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Abstract:Bigeye tuna (BET), Thunnus obesus is a large epi- and mesopelagic species distributed in tropical and subtropical waters of the Indian Ocean. Its stock status has been the focus of regional tuna fisheries management organizations. Because of a variety of fishing gear and fishing fleet structures, there have some statistical biases in the historical nominal catches of the Indian Ocean BET. However, the impact of this bias on stock assessment has been neglected in recent years. This paper aimed to assess the impact of observation error and statistical bias of catch on the stock assessment of Indian Ocean BET, using Age-Structured Assessment Program (ASAP) based on fishery-specific catch, catch-at-age, and standardized catch-per-unit-effort data. The results showed that the current stock of BET in the Indian Ocean was not overfished. The results of base model and eight sensitivity analysis models showed that the observation error of catch had influences on the stock status evaluation. When the bias of nominal catch was assumed to be 15% (i.e., the historical catch was underestimated), the assessment result was consistent with the base model (i.e. not overfished). When the bias of nominal catch was assumed to be 20%, the stock might be overfished. Therefore, both the observation error and the statistical bias associated with catch data can have significant impacts on the assessment results, with the latter having a greater effect. This study highlights the importance of considering both the assumption of observation error and statistical bias in catch data in tuna fishery stock assessment, which has been neglected recently.

Abstract:In this study, fuzzy synthetic evaluation was used to build a reef ecosystem-health assessment model. To eliminate the dimension effect of sub-indexes, normalization processing was used. Analytic hierarchy process was used to assign a threshold of ecological indicator. The synthetic health index was used to assess the health status of artificial reef areas and contrast areas. Sub-indexes of artificial reef ecosystem health assessments included physical, chemical, and biological factors. Assessment standards were based on seawater quality standards, marine sediment quality, and guidance for the assessment of coastal marine ecosystem health. Data and survey results of a background investigation in July 2013, and a tracing investigation in June 2014 and May 2015, were processed using the ecosystem-health synthetic assessment model. The results indicated that both reef area and contrast area are in a healthy state. The health state of the contrast area was better than that of the reef area in July 2013 before reef construction. Data from June 2014 was consistent with that from the background investigation; the health status of the contrast area was also better, which is related to the disturbance of reef construction and the evolution of the reef system. In May 2015, the health index of the contrast area declined, but that of the reef area increased to 0.838, indicating that the reef is very healthy. Health evaluation indicates that the health of the reef ecosystem in the Beidaihe demonstration area is gradually improving. According to the results obtained with the ecosystem health synthetic assessment model, the state of the reef improved with completion of construction and ecosystem stability; therefore, ecosystem health synthetic assessment is an important method for the management and development of reefs. Researching reef ecosystem health and scientifically assessing artificial reef ecological effects are significant for the construction of reef projects, environment protection, stock enhancement, efficient administration, and sustainable development of fisheries.

Abstract:β-defensins are cysteine, cationic, antibacterial peptides that play very important roles in the vertebrate immune system. In this study, the cDNA fragment of β-defensin gene from Acipenser gueldenstaedti was cloned using a PCR method based on the Solexa sequencing results of gonad transcriptome, and gene expression was performed by qRT-PCR. The results showed that cDNA of β-defensin was 333 bp in length and contained an open reading frame (ORF) with 213 bp, which was deduced to encode a precursor peptide of 71 amino acids consisting of a signal peptide of 23 amino acid residues and a mature peptide of 48 amino acid residues. Russian sturgeon β-defensin possessed six conserved cysteine residues, forming three disulfide bridges at C38~C66, C44~C60 and C48~C67, and containing 1 defensin-β-2 domain. This structure is very similar to other species and conserved in evolution. Homologous alignment revealed that Russian sturgeon β-defensin had the highest sequence identity with fish species, about 54%~60%. qRT-PCR analysis showed the β-defensin transcript is constitutively expressed in 11 tissues (liver, intestine, spleen, head-kidney, gill, blood, brain, skin, muscle and gonad, stomach) in healthy fish, with highest expression in the gonads and skin. Artificial infection of A. gueldenstaedti with Aeromonas hydrophila resulted in a significantly upregulated expression in the intestine, spleen, head kidney, blood, and gill while downregulating expression in the liver. The most significant upregulated expression and the peak level at 72 h reached 700 times baseline in the head-kidney. The results indicate Acipenser gueldenstaedti β-defensin genes are involved in the immune response.

