In order to research the genetic diversity of different populations of Paramisgurnus dabryanus, four populations from Liaoning, Henan, Hubei and Taiwan were analyzed by sequencing the mitochondrial cytochrome oxidase subunit Ⅰ(COⅠ) genes. The results showed that 642 base pairs (bp) fragments was consisted of A, T, C and G base with 23.5%, 30.6%, 26.7% and 19.2%, respectively, indicating a preference for A and T bases. A total of 44 mutations of nucleotide and 23 haplotypes were identified in 4 populations. The haplotype diversity index ranged from 0.424-0.855, and the nucleotide diversity ranged from 0.000 84-0.016 59. The results indicated that Paramisgurnus dabryanus from Taiwan population had a higher genetic diversity than Liaoning, Henan and Hubei populations. The genetic differentiation index (Fst) and genetic distance showed that genetic differentiation between Taiwan and other populations represents extremely significant difference, also had a further genetic distance with other populations. In general, the results showed that there was a certain genetic difference between the different geographical groups, AMOVA analysis revealed that 52.11% of genetic variations derived among populations and 47.89% of genetic variations occurred within populations. In terms of the negatively selective neutrality test, the results indicated that a population expansion occurred in the populations of Liaoning, Henan and Hubei population, and provided a reference for protection of the genetic diversity and breeding work of Paramisgurnus dabryanus.
Using magnetic-bead enrichment and PCR screening method, we obtained 19 pairs of microsatellite primers from Tridacna crocea. Comparison between Qilianyu and Yongxing Island wild populations with these markers shows: the mean allele number and the effective allele number were 11.105, 11.895 and 6.274, 6.173 respectively; the values of average expected heterozygosity were 0.776 and 0.788; the mean PIC was 0.730 and 0.744; high genetic diversity was maintained in those wild populations. Four loci significantly deviated from Hardy-Weinberg equilibrium after Bonferroni correction in each population. In addition, the transferability of all polymorphic short sequence repeats (SSRs) was assessed in five closely-related species. The test resulted in 7 loci amplifying and 6 loci being polymorphic in T. squamos; 3 loci amplifying and 1 loci being polymorphic in T. derasa; 5 loci amplifying and 5 loci being polymorphic in T. noae; 9 loci amplifying and 8 loci being polymorphic in T. maxima; 2 loci amplifying and 2 loci being polymorphic in H. hippopus.
The Pacific oyster Crassostrea gigas is cultured worldwide due to the advantages of rapid growth and good adaptability, and has become one of the most commercially important bivalve species. In our successively selective breeding, three strains of C. gigas with black, white and gold shell color traits have been developed. Tyrosinase (Tyr) is known as one of the most important enzymes in the regulation and production of melanin in animals. In this study, exons in the C. gigas TYR gene (CgTyr1) were sequenced. Mutations of the CgTyr1 gene and its association with shell color were analyzed in the three shell color strains of C. gigas. A total of 23 single nucleotide polymorphisms (SNP) were detected using single-strand conformation polymorphism (SSCP) and sequencing analysis, of which 11 SNP loci had highly significant differences between the three shell-color strains. In the SNP loci with significant difference, mutation c.591C/T, c.632G/A and c.1155T/C are non-synonymous which lead to amino acid changes Ala122Val, Gly136Ser and Phe310Ser. For further analysis, 11 SNP loci with a highly significant difference were selected for haplotype construction. One specific haplotype for every shell color strain was constructed and confirmed in the validating group. The mutations and haplotypes that are strongly associated with the shell color phenotypes in this study could be useful in understanding the molecular mechanism of pigmentation, and could also be potentially applied to marker-assisted selection breeding programs for C. gigas.
Spotted mackerel (Scomber australasicus) is an important economic species in northwest Pacific. The paper evaluates the Pacific stock biomass by age structure virtual population analysis (VPA) and yield per recruit model, and analyses the utilization of stock biomass by using the mackerel catch and CPUE data of the Pacific group which is supplied by Japan’s central fisheries research institute from 1995 to 2015. The results show that the biomass of spotted mackerel remains at a high level. The stock biomass is about 650000 tons in 2015, the average fishing mortality shows a trend of falling volatility and fishing mortality of 2015 is 0.15, the current fishing mortality (Fcur) is 0.33, the spawning potential ratio (SPR) remains 36.5%. There is no growth overfishing and recruitment overfishing. It is also discussed that the fluctuations of natural mortality caused by water temperature change and different fishing age affect the resources of spotted mackerel. The study suggests that the fishery is currently utilized in the sustainable way and has great potential for development. It also suggests that the fishery resources could be developed and utilized by using the management reference point F0.1.
