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Abstract:Fishery resource surveys are an important basis for the scientific management and conservation of fishery resources. The representativeness of resource survey stations is particularly important because there is limited sampling time and space. Optimizing the design of fishery resource survey stations under limited conditions is always the focus of fishery resources research. Taking the Bohai Sea as a study area and using a numerical simulation method, this paper analyzed the impact of species occurrence rate (17 main species were divided into three categories: classⅠwith an occurrence rate of more than or equal to 70%, class Ⅱ with an occurrence rate of 50%~70%, and class Ⅲ with an occurrence rate of lower than 50%) and habitat water layer on the relative error (REE) of single species biomasses, explored the influence of the number of stations (48 and 60) on the results of stationary sampling and stratified random sampling, and further optimized the design of multi-objective fishery resource surveys in the Bohai Sea. Results showed that the relative errors of biomass for five species in class Ⅰ, three species in class Ⅱ, and six species in class Ⅲ were less than 20%, less than 30%, and less than 35%, respectively. This indicated that the relative error for single species biomasses increased with a decline in the species occurrence rate, while the habitat water layer had no significant influence on the REE of species biomasses. As the number of survey station decreased, the accuracy value for stationary sampling decreased (the REE of biomass indexes for fish, shrimp, crab, and cephalopod increased by 1.1%, 2.5%, 8.4%, and 4.4%, respectively, and the REE of Margalef richness index increased by 3.3%). Stratified random sampling could compensate for the declining accuracy due to a decrease in the number of survey stations. For example, the accuracy of the fish biomass index (REE is 4.6%) obtained by stratified random sampling with 48 survey stations is higher than that obtained by stationary sampling with 60 survey stations (REE is 7.7%), which could help reduce survey costs and protect low-abundance species. However, each sampling method could not fully satisfy the multi-objective optimization, and the accuracy of stratified random sampling was affected by station allocation schemes. Setting survey stations based on the optimal criterion of sampling cost can obtain a higher accuracy of biomass assessment for fish, shrimp, crab, cephalopod, and main species such as Setipinna taty, Oratosquilla oratoria, Loligo japonica, Engraulis japonicus, Johnius grypotus, Carcinoplax vestita, and Fenneropenaeus chinensis, and could be used as a station design scheme for multi-objective fishery resource surveys in the Bohai Sea.

Abstract:In recent years, environmental DNA (eDNA) technology has developed rapidly as a new method for investigation of aquatic organisms and has been widely used in the field of aquatic ecosystem research for species detection, biodiversity evaluation, and biomass assessment. However, few studies have specifically evaluated the effects of different eDNA enrichment and extraction methods on the results of the eDNA technology operation process to establish a set of optimal eDNA technology operation procedures for specific research subjects. In addition, because of differences in living habits between species, the amount of DNA released from different species and the size of DNA fragments are different. Therefore, different eDNA enrichment and extraction methods should be adopted for different research subjects. In this study, Fenneropenaeus chinensis was used as the research object, and eDNA was enriched by the membrane method, and eDNA was extracted by combining a blood and tissue DNA extraction kit. A filter membrane with a diameter of 47 mm, a nitrocellulose membrane, a polycarbonate membrane, and a nylon membrane were used, and each membrane was set to 0.45 μm, 0.8 μm, 1.2 μm, and 5 μm according to pore size. With 4 gradients, the sampling water volume was set to three gradients of 500 ml, 1 L, and 2 L. The experimental results showed that the membrane material size, pore size of the membrane, and the volume of the sampled water had certain influences on the qualitative and quantitative analysis of F. chinensis. The 0.45 μm glass fiber membrane filter and 2 L water sample detected the greatest DNA copy number. Based on this, a set of operational procedures for F. chinensis prawn eDNA technology was established to improve the detection rate of F. chinensis, which provided the basis for subsequent distribution monitoring and biomass assessment of F. chinensis.

Abstract:In order to calibration the long-term change of heavy metal content and the heavy metal background in the muddy area of the middle Yellow Sea sediment of China, we combined with the contents of heavy metals (Cu, Pb, Cd, Zn), 210Pb data and the specific surface area (SS) of core A03 from the Huanghai Sea to structure the function models of the heavy metals (Cu, Pb, Cd, Zn) background. With the function models, we calculated the long-term change of the content of the heavy metals in the same SS in core A03. We compared with the content of the heavy metals in the same SS versus the content of heavy metals, showed a obvious demarcation point between them, in 100 cm. The trend is similar with a ideal sediment profile, which means that the starting point of contamination is around 100 cm. Calculated by deposition rate, the sediment age in the 100cm of the core is much older than 200 years. The average value of heavy metal contents under a single specific surface area (SS) below 100 cm without human pollution is taken as the background value of heavy metal Cu, Pb, Cd and Zn at station A03. The background of them are 26.31 mg/kg, 22.11 mg/kg, 0.20 mg/kg and 63.21 mg/kg, respectively. Compared with the nearby sea area, the background of heavy metals is similar. Compared with historical events, the variation characteristics of the heavy metals content under a single specific surface area correspond to major coastal historical events, which means that the estimation reduced the impacts from the provenance and particle characteristics of the sediments on the heavy metals content and that it provides a reference useful for increasing the veracity of evaluation of heavy metals contamination.