Abstract:Based on 1022 bp and 1322 bp sequences, partial nucleotide sequences of Cyt b and D-loop segments of mtDNA were analyzed for Coilia nasus, C. nasus taihuensis, C. grayii, and C. mystus to develop a reference dataset for species identification in the genus Coilia. The genetic distances were 0.00360±0.00083 and 0.00378±0.00082 between C. nasus and C. nasus taihuensis, 0.12146±0.01106 and 0.10753±0.01076 between C. mystus and C. nasus, 0.12207±0.01111 and 0.10674±0.01074 between C. mystus and C. nasus taihuensis, 0.03418±0.00561 and 0.05267±0.00686 between C. nasus and C. grayii, 0.03506±0.00568 and 0.05287±0.00698 between C. nasus taihuensis and C. grayii, as well as 0.11585± 0.01109 and 0.11233±0.01114 between C. grayii and C. mystus, respectively. Based on the sequences of the Cyt b gene and D-loop and the neighbor-joining (NJ) molecular dendrogram using Kimura 2-parameters, individuals of C. nasus and C. nasus taihuensis were combined in a cluster while C mystus and C. grayii formed another two clusters. First, based on sampling sites, the identities of the unknown larvae from the estuary of the Yangtze River and unknown juveniles from the Taihu Lake could be deduced preliminarily as C. nasus and C. nasus taihuensis, respectively. Then, nucleotide sequences of Cyt b and D-loop of larvae and eggs of genus Coilia were analyzed and compared with the reference established above. Genetic distance ranged from 0.0024 to 0.0032 and 0.0025 to 0.0082, respectively, within the larva and egg individuals collected from different places. The NJ molecular dendrogram showed that all the unknown larvae, the eggs of C. nasus, adult C. nasus, and C. nasus taihuensis clustered together. In contrast, individual eggs of C. mystus and adult C. mystus formed a separate cluster. Thus, Cyt b and D-loop sequence analysis could be used to identify unknown Coilia nasus, C. grayii, and C. mystus at early life stages, but could not effectively be used to identify unknown individuals at early life stages between C. nasus and C. nasus taihuensis.

Abstract:Grass carp haemorrhage is an epidemic disease, which is caused by grass carp reovirus (GCRV). Grass carp fingerlings and yearlings are vulnerable to GCRV. A virulent reovirus, tentatively named GCRV-AH528, was isolated from a diseased grass carp exhibiting typical hemorrhage symptoms in Hefei, Anhui Province. Intraperitoneal injection with the virus suspension led to hemorrhage similar to the observed clinical symptoms of blackened dorsum and hemorrhage around the mouth cavity, gill, intestine, belly, and base of the fin ray. The GCRV-AH528 genome contained 11 double-stranded RNA segments ranging from 1027 to 3925 bp, and its average content of AT and GC was 50.2% and 49.8%, respectively. Compared with registered GCRV Ⅱ strains, the GCRV-AH528 L1 segment had lost three continuous nucleotides (TAT) at nt 701~702, leading to the loss of a tyrosine, and the M4 segment mutated two ORFs encoding two non-structural proteins (NS9 and NS69). All segments had 6 bp-conserved terminal nucleotides 5'-GUAAU/CUžžžUU/GCAUC-3'. Except for the L1 and M6 segments, the two ends in the coding region of the other nine segments existed at 5~9 bp short specific inverted complementary sequences. GCRV-AH528 showed 37.1%~98.1% nucleotide and 24.3%~98.3% AA sequence identities with other reovirus. A phylogenetic tree based on VP1 protein revealed that GCRV-AH528 clustered with members of the genus Aquareovirus and was far from GCRV-873. These results indicated that the GCRV-AH528 isolate was a new GCRV Ⅱ type virulent strain.

Abstract:Cytogenetic analysis is a valuable tool for germplasm study and protection. In order to clarify the Germplasm resources of Epinephelus moara, the metaphase chromosomes of E. moara of Tokyo Bay population were obtained from the head kidney tissue by the method of PHA and colchicine injection and the karyotype was analyzed by using microscopes. We observed 464 mitotic metaphases from 6 individuals, and 356 of all observed mitotic metaphases (76.72%) showed 2n = 48 chromosomes, which indicated that there were 48 chromosomes in the diploid. We then analyzed 10 mitotic metaphases and calculated the relative length and arm ratio of the chromosomes. The relative length of the chromosomes in E. moara of Tokyo Bay population varied from (5.13±0.15) % to (2.81±0.13) %. The arm ratio of the 14th chromosome was 2.35±0.14, and the arm ratio of the 16th chromosome was 2.71±0.24. The 24th chromosome had the minimum arm ratio, which was 1.99±0.13. According to arm ratio, the diploid consisted of 6 submetacentric, and 42 telocentric chromosomes, and the fundamental number of chromosome arms was 54, which indicated karyotypic formula in E. moara of Tokyo Bay population was 2n = 48，6sm + 42t，NF = 54. The form of the 24th chromosome was changed in different metaphase from the same individual (the shorter arm changed among filiform, T type, and SM type), which may be related with the expressed activity of the rDNA in nucleolar organizing region. By comparison with the chromosome karyotypes of other Epinephelus species, E. moara has the evolutive characteristics of karyotypic pattern and belongs to the specialization group. As detected, the karyotypes of E. moara were different between Japan (Tokyo Bay) and China (Xiamen coastal waters) samples, which may be caused by geographic isolation. There were abundant genetic diversity among different geographic populations, and the germplasm improvement of E. moara could be accelerated by heterosis among different geographic groups.

