. Based on conserved sequences of other cathepsins B and the N-terminal amino acid sequences of 29 kDa bands, we cloned carp cathepsin B cDNA. The nucleotide sequence of carp cathepsin B cDNA consists of 1470 bp including a 993 bp open reading frame, encoding a deduced protein of 330 amino acids. The deduced amino acid sequence of carp cathepsin B has similarity of 80% to rainbow trout cathepsin B and of 76–78% to other vertebrate cathepsins B. The sequence of its isoform was also determined during molecular cloning, which has 94.8% similarity with first cloned cathepsin B. They are completely same in N-terminal amino acid sequence of heavy chain, active site and potential N-glycosylation site. This indicates there are at least two kinds of cathepsin B functioning in vivo in carp.     

Universal PCR primers for ribosomal protein gene introns of fish

Human ribosomal protein (RP) gene sequences with respect to intron/exon structures and corresponding cDNA or genomic data of fish species were obtained from the GenBank database. Based on conserved exon sequences, 128 primer pairs for 41 genes were designed for exon-primed intron-crossing (EPIC) polymerase chain reaction (PCR). In reference to the draft genome sequences of the Pacific bluefin tuna (Thunnus orientalis), 12 primer pairs expected to amplify introns of the bluefin tuna with lengths of 500–1000 bp were selected and applied to six distantly related fish species belonging to the Orders Clupeiformes, Tetraodontiformes, Pleuronectiformes, Perciformes, Scorpaeniformes, and Anguilliformes. PCR amplification was observed for at least four species in each primer pair, and all fragments were larger than those expected for intronless amplification. Single fragment amplification was observed for at least seven primer pairs per species. Fragment sizes of the bluefin tuna for nine primer pairs corresponded to those expected from the genomic data. Thus, our primer pairs are potentially applicable to a wide variety of fish species and serve as an initial step for isolating single-copy nuclear DNA sequences.     

Inclusion of prey data improves prediction of bluefin tuna (Thunnus thynnus) distribution

We examined the distribution of Atlantic bluefin tuna ( Thunnus thynnus ) in the Gulf of Maine, Northwest Atlantic Ocean, from 17 to 23 August 1995, in relation to physical and biological parameters. Specifically, we fit a binomial GLM to the bluefin tuna presence–absence data and predictor variables that include: sea surface temperature (SST), ocean depth, distance to an SST front, time-lagged density of SST fronts, and an interpolated surface of Atlantic herring ( Clupea harengus ) density. In addition, we use simple and partial Mantel tests to examine whether bluefin tuna presence–absence data are significantly associated with these predictors, once spatial autocorrelation is accounted for. Results suggest that the distribution of bluefin tuna significantly correlated with herring density ( z  =   3.525, P  =   0.000424), and that inclusion of biological variables results in a more parsimonious model. Mantel tests results indicate that bluefin tuna abundance is significantly correlated with herring density after the effect of spatial structure is removed (Mantel r  =   0.043, P  <   0.019).     

Turbulence effect on survival and feeding of pacific bluefin tuna <Emphasis Type="Italic">Thunnus orientalis</Emphasis> larvae,on the basis of a rearing experiment

ABSTRACT:   Physical conditions such as oceanic turbulence related to food availability are considered to be important factors affecting fish larval survival. Rearing experiments were conducted to elucidate the effects of turbulence on the survival and feeding rates during the initial feeding period of Pacific bluefin tuna Thunnus orientalis . Six levels of turbulence intensity were provided by changing flow rates from pipes set on the bottom of rearing tanks. The result showed a dome-shaped relationship between turbulence level and survival rate, in which the feeding rate appeared higher at a logged turbulence energy dissipation rate of −6.32, and decreased at both higher and lower turbulence levels. Compared with the turbulence intensity in the ocean, the optimal turbulence level for Pacific bluefin tuna larvae corresponded to the turbulence caused by sea surface winds with speeds of 4–12.5 m/s. The estimated optimal turbulence intensity for Pacific bluefin tuna larvae is comparable to that for yellowfin tuna Thunnus albacares .     

Three cone opsin genes and cone cell arrangement in retina of juvenile Pacific bluefin tuna <Emphasis Type="Italic">Thunnus orientalis</Emphasis>

ABSTRACT:   In bluefin tuna aquaculture, collision of juveniles with the tank or net walls is a major cause of high mortality. This problem may be related to color sensibility of the visual mechanisms of this species. As a first step in understanding of color vision of Pacific bluefin tuna Thunnus orientalis , we applied a molecular technique and histology to study cone cell distribution in the retina of juvenile fish. We isolated three cone opsin genes encoding one blue-sensitive ( SWS2 ) and two green-sensitive ( RH2 ) visual pigments. In situ hybridization revealed that SWS2 mRNA is localized in the single-cone photoreceptors. The localization of the two RH2 signals in distinct cone cells was not determined, probably because of the high homology between the two RH2 genes. Single-cone photoreceptors appeared frequently in the ventral–temporal retina in approximately 80-mm fish and in the temporal retina in approximately 230-mm fish. These cone distributions may define a visual field with best color contrast vision in front and above the fish with a short wavelength (blue) reflecting target (sensed by single cones), and may be enhanced against the longer wavelength (green) background when fish see a target below them (sensed by double cones).     