Abstract:Ocean acidification caused by the rising atmospheric CO2 concentration has been paid attention worldwide, the response process and mechanism of marine phytoplankton to ocean acidification are still not very clear. In this paper, we studied four kinds of microalgae Chrysophyta: Dicrateria sp., Bacillariophyta: Phaeodactylum tricornutum, Chlorophyta: Chlorella vulgaris and Platymonas subcordiformis to assess the response of microalgae to CO2-driven ocean acidification (the future level of the year 2300), and by the variation of quality and quantity of phytoplankton, to predict the potential influence of future global climate change on secondary consumers. The results indicated that compared with the control group, the average growth rates (μ) of the four kinds of microalgae were promoted by elevated CO2 concentration (P<0.05); for the value of μ, P. subcordiformis was the highest, 13.5% higher than the control group, followed by C. vulgaris (μ=5.9%), and then Dicrateria sp. and P. tricornutum (μ=2.2%). High CO2 concentration could increase carbon content and/or decrease nitrogen or phosphorus content, and then increase C/N or C/P ratio of phytoplankton. However, there were species different, both of the C/N, C/P ratio for P. subcordiformis were significantly increased (P<0.05), and C/P ratio of C. vulgaris and C/N ratio of P. tricornutum were significantly increased (P<0.05). The cellular chlorophyll a contents of C. vulgaris was increased significantly by elevated CO2 concentration. However, there were decreasing trends of the others. The maximal efficiency of PSⅡ in a dark-adapted state (Fv/Fm) of P. tricornutum elevated remarkably in the beginning of the experiment, the initial slope of rapid light curves (α) of Dicrateria sp. improved, non-photochemical quenching (NPQ) decreased significantly, and the maximum relative electron transport rate (rETRmax) of P. tricornutum and P. subcordiformis increased significantly (P<0.05). But high CO2 concentration has no remarkable effect on photochemical quenching (qP) of the four phytoplankton (P>0.05). Therefore, the growth rate of P. subcordiformis, C. vulgaris and P. tricornutum accelerated under the high CO2 concentration, whereas nutrition quality declined. Different kinds of phytoplankton have different responses to ocean acidification, which may change oceanic phytoplankton community structure in the future. In addition, the change of C/N and C/P ratio of phytoplankton could influence the primary consumer, such as zooplankton and filtering shellfish, through the food chain.

Abstract:In order to further understand the dynamics of phytoplankton and its physio-chemical variables during the development of cyanobacterial blooms in aquaculture ponds, water samples were collected and evaluated in an aquaculture pond during the cyanobacterial bloom (July to September, 2017) in Henan Normal University, Xinxiang, Henan (351938.363N, 1135409.482E). The results showed that during the study periods, in total, 25 phytoplankton species belonging to six phyla were identified, and the total phytoplankton abundance and biomass were (0.883~12.666) × 108 cells/L and 9.740~ 70.020 mg/L, respectively. Furthermore, the Shannon-Wiener index was determined, ranging between 0.05 and 1.15. Additionally, the total phosphorus (TP) and total nitrogen (TN) concentrations were 0.32~0.51 mg/L and 4.18~7.09 mg/L, respectively; the water temperature was 22.1~30.6℃. It was suggested that the high amount of TN and TP contents, combined with the high water temperature, might play important roles in this occurrence of cyanobacterial blooms and the low biodiversity in ponds. To further understand the key factors affecting phytoplankton composition, Redundancy analysis was applied. The results showed that cyanobacteria were closely related to the TP and TN concentrations and water temperature. In addition, in consideration of microcystin release due to the occurrence of Microcystis blooms, microcystin content was also determined in this study. At the end of the study periods, according to the ELISA, microcystin content was detected in algal cells and pond water, which were 0.686 µg/L and 0.040 µg/L, respectively. The World Health Organization suggested that the contents of microcystin should be no more than 1.0 µg/L in water samples. Thus, according to the present study, it suggested that the microcystin content was within the safety range in this bloom-occurring pond.

