• Volume 44,Issue 1,2020 Table of Contents
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    • Isolation and identification of hepatocellular exosomes and their effects on the expression of miR-122/33 and immune-related genes in grass carp(Ctenopharyngodon idella)

      2020, 44(1):1-10. DOI: 10.11964/jfc.20181211575

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      Abstract:Exosomes are nano-scale vesicles with a phospholipid bimolecular membrane, which can participate in many physiological processes. The aim of this study was to investigate the isolation and identification methods of extracellular exosomes from grass carp hepatocyte and to conduct preliminary study on the effects of exosomes secreted from fatty hepatocyte on the expression of miRNAs and immune-related genes in grass carp hepatocytes. In this study, The exosomes of grass carp hepatocytes L8824 were extracted by ultracentrifugation. The morphology of exosomes was observed by Electron Microscopy, the parcicle size and quantity of exosomes were determined by Nano-particle Tracking Analysis. Meanwhile, we analyzed the expression of the specific protein CD63 using Western blot. Then, we used exosomes,which wwew secreted from normal hepatocytes and fatty hepatocyte induced by oleic acid to incubate grass carp hepatocytes, and the effects of two different exosomes on the transcription levels of miR-122/33 and immune-related genes, which are TNF-α, NF-κB, IL-1β, IL-6 and IL-10, were detected by real-time qPCR. Our results indicated that the exosomes secretion of grass carp hepatocytes culture medium were uneven vesicles of 30-150 nm, round or oval, and had a complete membrane structure. The expression of exosomes marker protein CD63 was positive, and the NTA results showed that the exosome vesicles accounted for more than 50 % of all the vesicles. Additionally, the exosomes of fatty liver cells significantly increased the expression of miR-122 and the immunogenic genes (TNF-α, IL-1β and IL-6) in hepatocytes (P<0.05). These results suggest that the exosomes of grass carp hepatocytes can be isolated successfully by ultracentrifugation. Moreover, exosomes may play a key role in the regulation of immunity of grass carp hepatocytes.

    • Identification and characterization of the calcineurin B in Fenneropenaeus chinensis: its implications in competitive ability

      2020, 44(1):11-20. DOI: 10.11964/jfc.20190111623

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      Abstract:Competitive behavior exists widely in animal and plant groups. When groups or individuals have the same needs for the limited resources, competition will occur in intra-species or inter-species. The existence of competitive behavior can significantly affect the growth performance of individuals. There is strong competitive behavior in the breeding of Fenneropenaeus chinensis, and we have proved that the competitive behavior among individuals of F. chinensis has a significant effect on individual growth traits, and this effect can be inherited. However, little information was available for the molecular mechanism of competition behavior in shrimp. Candidate genes involved in competitive behavior of F. chinensis have been identified by comparative transcriptome analysis in our previous study, among which there is a calcineurin (CN) B gene. CN is a highly conserved Ca2+/calmodulin (CaM)-dependent serine/threonine phosphatase, which is composes of a catalytic subunit (CNA) and a regulatory subunit (CNB). Previous studies reported that CN-B played an important role in the central nervous system mediated by Ca2+/CaM. In order to further prove the role of CN-B gene in the competitive behavior, in the present study, the full-length cDNA of CN-B of F. chinensis (FcCN-B) was cloned by RACE technology, and its expression in different tissues (ganglion, heart, stomach, hepatopancreas, and intestine) between high competitive group (HCG) and low competitive group (LCG) was analyzed by Real-time PCR. The results show that the full-length cDNA of FcCN-B is 2 867 bp, containing 95 bp of 5' untranslated regions (UTR), 540 bp of open reading frame (ORF), and 2 232 bp of 3' UTR. There are four conserved EF-hand Ca2+ binding domains in the ORF of FcCN-B. Protein homology analysis indicated that the amino acid sequence of FcCN-B has high homology with other species (78.8%-93.8%), among which FcCN-B has the highest homology with Eriocheir sinensis (93.8%) and then with Drosophila. melanogaster (90.5%). The phylogenetic relationship analysis showed that vertebrates and invertebrates were independently clustered into two branches. Furthermore, F. chinensis was clustered with E. sinensis into an independent branch, and then it was clustered to D. melanogaster, which suggesting that FcCN-B might have similar functions as it did in D. melanogaster. The results of Real-time PCR showed that the expression of FcCN-B in the ganglion was significantly higher in HCG than in LCG, but its expression in heart was significantly lower in HCG than in LCG. This study preliminarily proved that the calcineurin B may play a certain role in the competition behavior of F. chinensis, and will lay an important foundation for investigating the molecular mechanism of the competition behavior of F. chinensis.

