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Abstract:A 67-days feeding experiment was conducted to investigate the effects different levels on the growth performance and biochemical parameters of tiger puffer [average body weight: (13.83±0.63) g], and to determine the dietary methionine requirement for tiger puffer. There were six experimental isonitrogenous and isolipidic semipurified diets which were formulated with the graded levels of methionine (0.61%, 0.85%, 1.10%, 1.39%, 1.60%, 1.84%, respectively). Each diet was randomly assigned to triplicate groups, and each group contained 30 individuals. Fish were fed fourth daily(07:00, 12:00, 16:00, 20:00) to apparent satiation. The result showed that no significant differences in protein productive value (PPV) were found among dietary treatments (P>0.05). Specific growth rate (SGR), feed conversion ratio(FCR) and protein efficiency ratio (PER) increased with increasing dietary methionine levels from 0.61% to 1.60%(P<0.05), the higest group was observed in methionine levels 1.60% group, and there after showed a declining tendency with increasing dietary methionine levels from1.60% to 1.84%. Hepatosomatic index (HSI) and viscerosomatic index (VSI) of the 1.60% methionine diets was significantly higher than that of 0.61% groups (P<0.05), with further increase from 1.60% to 1.84%, HSI and VSI decreased. The crude lipid contents of whole body were significantly affected by dietary methionine levels (P<0.05), the 1.60% methionine diets was significantly higher than 0.61% groups (P< 0.05), with further increase from 1.60% to 1.84%, crude lipid decreased. while moisture, crude protein and ash showed no significant differences (P>0.05). Dietary methionine levels significantly influenced the contents of triglyceride, bile acid and blood glucose in serum (P<0.05), There were no significant difference in total cholesterol, high density lipoprotein cholesterol, low density lipoprotein cholesterol, total protein, activity of glutamic-pyruvic transaminase and glutamic-oxaloacetic transaminase (P>0.05) in serum. Second-order regression analysis on the basis of SGR indicated that the optimum dietary methionine requirements of tiger puffer were estimated to be 1.38% of diet (2.71% of dietary protein).

Abstract:A 12-week feeding trial was conducted to evaluate the effects of replacement of fish meal with alternative proteins on growth performance, body composition, and physiological and biological indices of juvenile turbot (Scophthalmus maximus L.). Five isonitrogenous and isolipidic practical diets were formulated to contain graded levels (100%, 60%, 50%, 40%, and 30%) of fish meal (D1~D5); the D1 group was used as the control. A protein blend of four ingredients (soybean meal, peanut meal, stickwater meal, poultry by-product meal = 2:1:3:2) were used to replace the fish meal; subsequently, amino acids were supplemented in the low fish meal diet to obtain similar amino acid profiles to the high fish meal diet. Each diet was randomly assigned to triplicate groups of 30 fish [(initial weight, (53.0±0.2) g)] per aquarium. The results of the survival rate, feed conversion ratio, feed intake, whole body moisture and crude protein, dorsal muscle moisture, crude protein, crude lipid, and ash content did not show any significant differences between any of the groups (P>0.05), whereas the diets with low fish meal content (D4/D5) significantly reduced the weight gain rate of fish compared with the control diet (P<0.05). The condition factor was significantly enhanced in the D2 treatment compared with the D3, D4, and D5 groups (P<0.05). However, the viscerosomatic and hepatosomatic indices were significantly reduced in the D2 treatment compared with the D5 (P<0.05). The dietary inclusion of the animal and plant protein mixture significantly elevated the whole-body crude lipid content (P<0.05). The whole-body ash content in the D5 treatment was significantly lower than the control diet (P<0.05). The essential amino acid compositions of the muscle were not significantly affected by the replacement of dietary fish meal with animal and plant protein mixtures (P>0.05). The dietary inclusion of animal and plant protein mixture significantly elevated serum aspartate aminotransferase and alanine transaminase activities (P<0.05). All the substituted protein diets caused significantly reduced serum cholesterol and low-density lipoprotein cholesterol (P<0.05), but only D3~D5 diets caused a significantly decreased serum high-density lipoprotein cholesterol (P<0.05). The dietary inclusion of animal and plant protein mixture had no strong effects on the total protein content and the alkaline phosphatase content (P>0.05). The replacement of up to 50% of dietary fish meal with an animal and plant protein mixture did not impair growth or reduce the feed efficiency of juvenile turbot.