Abstract:To assess the genetic background of Liza affinis in the coastal waters of China, the sequence variation in 712 bp of the partial mitochondrial DNA COⅠgene of 84 individuals from 4 provinces was analyzed. In total, 18 polymorphic sites defined 21 haplotypes and low levels of genetic diversity (Hd = 0.8230±0.0400, π=0.0023±0.0002) were detected. Individuals from different sites were intertwined together in the parsimony network, suggesting no geographical clustering. AMOVA analysis of various groupings (sampling sites, seas, division of the Taiwan Strait and Qiongzhou Strait) detected no significant genetic differentiation, as evidenced by pairwise fixation indexes (Fst = –0.1940 ~ 0.0141, P > 0.05), and the percentage of variation (–0.6%~0.46%) among groups. In the SAMOVA analysis, the Dongtou population was treated as one group, and the others were treated as another group; the percentage of variation among groups was 2.87%, congruent with the results of Fst and AMOVA analysis. The pattern of no lineage structure and no obvious genetic differentiation in L. affinis populations in coastal waters of China might have been caused by: 1) Passive transport of larvae and juveniles by marine currents, or active feeding migration and spawning aggregation of adults. 2) Recent population expansion. Significant negative Fu’s Fs for all populations (FS = –20.3900, P = 0), and obvious unimodal nucleotide mismatch distribution suggested that L. affinis in coastal waters of China had experienced population expansion and the deduced expansion time was approximately 134.199~14.911 kaBP; i.e., in the late Pleistocene, which might have been caused by the rise and fall of sea-levels with dramatic climatic changes during glacial-interglacial cycles. The Dongtou population should be given priority protection as it has the highest genetic diversity (Hd = 0.8080±0.1130, π = 0.0021±0.0006) among the populations that were studied. Because the study only involved limited L. affinis populations in China, COⅠ might be a more conservative molecular marker than evolutionarily faster control region. Furthermore, maternally inherited mtDNA is independent of nuclear inheritance, and may not completely represent the evolutionary history of a species. Thus, future studies should incorporate more sampling sites across the species range using both nuclear and mtDNA markers together. A better understanding of the genetic background of L. affinis will provide a sound scientific basis for the preservation and sustainable utilization of its germplasm resources.

Abstract:This study aimed to develop a universal genotyping tool for common salmonid species cultured in China. Using an Affymetrix rainbow trout (Oncorhynchus mykiss) 57 K single nucleotide polymorphism (SNP) array, four representative aquaculture salmonid populations were genotyped, including Oncorhynchus masou masou, Oncorhynchus kisutch, Salvelinus fontinalis, and Salvelinus leucomaenis. A total of 96 shared polymorphism markers was screened out of 57,501 SNPs, then a low-density SNP array was constructed based on a Fluigidm 96.96 dynamic array, which is universal for the genera Oncorhynchus and Salvelinus. Reliability of the genotyping results was high because the identity of results with those of the Affymetrix 57 K array reached 96.55%. Parentage assignment for 48 Oncorhynchus kisutch offspring was performed with Cervus 3.0.7 software, based on genotyping data by the 96 SNP array. The results matched perfectly with the true pedigrees among complicated families, proving high accuracy in the pedigree assignment application of the 96 SNP array. The non-exclusion probability for the first parent (NE-1P) was 4.120×10–4 during single parent assignment, whereas the non-exclusion probability for the parent pair (NE-PP) was as low as 6.219×10–12. Genetic structure analysis for four salmonid populations was also performed using genotyping data of the 96 SNP array. Clustering results were in accordance with the taxonomic categories of the samples, whereas certain degrees of genetic introgression were observed among the populations, indicating that the genetic composition and relationships among populations were properly demonstrated in the analysis. All these results suggested that the developed 96 SNP array has great potential in genetic analyses for salmonid populations, which might provide helpful genome-wide information for plasm resource evaluation, selective breeding, high-quality fry production, and reasonable foreign strain introduction in the Chinese salmonid aquaculture.

Abstract:White spot syndrome virus (WSSV) is one of the most harmful pathogens in shrimp farming, which has great impacts by causing economic losses in aquaculture industry. It belongs to the Whispovirus genus of the Nimaviridae family. However, how the virus enter the cell is not very clear. Interaction between the virus and host is important for viral infection. Recently, some researchers found that GTP-binding proteins, such as Rab5, Rab6, and Rab7 interacted with infectious hypodermol and hematopoietic necrosis virus (IHHNV) and WSSV. However, the studies did not reveal the interaction between RAS protein and WSSV. The RAS protein belongs to the GTP-binding proteins and exists in eukaryotes from yeast to humans. Being a membrane protein, RAS becomes post-translationally modified after synthesis and is transported via the endoplasmic reticulum and Golgi complex route. Thus, in this study, the RAS of Litopenaeus vannamei was cloned. Then, it was ligated with prokaryotic expression vector pBAD/gⅢA using T4 DNA ligase, transformed into E.coli TOP10, and induced with L-arabinose. Pure RAS protein was acquired using Co2+ affinity chromatography purification. Mass spectrometry analysis showed the recombinant protein was RAS of L. vannamei. Far-western and ELISA assays were applied to determine the interaction of RAS with WSSV-VP26, WSSV-VP28N, and WSSV-VP37. The Far-western assay showed that the RAS protein interacted with VP26. The ELISA assay indicated that the interaction between RAS and VP26 grew stronger with the increasing concentration of RAS. Taken together, the RAS protein was responsible for WSSV infection. This study could provide the basis for research on the mechanism of WSSV invasion.