    • Cloning, expression and biological functional characteristics of Cactus from Scylla paramamosain

      2020, 44(1):21-32. DOI: 10.11964/jfc.20181211604

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      Abstract:In this study, we obtained the full length of the SpCactus gene by RACE technology from Scylla paramamosain(SpCactus). And we analyzed the biological information of the SpCactus and its immune response under pathogen stimulation. The full length of SpCactus mRNA contains a 5' untranslated region (UTR) of 228 bp, an open reading frame of 1 311 bp and a 3' UTR of 496 bp. The SpCactus protein contains five characteristic ANK homology domains and showed 62% identity (73% similarity), 24% identity (36% similarity), and 9% identity (18% similarity) to the Litopenaeus vannamei Cactus (LvCactus) protein, the Drosophila melanogaster Cactus (DmCactus), and the Homo sapiens IκB protein, respectively. Prediction of the protein physicochemical properties of SpCactus revealed that it is a hydrophilic protein. By physical and chemical properties analysis, SpCactus was a hydrophilic protein. And its isoelectric point (pI) is 4.91. The mRNA of SpCactus showed high expression in the muscle, eyestalk and heart, but low expression in the hepatopancreas. Moreover, the expression of SpCactus were significantly upregulated by stimulation with Staphylococcus aureus and lipopolysaccharide. In this study, SpCactus gene was successfully cloned and characterized for the first time, and its bioinformatics analysis, physical and chemical properties were predicted. The biological functions of SpCactus gene were preliminarily explored, providing a basis for further study of its biological functions in immune response.

    • Estimates of genetic parameters of growth-related traits for the selective breeding of deep-cupped shell Crassostrea hongkongensis

      2020, 44(1):33-42. DOI: 10.11964/jfc.20181111538

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      Abstract:In order to breed a new strain with deep-cupped shell of Crassostrea hongkongensis and better growth traits, a breeding program of truncation selection was initiated based on the 2-year-old culture population of wild C. hongkongensis collected naturally from Taishan Town Bay, Guangdong Province. The index of shell depth was used as an indicator, with 10% retention rate and 1.755 selection strength. The genetic parameters including selection reaction, genetic gain and realized heritability in larval stage, intermediate cultivation stage and later grow-out stage were estimated. The results showed that the shell height and index of shell depth in selected group were higher than those in control group, and the genetic parameters increased with the growth of individuals, showing a favorable genetic improvement potential. The average selective reaction, genetic gain and realized heritability of shell height at larval stage was 0.363±0.167, 1.678%±0.416% and 0.207±0.095, respectively; at intermediate cultivation stage was 0.639±0.115, 7.618%±2.666%, and 0.364±0.065, respectively; and later grow-out stage was 0.668±0.179, 8.861%±3.072% and 0.381±0.102, respectively. At the later grow-out stage, the average selective reaction, genetic gain and realized heritability of index of shell depth was 0.748±0.066, 9.090%±0.565%, and 0.426±0.038, respectively. The research provided useful data for the breeding of a new strain with deep-cupped shell and good growth traits in C. hongkongensis.

