2019 Vol. 15 No. 3
We analyzed 15 persistent organic pollutants (POPs) residues such as organochlorine pesticides and polychlorinated biphenyls in cultured shellfish (Mactra veneriformis, Ruditapes decussatus and Ostrea gigas, etc.) in Shandong coastal areas by gas chromatography (GC), comparing with the data at home and abroad, and investigated their residual level, spatial distribution and seasonal distribution characteristics. The risk assessment was evaluated by EPA method. The results show that w(HCHs), w(DDTs) and w(PCBs) were n.d.−17.6 μg·kg–1, 2.68−66.7 μg·kg–1 and n.d.−36.8 μg·kg–1 (wet mass, the same below). HCHs residues accorded with the first-class biological quality evaluation standard in Marine Biological Quality. Besides, 73% of samples had DDTs between the first and the second standards. OCPs and PCBs residues in all samples did not exceed the standard limits in National Food Safety Standard−Maximum Residue Limits for Pesticides in Food, Contaminant. There were no exposure risks and cancer risks were at acceptable level. Compared with the shellfish in other coastal waters, the residues of HCHs, DDTs and PCBs in the shellfish in Shandong coastal waters are at medium level.
A sensitive and reliable ultra-high performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) method was eatablished for the determination of olaquindox (OLA), carbadox (CBX), cyadox (CYA), quinocetone (QCT), mequindox (MEQ) and their main metabolites (QCA and MQCA) in fishmeal. The samples were extracted by acetonitrile-ethyl acetate mixture (1∶1, V∶V) and hydrochloric acid solution (1 mol·L–1), then the analyte which dissolved in the aqueous phase was re-extracted by ethyl acetate. The extraction was concentrated and reconstituted with acetonitrile. After purification with the Oasis PRiME HLB SPE cartridge, the sample was subjected to the following analytical procedure. The mobile phase containing acetonitrile and 0.1% formic acid solution with a linear gradient elution was utilized to separate all compounds on a Phenomenex Kinetex C18 column. The quantitative analysis of metabolites was carried out with an internal standard method and the others with an external standard method in the multiple reaction monitoring mode using positive electrospray ionization. The calibration curves for all compounds were linear (R≥0.994) within their corresponding concentration range. The recoveries were 64.4%−102.2% at different spiking levels with RSDs of 3.2%−10.2%. The limits of detection and quantification of MQCA and QCA were 2 μg·kg–1and 5 μg·kg–1; MEQ was 10 μg·kg–1 and 20 μg·kg–1; the others were 1 μg·kg–1 and 2 μg·kg–1, respectively. This method with high sensitivity and good precision can be applied to the simultaneous determination of quinoxaline drugs and their major metabolite in fishmeal samples.
We analyzed the resource dynamic, diversity and community structure of nekton based on the data collected by demersal trawl at Daya Bay mouth in 2015. The results show that the density of nekton in summer was significantly higher than that in the other seasons (F=4.556, P=0.01), and it was mainly affected by the seasonal alternation of nekton dominants. The first dominant species in each season were Paerargyrops edita (in spring), Thamnaconus hypargyreus (in summer), Leiognathus brevirostris (in autumn) and Portunus gladiator (in winter). The Shannon-Wiener diversity index of nekton community varied between 2.35 and 2.66, and the seasonal difference was insignificant (F=0.597, P=0.623). However, the cluster and non-measured multi-dimensional scale sequencing (NMDS) analyses show that the dissimilarity of nekton community in different seasons varied from 69.13% to 80.77%. Therefore, the single α diversity index can not reflect the characteristics of the nekton community of the survey area effectively. The results of redundancy analysis (RDA) indicate that there is a clear relationship between seasonal heterogeneity of nekton community and environmental factors such as water temperature, dissolved oxygen, salinity, transparency, pH and chlorophyll a.
We observed and determined the sagittae morphology of 105 individuals of genus Decapterus (Decapterus macrosoma, D. tabl and D. kurroides) collected from the Dongsha Islands in the South China Sea. The sagittae shape of genus Decapterus was long oval with developed rostrum, and the difference between species was distinct. The front end of D. macrosoma rostrum was slightly bent downward. The knob of otolith dorsal of D. tabl was obviously developed with serrated shape, and the front end of rostrum was slightly broad and thick. The overall outline of D. kurroides otolith was smoother, and antirostrum was poorly developed. According to the morphological indices (otolith length, otolith width, rostrum length, rostrum width, otolith mass and otolith area), the classification of different species of genus Decapterus (D. macrosoma, D. tabl and D. kurroides) achieved success rates of 100%, 100% and 91.7%, respectively. It is shown that sagittae can identify genus Decapterus.