Abstract:The white spot syndrome virus (WSSV) is a highly virulent pathogen in crustaceans. The purpose of this study was to compare the WSSV-resistant traits of Pacific white shrimp Litopenaeus vannmei from different commercial brands. After detecting the infection status of eight different pathogens, including WSSV, infectious hypodermal and hematopoietic necrosis virus (IHHNV), taura syndrome virus (TSV), acute hepatopancreatic necrosis disease causing-Vibrio parahaemolyticus (VPAHPND), Yellow head virus (YHV), infectious myonecrosis virus(IMNV), Enterocytozoon hepatopenaei (EHP), and covert mortality nodavirus (CMNV), a quantitative feeding method to conduct a WSSV infection experiment on six commercial brands of juvenile L. vannamei (named Hainan Z, Hainan S, Guangzhou P, Guangzhou Z, Huanghua R, and Dongying M in the present study). The WSSV-resistant traits of each commercial brand were evaluated using the average survival time, survival rate, and cumulative mortality after infection. The results showed that no WSSV was found in shrimp of the six commercial brands, whereas some individuals were positive for EHP and CMNV. The average survival time for Hainan Z, Hainan S, Guangzhou P, Guangzhou Z, Huanghua R, and Dongying M was (150.18±25.92) h, (99.51±28.04) h, (122.32±43.59) h, (93.50±34.77) h, (100.99±26.84) h, and (71.70±23.79) h, respectively. The average survival time of Hainan Z was significantly higher than that of the other five brands. The average survival time of Guangzhou P was lower than that of Hainan Z, and was significantly higher than that of Dongying M, Guangzhou Z, Hainan S, and Huang Hua R. Dongying M had a significantly lower survival time than that of the other five brands. There was no significant difference between Guangzhou Z, Hainan S, and Huanghua R (P>0.05). The mortality peak for Hainan Z started on the 6th day and continued to the 7th day; whereas that of Dongying M was on the 4th day. The mortality peak for Hainan Z was 2 to 3 d later than that of Dongying M. At the end of the infection experiment, the survival rate of Hainan Z and Guangzhou P was the highest and was 72.5%. The survival rates for Dongying M and Huanghua R were the lowest, at 39.6% and 38.7%, respectively. The results suggested that Hainan Z and Guangzhou P have the strongest resistance against WSSV. The results could provide basic data for the selective breeding of L. vannamei.

Abstract:Intermediate cultivation of shrimp postlarvae (ICSP) is an important part of industrial shrimp farming. Full-scale ICSP experiment in an aquafarm (21 d) was conducted to investigate the effect of stocking density (15000~22500 pieces/m3) on the shrimp growth performance, aquaculture water quality, and bacterial community. The results indicated that the shrimp growth performance (Expressed as yield rate, special growth rate, survival rate, and feed conversion rate) was enhanced with the increasing stocking density when the stocking densities were 15000~22500 pieces/m3. For all aquaculture ponds, the pH values and the concentrations of ammonia, nitrite and COD gradually decreased and increased，respectively over the ICSP period, while Vibrio concentrations fluctuated in the range of (0.3~7.5)× 104 CFU/ml. Furthermore, higher stocking density resulted in lower pH and higher concentrations of ammonia and COD in the water. However, little effect of stocking density on Vibrio concentration was observed in this study. At the end of ICSP, the aquaculture pond with higher stocking density had higher bacterial biodiversity. Proteobacteria (56.52%~71.22%) and Bacteroidetes (20.65%~38.23%) constituted the main taxonomic groups (Phyla) of the bacterial community in all aquaculture ponds, and Vibrio (2.3%~9.4%) was the dominant genus. During the ICSP period, daily-increasing amounts of water exchange could control the decrease in pH and the increase in COD concentration to some extent, but failed to stem the rise of ammonia and nitrite concentrations.

Abstract:Hemoglobin is the main component of haemolymph, which is widely distributed in vertebrates and some invertebrates. It has many biological functions, including transporting oxygen, storing energy, sustaining osmotic pressure, immunological functions, and so on. According to previous reports, the functions of hemoglobin from different organisms are varied, especially in invertebrate animals, because of less conservation in gene structure. However, the study of the immune function of hemoglobin in mollusks is limited. The ark shell, Scapharca broughtonii, is one of the important economic marine shellfishes in China, Korea, and Japan, and other places. In particular, previous studies showed that there were three types of hemocytes in S. broughtonii: red blood cells, white blood cells, and thrombus cells. To determine the structure, distribution, and immunological activity of the hemoglobin gene, the cDNA sequence of hemoglobin (named SbHbⅠ) was obtained using EST and RACE methods. Sequence analysis revealed that the cDNA was 867 bp in length, including an open reading frame (ORF) of 483 bp encoding a polypeptide of 160 amino acid residues. The predicted molecular weight was 17.5 kDa and isoelectric point (pI) was 9.68. The mRNA expression analysis of SbHbⅠin tested tissues by quantitative real-time PCR (qRT-PCR) revealed that the mRNA of SbHb I could all be detected in the foot, gill, mantle, adductor muscle, haemocytes, hepatopancreas, and the larvae aged from oosperm to 23 d. The expression level of SbHbⅠin haemocytes was more than 70 folds that in the adductor muscle. The expression of SbHbⅠin the larvae from the oosperm to 23 d was basically showed a declining tendency at the beginning and increased later, with a sharp increase at 12 d (P<0.05). After being stimulated in Vibrio anguillarum, SbHbⅠfirst showed an upgraded tendency and later descended, and the same tendency occurred during hypoxia stress, during which SbHbⅠreached the maximum (P<0.05) at 16 h at three dissolved oxygen levels (4.5 mg/L, 2.5 mg/L, and 0.5 mg/L). Basic data were provided from the study of hemoglobin in mollusks.