In order to complement the basic biological information of Oratosquilla oratoria and provide managers with scientific guidance and theoretical basis, based on the size composition data of O. oratoria obtained from fishery independent surveys from 2016 to 2017 in coastal waters of Shandong Peninsula, this study estimated the growth and mortality parameters of O. oratoria. A length-structure yield per recruitment model was used to assess the population dynamics and management strategy of the species. A total of 5 028 individuals were caught, from which the length-weight relationship was derived as W=0.014 5L2.88, showing a negative allometric growth pattern. Asymptotic body length and growth coefficient estimated by ELEFAN were L∞=19.87 cm and K=0.62 a−1, respectively. The growth rate of O. oratoria showed remarkable seasonal oscillation, the highest in October and the lowest in April, and the amplitude of the seasonal oscillation (C) was 0.76. The total mortality (Z) estimated by length-converted catch curve was 3.24 a−1, and the natural mortality (M) estimated by several estimators ranged from 0.75 to 1.27 a−1. Accordingly, the fishing mortality (F) ranged from 1.96 to 2.49 a−1, and the mean exploitation rate was 0.67. Results of YPR model showed that with the F increasing, YPR tended to increase at first, and then decrease. Biological reference points F0.1 and Fmax were 0.92 a−1 and 1.88 a−1, respectively. This study shows that O. oratoria stock is over-exploited. In order to maintain stock and fishery of O. oratoria, fishing pressure needs to be reduced and length at first capture should be increased.
The growth and maximum photon quantum yield of photosystem II (Fv/Fm) of the blades of two high-temperature resistant strains (YZ-4 and TM-18) and one wild-type strain (WT, used as control group) in Pyropia yezoensis were analyzed at high-temperatures or low-salinities, in order to screen out strains resistant to the two abiotic stresses. The results showed that YZ-4 and TM-18 blades grew faster than those of WT for 50-85 d under the optimal culture conditions (18 °C and salinity of 26). The ratios of Fv/Fm and contents of photosynthetic pigments were also higher than those of WT. The absolute growth rate and Fv/Fm of every strain showed high positive correlations. In addition, WT blades began to form spermatangia around 70 d, their absolute growth rate and Fv/Fm decreased significantly. When cultured at high-temperatures, Fv/Fm and absolute growth rates of the blades of every strain decreased. The higher the temperature or the longer the stress lasted, the greater the decline would be. Fv/Fm of WT, YZ-4 and TM-18 decreased by 56.7%, 43.2% and 28.7%, respectively, after 35 d of high-temperature stress at 24 °C. The growth of WT was completely stagnated after 15 d of high-temperature stress at 25 °C. Meantime, growth rates of YZ-4 and TM-18 were 0.51 and 0.84 cm/d, respectively, indicating that they were veritable high-temperature resistant strains. The effects of low-salinity stress on the blades were similar to those of high-temperature stress. After 35 d of low-salinity stress at salinity of 9, Fv/Fm of WT, YZ-4 and TM-18 decreased by 46.2%, 42.0% and 32.0%, respectively. The absolute growth rates of WT, YZ-4 and TM-18 were 0.12, 0.10 and 0.90 cm/d, respectively. Besides, growth of the blades of WT and YZ-4 cultured at salinity of 5 was greatly affected, in that the blades were deepened in color, curled and rotted after 15 d under stress, while TM-18 did not rot at the same salinity of 5 for 25 d. These results indicated that TM-18 was more tolerant to high-temperature and low-salinity stress than WT and YZ-4, and the internal reason was that the decrease of Fv/Fm of TM-18 blades was small. It indicated indirectly that Fv/Fm could be used as a new index for breeding of stress-resistant strains in P. yezoensis.