Abstract:A 70-day trial was conducted to investigate the effect of replacement of dietary fish meal by Nannochloropsis sp. meal on growth performance, body composition, and serum biochemical indices of juvenile turbot Scophthalmus maximus L. with an initial weight body weight of (24.60±0.02) g. Five isonitrogenous and isoenergetic diets (N0, N3.88, N7.76, N11.64, and N15.52) were formulated with 0%, 3.88%, 7.76%, 11.64%, and 15.52% fish meal replaced by Nannochloropsis sp. meal, respectively. Each diet was randomly fed to three replicates of fish, with 40 fish per replicate. The following results were obtained. 1) There were no significant differences (P>0.05) in weight growth rate, specific growth rate, protein efficiency ratio, feed coefficient ratio, feed intake, condition factor, and survival rate among the groups. 2) As the meal content of Nannochloropsis sp. increased, the crude lipid content in the whole body and muscle was significantly decreased (P<0.05); however the crude protein, crude ash, and moisture content were not significantly different (P>0.05). 3) Lysozyme, complement protein C3, complement protein C4, and acid phosphatase activities showed a rising trend followed by a decline, with the highest point reached in the N7.76, N7.76, N11.64, and N7.76 groups, respectively, and were significantly higher than that in the N0 group (P<0.05). The alkaline phosphatase activity of the N15.52 group was significantly lower than that the other groups and there were no significant differences among other groups (P>0.05). 4) The total superoxide dismutase, total antioxidant capacity, and glutathione peroxidase activities in the serum of the algae meal groups first increased and then decreased, and reached a maximum in the N7.76 group, which was significantly higher than that in the N0 group (P<0.05). 5) The triglyceride content in the serum of the N7.76 group was significantly lower than that in the other groups (P<0.05), and there were no significant differences among the other groups (P>0.05). The total cholesterol content of the algae meal groups was significantly lower than in the N0 group (P<0.05), but there were no significantly differences among the algae meal groups. 6) The aspartate aminotransferase activity in the serum of the algae meal groups followed a decreasing to increasing trend, reaching the lowest point in the N11.64 group and was significantly lower than that in the N0 group (P<0.05). The alanine aminotransferase activity of the algae meal groups was significantly lower than that in the N0 group (P<0.05). In conclusion, under the experimental conditions tested, Nannochloropsis sp. meal could replace 15.52% of fish meal in juvenile turbot feed without any effects on the growth performance, and the replacement of 7.76% fish meal significantly improved the nonspecific immunity and reduced the blood lipid levels in juvenile turbot.

Abstract:This experiment evaluated the effects of fish meal (FM) replacement by silkworm powder (SP) on the growth performance and body composition of juvenile black bass (Micropterus salmonides). Six isonitrogenous and isoenergic diets were formulated by the replacement of 0 (control), 10%, 20%, 30%, 40%, and 50% FM with SP. Each diet was fed to three replicate groups of 30 juvenile black bass (total, 540 fish) with an initial body weight of (3.13±0.04) g. Owing to the decline in the temperature, the breeding cycle was 45 days. The results show that weight growth rate (WGR), special growth rate (SGR), and the protein efficiency ratio (PER) of black bass increased first and then decreased as along with the replacement rate increased, and reached maximum when replacement level was 30%, with increases of 227.87%, 1.98%/day, and 164.55%, respectively (P<0.05). The feed conversion rate (FCR) decreased first, then increased as along with the replacement rate increased, and reached a minimum when replacement level was 30%, with a decrease of 1.43 (P<0.05). The condition factor (CF), hepatopancreas somatic index (HSI), and viscera somatic index (VSI) increased first, then decreased as the replacement rate increased, and finally tended towards stability (P<0.05). The survival rate (SR) of the six groups of black bass was not significantly different (P>0.05). Through quadric regression analysis, when WGR, SGR, FCR, and PER reached the optimum values, the replacement rates were 19.90%, 22.00%, 18.11%, and 17.90%, respectively. The content of crude lipid decreased first and then increased as the replacement rate increased (P<0.05); whole body crude protein did not change significantly as the replacement rate increased (P>0.05); moisture content in the whole body was not significantly different in any group (P>0.05) except the 10% and 40% groups (P<0.05); and the ash content of the whole fish decreased significantly when the replacement rates were above 20% (P<0.05). Therefore, under the experimental conditions, the measurement of the growth performance and body composition of black bass indicated that the optimum substitution rate of SP with FM was between 17.90% and 22.00%.