    • Primary culture and identification of brain neurons in Chinese perch (Siniperca chuatsi)

      2020, 44(1):43-48. DOI: 10.11964/jfc.20190111630

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      Abstract:The diversity of species makes the research at cell level of Chinese perch extremely limited. Establishing a simple and feasible method for primary culture of brain neurons from Chinese perch (Siniperca chuatsi) in vitro is beneficial to further study on fish nervous system. Our laboratory combined common methods of cell culture, Chinese perch brain neruons (born 3 months) were isolated by collagenase digestion and mechanical blow. L15+20%FBS suspended brain neruons were inoculated in the cell culture vessels, at 28 ℃ without CO2 incubator. The medium was replaced after 3 days. Subculture was carried out after the cells covered the bottom of the cell culture vessels. The morphological changes of the neurons were observed under inverted phase-contrast microscope; Immunofluorescence staining for NeuN or β-tubulin was performed to identify the purity of neurons. The results showed that the cells began to adhere to the culture bottle and develop small neurites and form network gradually after primary culture for 2 days. On the 5th day, many neurites extended to form dense network and Soma of neurons became well. Fluorescence staining with Neu-N or β-tubulin showed that the purity of neurons can reach above 95%. The present protocol is a simple and efficient method for culturing brain neurons of Chinese perch with high purity, which is of great significance for the further study of fish growth and development, expression regulation of various receptors and proteins, cell apoptosis and cell signal transduction.

    • Statolith morphology of Gonatopsis borealis in the northwest Pacific Ocean

      2020, 44(1):49-60. DOI: 10.11964/jfc.20190411772

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      Abstract:The morphology and microstructure of statolith of Gonatopsis borealis were studied for the 339 samples, which were collected by Chinese jigging fleets in the northwest Pacific Ocean from September to November 2018. The study indicated that the statolith has the structure with a wide wing and long, narrow rostrum domes. The principal component analysis showed that the total statolith length (TSL), rostrum lateral dome length (RLL), lateral dome length (LDL), wing length(WL) and maximum width (MW) could be used as the morphological characteristic parameters to study the morphological changes of statolith. The analysis of covariance (ANCOVA) showed that there were significant differences in the relationships between MW, LDL and WL versus mantle length (ML) between males and females, but no significant difference was found in other characteristic parameters. The relationships between all of the statolith characteristic parameters and body weight (BW) differed significantly. The relationships between TSL, RLL and ML were optimally described by the linear growth models, and power functions were optimal for WL, MW of males and LDL of males, however, exponential function was optimal for LDL of females. The exponential functions were optimal for the relationship between the LDL, MW of males, and linear functions were optimal for LDL, MW, TSL of females, and power function for the other parameters. With the squid growth, the size of statolith gradually increased, but the ratios of DLL, LDL, RLL, WL to TSL almost remain at the same level, corresponding to 35.66%, 55.84%, 75.23% and 85.82%, respectively.

    • Relationship between the recruitment of the Pacific-cohort of chub mackerel (Scomber japonicus) and the influence factors on the spawning ground based on GLM and GAM

      2020, 44(1):61-70. DOI: 10.11964/jfc.20180811426

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      Abstract:Chub mackerel (Scomber japonicus) is an important pelagic fish in the Northwest Pacific Ocean. It is necessary for us to find the relationship between the abundance and influence factors, which are beneficial to exploit and utilize this resource. In this study, based on the recruitment data and the spawning stock biomass (SSB) data of the Pacific-cohort of S.japonicus during 1980—2016 obtained from Japan fisheries institution, the normality test of natural logarithm of recruitment was finished and the time period that passed the normality test was during 1980—1999, with the environmental data of spawning ground, we analyzed the relationship between the sea surface height (SSH), sea surface salinity (SSS), sea surface temperature (SST) and the natural logarithm of SSB [ln(SSB)] and the recruitment during 1980—1999 with generalized linear model (GLM) and generalized additive model (GAM). The GLM results revealed the order of importance of variables ranked by decreasing magnitude was ln(SSB)×Year, ln(SSB), SSS×Year and SSS, which were significant (P<0.05) and considered the combined effects of factors. Considering the single factor in GLM models affecting the recruitment, order of the importance of variables ranked by decreasing magnitude was SST, SSS, Year, ln(SSB) and SSS. The GAM results indicated that the model which contained Year, SST and SSH was the optimal model based on Akaike’s Information Criterion (AIC), the importance ranked by decreasing magnitude was Year, SST and SSH. However, considering the single factor in GAM models affecting the recruitment, the importance of variables ranked by decreasing magnitude was Year, SSS, ln(SSB), SST and SSH. The suitable range of SSH was 62—65 cm, the suitable range of SSS was 34.72—34.74 and 34.78—34.83 and the suitable range of SST was 20.2—20.6 ℃. When ln(SSB)>6.0,the recruitment was at a high level, based on GAM analysis.