Abstract:Thamnaconus septentrionalis is a common economic marine fish species that can be found in coastal waters of China. The artificial breeding technique has been established for T. septentrionalis. In order to optimize artificial culture techniques and to promote industrialization of T. septentrionalis, path analysis was conducted to investigate the effects of morphological traits on body weight. According to body weight, the samples were divided into 3 groups (small size, medium size, and large size). Full length (FL), body length (BL), body height (BH), head length (HL), snout length (SL), eye diameter (ED), caudal peduncle length (CL), caudal peduncle height (CH), and body weight(BW) of T. septentrionalis were measured. We calculated the correlation coefficients, path coefficients and determination coefficients. The regression equations with morphological traits as variables and body weight as the dependent variable at each size were established. The results showed that the morphological traits affecting body weight at different sizes were different. Body height, body length, eye diameter, and caudal peduncle length were the significant traits affecting body weight at small sizes; for medium T. septentrionalis, the morphometric traits that significantly affected body weight were body length, caudal peduncle height, head length, body height, full length, and caudal peduncle length; for large T. septentrionalis, the significant traits were body height, full length, and caudal peduncle height. The regression equations for small, medium and large size: Y=–84.753+17.263XBH+6.572XBL–28.928XED–11.353XCL; Y=–325.824+20.321XBL+94.047XCH–11.425XHL +15.713XBH–7.218XFL+15.744XCL; Y=–516.433+38.012XBH +10.545XFL+82.752XCH. The sums of the coefficients of determination at each size were all greater than 0.85. This indicates the main morphological traits affecting body weight were found. This study can serve as a valuable tool to improve the culture industry of T. septentrionalis.

Abstract:To investigate the effect of starvation on amino acids and fatty acids of muscle in juvenile large yellow croaker Larimichthys crocea, 540 healthy fish with similar body weight (40.80±3.40 g) were selected for the experiment. The muscle was sampled from experimental fish after 0, 4, 8, 12, 16, and 20 days of starvation for chemical analysis, respectively. Results showed that starvation significantly affected methionine content (P<0.05), which reached the lowest content (0.48%±0.12%) after 8 days of starvation and obtained the highest content (1.62%±0.23%) after 16 days of starvation. However, the other 15 kinds of amino acids showed no significant difference among the different groups (P>0.05), with the same trends of increased at first and then decreased when starvation time was prolonged. Also, the non-essential amino acids, essential amino acids, flavored amino acids, essential amino acids and total amino acids showed the same changing trends as the 15 kinds of amino acids. As for fatty acid, starvation had significant effects on the fatty acid content in muscle of juvenile L. crocea (P<0.05). Among them, the saturated fatty acids (SFA) increased first and then decreased with the prolongation of starvation, and reached the maximum content after 8 days of starvation (30.90%±0.28%); monounsaturated fatty acid (MUFA) content among different treatment groups was not significantly different (P>0.05), but all the treatment groups were slightly larger than that of the control group (starvation for 0 day); polyunsaturated fatty acid (PUFA) content showed an increasing trend after the first drop, and the lowest value was in the group of that was starved for 12 days (31.87%±0.65%). From the above research results, we know that the amino acid and fatty acid contents in the muscle related to meat quality can be changed by proper starvation treatment, which can improve the meat flavor of L. crocea. These results have important theoretical and directive significance for improving meat quality of L. crocea through proper starvation treatment in the process of L. crocea culturing.

Abstract:The pen shell, Atrina pectinata, has high commercial and scientific research value because of the lack of large-scale culture in China. In recent decades, many problems such as overfishing, deterioration of marine eco-environment, and failure of artificial seedling breeding, have drastically reduced the amount of wild sources. However, there are few reports on A. pectinata systematic classification and no karyotype reports until now. To explore the cytogenetical characters and identify the status of germplasm resources of A. pectinata in northern China, by adjusting the water temperature of temporary rearing, and using the healing hyperplasia tissue of gills for chromosomal investigation, we improved the methods of chromosome preparation, and obtained many well-spread mitotic chromosomal plates of seven male and five female A. pectinata. The results revealed that a certain degree of temperature-change stress during temporary rearing could promote the proliferation of somatic cells effectively in the short term; gill-healing hyperplastic tissue has mass proliferation of cells in mitotic metaphase. The karyotypes of both sexes were examined separately. A. pectinate has 17 pairs of chromosomes (i.e. n=17, and 2n=34); there were obvious differences in karyotype between males and females; male karyotype formula is 2n=8m+10sm+16st；female karyotype formula is 2n=6m+10sm+18st. The first pair of the particularly larger chromosomes were heterotypic in male somatic cells; the No.14 pair of chromosomes was metacentric in males but telocentric in female metaphase somatic cells; the primary sex-determination mechanism is XX/XY type. The newly improved methods for chromosome preparation could be beneficial to large-scale shellfish germplasm identification and classification.