To investigate the antibacterial substances from Bacillus amyloliquefaciens HE and antibacterial characteristics, we synthetized the genes related to lipopeptides synthesis by PCR detection according to the stability of B. amyloliquefaciens fermentation broth, and further analyzed the active substances by UPLC-Q-TOF-MS. Then we applied the inhibition zone method to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of lipopeptide against Aeromonas hydrophila, whose antimicrobial activity was discussed from the microscopic structure. The antimicrobial active substances were stable against high temperature, acid, acid-base and protease. The PCR with four primer pairs were cloned and sequenced. The BLAST analysis shows that the ituA, fenB, sfP and mycB genes existed in the genome of B. amyloliquefaciens HE. Three kinds of lipopetides (Surfactin, Iturin and Fengycin) were identified in the antimicrobial extract by mass spectrum analysis. The MIC and MBC of antimicrobial lipopetides against A. hydrophila were both 137.97 μg·mL−1 by inhibition zone method. The microscopic characteristics indicate that lipopeptides can cause membrane collapse and holes in A. hydrophila cells. The results provide references for the structural identification and safety evaluation of active metabolites of B. amyloliquefaciens.
We investigated the anaesthetic effect of eugenol at different mass concentrations on juvenile Siganus oramin with body mass of (4.83±1.30) g at natural seawater at temperature of (29.3±1.1) ℃. The results show that when the mass concentration of eugenol was 30−80 mg·L–1, the anesthesia time reduced but the recovery time increased with the increase of mass concentration of eugenol. During the anesthesia, the respiratory rate of juvenile S. oramin increased at light anesthesia state, but decreased rapidly during deep anesthesia until the respiratory arrested due to excessive anesthesia. When the mass concentration of eugenol was 30−100 mg·L–1, the juvenile S. oramin could reach Stage 4 anesthesia within 3 min and recovered less than 4 min with a survival rate of 100%. When the air exposure time was within 8 min, the recovery time slightly reduced with extension of air exposure time; when the air exposure time was longer than 8 min, the recovery time increased but the recovery rate decreased with the extension of air exposure time. It is indicated that eugenol, with advantages of fast anesthesia, short recovery time and high recovery rate, is an ideal anesthetic for juvenile S. oramin. The concentration of eugenol is suggested to be 30 mg·L–1 in production, and the artificial operation time should not exceed 6 min after anesthesia. The anesthesia concentration should not exceed 80 mg·L–1 even for faster anesthesia.
Chinese sea bass (Lateolabrax maculatus) with initial body mass of (25.49±0.20) g were fed for eight weeks with six diets containing 0 (control), 1% (T1), 2% (T2), 3% (T3), 4% (T4) and 5% (T5) Spirulina platensis, so as to investigate the effect of S. platensis on the growth performance, digestive enzymes, haematological indices and antioxidant capacity of L. maculatus. The results show that the dietary supplementation with S. platensis improved the weight gain rate (WGR) and specific growth rate (SGR) of L. maculates (P<0.05) significantly. The feed conversion ratio (FCR) decreased significantly (P<0.05). The protease activities in S. platensis supplemented groups were significantly higher than that of the control (P<0.05), while no significant change was observed in lipase and amylase activities among different groups. Compared with the control, the levels of red blood cell (RBC) increased in T4 and T5 groups significantly (P<0.05), and levels of white blood cell (WBC) and hemoglobin (Hb) in T3, T4 and T5 groups were significantly higher than those of the control (P<0.05). The levels of triglyceride, cholesterol, low-density lipoprotein cholesterol (LDL-C) in T4 and T5 groups decreased significantly than those of the control (P<0.05). The total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) activities in T3, T4 and T5 groups were significantly higher than those of the control (P<0.05). The malondialdehyde (MDA) contents in T3, T4 and T5 groups were significantly lower than those of the control (P<0.05). It is suggested that the suitable S. platensis supplementation level in diet of L. maculates is 4%−5%.