Abstract:Methyltransferase (DNMT1) is an important gene that maintains the methylation state of the genome. This study uses the SMART-RACE technique to clone the DNMT1 gene of Portunus trituberculatus (PtDNMT1). The cDNA sequence of PtDNMT1 gene is 5919 bp, including the open reading frame of 4832 bp, and 1610 amino acids are encoded. The predicted molecular weight is 148.15 kDa, and the theoretical isoelectric point is 4.68. The structure prediction found that there are two special PtDNMT1 domain structures: the zf-CXXC zinc finger domain and the methyltransferase familial Dcm structure domain. The analysis of the evolutionary tree showed that PtDNMT1 gene and insects DNMT gather into one. PtDNMT1 was expressed in the hepatopancreas, gill, ovary, muscle, stomach, heart, and blood. The highest expression was found in the hepatopancreas, followed by the ovaries and gills. Then we get a further research about the PtDNMT1 gene expression during low salinity stress in gill, l hepatopancreas and muscle tissue. In gill tissue, we found that PtDNMT1 gene expression peaked (5 folds) after 6 h, and continued to 12 h (6 folds), then declined gradually, but was still significantly higher than the control group (3 folds). The expression of PtDNMT1 in the hepatopancreas was similar to the gills, however the peak time was later than in gill tissue (24 h), and the peak was higher than in the gills (8 folds). After low salinity stress, the expression of the PtDNMT1 gene in muscle was firstly reduced, then (24 h) increased to a peak (2.2 times), and the expression was raised to 72 h. This study is the first to clone the PtDNMT1 gene. According to its distribution characteristics and occurrence in various organizations, expression changes under salinity stress. We speculate that low salinity adaptations in DNA methylation in P. trituberculatus played an important role.

Abstract:Based on the high-density genetic linkage map and QTL of the sea cucumber (Apostichopus japonicus), we selected 26 candidate SNPs associated with body length, body weight, body breadth, pallet number, and disease resistance. Thirteen pairs of primers were successfully designed, which could be used for high resolution melting (HRM) detection. The 13 candidate SNPs associated with important economic traits were validated and analyzed with phenotypic data using genotyping of HRM in the expanded population. Polymorphic analysis results showed 3 loci were monomorphic sites and the other 10 loci possessed polymorphic minor allele frequency (MAF) at the 10 polymorphic sites, which ranged from 0.016 (SNP113) to 0.332 (SNP160), with an average of 0.173. Observed heterozygosity (Ho) ranged from 0.031 (SNP113) to 0.818 (SNP9), with an average of 0.433. Expected heterozygosity (He) ranged from 0.031 (SNP113) to 0.834 (SNP160), with an average of 0.402. The polymorphism information content (PIC) value ranged from 0.030 to 0.393, with an average of 0.284. Six loci departed from the Hardy Weinberg equilibrium. The results of QTL verification indicated that loci SNP40 and SNP160 associated with growth traits (body length, body weight, body breadth) with the dominant genotypes SNP40 (CC), SNP160 (AA). SNP88, SNP112, and SNP126 were associated with disease resistance. The dominant genotypes were SNP88 (CC), SNP112 (AA), and SNP126 (TT). Diplotypes were constructed based on the five SNPs and association analyses revealed that K1 (CC AA TT) was best for disease resistance, and S1 (CC AA) and S3 (CC AC) were dominant diplotypes for growth traits. These results provide basic data for marker-assisted selection in sea cucumber breeding.