Abstract:An 8-week experiment was conducted to investigate the effects of the replacement of dietary fish oil (FO) with soybean oil (SO) on growth, lipid deposition, and fatty acid composition of juvenile spotted halibut. Four isonitrogenous and isolipidic practical diets were formulated with SO replacing 0, 25%, 50%, and 75% FO, respectively. In total, 360 juvenile spotted halibut, with an initial body weight of (65.47±1.57) g were randomly distributed into four groups; three replicates were used per group and 30 fish were used per replicate. The fish in each group were randomly fed one of the four experimental diets. The following results were obtained: (1) Compared with 100% FO group, the feed conversion rate of spotted halibut was significantly increased (P<0.05), whereas the weight gain rate was significantly decreased (P<0.05). (2) SO replacement levels had no significant effects on body composition and muscle lipid content of juvenile spotted halibut (P>0.05), but had a significant effect on the liver lipid content of juvenile spotted halibut (P<0.05). The liver lipid content in the 50% and 75% SO groups was significantly higher than that in 100% FO group (P<0.05). (3) The fatty acid composition in the muscle was positively correlated with dietary fatty acid composition. Compared with the 100% FO group, the content of linoleic acid (C18:2n-6) and linolenic acid (C18:3n-3) in the muscle were significantly increased (P<0.05), whereas the content of eicosapntemacnioc acid (EPA) and docosahexaenoic acid (DHA) in the muscle were significantly decreased (P<0.05). This study suggested that there were no significant effects on the growth of juvenile spotted halibut when dietary fish oil was replaced by soybean oil. However, the fatty acid composition of muscle was significantly altered.

Abstract:A 2×3 two-factorial experiment was designed to investigate the effects of dietary administration of Bacillus subtilis and yeast culture on growth, serum biochemical indices, antioxidant capacity, and disease resistance of juvenile hybrid grouper (Epinephelus fuscoguttatus♀×E. lanceolatus♂) [initial weight: (23.41±0.47) g]. Nine isonitrogenous and isoenergetic diets were formulated with three levels of B. subtilis [0 (control), 0.5%, and 1.0%], and each B. subtilis level was formulated with three yeast culture levels [0 (control), 0.5%, and 1.0%]. Juvenile hybrid groupers were fed for 8 weeks. The results showed that the interaction between B. subtilis and yeast culture had no significant effects on the survival (SR) and weight gain rate (WGR) of grouper (P>0.05). The WGR of Y1B1 and Y1B2 groups was significantly higher than that of the control and Y2B2 groups (P<0.05).The interaction between B. subtilis and yeast culture had a significant effect on the aspartate aminotransferase (AST) and alkaline phosphatase (AKP) in the serum of grouper (P<0.05); the AST and AKP activities in the serum of Y1B1 and Y1B2 groups were significantly lower than those in the control and Y2B2 groups (P<0.05). The interaction between B. subtilis and yeast culture had a significant effect on the superoxide dismutase (SOD) and malondialdehyde (MDA) levels in the intestinal tract of grouper (P<0.05), but no significant interaction with catalase (CAT) or total antioxidant capacity (T-AOC) in the intestinal tract (P>0.05). The activities of SOD, CAT, and T-AOC in the intestinal tract of the Y1B1 and Y1B2 groups were significantly higher than those in the control and Y2B2 groups (P<0.05), but the MDA content in the intestinal tract showed an opposite trend to SOD (P<0.05). There was no interaction between the effects of dietary B. subtilis and yeast culture on the cumulative survival rate of juveniles challenged with Vibrio anguillarum (P>0.05). The highest value of cumulative survival rate of juveniles was found in the Y1B1 group, which was significantly higher than that in the control group (P<0.05). In conclusion, supplementation of 0.5% or 1.0% B. subtilis and 0.5% yeast culture in diets could promote WGR, antioxidant capacity, and disease resistance in juvenile hybrid grouper.

Abstract:Largemouth bass (Micropterus salmoides) is a very important fish in commercial farming. As it is a carnivorous teleost, a large amount of forage fish and fish meal were used as food for largemouth bass every year. Residual forage fish and fish meal not only increase the costs of largemouth bass aquaculture, but also seriously pollute water bodies and farm lands. In order to reduce farming costs and protect the natural environment, selective breeding suitable for formulated feeds was carried out. In 2012, our research team selected the breeding population contain Youlu No.1 largemouth bass and introduced north largemouth bass. The genetic structure of largemouth bass breeding population was analyzed based on microsatellite DNA markers. A total of 240 individuals, from the foundation population (Sp0), the second generation group (Sp2), the third generation group (Sp3), and the fourth generation group (Sp4) (60 individuals per population) were assayed with 18 microsatellites. The results showed that a total of 44 alleles were identified, the numbers of alleles (Na) were 1~4 in four selected populations, respectively. The observed heterozygosity (Ho) of Sp0, Sp2, Sp3 and Sp4 was 0.4895, 0.4802, 0.4579 and 0.4206, respectively; the expected heterozygosity (He) was 0.4615, 0.4454, 0.4621 and 0.3916, respectively; the polymorphism information content (PIC) of Sp0, Sp2, Sp3 and Sp4 was 0.3791, 0.3659, 0.3764 and 0.3257, respectively. The F-statistics (Fst) value is between 0.01612 and 0.16162, and the genetic distance (Da) is between 0.0249 and 0.1434. Partitioning of the genetic variation revealed that only 8.38% genetic variation was among the populations, and the other genetic variation was within the populations. The results showed that the artificially selected population using formulated feeds showed moderate genetic diversity, and still had great potential for future selective breeding.