    • Growth and feeding behavior development of a marine ecotoxicology mode fish-medaka (Oryzias melastigma)

      2020, 44(1):71-78. DOI: 10.11964/jfc.20181211603

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      Abstract:Aiming at the low survival rate of marine medaka (Oryzias melastigma), the experiment was carried out to understand the feeding behavior development of O. melastigma in early stage, so as to find a theoretical basis for breeding. The growth characteristics and feeding behavior of O. melastigma were observed and analyzed by feeding with Artemia nauplii during the first 90 days after hatching, by using a single camera, a waterproof-mirror and fish behavior analyze system. The results were as follows: its average growth rate in body length was 2.579%/d, and the relationship between body length and day-old age was y=3.132+0.383x−0.004x2+0.000 03x3, R2=0.98. Its larvae period could be divided into 3 stages (larvae stage was 0~10 d, juvenile stage was 11-30 d and young stage was 31-65 d) and 6 phases (pre-larvae was 0-4 d, post-larvae was 5-10 d, pre-juvenile was 11-18 d, mid-juvenile was 19-24 d, post-juvenile was 25-30 d, and young was 31-65 d) according to several indicators of the different feeding behavior. On the 4th day after hatching (DAH), O. melastigma were fed with Artemia nauplii that hatched within 12 hours. And all larvae can feed on Artemia nauplii on 10th day after hatching (DAH). There was high mortality rate in larvae stage but the feeding ability was improved steadily. By the end of the period, the feeding success rate increased to 45%-55%, and the feeding efficiency reached 0.5-0.6 ind./min. At the juvenile stage (26-30 d), all indicators of feeding behavior greatly improved, such as the response time to Artemia was shortened and the distance was elongated, the feeding rate increased, and food intake also increased. At the same time, the success rate of feeding rose to a high level, up to 90%-95%. At the young stage, all indicators of feeding behavior were very close to the adult fish. The feeding efficiency reached more than 9 ind./min. Its feeding behavior development was mature, and feeding function tends to prefect at this stage. Experiments show that O. melastigma had a low feeding ability with a short response distance to Artemia nauplii, and the feeding success rate and feeding efficiency were still at a low level. The feeding success rate and feeding efficiency were gradually increased with its own development and feeding ability improving, and the survival rate tended to be stable. It may be an effective way to increase the survival rate of breeding when paying close attention to the changes in feeding behavior of O. melastigma through changing the feeding quantity and frequency to make a faster larvae fish transition to the juvenile period.

    • Behavioral characteristics of large yellow croaker (Larimichthys crocea) in tank by ultrasound pinger system

      2020, 44(1):79-84. DOI: 10.11964/jfc.20181211582

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      Abstract:In order to understand the behavioral characteristics of large yellow croaker (Larimichthys crocea), the ultrasound pinger system has been used to track 4 tails of test fish for 24 hours to access the fish movement data by implanted methods on Aug. 27th and 28th, 2018. The results show that: ① the vertical movement depth of the test fish is (0.89±0.51)m (18:00—24:00), (0.73±0.50)m (0:00—6:00), (1.04±0.50)m (6:00—12:00), (1.00±0.45)m (12:00—18:00), and keep in 0.50~1.25 m; ②the horizontal movement of test fish appeared on the inside of tank about 159.0±9.50 times, and 27% in total data, and about 489.0±12.5 times appeared around tank, 73% in total data. This means that the fish are assembling inside the tank and do the random motion, occasionally the motion appeared around the tank wall. This experiment is the first time to study the behavioral characteristics in tank-farming by ultrasound pinger system, and it aims to provide a scientific theoretical basis and data support for the fishing management for production and behavior monitor of L. crocea.