Abstract:A feeding trial was conducted to evaluate the effects of vitamin D3 (VD3) on growth performance, body composition, and antioxidant capacity of the juvenile sea cucumber Apostichopus japonicus. Six isonitrogen and isoenergetic diets were formulated by adding 0 (D1), 250 (D2), 500 (D3), 1000 (D4), 2000 (D5), or 3000 (D6) IU/kg VD3 into the basal diet (measured value: 95, 334, 570, 1076, 2063, and 3081 IU/kg). Juvenile sea cucumbers with initial body weights of (15.43±0.14) g were fed for 56 days. Results showed that VD3 had no effect on survival rate (P>0.05) of the sea cucumber. Further, both the weight gain rate and specific growth rate were significantly improved, and those of the D1 group were significantly lower than the other groups (P<0.05). The ratios of intestine weight to body weight of the D3, D4, and D5 groups were significantly higher than that of the other three groups (P<0.05), but there were no differences among them (P>0.05). With increasing VD3, moisture, crude protein, and crude lipid contents of the body wall were not affected significantly (P>0.05), but crude ash contents of the body wall increased and thereafter declined, reaching the maximum in the D5 group. The contents of hydroxyproline of the body wall decreased initially and then increased, reaching the minimum in the D4 group (P<0.05). The deposition amount of VD3 in the body wall increased initially and then decreased. The D4 group showed the highest value. VD3 significantly increased the contents of calcium in the body wall (P < 0.05). Additionally, the phosphorus content increased at first and was maintained. The contents of phosphorus of the D4, D5, and D6 groups were significantly higher than that of the D1 group (P<0.05), but there were no differences among them (P>0.05). The contents of magnesium in the body wall increased in all groups by dietary VD3, and all experimental groups had significantly higher magnesium contents than that in the D1 group (P<0.05). The contents of iron increased initially and then decreased, and the contents in the D5 and D6 groups were significantly lower than those in the other groups (P<0.05). The contents of manganese decreased, but the difference between each group was not significant (P>0.05). With increases in dietary VD3 level, both the activities of alkaline phosphatase and total superoxide dismutase in the intestines of the experimental groups first increased and then declined, while the contents of malondialdehyde decreased. The total antioxidative capacity and catalase activity in the intestines of all groups were not significantly affected (P>0.05) by VD3. In conclusion, analysis by a WGR linear regression equation indicated that the optimal demand for VD3 in the sea cucumber (initial body weight 15.43 g) was 1587.5 IU/kg.

Abstract:In July 2016, a severe disease outbreak occurred in offshore-cage cultured black rockfish (Sebastes schlegelii) in Changdao County, Shangdong Province, which resulted in significant mortality. The main symptom of diseased rockfish was skin ulcers on the body. In order to determine the cause of the disease, pathogenic bacteria were isolated from diseased fish and identified by various methods including morphological observation, traditional physiological and biochemical tests, and gyrB gene sequence alignment. A dominant bacterial strain was isolated from ulcerative skin and was marked as BZ01. The strain formed a subtransparent colony on TSB solid medium plate and a green colony on TCBS selective medium plate. Its shape was short and rod-like with a single polar flagellum when observed by transmission electron microscopy (TEM). An artificial infection experiment was carried out to verify its pathogenicity to black rockfish. The results demonstrated that this strain of BZ01 had strong pathogenicity to black rockfish and could cause death. The artificial infected black rockfishes showed the same symptoms as those of natural infection. The value of LD50 was calculated as 2.07106 CFU/ml. Analysis of the gyrB gene sequence suggested that the strain of BZ01 was most closely related to Vibrio. In gyrB gene sequencing, it clustered into Vibrio rotiferianus in the phylogenetic tree with 100% confidence. Based on these results, this strain was identified as V. rotiferianus. At the same time, the drug sensitivity test showed that this strain of BZ01 was highly sensitive to tetracyclines (tetracycline, doxycycline), quinolones (norfloxacin, ofloxacin, fleroxacin, nalidixic acid), coumarin (streptonivicin), and peptidyl transferase (florfenicol), while medium sensitive or resistant to macrolides, peptides, sulfonamides, β-lactams, and aminoglycosides. This is the first report that V. rotiferianus could infect offshore-cage cultured rockfish in China, which would provide an effective reference for disease control in cultured S. schlegelii.

Abstract:To study the absorption, distribution, and accumulation of arsenobetaine in Antarctic krill (Euphausia superba) oil, Wistar rats were randomly divided into 3 groups and treated with 2.625 g/kg·bw of soybean oil, krill oil, or 100 mg/kg arsenobetaine added to krill oil. The plasma, liver, kidney and spleen of the rats were collected at 1 h, 2 h, 4 h, 6 h, 12 h, and 24 h after gavage, whereas the urine was also collected at 12 h and 24 h. The total arsenic content in the plasma, organs, and urine was detected by ICP-MS. The total arsenic contents in liver and kidney of rats were also measured at 7 d and 30 d after continuous gavage with the same dose of the lipids. The total arsenic contents in the plasma and organs showed no obvious changes at each time point between the krill oil group and the control group. The maximum content of total arsenic in plasma occurred at 1 h after gavage, whereas the total arsenic content in the liver, kidney, and spleen were all the highest at 6 h after gavage. At 24 h after gavage, the content of total arsenic in the liver, kidney, and spleen decreased to below 30% of the maximum. Within 24 h, 85% of the arsenic in the krill oil was passed out of the body in the urine. After continuous gavage for 7 days and 30 days, the total arsenic contents in the liver and kidney of the rats in each group showed no significant changes when compared with those of the control group. Arsenobetaine in the krill oil was rarely absorbed in rats and most passed out of the body in the urine within 24 h. No cumulative effects appeared after long-term feeding with the krill oil and the arsenobetaine.