Abstract:In the management of stock enhanced Japanese flounder, it is often necessary to distinguish those that are hatchery-reared, from wild stocks. This study examines the feasibility of marking Japanese flounder by feeding them a diet enriched with strontium. To achieve nominally different concentrations of strontium in the fish food, 1, 8, and 64 g of strontium chloride crystals (SrCl2∙6H2O) were each dissolved in 1 L of distilled water and sprayed onto 1 kg of pelleted feed. The treatments groups of fish were fed the pellets for 10 d, then farmed for 30 days, and then sampled. Our data shows that both the 8 g SrCl2/kg feed and 64 g SrCl2/kg feed resulted in a mark on the otoliths of the Japanese flounder. The peak concentration of strontium (8 g SrCl2/kg), often resulted in otoliths with concentrations two orders of magnitude higher than those found in the control fish. The concentration of strontium in the 64 g SrCl2/ kg feed, resulted in a 6-fold or 18-fold increases in the otoliths. The strontium value increased with the increased concentration of strontium in the pellet feed. The results also showed that strontium was safe for the fish and did not negatively affect their mortality. The total body length of the treatment groups with 1 and 8 g SrCl2/kg, were (11.11±1.32) cm and (10.88±1.07) cm, respectively, in which the length growth rate was faster than that of the control group and the treatment group with 64 g SrCl2/kg feed (P≤0.05). With appropriate strontium concentrations, the growth of the Japanese flounder youth was enhanced. Our results suggest that feeding a diet enriched in strontium could be an effective means of marking hatchery-reared Japanese flounder for stock enhancement.

Abstract:Seven countable traits and 20 measurable traits of three populations of the hybrid “Epinephelus moara ♀ ´ E. lanceolatus♂” and its parents, E. moara (♀) and E. lanceolatus (♂) were measured, and the comparative analysis of morphological characteristics among the three groups were performed by virtue of three kinds of multivariate statistical analysis methods, including cluster analysis, principal component analysis and discriminant analysis and variance analysis methods. Chi- square test results showed that there was no significant difference in the countable traits among the three grouper populations. The result of cluster analysis revealed that, characters of “E. moara ♀ ´ E. lanceolatus♂” was more similar to E. moara than E. lanceolatus. Six comprehensive indexes reflecting morphological characteristics were constructed by principal component analysis. The contribution rates of the six principal components were 24.585%, 17.985%, 10.410%, 9.970%, 6.542%, and 5.542% respectively, and the cumulative contribution rate of those was 74.945%. There were obvious deviations in morphological characteristics among the three groupers, forming three different groups and hybrids. “E. moara ♀ ´ E. lanceolatus♂” has its own unique morphological features. By discriminant analysis, four morphological characteristics that have the largest contribution to discriminating grouper, X3 (tongue length/head length), X6 (height at withers/body length), X8 (height at caudal peduncle/body length), X14 (ventral fin basal to the front of anal fin basal/body length), were selected to fit the typical discriminant function (F1=21.108X3+38.413X6-97.206X8+2.223X14-6.849, F2=-10.344X3+21.570X6+21.931X8+26.901X14-17.406) and the classification discriminant function (Y1=217.572X3+750.765X6+1425.499X8+413.410X14-336.464, Y2=318.678X3+839.713X6+1020.377X8+358.203X14-319.723 and Y3=258.526X3+672.660X6+1334.033X8 +311.962X14-269.278) that can distinguish the above three groupers, the comprehensive discriminant rate was 96.43%. It can be considered that the four morphological characteristics selected by stepwise discriminant analysis are feasible for the preliminary identification of three groupers. The results of variance analysis showed that the morphological differences of the three groupers reached significant or extremely significant levels in all characteristics except for X11 (The pectoral fin base to the pelvic fin/body length). Among them, there were 14 morphological traits between “E. moara ♀ ´ E. lanceolatus♂” and E. lanceolatus reached extremely significant levels, and 12 morphological traits between “E. moara ♀ ´ E. lanceolatus♂” and E. moara were extremely significant. The results of this study provide biological basic data for the germplasm identification of “E. moara ♀ ´ E. lanceolatus♂”, E. moara and E. lanceolatus.