    • Effect of dietary vitamin A on growth, serum biochemical index, digestive enzyme activities and glucose and lipid metabolism in juvenile Mylopharyngodon piceus

      2020, 44(1):85-98. DOI: 10.11964/jfc.20190511785

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      Abstract:An 8-week feeding trial was carried out to investigate the effect of dietary vitamin A (178.2, 2 058.9, 18 436.2 IU/kg) on growth performance, serum biochemical index and hepatic glucose and lipid metabolism key enzyme activities and gene expression levels of juvenile black carp (Mylopharyngodon piceus). Using a one-factor experiment design, three isonitrogenous and isoenergetic purified experimental diets (diet 1, diet 2 and diet 3) containing 0, 2 200 and 20 000 IU/kg VA were formulated. Total vitamin A contents in the diets were measured by HPLC (Agilent-1100, Agilent, USA), actual content of vitamin A in each diet were 178.2, 2 058.9 and 18 436.2 IU/kg, respectively. Casein (vitamin free) and gelatin were used as protein sources, rapseed oil was used as lipid source, dextrin was used as carbohydrate source. Total 360 juvenile black carp with initial weight (6.10 ±0.10) g were randomly divided into 3 groups with 3 replicates per group and 40 black carp per replicate. The results showed as follows: weight gain rate (WGR) and specific growth rate (SGR) were significantly reduced for fish fed diet with 178.2 IU/kg VA. When VA is deficient, concentrations of serum glucose (GLU), triglycerides and low-density lipoprotein (LDL) were decreased, but total cholesterol (TCH) concentration was increased. The activities of hexokinase (HK), phosphofructokinase (PFK) and pyruvate (PK) increased for fish fed diet with 2 058.9 IU/kg VA. Gene expression of liver glucose transporter-2 (GLUT-2), HK, GK, PFK and G6Pase were increased in fish fed diet with 2 058.9 IU/kg VA. However, there were no significant differences in the gene expression of fatty acid transporter protein-1 (FATP-1) for fish fed diet with 2 058.9 IU/kg VA. But the gene expression of Carnitine O-palmitoyltransferase-1 (CPT-1) and Carnitine O-palmitoyltransferase-2 (CPT-2) were significantly influenced. The gene expression of CPT-1 and CPT-2 were inhibited for fish fed VA deficient diet. When fish fed diet with 18 436.2 IU/kg VA, the expression of acetyl-CoA carboxylase-2 (ACC-2) and lipoprotein lipase (LPL) genes were inhibited. Besides, the gene expression of peroxisome proliferator-activated receptor-γ (PPAR-γ) decreased when the VA content was 18 436.2 IU/kg in feed. Overall, fish fed diet with 2 058.9 IU/kg VA could improve growth performance and ability of liver cells to transport glucose of juvenile black carps. Diet with 2 058.9 IU/kg VA could keep a balance between glycolysis and gluconeogenesis, and it also could promote fatty acids synthesis and transport.

    • Dietary protein requirement of juvenile giant salamander (Andrias davidianus)

      2020, 44(1):99-110. DOI: 10.11964/jfc.20181111533

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      Abstract:In order to determine the dietary protein requirement of juvenile giant salamander, 6 isolipidic diets were formulated to contain graded levels of D1(43.7%), D2(47.1%), D3(51.3%), D4(55.7%), D5(59.9%) and D6(64.4%) crude protein(dry matter) to feed juvenile giant salamander with initial weight (20.99±0.15)g for 92 days. The results show that: ① Dietary protein levels had significant effects on weight gain rate of giant salamander, and it reached the highest in the D4 group, which is 276.4% higher than group D1, and the whole body protein deposition rate and muscle RNA, RNA/DNA ratio, pepsin, H+-K+- ATPase in stomach, trypsin, lipase and Na+-K+- ATPase in intestine, liver superoxide dismutase (SOD) reached the best in the group D4, and the malondialdehyde (MDA) in liver and intestine were both the lowest in group D4. ② The content of muscle crude protein increased linearly with the increase of dietary protein level, while the fat content in giant salamanders declined linearly, and there was no significant difference in moisture and crude ash of whole body between different groups, and whole body crude protein increased first and then tended to be stable, reached the highest in the group D4. ③ The skin collagen content in group D4 reached the highest, increasing by 27.8% compared with D1. With weight gain rate, muscle RNA/DNA ratio, protein deposition rate, and skin collagen content as evaluation indexes, the optimum dietary protein level of the giant salamander was 55.9%-58.3%(based on dry mater), and this dietary protein level can significantly improve gastric acid secretion, the digestion and absorption, and antioxidant capacity, and increase the nutrient deposition, thus promote growth and feed conversion; While low protein level diet significantly inhibited the growth of A. davidianus.