Abstract:Bacillus hwajinpoensis SLWX2 is a bacterium strain isolated from maricultural environments that can effectively remove NH4+-N, NO2–-N, and NO3–-N from water. In this study, the effects of different nitrogen sources (NH4+-N, NO2–-N, and NO3–-N) and environmental factors (temperature, pH, C/N, and salinity) on growth and nitrogen removal were studied. The results showed that the growth of Bacillus hwajinpoensis SLWX2 was consistent with the removal of the three forms of inorganic nitrogen, which occurred mainly in the logarithmic growth phase. There was no delay in growth of the bacterial cells and NH4+-N removal, and no NO2–-N nor NO3–-N was produced when NH4+-N was used as the sole nitrogen source. Further, there was no accumulation of NH4+-N and NO3–-N when NO2–-N was used as the sole nitrogen source, but there was a long delay in growth and NO2–-N removal. NH4+-N was not detected, but NO2–-N first accumulated and then decreased when NO3–-N was used as the sole nitrogen source. The experimental results showed that the effects of the environmental factors on the growth of SLWX2 correlated with its ammonium removal function. The strain showed good growth and ammonium removal performance under the conditions of weak acids, neutral, and weak bases (pH 6~8.5), temperature 28~40℃, C/N 5~25, and NaCl 0~30 g/L. The optimal conditions for SLWX2 were temperature of 30℃, C/N 25, pH 8.0, and salinity 25. The strain SLWX2 showed high efficiency in removing NH4+-N, NO2–-N, and NO3–-N in water, and it has a wide adaptation range to environmental conditions, which suggests its huge potential in bio-removal of nitrogen in waste water from industries and aquaculture.

Abstract:To develop novel antiparasitic agents, five 1-methyl-1,2,3,4-tetrahydroisoquinoline derivatives were designed and synthesized. Cyclohexanecarboxylic acid chloride, benzoyl chloride, thiophene formyl chloride, acetylchloride, and chloroacetyl chloride were introduced into the 2-position of amido in the initial compound-1-methyl-1,2,3,4-tetrahydroisoquinoline. The chemical structures of all five synthesized compounds were identified by comparing spectral data (EI-MS, 1H NMR, and 13C NMR) with literature values, and their antiparasitic activity against Ichthyophthirius multifiliis theronts and encysted tomonts in vitro were also evaluated. Additionally, the acute toxicity of the five synthesized compounds against Erythroculter ilishaeformis was evaluated. The results indicated that all five synthesized compounds showed promising antiparasitic activity against I. multifiliis theronts and encysted tomonts; however, compound 1 [cyclohexyl-(1-methyl-3,4-dihydroiso-quinoline)-ketone] with the cyclohexanecarboxylic acid chloride group showed better antiparasitic activity than the other four compounds. The in vitro tests revealed that compound 1 could be 100% effective against theronts at a concentration of 24.0 mg/L. The median lethal dose (LD50) of compound 1 against theronts at 4 h was 16.4 mg/L. All encysted tomonts were killed when the concentration of compound 1 was 60.0 mg/L. The LD50 of compound 1 for E. ilishaeformis was 234.3 mg/L with a safe concentration of 64.1 mg/L. This study firstly demonstrated that compound 1 has potent antiparasitic efficacy against I. multifiliis, and it could be a good candidate drug for chemotherapy and control of I. multifiliis infections.

Abstract:The aim of this study was to evaluate the taste characteristics of Porphyra yezoensis during different harvest periods. Electronic tongue (INSENT taste analysis system) was applied to characterize the taste of lavers. The contents of free amino acids and flavor nucleotides in the first, second, fourth, and sixth harvest of lavers were measured. The electronic tongue data showed that the taste of lavers mainly consisted of umami, aftertaste of umami, saltiness, and bitterness. The umami value decreased successively in the first, second, fourth, and sixth harvest of lavers. The intensity of aftertaste of umami was the greatest in the second harvest of lavers, followed by the first and fourth harvest, and was significantly decreased in the sixth harvest (P<0.05). The saltiness intensity was higher in the first and second harvest of lavers. The intensity of bitterness was similar between the first and second harvest of lavers. The bitterness value was significantly lower in the sixth harvest of lavers (P<0.05). The astringency value was small, and there was no significant difference among the four samples (P>0.05). The contents of taste compounds and their contributions to the flavor were significantly different among the four laver samples. The contents of total free amino acids in the first, second, fourth, and sixth harvest of lavers were 4291.54 mg/100 g, 3485.29 mg/100 g, 3277.98 mg/100 g, and 1148.12 mg/100 g, respectively. Alanine and glutamic acid made the biggest contribution to the flavor. The content of inosine monophosphate (IMP) was relatively high in all flavor nucleotides. The taste active values (TAV) of IMP in the first, second, fourth, and sixth harvest of lavers were 4.13, 4.56, 2.19, and 2.33, respectively. The equivalent umami concentrations (EUC) of the first, second, fourth, and sixth harvest of lavers were 223.89 g MSG/100 g, 222.13 g MSG/100 g, 118.54 g MSG/100 g, and 47.19 g MSG/100 g, respectively, which means the umami of the earlier harvest of lavers was more intense. The electronic tongue analysis results are consistent with the experimental data of the flavor substances measurements. Electronic tongue technology could be used as a quick method to evaluate the taste of lavers and other aquatic products.