In order to explain the influence of temperature and light intensity on the growth process of Saccharina japonica sporophyte and explore the physiological response mechanism of the temperature and light environment from the perspective of physiological ecology. In this study, based on the growth parameters of S. japonica sporophyte, we set up four water temperature gradient (6, 10, 14 and 18 °C) of S. japonica sporophyte in a laboratory experiment, and determined the chlorophyll fluorescence parameters at night with 9 photochemical light gradients (0, 25, 70, 133, 230, 317, 421, 582, 786 μmol photons/(m2·s)). Results showed that: ① under the condition of 6 °C, S. japonica sporophyte fluorescence parameters (Fv/Fm) and (Fv/F0) maximum value was 0.71 and 2.40 respectively; when water temperature 18 °C, its (Fv/Fm) and (Fv/F0) minimum value was 0.65 and 1.85 respectively. ② The maximum values of photochemical quenching and non-photochemical quenching of S. japonica sporophyte occur at 18 °C (0.92 and 3.29, respectively). ③ The light response curve of kelp first increased and then decreased with the enhancement of PAR. ④ The maximum leaf length growth rate, leaf width growth rate and dry weight growth rate of S. japonica sporophyte were 1.34 cm/d, 0.33 cm/d and 1.01 g/d, respectively. These results indicated that the change of dry weight growth rate was consistent with the change of light response curve under different water temperature conditions, and high temperature inhibited the photosynthetic efficiency of S. japonica sporophyte. When the ambient photosynthetic effective radiation was larger than the light saturation point (Em), the relative electron transfer rate of kelp decreased and photosynthesis was inhibited.
The emergence of novel variety “All-male NO.1” dramatically promoted the development of the Pelteobagrus fulvidraco industry, of which the cultivation of YY super-male was a critical step. Recently, YY super-male P. fulvidraco with intersexual gonad emerged largely, which hindered the development of P. fulvidraco industry. We only found the problem of gonad development in YY super-male breeding with Tubificidae by checking the production and breeding records for nearly 10 years. Therefore, four different baits including Artemia salina, zooplankton, Chironomus plumosus, Tubificidae were used to treat YY super-male for 20 days (11 to 30 days of age post hatching). The survival rate, body length, and body weight of each group were measured at 60 dph (day post-hatching). Tubificidae treatments significantly increased the body length and weight compared with other baits, and the survival rate of fish fed with Artemia salina was significantly lower than fish fed with the other three baits. In addition, we performed histology analysis on gonadal structure at 60 dph and 1 year old and statistics of fertilization rate at 1 year old. As a result, the fish group fed with Tubificidae displayed 75% intersexual gonad and 25% testis without seminiferous lobule, and the fertilization rate was only 36.70%±4.05%, which was significantly lower than that of the other groups. In order to study the reasons for the feminization of YY P. fulvidraco, we measured the estradiol content and found that the estradiol content was low in all four different animal baits. It is speculated that the feminization of YY super-male P. fulvidraco may be caused by the environmental endocrine disruptors (EDCS) enriched by Tubificidae. Therefore, zooplankton or Chironomus plumosus should be fed in the early stage of large-scale breeding of YY-supermale P. fulvidraco, whereas Tubificidae should not be fed.
In April 2018, a large number of largemouth bass (Micropterus salmoides) were infected with lethal sarcoidosis in a farm in Qionglai, Sichuan Province. In order to define the potential pathogens, a bacterial strain named HSY-NS02 was isolated from skin ulcers, effusions from fish swimming bladder’s cavity and liver of the diseased M. salmoides by traditional pathogen isolation methods. The strain HSY-NS02 was identified as Nocardia seriolea by means of morphologic structure and staining observation, PCR amplification and sequence analysis of 16S rRNA gene and specific primer, constructing phylogenetic tree, physiological and biochemical test. Furthermore, the pathogenicity of the strain HSY-NS02 was confirmed by the infection experiment in healthy M. salmoides. The results showed that the strain HSY-NS02 was cofirmed to be pathogenic to healthy M. salmoides and the same bacterium could be recovered from these infected M. salmoides. Histopathological observation and analysis of diseased M. salmoides showed that there were different degrees of chronic granulomatous lesions in skin ulcers, heart, liver, spleen, kidney and gill, of which spleen lesions were the most serious. The antibiotic susceptibility of the strain HSY-NS02 was carried out to guide clinically choosing medicine, and the results showed that the strain was sensitive to gentamicin, neomycin and nystatin, but resistant to other 18 kinds of antibiotics, which implied the strain HSY-NS02 was resistant to multiple antibiotics.