Abstract:In order to study the effects of environmental factors on the intestinal flora structure of turbot (Scophthamus maximus), we used high-throughput sequencing to explore the bacterial community structure and diversity in juvenile turbot intestines, the culture environment, and biological baits. The results showed that 547621 effective sequences were detected in nineteen samples, and they could be classified into 3771 operational taxonomic units (OTUs), among which 3038, 1090, 87, and 777 originated from the aquaculture water, the biological baits, healthy juvenile turbot intestine, and diseased juvenile fish intestine, respectively. There were 57 OTUs shared between the healthy juvenile turbot intestine and the biological baits, 0 OTU shared between the healthy juvenile turbot intestine and aquaculture water, 481 OTUs shared between the diseased juvenile fish intestine and the biological baits, 31 OTUs shared between the diseased juvenile fish intestine and the aquaculture water. The effect of biological bait on microbial diversity of intestinal tract of juvenile fish was much greater than that of environment. In total, the predominant phyla in the turbot intestine were Bacteroidetes, Firmicutes, and Proteobacteria. The intestinal microflora of healthy juvenile turbot can be clustered into 8 phyla, and the intestinal microflora of the diseased juvenile fish could be clustered into 19 phyla. Compared with the healthy juveniles, the community structure of the predominant phyla was imbalanced at the intestinal level of the diseased juvenile fish. Furthermore, analysis of the 100 most abundant bacterial OTUs in the different samples revealed that the species dominant in the intestinal bacteria of juvenile fish was closely related to the dominant species in the biological baits. Meanwhile, the intestinal dominant bacteria species of each diseased juvenile are different. This study provided the basis for healthy culture and micro-ecological regulation of turbot.

Abstract:Interleukin 15 (IL-15) is an important cytokine of fish immune system. In the present study, the IL-15 cDNA named as TfIL-15 was cloned from roughskin sculpin, Trachidermus fasciatus. The full-length of TfIL-15 cDNA is 1140 bp, which contains a 5¢-UTR (untranslated region) of 165 bp, a 3¢-UTR of 453 bp and an open reading frame (ORF) of 522 bp, encoding a polypeptide of 173 amino acids (aa) with a putative 59 aa-long signal peptide. Four out-of-frame AUG initiation codons, the negative translational regulators of mammalian IL-15 genes were also detected in the 5¢-UTR of TfIL-15. The protein sequence shared 23%~61% identity with reported fish IL-15s, displaying relatively high degree of variation. TfIL-15 homologues also contained four highly conserved cysteine residues allowing the formation of two disulfide bridges along with four predicted α-helices. Phylogenetic analysis grouped roughskin sculpin with other fish on a separated branch, excluded from mammalian and avian IL-15s. Quantitative real-time PCR (qRT-PCR) analysis showed that TfIL-15 was widely expressed in all detected tissues, with the highest expression in the heart. Post LPS challenge, TfIL-15 increased rapidly to the maximum of 74 folds and 41 folds compared with that of the control group at 2 h post challenge (hpc) in the skin and blood. The induction of TfIL-15 mRNA in the spleen and liver was 3 folds and 18 folds at 12 hpc. Interestingly, at 96 hpc, the expression of TfIL-15 in the liver was up-regulated again to the 86 folds higher than that of the control group. These results indicate that TfIL-15 may play an important role in fish innate immune response against microbial infections. Furthermore, the mature peptide of TfIL-15 was expressed in E. coli BL21 (DE3) cells successfully, laying a foundation for further research on the function of TfIL-15 protein.

Abstract:In order to clarify the shrimp disease occurred currently in the shrimp farms, shrimp pathogens including vibrio, Enterocytozoon hepatopenaei (EHP) and four viral pathogens(WSSV、IMNV、CMNV and IHHNV) were detected using molecular biological detect methods under a regular sampling pattern, meanwhile the concentration of ammonia nitrogen and nitrogenous nitrogen in water body were also detected. The results showed that the main pathogens in the shrimp farm comprised various pathogenic vibrio and EHP. However the shrimp virus WSSV、IMNV、CMNV and IHHNV were not detected. The vibrio species isolated and identified reached 16 totally, and the main species were Vibrio parahaemolyticus, V. alginolyticus, V. owensii, V. vulnificus, V. harveyi. Furthermore the V. parahaemolyticus that could cause acute hepatopancreatic necrosis disease (VAHPND) was found out. The concentration of ammonia nitrogen and nitrate nitrogen in the indoor pond reaching (3.5±2.0) mg/L and (8.2±0.7) mg/L, were significantly higher than that in the outdoor pond respectively (P < 0.05) in the medium-term (July 24) of crop cycle. The intensity of infection of VAHPND and EHP show a growing trend in the culture period. In the seven ponds monitored, the ponds indoor had a higher incidence rate (100%), while the ponds outdoor had a lower incidence rate (25%). The results indicated that shrimp disease onset in the pond indoor might be caused by the infection of various vibrio and EHP, also the higher stocking density, higher concentration of the ammonia nitrogen and nitrate nitrogen in the water body played a synergy role in outbreak of shrimp disease. The results of this study could provide theoretical support and scientific data for the prevention and control of shrimp diseases.