Abstract:The surimi was prepared with sturgeon as raw material. An optimal rinsing process suitable for sturgeon was obtained through orthogonal optimization experiments. Orthogonal experiments were carried out using water rinse times, salt rinse times, salt concentration, and rinsing time as factors and gel strength as the detection index. To determine the quality changes before and after rinsing of sturgeon surimi, in combination with the optimal rinsing process, the protein content (biuret method), moisture distribution (nuclear magnetic resonance), breaking force, deformation, gel strength (texture analyzer), water holding capacity, and whiteness (colorimeter) were measured. The results showed that the optimal rinsing process for the sturgeon surimi was as follows: rinse once in distilled water (1 min each), rinse once in 0.25% salt water (1 min each), and the sturgeon surimi gel had the best gel strength. After the optimal rinsing process, the salt-soluble protein content increased significantly, while water-soluble and insoluble protein content decreased. The relative increase in protein content indicated that extra impurities were removed and myofibrils proteins aggregated by rinsing. Breaking force, deformation, and gel strength all increased significantly after rinsing, indicating that the surimi after rinsing had better gelling properties. In addition, the sturgeon surimi had better water retention and whiteness after rinsing, which indicated that rinsing facilitated the formation of a dense network of surimi and removed some of the colored impurities. Concurrently, the NMR spin-spin relaxation time measurement results showed that the water distribution of the rinsed sturgeon surimi tended to have a stronger binding force and a smaller degree of freedom. This indicated that after the best process rinsing, the sturgeon surimi formed a dense gel network structure. In summary, rinsing could significantly improve the quality of sturgeon surimi. The research results provide references for improving the rinsing process of sturgeon surimi and enhancing the quality of surimi.

Abstract:Saccharina japonica is one of the most economically important seaweeds in China. Because of both internal and external forces, S. japonica farming in offshore waters has begun to emerge in China. Focusing on offshore aquaculture of S. japonica, this paper contains a discussion of the definition of inshore and offshore aquaculture of S. japonica, compares the advantages and disadvantages of these two aquaculture methods, and analyzes the driving forces of S. japonica offshore aquaculture. We also summarized the current situation and problems involved in S. japonica offshore aquaculture and proposed the development of a strategy, with the hopes that the discussion herein will provide new ideas and concepts for the healthy and sustainable development of S. japonica offshore aquaculture.

Abstract:The ovate pompano Trachinotus ovatus is a marine fish species with important economic value, whose culture was developed rapidly in recent years in China, and the annual cultured output has reached about 120,000 tons, which has become one of the most potential cultured marine fish in southern China. Up to now, there have been a lot of reports on its nutritional requirements and feed, however studies on some nutrients are still absent. This paper summarized the requirements of pompano on protein, amino acids, lipid, fatty acids, carbohydrates and micronutrient, dietary replacement of fish meal with other protein resources and fish oil with other lipid resources, as well as the application of functional feed additives. Available data showed that the optimum dietary requirements of juvenile golden pompano for protein, lipid and carbohydrate are 41%~49%, 6.5%~12% and 11.2%~16.8%, respectively. The optimal requirements for lysine, arginine, methionine, isoleucine, leucine, linolenic acid and arachidonic acid have also been determined. Preliminary studies on the replacement of dietary fish meal (FM) and fish oil (FO) with other protein or lipid resources have been conducted, and the results showed that the suitable substitution levels of FM by soybean meal (SBM) or fermented soybean meal (FSBM) are 20%~50%. When certain amount of FM was added in diet, it is feasible to replace FO with other animal or vegetable oils. The optimum dietary addition levels for soy isoflavone, chitosan and fermented product of Rhodotorula mucilaginosa are respectively 40 mg/kg, 4.0 g/kg and 500 mg/kg. However, studies on the requirements of vitamins and minerals are not reported. Besides, fewer studies were conducted on the nutritional requirements of fish during different growth stages or in different cultured patterns. We hope that this review can provide reference for the study of accurate nutrition and development of high-efficient, low cost and environment-friendly formula feed, and thus promote the healthy and sustainable development of T. ovatus aquaculture industry.