In order to determine the pathogen, which can cause the death of cultured Paralichthys olivaceus from ascites disease in Beidaihe area, Hebei Province, three dominant bacteria were isolated from P. olivaceus infected with ascites. The biological status of the isolates was determined by physiological and biochemical identification and 16S rRNA sequence alignment. The pathological characteristics of the isolates were further identified by virulence genes (toxR, vhhA, vhhB) and histopathological analysis. The results showed that the three isolates were Vibrio harveyi and the virulence genes of the three isolates were all positive. Pathological sections showed that the isolates could be used to damage multiple organs (intestine, kidney, spleen and liver) of P. olivaceus. The LD50 of strain BDHYPFS-Y1G to P. olivaceus was 5.88×106 CFU/g, which was lower than the toxicity of natural state. Drug susceptibility test indicated that all three isolates were highly sensitive to the nitrofurantoin. This study confirmed the pathogen of this ascites disease, and preliminarily studied the pathogenicity and drug sensitivity of the pathogen, which can provide scientific basis for the prevention and control of the disease in industrial culture of flounder.
Recently, tilapia lake virus (TiLV) has been epidemic in many countries and posed a serious threat to Oreochromis spp. aquaculture industry. China has contributed the most amount of cultured Oreochromis spp. in the world. Up to date, there is no report of the TiLV epidemic in Oreochromis spp. in the mainland of China. However, since GIFT O. niloticus is one of the most cultured Oreochromis spp. species in the mainland, therefore it is necessary to characterize the features of the GIFT strain infected with TiLV. Taking the advantage of the TiLV which was kindly given as a gift by Dr. Sven Bergmann from Institute of Infectology, Friedrich Loffler Institute, we performed the infection of TiLV in the GIFT strain. The whole nucleotide sequences of the sixth genomic segment of TiLV from the experimental infected O. niloticus were determined. The length of the cDNA of the sixth genomic segment was1 044 bp containing an open reading frame of 954 bp encoding a protein with 317 amino acids with predicted molecular weight of 36.38 ku. There is 5′ end non-coding region of 19 bp and 3′end non-coding region of 972 bp. The sequences and phylogenetic tree analysis showed that the sixth genomic segment encoded TiLV nucleoprotein (NP). Subsequently, GST fusion NP was expressed in Escherichia coli and purified, and it was used to immunize New Zealand white rabbit (Albus lepus) according to the conventional method to prepare rabbit anti-NP polyclonal antibody. The results showed that the antibody titer obtained by ELISA was higher than 1:51 200, and the antibody could specifically recognize the NP protein from the tissues of O. niloticus infected with TiLV. Hematoxylin-eosin staining (H.E) was performed on different tissues of O. niloticus. The results showed that there were apparent pathological changes in the observed tissues, including hepatic necrosis and syncytium; vacuolization, necrosis and increased amount of hemosiderin in the spleen; necrosis and inclusion body in the head kidney; dissociation and shedding of the epithelial cells of the gill filament, small pieces adhered to each other; vacuoles of nerve cells in the brain tissue. Western blot and immunohistochemistry (IHC) were used to detect the expression of the NP protein in different tissues of O. niloticus infected with TiLV. The results showed that the highest amount of NP protein was expressed in the liver, followed by the brain, trunk kidney and head kidney. In order to elucidate the immune responses of O. niloticus to the TiLV infection, real-time quantitative PCR (qRT-PCR) was used to measure the mRNA expressions of TNF-α and TGF-β in the spleen and head kidney which are the two major immune tissues of fish. The results showed that during the early period of the infection (12-24 h post infection), the expression of both TNF-α and TGF-β was significantly inhibited by the viral infection, indicating that TiLV might inhibit these cytokines so as to facilitate its early replication in the host. The current study will shed new light on the pathogenesis of TiLV infection and will pave a new way for the development of effective prevention and control strategy against the epidemic of TiLV in O. niloticus.
The four experimental groups were set up to test the response of Carassius auratus and Ctenopharyngodon idella to ammonia toxicity and taurine within 96 h. Group 1 was injected with NaCl, group 2 was injected with ammonium acetate (C. auratus 7 mmol/g; C. idella 9 mmol/g), group 3 was injected with ammonium acetate and taurine (100 μg/g), and group 4 was injected with taurine. C. auratus in group 2 had lower mRNA expression of SOD, CuZnSOD and CAT in liver than those of tested fish in groups 1 and 3; tested fish in groups 2 and 3 had lower mRNA expression of GPx in liver than that in group 1; the highest mRNA expression of SOD, CuZnSOD, CAT and GPx in brain were found in group 1; C. idella in groups 2 had higher mRNA expression of SOD, CuZnSOD, CAT and GPx in liver; fish in groups 2 and 4 had lower mRNA expression of SOD and CuZnSOD in brain than those in groups 1 and 3; fish in group 3 had the highest mRNA expression of CAT and GPx in brain. C. auratus and C. idella in group 2 had the highest mRNA expression of TNF and IL in the brain and liver. This study indicates that defensive strategies are more effectively on C. idella in dealing with the ammonia challenge compared with C. auratus; the taurine could more effectively mitigate the adverse effect of oxidative stress on C. auratus and C. idella, but the inflammatory response in hyperammonemia C. auratus and C. idella was not alleviated by taurine.