Abstract:There still exists considerable controversy regarding the taxonomic status of different geographical populations of Scapharca broughtonii in Northeast Asia. To study the taxonomic status of the Chinese population of S. broughtonii, the mitochondrial genomic COI and 12S rRNA sequences of the Chinese population of S. broughtonii were determined using PCR amplification and sequencing. To analyze the molecular phylogenetic relationships and classification, the homologous sequences of 26 species of shellfish were downloaded from GenBank, the molecular phylogenetic trees of COI and 12S rRNA of Arcidae molluscs were constructed using the neighbor-joining and maximum-parsimony methods with Mytilus edulis as the outgroup taxa. The results showed that the base composition, codon utilization, and mutation locus of COI and 12S rRNA genes were similar to those of the Japanese population of S. broughtonii and A. sativa, while the genetic distance was small, showing the genetic relationship was close. The classification status of the Chinese population of S. broughtonii was consistent with that of S. broughtonii. The purpose of this research was to better understand the arkshell genetic background and resources, to protect the natural resources, and promote the development of aquaculture.

Abstract:This experiment was conducted to evaluate the effects of replacing soybean oil with rubber seed oil on growth performance, digestive enzyme activities, serum lipoprotein levels, protein metabolism indices and antioxidant capacities of grass carp (Ctenopharyngodon idellus). Five isoproteic and isolipidic experimental diets were formulated with rubber seed oil replacing 0%, 25%, 50%, 75%, and 100% of soybean oil (R0, R25, R50, R75, and R100), respectively. The results showed that the weight gain, daily gain coefficient in fish fed R0 and R50 diets were significantly higher than those in fish fed the R100 diet (P < 0.05). With increasing rubber seed oil replacement level, the feed coefficient first decreased and then increased, and the lowest value was observed in fish fed the R25 diet, which was significantly lower than that in fish fed the R50, R75, and R100 diets (P<0.05). Conversely, protein efficiency first increased and then decreased, and the highest value was observed in fish fed the R25 diet, which was significantly higher than that in fish fed other diets (P<0.05). With increasing rubber seed oil replacement level, intestinal trypsin and lipase (LPS) activity first increased and then decreased, and the highest values of intestinal LPS activity were observed in fish fed the R25 diet, which were significantly higher than that in fish fed other diets (P<0.05). The highest values of intestinal trypsin activity were observed in fish fed the R50 diet, which were significantly higher than that in fish fed R0, R75, and R100 diets (P<0.05). Hepatopancreas trypsin activity in fish fed R0 and R50 diets was significantly higher than that in fish fed R75 and R100 diets (P<0.05). The serum total cholesterol (TC) level in fish fed R25, R50, R75, and R100 diets were significantly lower than that in fish fed the R0 diet (P<0.05). With increasing rubber seed oil replacement level, hepatopancreas hepatic aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities first increased and then decreased, and the lowest value was observed in fish fed the R25 diet, which was significantly lower than that in fish fed R0 and R100 diets (P < 0.05). Further, plasma total antioxidant (TAC) capacity in fish fed R0 and R25 diets was significantly lower than that in fish fed R75 and R100 diets (P<0.05). Conversely, plasma and hepatopancreas malondialdehyde (MDA) levels in fish fed the R0 diet were significantly higher than in fish fed the R100 diet (P<0.05). These results indicated that replacing 25% to 50% of soybean oil with rubber seed meal had no obvious negative effects on growth performance, feed utilization rates, serum lipoprotein levels, protein metabolism indices, or antioxidant capacities, whereas replacing 50% of soybean oil with rubber seed oil may have depressed the growth performance and digestive enzyme activities of grass carp.