    • Effects of south-north relay mode on the nutritive compositions of Haliotis discus hannai

      2020, 44(1):111-117. DOI: 10.11964/jfc.20181011475

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      Abstract:The south-north relay mode of abalone culture, a common practice in China, was designed to increase the survival rate of abalone in the south in summer. The abalone was usually transported to the north(Rongcheng, Shandong) in April from southern China(Lianjiang, Fujian)and returned to the south in November. In this study, the impact of this mode on the nutritive compositions of abalone (Haliotis discus hannai) was investigated. The main method was the series of GB.5009. The determined indexes include moisture, cholesterol, ash, protein, lipid, glycogen, collagen, mineral elements(Mg, Al, Ca, Mn, Fe, Cu, Zn, Se),delicious amino acids(aspartic acid, glutamic acid, glycine, alanine, arginine, methionine, lysine),taurine and fatty acid. SPSS 24.0 statistical software(independent sample t-test)was used to determine the significance of the differences between two groups. The experimental results were expressed as mean ±standard deviation.Two groups of adult abalone(two years old)with the same algae diet (Gracilaria lemaneiformis) were sampled for the analysis in December, 2017. One group was cultured in Lianjiang, Fujian (Southern China) without migration (LJ-DD) and the other was moved to Shandong (Northern China) before the summer and returned to Lianjiang in November (SD-DD). Live samples were moved to laboratory within 24 hours, taking abalone foot muscle to freeze drying after dissection and grinding into powder. These powder were saved in −80 ℃ freezer. The results showed that contents of ash and collagen were higher in LJ-DD, but not significant. The cholesterol, lipid and glycogen content were higher in SD-DD, but not significant. Moisture content was significantly higher in SD-DD (76.50%WW) than in LJ-DD(73.70%WW) while total protein content was significantly lower in SD-DD(48.40%DW). For mineral profile, Se content was significantly higher in LJ-DD(0.07 mg/100g)than SD-DD(0.05 mg/100g). Besides, Mg and Al contents were higher in SD-DD and Ca, Fe, Cu and Zn contents were lower in SD-DD, but the differences were not significant. For amino acid profile, the contents of glutamic acid, arginine, lysine and total delicious amino acid were significantly higher in LJ-DD than in SD-DD. Besides, taurine content was significantly lower in SD-DD than in LJ-DD. For fatty acid profile, C20:3n6 content was significantly higher in LJ-DD than in SD-DD. SFA and N-3 fatty acid were higher in SD-DD,and MUFA, PUFA and EPA+DHA contents were higher in LJ-DD,but not significantly. The value of N6/N3 was lower in SD-DD than in LJ-DD, but the differences were not significant. Overall,two groups had similar composition, but SD-DD had higher nutritional values than LJ-DD. To summarize, the south-north relay mode has both some negative influences and some positive influences on the nutritional quality of H. discus hannai, but such difference is not significant as a whole.

    • Effects of dynamic succession of Vibrio biofilms on settlement of the mussel Mytilus coruscus