Abstract:Laminaria hyperborea is an important commercial seaweed, which has great potential for seaweed bed construction and artificial cultivation in China. To verify the suitable light for the growth of L. hyperborea young seedlings and clarify the biochemical response mechanism under light stress, the relative growth rate and biochemical responses under different light intensities were studied. Under these experimental conditions, the main conclusions are as follows: Light was beneficial for the growth of L. hyperborea young seedlings in the range of 40~80 μmol photons/(m2·s). The relative growth rate showed significant negative interrelation with the content of malondialdehyde (MDA), which indicates that membrane lipid peroxidation may be one of the reasons contributing to the low relative growth rate of L. hyperborea young seedlings. An extremely significant positive correlation was found between the content of superoxide anion (SA) and the content of MDA of L. hyperborea young seedlings. The specific activity of superoxide dismutase (SOD) and catalase (CAT) were negatively correlated with the content of SA and MDA, respectively. And the negative relationship between the specific activity of CAT and the content of SA was significant. Moreover, an extremely significant and positive correlation was found between specific activity of peroxidase (POD) and SOD. It can be inferred from the above correlation analysis results that SOD, CAT and POD play important roles in removing reactive oxygen species (ROS) and there is a significant synergy between SOD and POD in L. hyperborea young seedlings. The light stress environment was not conducive to the accumulation of soluble protein resulting in the accumulation of MDA in L. hyperborea young seedlings, which indicates that the total metabolic level was reduced and the ROS level was increased in this environment. The content of carotenoid and the specific activity of SOD and POD in L. hyperborea young seedlings will increase so that the ROS level can be reduced under high light stress, thus relieving light oxidative damage. The content of chlorophyll-a of L. hyperborea young seedlings will increase so that it can capture the limited light energy more effectively under low light stress. However, the antioxidant enzyme activity was significantly decreased as a result that led to a serious imbalance of ROS metabolism under low light stress, which resulted in more severe membrane lipid peroxidation damage. These results can provide the theoretical basis for the artificial breeding and cultivation of L. hyperborea in the future.

Abstract:We studied the effects of temperature and light intensity on growth, photosynthetic pigment content, and nutrient components in Sargassum horneri. These factors had significant effects (P<0.05) on all three aspects. S. horneri survived at 5℃~20℃ and 50~300 μmol/(m2·s) The optimal conditions, at which specific growth rate was the highest, were 20℃ and 200~300 μmol/(m2·s). At temperatures above 25℃, the blades stopped growing, became pale, and rotted. S.horneri accumulated the most photosynthetic pigment at 25℃ and 50 μmol/(m2·s). The effects of light intensity were not significant. Soluble sugar content and soluble protein content were the highest at 10℃~20℃, and ash and crude protein content were the highest at 5℃~10℃ and 50 μmol/(m2·s). Ether extract yield was low at 10℃ and 25℃, peaked between these values, and did not vary significantly with light intensity. Alginate content was the highest at 10℃ and 100 μmol/(m2·s), and fucoxanthin content was the highest at 10℃ and 50 μmol/(m2·s). Phlorotannin content increased with increasing light intensity from 200 to 300 μmol/(m·s), where it reached a maximum; it reached local maxima at 5℃ and 25℃, and was lower between these values. S. horneri grew faster atc. 20℃ and 200~300 μmol/(m2·s), whereas color and vigor were better at 10℃ and 100 μmol/(m2·s). Its long body length and fast growth make it ideal for large-scale culturing. The nutrient components varied with culture conditions. S. horneri is a health food of moderate protein and low fat content, with high levels of functional components such as alginate, fucoxanthin, and polyphenols. It can therefore be used to provide raw material for food additives and to obtain its active components. These findings provide reference values for culturing S. horneri and extracting its components.