cDNA cloning and complete primary structures of myosin heavy chains from brushtooth lizardfish Saurida undosquamis and wanieso lizardfish S. wanieso fast skeletal muscles

ABSTRACT:   The complete cDNA sequences encoding predominant types of myosin heavy chain (MYH) in the fast skeletal muscle were determined for brushtooth lizardfish Saurida undosquamis and wanieso lizardfish S. wanieso , which are used as materials for preparing high-quality surimi-based products. The cDNA consisted of 5973 and 5987 bp, respectively, and both encompassed an open reading frame encoding a polypeptide of 1936 amino acid residues. Brushtooth and wanieso lizardfish MYH showed the amino acid sequence identity of 92–93% to white croaker MYH, which was higher than that of 90% to walleye pollack MYH. The putative binding sites for ATP, actin, and regulatory and essential light chains in the subfragment-1 region of brushtooth lizardfish MYH exhibited a high identity with white croaker counterparts as well as the sequences of subfragment-2 and light meromyosin. In contrast, phylogenetic tree, constructed by the neighbor-joining method based on mitochondrial 16S rRNA gene, revealed that the two lizardfish species formed a cluster with walleye pollack, which was paraphyletic with white croaker. Therefore, a good reputation for lizardfish and white croaker to have a high thermal-gel forming ability seemed to be reflected by MYH rather than biological similarity as revealed by the mitochondrial 16S rRNA gene.     

Bluefin tuna (Thunnus thynnus) distribution in relation to sea surface temperature fronts in the Gulf of Maine (1994–96)

Fishery‐linked aerial surveys for bluefin tuna (Thunnus thynnus) were conducted in the Gulf of Maine (GOM) from July through October, 1994–96. Each year, from 507 to 890 surface schools were detected and their locations examined in relation to oceanographic conditions. Correlations between bluefin tuna presence and environmental variables were explored for sea surface temperature (SST), distance to a SST front, frontal density (relative density of all SST fronts seen in a given 1 km area for 2 weeks prior to each tuna sighting), and bottom depth and slope. Mean SST associated with bluefin schools was 18.1°C (±2.8). Schools were located at a mean distance of 19.7 km (±19.6) from SST fronts, and in water masses with an average frontal density of 28.2 m km^?2 (±35.7). Mean bottom depth of detected schools was 139.0 m (±70.3), and mean bottom slope was 0.7% rise (±0.7). A binomial generalized linear model fit to these variables indicated that bluefin are seen closer to fronts than locations in which no tuna were seen. Using simple and partial Mantel tests, we investigated the spatial correlation between bluefin tuna presence and the environmental variables, controlling for spatial autocorrelation. For each day that schools were sighted, we performed 24 Mantel tests, on a combination of response and predictor variables. The spatial relationship between bluefin tuna and SST fronts was inconsistent. Our analysis identified significant spatial structure in the bluefin school locations that had no significant correlation with any of the measured environmental features, suggesting that other untested features, such as prey density, may be important predictors of bluefin distribution in the GOM.     

Recruitment abundance index of Pacific bluefin tuna using fisheries data on juveniles

ABSTRACT:   The recruitment abundance index of Pacific bluefin tuna Thunnus orientalis was estimated from 1980 to 2003 fishing year by using the troll fishery data in Nagasaki Prefecture, western Japan. It has been shown that the troll fishery in Nagasaki Prefecture operates with good time–area coverage of the species habitat, and that the fishing power slightly changed during the period analyzed, based on fisheries statistics, published information, and interviews with the fishers. Average catch per unit effort (CPUEs) were standardized by a generalized linear model (GLM) considering the effects of fishing year, season and landing area. Standardized CPUE of age-0 bluefin tuna showed larger fluctuations year by year than the nominal CPUE combined for all ages. High CPUEs in fishing years of 1981, 1994, 1996 and 1999 were observed. Data from these years agreed with the higher recruitments estimated by virtual population analysis (VPA) or higher catch of age-0 fish reported for the Pacific side. The age-specific standardized CPUE of age-0 bluefin tuna in this study was judged to be a useful indicator of recruitment.     

Comparative intestinal absorption of amino acids in rainbow trout (Oncorhynchus mykiss), totoaba (Totoaba macdonaldi) and Pacific bluefin tuna (Thunnus orientalis)

The kinetics of amino acid absorption in the proximal section of three fish species were studied using an everted intestine technique and a pancreatic digest of casein (Tryptone), as the model amino acid mixture. Fresh intestine of rainbow trout (Oncorhynchus mykiss) was used to set up the experimental system, and results were compared with those obtained using intestines from totoaba (Totoaba macdonaldi) and bluefin tuna fish (Thunnus orientalis). The kinetics of amino acid absorption was sigmoidal for all amino acids and for each species of fish tested. In general, specific absorptions of essential amino acids were higher than those of non‐essential ones. No correlation between the concentration of amino acids in the Tryptone solution and the corresponding absorptions was found. The maximum specific absorption rates of all amino acids for trout were 10 times higher than those determined for totoaba and bluefin tuna. The relative amounts of the different amino acids preferentially absorbed in all three species were different. Results obtained from the everted technique may be applicable to the design of formulated diets for large fish species with commercial value.