      2020, 44(1):118-129. DOI: 10.11964/jfc.20190111652

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      Abstract:Vibrio, which are widely distributed in marine environments, have ability to form biofilms for the purpose of adaptation to harsh environmental conditions. Although biofilms formed by Vibrio have been known to promote larval and plantigrade settlement of the mussel Mytilus coruscus, how the dynamics succession of these biofilms during the formation impacts mussel settlement remains unknown. In this study, V. cyclitrophicus, V. chagasii and Vibrio sp. 22 with different settlement-inducing activities on plantigrades of the mussel M. coruscus were used to observe the changes of biofilm characteristics such as bacterial density, biofilm thickness and extracellular polymeric substances during the dynamic succession, and to explore subsequent effects of biofilm characteristics on mussel plantigrade settlement. The results showed that during the dynamic evolution of three Vibrio species, the bacteria on the biofilm were aggregated over time. The bacterial density and thickness of the Vibrio biofilms firstly increased with time and finally decreased. Except for Vibrio sp. 22, the bacterial density and biofilm thickness of V. cyclitrophicus and V. chagasii were correlated to the settlement of plantigrades. During the dynamic succession of Vibrio biofilms, the extracellular polysaccharides increased first with time and then began to decrease. In contrast, there was no change in the proteins and lipids on biofilms. The change trend of extracellular polysaccharide was similar to the settlement-inducing activity of biofilms, suggesting that extracellular polysaccharides play an important role in succession of biofilms regulating mussel settlement. Thus, the present finding is important to understand the interaction between biofilms and marine invertebrate settlement, and to clarify settlement mechanism of micro- and macro-organisms on artificial reefs.

    • Analysis of the effect of adding Bacillus on the physicochemical factors and bacterial community structure in ponds

      2020, 44(1):130-141. DOI: 10.11964/jfc.20181111520

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      Abstract:In order to study the effect of adding Bacillus in the pond on the physicochemical factors and bacterial community structure, high throughput sequencing technology was used to analyze the bacterial community structure in water and sediment of the experimental group (adding Bacillus in pond) and the control group (without adding). At the same time, the physicochemical parameters in water and sediment of the two groups were collected. The results indicated that the contents of TN, ${\rm{NH}}_4^ + $-N, ${\rm NO}_3^-$-N in water were significantly lower than those of the control group (P<0.05), but the contents of ${\rm{NO}}_3^ - $-N, ${\rm{NH}}_4^ + $-N, TN and TP in sediment were significantly higher than those of the control group (P<0.05). The abundance of Chlamydomonas debaryana, Zymomonas, Roseomonas, Dechloromonas and Chitinophaga in water of the experimental group was significantly higher than those in the control group (P<0.05). Among these bacterial communities, Dechloromonas and Chitinophaga have the function of removing ${\rm{NO}}_3^ - $-N, and Zymomonas and Roseomonas have the function of denitrification. The Chao1 and Shannon indexes in water of the experimental group were significantly higher than those in the control group (P<0.05). Hydrogenophaga, Geothermobacter and Haliscomenobacter were negatively correlated with ${\rm{NO}}_3^ - $-N and TP, and positively correlated with ${\rm{NH}}_4^ + $-N and TN. Above all, these results showed that adding Bacillus to the pond would lead to the variations of bacterial community structure, thus realizing the regulation of physicochemical factors in the pond. And the results are useful for reducing the pollution of aquaculture tail water to water environment.

    • Infection of tilapia lake virus in GIFT Oreochromis niloticus and E-11 cell

      2020, 44(1):142-155. DOI: 10.11964/jfc.20190111649

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      Abstract:In order to study the infection characteristics of TiLV in the cultured tilapia species and susceptible cells, GIFT Oreochromis niloticus and E-11 cells were chosen as models. For the present study, first of all, the whole nucleotide sequences of the fourth genome segment of TiLV from the experimental infected GIFT O. niloticus were determined. The length of the cDNA of the fourth genome segment was 1 250 bp containing an open reading frame of 1 065 bp, encoding a protein with 354 amino acids. The sequences and phylogenetic tree analysis showed that the fourth genome segment encoded TiLV Hemagglutinin-esterase-fusion (HEF) protein. Subsequently, GST fusion HEF was expressed in Escherichia coli and purified, and it was used to immunize New Zealand white rabbits according to the conventional method to prepare rabbit anti-HEF polyclonal antibody. The results showed that the antiserum titer obtained by ELISA was higher than 1∶51 200, and the serum could specifically recognize the HEF protein from the spleen of TiLV infected GIFT O. niloticus. Through artificial infection experiments, it was found that TiLV infected juvenile GIFT O. niloticus severely and caused surface ulceration, systemic bleeding and ocular lens opacity. Furthermore, hematoxylin and eosin (HE) stain showed the syncytium in liver, hemosiderin and vacuolar degeneration in spleen, necrosis in head kidney lymphocytes, protein precipitation and glomerulus necrosis in trunk kidney. Western blot and immunohistochemistry results showed that the virus was distributed in all the tissues with the higher abundance in the spleen, head kidney and gill than that in the liver, trunk kidney and brain tissues. Through indirect immunohistochemistry assay, it was found that HEF protein was mainly distributed in the cytoplasm in E-11 cells infected with TiLV. Our results demonstrate that TiLV infection could cause disease by targeting liver, spleen, kidney, gill and brain tissues of GIFT O. niloticus.