Abstract:In order to explore the molecular mechanisms of the Hsp70 gene in Pyropia guangdongensis, we have stimulated it with high temperature stresses. The aim was to provide technical references for the cultivation of P. guangdongensis. The full-length of PgHsp70 was obtained by rapid amplification of cDNA ends technical (RACE). On this basis, the expression of PgHsp70 with the different temperature (22℃, 27℃, and 31℃) stresses after 0, 1/6, 1/2, 1, 6, 12, 24, and 36 h were detected using quantitative real-time PCR. The results showed that the full-length cDNA of PgHsp70 was 2004 bp, including an open reading frame (ORF) of 1866 bp, encoding a polypeptide of 621 amino acids. The molecular mass of the deduced amino acid sequence was 67.7 kDa, with an estimated pI of 4.87. The expression of the PgHsp70, as measured by qRT-PCR, can be significantly induced by high-temperature stress. The three kinds of expression profiles for PgHsp70 were similar at different times, and all of them significantly increased first and then reached their maximum levels after one hour, and then dramatically decreased. Compared to the temperature stress of 22℃ and 27℃, the expression of the P. guangdongensis with the 31℃ temperature stress reached the highest level after being challenged for 1 h and was 11-fold higher than the normal. These results suggested that PgHsp70 plays an essential role in response to high-temperature stresses.

Abstract:Green spot disease is one of the major disease considerations in the farming of Pyropia yezoensis. Green spot disease could be found in all the periods of thallus, and outbreaks usually occur in November and December. At the start of infection, small light red spots could be found on the thallus, and then the spots gradually turned green and expanded quickly. Holes from lesions formed later in the surface of thallus. At last, the entire thallus became green and fell off. In this research, bacteria strains were isolated from P. yezoensis with green spot disease farmed in Rizhao. Experimental infection showed that the strain Y1 could cause P. yezoensis green spot disease. Biochemical characterization and genes analysis of 16S rRNA, dnaA and dnaN indicated that the pathogen Y1 was Pseudoalteromonas marina. Influence of environmental factors on the outbreak of disease were also characterized by experimental infection with different temperature, stocking density and gravity of sea water. The results showed that higher temperature and stocking density will accelerate the spread of the disease, but gravity of sea water within a certain range did not affect the occurrence of the disease. In this study, we described the green spot disease caused by P. marina, which provides information for disease control in P. yezoensis cultivation.

Abstract:LuxR family proteins play a key role in various important physiological activities of gram-negative bacteria. In this study, RNAi technology was used to construct a stable silenced strain Aeromonas hydrophila luxR05735-RNAi. The expression of luxR05735 in wild-type strain and luxR05735- RNAi was detected by qRT-PCR. The results showed that the expression of luxR05735 in strain luxR05735- RNAi was reduced by 96.8% compared to that of the wild-type strain. Drug sensitivity tests showed that luxR05735-RNAi significantly reduced bacterial resistance to gentamicin, norfloxacin, kanamycin, and pipemidic acid compared with the wild-type strain. The transcriptome analysis of the wild-type strain and luxR05735-RNAi revealed that there were 1286 genes with significant differences in expression, of which 353 genes were up-regulated and 933 genes were down-regulated. There are four significantly enriched pathways, which are ribosome, arginine biosynthesis, sulfur metabolism and selenocompound metabolism. In the four pathways, some important functional genes, including metE, glnA, rplB, rplX, rpsA, rpsJ were found to be associated with drugs resistance of A. hydrophila. These genes encode the proteins that were mainly involved in bacteria biofilm formation and ribosomal protein synthesis. It can be speculated from the above results that the luxR05735 of A. hydrophila regulates the expression of the genes that encode bacteria biofilm formation and ribosomal protein synthesis, and then further regulates the drugs resistance of A. hydrophila.

Abstract:The ORF wsv112 of white spot syndrome virus (WSSV) encodes a dUTPases. It plays an essential role in nucleotide biosynthesis. Hydrolysis of dUTP by UTPase produces dUMP, required for the de novo synthesis of dTTP, and maintains low cellular ratios of dUTP/dTTP, thus preventing the mis-incorporation of uracil into chromosomal DNA. In order to identify the host interactors of wsv112，wsv112 was cloned into the bait vector pGBKT7 and used to screen an intestine cDNA library of Litopenaeus vannamei, which had previously been constructed by yeast two-hybrid sequencing transformation. The positive clone was identified through different culture media, color change, polymerase chain reaction (PCR), and sequencing. The result showed that the bait plasmid pGBKT7-112 showed no virulence or self-activating effect on yeast strain Y2H Gold. A total of 526 blue clones were screened, which were analyzed by PCR and homology analysis using the BLAST in NCBI, and 6 possible interaction proteins of Litopenaeus vannamei were obtained. Then through the Yeast two-hybrid reply hybridization experiment, only two gene interactions were confirmed with the wsv112. They were identified as lectin C gene of Marsupenaeus japonicas (AGW27416.1) and 40S ribosomal protein S20 gene of Procambarus clarkia (ALE99171.1) with 37% and 98% identity. This study may provide a theoretical basis for further study of the wsv112 interaction mechanisms.