To study the role of TGF-β/Smad signaling pathway in zebrafish (Danio rerio) ovary under different photoperiod conditions, we detected the relative expression of relevant genes in TGF-β/Smad signaling pathway of zebrafish ovary under continuous dark (0 L∶24 D, DD), natural photoperiod (14 L∶10 D, LD) and continuous light (24 L∶0 D, LL) using quantitative real-time PCR. The location of p-Smad2 in zebrafish ovary was detected by immunohistochemical technology. The results show that the expression patterns of genes involved in TGF-β/Smad signaling pathway were different in zebrafish ovary under different photoperiod conditions, and the expression profiles of ligand (tgfb3), receptor (tgfbr2a, tgfbr2b and tgfbr1b) and downstream protein kinase (smad2 and smad3a) were consistent. The relative expression levels of the above six genes were the highest in DD group and the lowest in LL group after 3-day treatment of different photoperiods, and an opposite trend was observed after 7 d. The p-Smad2 protein signal was detected in the development of zebrafish oocytes from previtellogenic stage (PV) to full-grown immature stage (FG). Photoperiod treatment did not affect the localization of p-Samd2 in zebrafish ovary. The results suggest that change of photoperiod might affect the ovarian development of zebrafish by changing the expression pattern of genes of tgfb3/receptor/protein kinase.
The elongases of very long chain fatty acids (Elovls), which are key rate-limiting enzymes in the long-chain polyunsaturated fatty acid (LC-PUFA) biosynthesis pathway, can extend the carbon chain of PUFAs. To investigate the function of Elovl4 in PUFA biosynthesis of Trachinotus ovatus, we cloned its cDNA sequence (ToElovl4-like), whose open reading frame (ORF) was 792 bp, encoding 263 amino acids. Bioinformatics analysis reveals that the encoded amino acids contained significant structural features of Elovl family such as histidine cassettes, multiple transmembrane regions and ER retention signals. The results of sequence alignment show that Elovl4-like gene was highly conserved, and the similarity between the protein sequence encoded by this gene and other fish was 73%−86%, among which Gadus morhua had the highest similarity of 86%. Phylogenetic analysis shows that Elovl4 mainly clustered into three categories (Elovl4a, Elovl4b and Elovl4-like). Among them, ToElovl4-like was closely related with Elovl4-like of other fish. Real-time PCR shows that ToElovl4-like gene was expressed in all tissues, with the highest mRNA expression in gonads and spleens and the lowest in brain. The heterologous expression of yeast shows that ToElovl4-like could extend 18:2n-6 and 20:5n-3 to 20:2n-6 and 22:5n-3, respectively. The study provides a theoretical basis for understanding the biosynthesis of long-chain polyunsaturated fatty acids (LC-PUFA).
Pellino protein is a member of highly conserved E3 ubiquitin ligase which plays an important role in ubiquitination and innate immunity. In this study, we cloned and identified a crustacean Pellino from Penaeus monodon (PmPellino). The full-length cDNA of PmPellino consisted of 1 961 bp with an 89 bp 5'UTR, a 573 bp 3'UTR and a 1 299 bp open reading frame encoding 432 amino acids. PmPellino mRNA was detected in all the tissues examined by real time PCR (highest in gill). The expression of PmPellino in hepatopancreas was significantly up-regulated after acute ammonia nitrogen stress (P<0.01), but was inhibited in gill (P<0.01). Vibrio harveyi could significantly activate the expression of PmPellino in gill and inhibit its expression in hepatopancreas. V. anguillarum could significantly inhibit the expression of PmPellino in hepatopancreas, but there was no sigficant difference in gill with the control group. Staphylococcus aureus could significantly activate the expression of PmPellino in both hepatopancreas and gill. The results indicate that PmPellino might play an important role in the immunity of P. monodon.
A total of 58 180 microsatellites were developed by RAD-Seq high-throughput sequencing technology in Decapterus macrosoma. One hundred and twelve microsatellite loci were randomly selected from di-/tri-nucleotide repeat microsatellite loci. Finally, twenty-seven highly polymorphic markers were developed. The population genetic analysis finds 18 dinucleotide-repeated microsatellite loci (9−14 repeated) and 9 trinucleotide-repeated microsatellite loci (6−10 repeated). The number of alleles (Na) ranged from 5 to 17 (mean 10.6). The observed and expected heterozygosities ranged from 0.342 9 to 0.857 1 (mean 0.631 7) and 0.538 3 to 0.911 8 (mean 0.796 8), respectively. The polymorphism information content (PIC) ranged from 0.497 to 0.886 (mean 0.780 9), and the PIC at all loci except for one was geater than 0.500, indicating that the microsatellites were highly polymorphic. The Hardy-Weinberg equilibrium (HWE) test shows that 19 microsatellite markers conformed to HWE. There is no linkage imbalance among the 27 loci. The 27 microsatellites developed by this study will be useful for further population genetic analysis.