Abstract:pH is an important environmental factor affecting the survival, growth, and immunity of aquatic animals. In general, alteration of environmental conditions often results in pH fluctuation in the aquaculture water of Apostichopus japonicus Selenka. However, little is known about the physiological response of sea cucumbers under pH stress. Therefore, the survival rate, growth, and antioxidant enzyme activities of sea cucumbers (7.02±0.81 g) were analyzed under different pH (control group, pH=8.4), low pH (pH 6.8, 7.0, 7.2, 7.4, 7.6, and 7.8), high pH (pH 8.6, 8.8, 9.0, 9.2, 9.4, and 9.6)) stress for 36 days. The survival rate of sea cucumbers decreased gradually with pH intension increasing and time extension, which in the groups ranging from pH 7.4 to pH 9.0 were 100%. There appeared to be a stress response and then death of individuals in the groups of pH 6.8, 7.0 and 9.6 from the third day, in which the whole group of sea cucumbers died after 30 days. Under different pH stress, the growth of the sea cucumbers differed significantly. The specific growth rate decreased with pH intension increasing and growth was negative growth in the group of pH 9.0. The superoxide dismutase (SOD) and catalase (CAT) activity of sea cucumbers among the pH treatment groups increased and was significantly higher than those of the control group, which increased at first and then decreased with pH intension increasing. The activity of SOD and CAT of sea cucumber in low pH groups increased to (74.92±2.24) U/ml and (14.99±2.38) U/ml peaking at pH 7.4, which was significantly higher than the control group. While in high pH groups, the activity of SOD and CAT of sea cucumber increased to (72.90±1.10) U/ml and (15.68±0.89) U/ml, peaking at pH 8.8 and pH 9.0 respectively. The results indicate that the pH range from 7.4 to 9.0 is appropriate for survival and growth of sea cucumbers, otherwise stress responses and even death of sea cucumbers are imminent.

Abstract:Peanut-worms Sipunculus nudus are exploited as a popular seafood in international and domestic markets due to high nutritional and pharmaceutical value. In recent years, over-exploitation of S. nudus has caused that natural resource to be degraded greatly, therefore study on S. nudus respiratory metabolism needs to be carried out to better understand the culture capacity and sustainable development of S. nudus resources. The effects of suspended feed concentration (5 mg/L, 10 mg/L, 15 mg/L, 20 mg/L, 25 mg/L) and body weight S(1.58±0.48 g), M(4.90±1.06 g) and L(7.96±0.94 g) on oxygen consumption rate (OR) and ammonia excretion rate (AR) rate of Sipunculus nudus Linnaeus were studied using experimental ecology methods. The results indicated that suspended feed concentration had significant effects on OR and AR (P<0.05), and interaction of the both factors had a significant effect on OR and AR (P<0.01). When feed concentration reached 15 mg/L, the OR and AR decreased significantly. Both the OR and AR decreased with the increasing feed concentration. The relationship between OR or AR and suspended feed concentration can be represented by the cubic function y＝ax3＋bx2+cx+d, where a values of OR or AR ranged in 0.003~0.033 and 0.0003~0.004, b values ranged in –1.698~–0.053 and –0.172~–0.003, c values ranged in –1.805~18.873 and –0.326~1.972, d values ranged in –15.675~79.940 and 3.634~14.008. Both the OR and AR increased with increasing body weight. The relationship between OR and body weight can be represented by the exponential function y=aebx, where a values ranged in 8.082~20.934 and b values ranged in 0.217~0.270. The relationship between AR and body weight can be represented by the power function y=axb, where a values ranged in 1.441~3.598, and b values ranged in 0.521~0.747. The O/N value ranged from 1.26 to 10.60. The study showed that high suspended feed concentration had a negative influence on S. nudus; once it reached 20 mg/L, the OR and AR fluctuated and declined significantly

Abstract:In this study, the effect of beta-glucan on survival rates and the mRNA expression of Lectin and TLR2 in both uninfected and infected Manila clams Ruditapes philippinarum were studied. The results showed that beta-glucan could effectively improve the survival rate of Manila clams infected by Vibrio parahaemolyticus. The survival rate was highest at a beta-glucan concentration of 1000 mg/L. In the gill tissues of the uninfected group (A1 and A2), the TLR2 peaked at 6 h, which was significantly higher than that of other times (P<0.05). In the infected group, TLR2 increased to a peak at 1.5 h and then decreased. Lectin expression in both the infected and the uninfected group increased first and then decreased. The relative expression of Lectin in the A1 group was significantly higher than that of the B1 group at 3 h (P<0.05). In the mantle, the expression of TLR2 in both infected and uninfected groups decreased gradually between 3~12 h. At 24 h, the expression of group A2 was highest. However, in the infected group, the expression of Lectin in 1000 mg/L beta-glucan was higher than at 100 mg/L in the mantle, but there was only a significant difference at 0 h and 24 h (P<0.05). The expression patterns of Lectin in the gill and in the mantle were different, but the feeding of beta-glucan promoted the expression of Lectin during the early stages of infection. From these results, beta-glucan soaking can increase the relative expression of the two genes, and TLR2 and Lectin are expressed more quickly after infection by V. parahaemolyticus when soaked with beta-glucan. The aim of this study is to understand the dose dependent effect conferred by beta-glucan on the immune system and the survival rate of Manila clams, which might provide some theoretical basis for the stock culture, seed breeding and disease control in pond culture of Manila clams.