    • Postmorten biochemistry and rigor development of pufferfish (Takifugu rubripes)

      2020, 44(1):156-165. DOI: 10.11964/jfc.20190111636

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      Abstract:Takifugu rubripes were instant killed, bled and gutted, two treatments for 6 days storage were designed including storage in ice and at ambient temperature, respectively. During storage, the progression of rigor mortis and body temperature were recorded. Sampling and assay were carried out at both different rigor stages and every storage day, and rigor index, ATP and related compounds, glycogen, pH and phosphoprotein were selected as indicators as postmortem biochemical changes. The results showed that the onset of rigor mortis for iced treatment started at around 7th hour after being slaughtered, and its maximum rigor index was up to 89% with 20-24 hours duration in stiff, the fish body maintained at rigor with rigor index at 9%-16% after 4 days icing storage. However, the maximum rigor index of 71% was observed fleetingly in the control treatment with higher temperature ambient, and the whole rigor progression or stages were undefined. Glycogen degraded sharply at the initial post-mortem period for all treatments. Comparing with the trace amounts was still detected in 5 days storage for the iced treatment, glycogen was almost depleted after 3 days storage in the control treatment. The pH decreasing patterns of two treatments reflected the different glycogen consumption rates between the two storage treatments. ATP was rapidly depleted in all treatments with accumulation of IMP exclusively. Maximum of IMP reached to 9.5 μmol/g on the 3rd day and remained at about 7.5 μmol/g on the 6th day of the iced treatment. As to the control treatment, IMP was degraded rapidly after the maximum of 9.2 μmol/g on the 1st storage day with higher temperature. Compared to the iced treatment with a trace Hx amounts, the control treatment presented a significant accumulation of Hx. Muscle protein phosphorylation indicated that the rigor mortis progression of T. rubripes was also related to the properties of muscle protein components. Notably, the protein components related to glycolytic enzymes had a significant difference in phosphorylation between the iced and ambient treatment. The ambient treatment presented higher phosphorylation level, which is consistent with the glycolysis properties observed in ambient treatment. In conclusion, T. rubripes advanced in icing stability and high accumulation of IMP after being slaughtered, and the association between protein phosphorylation level and the process of rigor mortis is worth exploring further.

    • >ROUNDUPS
    • Role of TGF-β signaling pathway in sex determination and differentiation in fish

      2020, 44(1):166-177. DOI: 10.11964/jfc.20190711882

      Abstract (865) HTML (0) PDF 1.90 M (810) Comment (0) Favorites

      Abstract:Although the molecular mechanisms of sex differentiation and gonad development are conserved across vertebrates, the most upstream sex-determining genes are quite different for different groups, especially for fish, whose sex-determining genes show obvious diversity. There are two types of sex determination in vertebrates, i.e. environmental sex determination and genetic sex determination. Environmental sex determination is mainly affected by temperature, light, hormones and pH, while genetic sex determination is generally determined by sex-determining gene located on sex chromosome. Transforming growth factor-β (TGF-β) signaling pathway is involved in a variety of biological processes. In recent years, many studies have shown that multiple sex-determining genes are members of the TGF-β signaling pathway, and this signaling pathway also plays an important role in sex differentiation of fishes. This paper summarizes the sex-determining genes or candidate genes that have been discovered and reported in fishes, and reviews in detail the role of TGF-β signaling pathway involved in sex determination and differentiation in fishes. We also discussed the mechanism of TGF-β signaling pathway involved in sex determination, which is important in understanding the role of TGF-β signaling pathway in fish sex determination, differentiation and sex control.

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