We observed and drawn the external morphology of 131 Thamnaconus septentrionalis collected from the Yellow Sea, and counted the number of dorsal fin rays, caudal fin rays, anal fin rays as well as pectoral fin rays, so as to conduct a regrassion analysis of their morphological features. Besides, the chromosome specimens were made by hot titration method, and the karyotype was analyzed. The results are: 1) when alive, the head and back of T. septentrionalis' body were in pewter and the abdomen was in whitish turquoise; after death, the color turned dark green. The fins' remaining turquoise was an obvious feature of its external morphology. 2) The body length was 1.5−3.3 times and 3.1−4.9 times as long as the body height and the head length; the head length was 1.1−1.6 times as long as the proboscis length. 3) The variation of total length/standard length was minimum with the smallest difference between female and male and had the best correlation coefficient (R2=0.96). The values of caudal length/caudal depth, standard length/caudal length, head length/eye diameter and head length/eye interal had the lowest correlation coefficient, and the difference between female and male was obvious. 4) The fin type was: dorsal fin II, 36−39; anal fin 34−37; tail fin 14−15, caudal fin 1+10+1, and the pelvic fin degenerated into pelvic fin spines. 5) The body was covered with a variety of fleece scales consisting of a base plate and a bony bulge, and the lateral lines were composed of trunk line and infraorbital line. 6) T. septentrionalis had 20 pairs of chromosomes (2n=40) and the karyotype formula was 2n=40t. All of them were telomeres without heteromorphic sex chromosomes.
During the pneumatic conveying process of automatic feeding systems for deep-water off-shore cage culture, blockages of pipeline and damage to feed pellets occur easily. To reveal the pneumatic transport characteristics of pellet feed in automatic feeding system, a numerical simulation of gas-solid two-phase flow of feed pellets in a pipeline was carried out. Based on gas-solid two-phase flow theory, both a computational fluid dynamics (CFD) model and discrete element model were established. The movement process of feed particles from initial stage to stable stage of pneumatic conveying pipeline was analyzed, and the position distribution of feed particles from initial stage to stable stage was obtained. Pressures at various locations in the pipeline were analyzed and compared to determine the pressure drop between the pipeline inlet and the instant at which the flow reached the steady state. The results provide references for the simulation and optimization design of automatic baiting equipment, which can better guide the movement of feed particles and reduce the collision energy loss of particles at elbow.
In this paper, the effects of ammonia-N (NH4-N) stress on the activities of lysozgme and three digestive enzymes of Babylonia areolata were studied to reveal its physiological changes. The results show that both the concentration and treatment time of NH4-N had a significant effect on the activities of digestive enzymes of B. areolata (P<0.05). Compared with the control group: 1) As the treatment time of NH4-N increased, the lysozyme generally demonstrated a suppression-induction trend. 2) The lower concentration of NH4-N (22 mg·L–1) showed an induction-suppression trend to the pepsin activity, while the other concentrations generally showed a suppression-induction trend. 3) The lipase activity of B. areolata was generally suppressed or similar with that of the control group with the prolongation of treatment time. 4) The NH4-N treatment time affected the amylase activity significantly, but in the case of short treatment time, the lysozgme activities of treatment groups did not have much induction or suppression. In conclusion, the stress of ammonia-N can influence the activities of these four digestive enzymes of B. areolata.
In order to study the rationality of Portunus trituberculatus release in Zhoushan fishing ground, Yangtze River estuary fishing ground and their adjacent waters in recent years, we analyzed the energy flow characteristics of the ecosystem in that sea area and estimated the proliferation capacity of P. trituberculatus based on the fishery resources survey data from 2006 to 2014 by constructing the Ecopath model. The results show that the ecosystem was mainly made up by benthos, and shrimps, Trichiurus leptures and P. trituberculatus had higher food utilization rates. The food utilization rates of Chelidonichthys kumu and Trachurus japonicus were relatively low. Four trophic levels can be divided in the survey area, and P. trituberculatus is a medium trophic organism. The total catch was 1.614 t·km–2 (the catch of P. trituberculatus was 0.057 8 t·km–2). The total consumption was 280.744 t·km–2. The total export was 790.396 t·km–2. The total production was 959.3 t·km–2. The biomass of P. trituberculatus was 0.125 t·km–2. The ecological capacity was 1.125 t·km–2. The proliferation capacity was 1 t·km–2. It is shown that the primary productivity of the survey area is high, and the marine organisms are distributed in the second and third trophic scales with relatively low ecosystem maturity. There is still a certain releasing space.