Abstract:Using the toxicity method, we studied the acute toxicity of ammonia nitrogen to Scylla paramamosain and its influence on phenol oxidase (PO), alkaline phosphatase (AKP), acid phosphatase (ACP), superoxide dismutase (SOD) , and lysozyme (LZM) in the conditions of different ammonia nitrogen concentrations of 0 (C0, control group), 10 (C10), 20 (C20), 30 (C30), 40 (C40), 50 mg/L (C50) and different times of 0, 6, 24, 48, and 72 h. The results showed that the semi-lethal concentration (LC50) at 24 and 48 h of ammonia-N were 104.793 and 66.124 mg/L respectively, and the safe concentration (SC) was 7.90 mg/L. The LC50 at 24 and 48 h of NH3-N were 8.396 and 5.298 mg/L respectively, and the SC was 0.63 mg/L. The LZM activity of each treatment group was significantly lower than that of C0 group (P<0.01) at 6, 24, 48, and 72 h. AKP and ACP activity of C10, C20, and C40 groups was significantly higher than that of control group at 24 h (P<0.05), while the SOD activity of the C20 group was significantly lower than that of other stress times (P<0.05). At 72 h, the PO activity of C30, C40, and C50 groups were significantly higher than that of C0 group (P<0.05). Under the experimental conditions, ammonia nitrogen that was lower than 40 mg/L could significantly enhance the AKP and ACP activity of S. paramamosain over a 24 h period, while 50 mg/L ammonia nitrogen had inhibitory effects. Ammonia nitrogen in all groups had significant inhibitory effects on LZM activity of S. paramamosain, which had no significant effect on SOD activity, but had an obvious enhancement effect on activity of PO by 72 h.

Abstract:Serine hydroxyl methyltransferase (SHMT) plays an important role in plant response to stress. Under adversity stress, the SHMT gene not only promotes the Calvin cycle to participate in the anti-stress reaction, but also participates in the synthesis of glycine, and then synthesis of glutathione, scavenging reactive oxygen species. Pyropia haitanensis, naturally grows in the coastal intertidal zone, is an important economic macroalgae in Fujian, Zhejiang and Guangdong provinces. Because of the special living environment, P. haitanensis has the unique ability to resist adversity. With global warming, the temperature of seawater increases, which causes damage to seedlings of P. haitanensis and leads to great economic losses in the aquaculture industry. As for the physiological and molecular response of P. haitanensis to high temperature and dehydration stress conditions, our previous studies found SHMT plays an important role in stress resistance. In this study, based on unigene sequences which were obtained from whole transcriptome sequencing of P. haitanensis, two full-length PhSHMT genes were obtained by ordinary PCR, and named PhSHMT-1 (GenBank accession No: MF687405) and PhSHMT-2 (GenBank accession No: MF687406). The full-length cDNA of the PhSHMT-1 gene comprised 1710 nucleotides and contained an open reading frame (ORF) of 1491 bp, encoding a protein of 497 amino acid residues, with the predicted molecular weight of 121.443 kDa and theoretical isoelectric point of 4.93; and the full-length cDNA of the PhSHMT-2 gene comprised 1957 nucleotides and contained an ORF of 1395 bp, encoding a protein of 465 amino acid residues with the predicted molecular weight of 113.969 kDa and theoretical isoelectric point of 4.95. Multiple sequence alignment and phylogenetic analysis indicated that the PhSHMT-1 and PhSHMT-2 belong to the plant SHMT enzyme family. The expressions of the two PhSHMT genes, as measured by real-time fluorescence quantitative PCR (qRT-PCR), were consistent under high temperature stress. During high-temperature stress, the expression levels of the two PhSHMT genes were significantly upregulated first and then decreased, but as the high-temperature stress continued, the expression levels were upregulated again. Results suggested that the gene of PhSHMT enzyme plays an important role in responses of P. haitanensis to high-temperature stress.

Abstract:Streptococcus agalactiae has become one of the most important emerging pathogens with strong infectivity and virulence, causing high mortalities and large economic losses in tilapia farming industries in China. There are no obvious symptoms in some tilapia infected by S. agalactiae, so it can be difficult to determine if a fish carried the pathogen. Thus, it was necessary to establish an effective method for detection of S. agalactiae in the breeding process of tilapia. Three pairs of specific primers were designed based on the conserved sequence of hylB, ponA, and cfb genes encoding hyaluronidase, penicillin binding protein, and CAMP factor of S. agalactiae published in GenBank, respectively. After the optimization of the reaction conditions and reaction system of multiplex PCR, the triple PCR method based on the three virulence genes was developed for detection of S. agalactiae isolated from tilapia. Furthermore, the established method was applied to detect tissue samples of tilapia collected from different farming areas in Guangdong Province. The detection result of S. agalactiae only amplified three specific bands, however there were no bands in the host and the other common bacterial pathogen strains in aquaculture, which indicated that the method had good specificity. The dynamic range of template concentration of S. agalactiae was 7.24×10-5~5.65 ng/μl. Sensitivity tests showed that the detection limit of the S. agalactiae in genomic DNA was 1.81×10-3 ng/μl, with a higher sensitivity. The positive rate for detection of 188 tissue samples isolated from tilapia using the triple PCR method was consistent with the positive rate by the conventional bacterial identification method. The triple PCR method was evaluated with the conventional method as the standard; diagnostic sensitivity (Dse) and diagnostic specificity (Dsp) were 100%. In summary, the results showed that the triple PCR method not only improved the accuracy and sensitivity of the detection, but also detected three virulence genes of S. agalactiae in the same reaction system. Therefore, this method provided a rapid, accurate, and efficient detection technology for monitoring S. agalactiae infection in harmless aquatic products and early warning of aquaculture diseases.