Abstract:To study the structure of organism attachment on artificial reefs made of different materials, we prepared artificial reefs with three types of materials: Ordinary portland cement(P), scallop shell portland cement (S), and iron plates(F). The artificial reefs were exposed to the marine environment near Shique Beach, Qingdao, in May 2017. We collected samples to analyze the effect of different reef materials on organism attachment on the artificial reef in July, September, November, 2017 and January 2018. The results showed that, in total, 69 species of attached organisms were identified. The numbers of species on ordinary portland cement, scallop portland cement, and iron plate reefs were 51, 53, and 31, respectively. The dominant species on the ordinary portland cement and scallop portland cement reefs were Caprella sp., Ostrea plicatula, and Mytilusgallo provincialis; the dominant species on iron plate reefs were Caprella sp., M. provincialis, and Stenothoe qingdaoensis. Biological attachment peaked between September and November. We found the highest average biomass of fouling organisms on scallop portland cement (4717.50 g/m2), followed by ordinary portland cement (2621.12 g/m2), and iron plates (163.85 g/m2). In July 2017, the Shannon-Wiener diversity index (H′) of fouling organisms was observed to follow the order P>S>F; in September 2017, the order was S>P>F; in November 2017, the order was P>S>F; and in January 2018, the order was F>P>S. In July 2017, the Pielou evenness index (J) followed the order F>P>S. In September 2017, the order was S>F>P; in November 2017, the order was F>S>P; and in January 2018, the order was F>P>S. Our study showed that scallop portland cement attracted the largest biological species and biomass, and was most suitable for organism attachment. This study provides a reference for the evaluation of the characteristics of biological communities attached to artificial reefs and the selection of artificial reef materials.

Abstract:In this study, the metaphase chromosomes of Collichthys lucidus were obtained from kidney tissue by the method of PHA and colchicine injection-air drying method. The experimental fishes were taken from Ningde coast of China. Because of their strong stress, we adjusted the drug treatment time (PHA: 6 h; Colchicine: 2 h, 5 h). In order to identify whether there were heteromorphic chromosomes, we separated the female from the male. The results showed that there were 48 chromosomes in C. lucidus and its karyotype formula was 2n=48t, NF=48. The relative length of chromosome was in the range of (5.694±0.514)~(3.039±0.161), which suggested that C. lucidus was in accordance with the original karyotype of perciformes and the higher group of fish evolutionary taxonomy. Sex chromosome, satellite chromosome and secondary constrictions were not found in C. lucidus. This study provides basic data for cytogenetics and germplasm identification for C. lucidus.

Abstract:Scallop skirts are by-products of scallop processing that are rich in nutrients such as proteins and lipids. Owing to the limitations of the existing processing technology, they have not been used on a large scale and are not currently of much value. In this study, to effectively utilize the scallop skirt, neutral protease, animal-complex protease, flavor protease, papain, and acidic proteinase were used to hydrolyze the scallop skirt. The free amino acid nitrogen content in the hydrolysate was used as an indicator for optimized reaction conditions. First, alcohol was used to degrease the scallop skirt, which increased the degree of enzymatic hydrolysis. Subsequently, single-factor experiments were performed on five enzymes to select the optimize enzymatic hydrolysis time, temperature, pH, and enzyme dosage. Subsequently, orthogonal experiments were performed to determine the optimum hydrolysis conditions for the five enzymes. The results showed that the amino acid conversion rate of the hydrolysate prepared under the optimum hydrolysis conditions was as high as 77%. In previous studies, calcium chloride was determined to be the most suitable calcium source. The scallop shell was used as a raw material to obtain calcium chloride by acid treatment and water flying, and then calcium chloride was mixed with compound amino acids in the hydrolysate to prepare complex amino acid-chelated calcium. The calcium ion content in the chelated calcium was chosen as an index; subsequently, single factor experiments were performed to determine the time, temperature, and pH of the chelation process. The optimum chelating conditions were screened by an orthogonal experiment. A chelation rate of up to 92% was obtained in the final products. The compound amino acid-chelated calcium not only functions as a calcium supplement, but also has a delicious taste; it can be used as a calcium supplement or functional condiment. Our findings will assist in achieving additional value from scallop-processing waste, and also provide us with novel insights and methods for high-value utilization of other aquatic processing waste.