Abstract:Vibrio anguillarum is a pathogen that can infect a variety of fish and causes highly fatal hemorrhagic septicemia. Its O1, O2, and O3 serotype strains are prevalent in the mariculture environment of China, causing great economic loss. However, the study of polyvalent vaccines of different pathogenic serotypes of V. anguillarum has not been undertaken. For this purpose, a trivalent inactivated vaccine V. anguillarum was made, and its antibody duration and immune protection period were evaluated. The trivalent vaccine was administered to turbot (80.2±4.7 g) at a dose of 108 cells/individual via intraperitoneal injection; booster vaccination was given at 60 days post vaccination (dpv), and the serum antibody titer and RPS of the vaccinated group and control group were compared. Fish vaccinated only once were observed for 150 days, and the results showed that during 14~150 dpv, the antibody level of the vaccinated V. anguillarum group was significantly higher than that in the control group (P<0.05), and rose to 1∶6.7 at 7 dpv, reached peak value at 28 dpv, and during 28~150 dpv hovered between 1∶106.7~1∶320. RPS of three serotype V. anguillarum were 16.7%~43.8% for 7 dpv, then reached 100% at 28 dpv; during 28~120 dpv was 70%~100%, 35%~100% for 150 dpv. The fish vaccinated twice were observed for 90 days; during 3~60 days post-second vaccination (dpsv), their antibody titer was significantly higher (P<0.05) than those vaccinated only once, then went down and was not significantly different (P>0.05) than that of those vaccinated only once. RPS of three serotype V. anguillarum were still higher than 70% and higher than those of fish only vaccinated once. All in all, antibody durability and efficient immune protection period (RPS>70%) of fish vaccinated with V. anguillarum trivalent inactivated vaccine once was longer than 150 days and 120 days respectively, and 150 days for fish that were vaccinated twice. Control group fish reached higher weight in comparison to the vaccinated fish, while SGR between groups for the whole period was not statistically significant (P>0.05).

Abstract:In order to obtain a higher yield of α-cyclodextrin conversion, we use a single factor experiment and response surface experiment method, to optimize the production of α-cyclodextrin conditions by α-cyclodextrin glucosyltransferase. The enzyme is derived from strain Y112. First, the single factor method was used to screen six factors including substrate species, substrate concentration, enzyme content, reaction time, temperature, and pH. Second, these main variables affecting the conversion of α-cyclodextrin were evaluated with the Plackett-Burman test. The results suggested that substrate concentration, temperature, and pH were the most impactful factors. Finally, the optimal levels of the three factors were determined with central composite design and Design-Expert 8.05 software. The optimum conversion conditions were shown as follows: the concentration of potato starch was 5%, the amount of enzyme 200 U/g (starch), pH 8.4, 200 r/min, reacted at 30℃ for 6 h. Under the optimized conditions, the average value of α-CD conversion was 28.67%, which was an increase of 2.48 times compared with the pre-optimization yield.

Abstract:The effects of reaction temperature, pH, and metal ions (Zn2+, Mg2+, Cu2+, Fe3+, Al3+, and Ca2+) on the protease activity of crude enzymes of Antarctic krill were studied, and the inhibitory effects of phenylmethanesulfonyl fluoride (PMSF), EDTA∙2Na, iodoacetamide (IAM), and toluenesulfonyl- phenylalanine chloromethyl ketone (TPCK) on protease activity also were investigated. The experimental results indicated the optimum reaction temperature of the protease in the crude enzyme of Antarctic krill was 40℃, and the optimum pH was 8.0; when the metal ion concentration was 0.5 mmol/L, the inhibitory rates of Zn2+, Mg2+, Cu2+, Fe3+, and Al3+ on the protease activity of Antarctic krill were 55.02%, 55.02%, 35.39%, 20.67%, and 41.12% respectively, while protease activity increased in the presence of Ca2+. The protease activity was significantly inhibited by PMSF and EDTA∙2Na. When the concentration of PMSF was 8.0 mmol/L, the inhibition rate was 60%; when the concentration of EDTA∙2Na was 0.6 mmol/L, the inhibition rate was 86.67%. IAM had a certain inhibitory effect at low concentrations, while TPCK showed no inhibitory effect. With the increase in temperature, the inhibitory effect of EDTA∙2Na on enzyme activity was gradually increased in the temperature range of 5–30℃. This study clarified the factors affecting the enzyme activity of Antarctic krill crude enzyme and developed an inhibitor of protease activity thereof, which provides the basic theoretical data for the development and utilization of Antarctic krill in the food industry.

Abstract:To explore the residual characteristics and sources of perfluorinated compounds (PFCs) in Monopterus albus within the middle and upper reaches of the Yangtze River, 150 M. albus were collected from 10 cities in Hunan, Hubei, Jiangxi, Anhui, Sichuan, and Chongqing. The source of PFCs in the M. albus was investigated in 2016 in the eel breeding base of Qianjiang City, Hubei Province. The samples were extracted and purified using C18 and carbon black (GCB). The contents of 19 PFCs were analyzed using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The results showed that there were 8 residual PFCs in the eels of the Yangtze River, and the main pollutants were perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS). PFCs in the Yangtze River basin were the most polluted in Hubei Province, followed by Jiangxi Province. The highest content of PFOS in Hubei was 5.42 μg/kg, and PFOA were the highest in Sichuan Province at 3.66 μg/kg. Meanwhile, in the entire farming process, combined with the special breeding methods of M. albus, we found that the pollution of PFCs in M. albus mainly originated from earthworm feeding and the breeding environment, including pond water and soil.

Abstract:In this experiment, the water temperature was raised at (26±2)℃. The experiment used high performance liquid chromatography tandem mass spectrometry(HPLC-MS/MS) to study thiamphenicol in pharmacokinetics of carp with 30 mg/(kg·bw) single dose of carp feed. Data were analyzed using DAS 2 dynamics software, and the experimental results suggest a two-compartment model of primary absorption. The peak of thiamphenicol (TAP) in muscles, kidneys, liver, skin, gills, spleen, and plasma were 16, 2, 16, 8, 0, 2, 16 h; the Cmax were 15.6, 35.3, 12.4, 9.0, 33.0, 11.6 mg/kg, and 21.0 mg/L; the area under curve (AUC) was 1084.5, 1578.1, 777.3, 541.1, 0.1, 478.1 mg/(kg·h), and 485.1 mg/(L·h); the t1/2β was 11.4, 100.2, 54.2, 41.1, 69.5, 38.0, and 71.9 h. The results showed that the distribution and elimination rate of TAP in carp tissues varied greatly; the peak time of renal drugs is shorter; the peak concentration is higher than in other tissues, and the elimination half-life is higher than that in other tissues, which suggested that kidneys are the main organ of TAP accumulation and metabolism. In accordance with the Ministry of Agriculture, Animal Food in Veterinary Drugs Maximum Residue Limit document provides that the maximum of TAP in aquatic animal residue limit (MRL) shall not be higher than 50 μg/kg. In this experiment, the amounts of TAP residues in muscle, kidney, liver, skin, spleen, and plasma were lower than MRL, starting from 16, 16, 12, 12, 12, 10, and 12 d respectively. It is recommended that the period of withdrawal should be no less than 16 d.

Abstract:The tropical South China Sea has profound development potential for its wide sea area, excellent marine environment, high biodiversity, and abundant bio-resource species. Recently, under the background of coastal fishery resource degradation, it has become vital to speed up the protection and sustainable utilization of tropical island fishery resources in this area. Marine ranching is a new type of modern fishery, which combines habitat remediation, bio-resource restocking, leisure fishery, and landscape ecology. It can help to develop the marine economy together with effective protection of the ecological environment. The development of marine ranching in the wide, tropical South China Sea area has been slow. There are several advantages to carrying out marine ranching in the tropical South China Sea, such as in political, natural habitat, bio-resources, and industrial economy aspects, etc. Two appropriate marine ranching types were identified, including stock enhancement and tourism and leisure types. The progress of tropical coastal and offshore island marine ranching construction in Guangdong, Guangxi and Hainan provinces were reviewed, especially the first successful construction of island marine ranching around Wuzhizhou Island in Sanya and the first coral reef marine ranching at Lingyang reef in Sanshan. Several potential risks affecting tropical offshore island marine ranching development were also documented, such as extreme weather (typhoon), predators, poor basic living facilities on islands, long distance for transportation, application difficulty on the using right of the marine ranching construction area, expensive fees for use of the sea area, etc. Based on considerations above, we provide detailed suggestions, including scientifically selecting sites, optimizing artificial facility design, developing new anti-corrosion reef material, deployment protocols, and anti-wave mooring techniques. We also give suggestions for: development of predator prevention techniques (to increase the recapture rate of released animals); development of automatic operation, monitoring, and management systems; development of feasible investment and operation modes; and more policy support from the government.