In order to study the function of serum amyloid A (SAA) in the immune defense of Macrobrachium nipponense, the full-length cDNA sequence of the SAA gene in M. nipponense, named MnSAA, was cloned using rapid amplification of cDNA ends (RACE) method. The expression levels of MnSAA in different tissues and different development stages of M. nipponense, and the expression profile of MnSAA in hemocytes and hepatopancreas of M. nipponense after Micrococcus lysoleikticus, Aeromonas hydrophila and white spot syndrome virus (WSSV) infections were also examined by quantitative real-time PCR (QPCR). At the same time, the expression changes of activator protein-1 (AP-1) and class B scavenger receptor (CD36) genes in hepatopancreas and cumulative mortality of prawn after MnSAA gene silencing plus A. hydrophila infection were further investigated using RNA interference (RNAi) technology. The results showed that the full-length cDNA of MnSAA was 649 bp, including a 21 bp 5′-untranslated region (UTR), a 232 bp 3′UTR, and a 369 bp open reading frame (ORF) encoding a deduced protein with 131 amino acids. Sequence and phylogenetic analyses revealed that MnSAA belongs to acute-serum amyloid A (A-SAA) and had the closest relationship with the invertebrate Crassostrea hongkongensis. mRNA expression analysis revealed that MnSAA gene was ubiquitously expressed in various tissues and growth stages of M. nipponense, and abundantly expressed in the hepatopancreas and adult prawn stage. Pathogen infection experiment showed that the expressions of MnSAA in hemocytes and hepatopancreas were all significantly increased compared with the control group after M. lysoleikticus, A. hydrophilahe and WSSV infections for 12-72 h. The RNAi experiment showed that after MnSAA silencing, AP-1 and CD36 gene expressions in hepatopancreas of M. nipponense were all significantly decreased compared with the control group after 12-72 h and 6-72 h, respectively, of A. hydrophila infection, and the cumulative mortality of prawn was increased compared with the control group after A. hydrophila infection. These results suggested that MnSAA is involved in the activity of anti-pathogen infection in M. nipponense and plays an important role in the immune defense system of M. nipponense.
Different thermal processing methods have great influence on the quality of Litopenaeus vannamei. In this paper, microwave-infrared combined heating technology was used to treat L. vannamei. The effects of combined heating conditions on the quality of L. vannamei were discussed, in order to lay a theoretical foundation for the development of shrimp products. Firstly, the change of center temperature of shrimp during microwave-infrared heating was clarified, and then the weight loss rate, color, texture and other quality changes of shrimp meat under different heating conditions were discussed. The results showed that at the same microwave power and different infrared temperatures, the weight loss rate of L. vannamei was the highest at 75 °C, and the lowest at 179 °C. Comparing the texture characteristics of the shrimp, the elastic fluctuation range of the shrimp was not large, about 3.0 mm. The chewiness of shrimp was not significantly different under different heating conditions, but positively correlated with its corresponding hardness, cohesiveness and resilience; the hardness, cohesiveness and resilience of shrimp meat changed regularly under different combined heating conditions; in terms of color difference, L*, a*, b* and ΔE were negatively correlated with infrared temperature. These results may lay a foundation for the application of combined heating technology in shrimp processing.
Germ cell transplantation is a technique that donor germ cells successfully colonized into genital ridges of recipient, proliferate, differentiate and finally develop into functional donor gametes after they are transplanted into the body cavity of allogeneic or xenogeneic recipients. It is a powerful assisted reproductive technique for the conservation and propagation of the rare and endangered animals; meanwhile, it also provides an effective assay system for the functional analysis of germline stem cell. Germ cell transplantation in fish was first successfully performed in the model fish, zebrafish (Danio rerio); during the past decade, a series of breakthroughs have been achieved for this technique. Firstly, three types of fish germ cell transplantation have been established to produce donor-derived gametes by transplanting germ cells into recipients at diverse developmental stages, such as the blastula-stage embryos, newly hatched larvae, and adult fish. Secondly, the discovery of spermatogonia and oogonia stem cells has expanded the choice of donor germ cells. Thirdly, the method for selection and preparation of recipient has been improved. This technique has powerful applications in shortening the time of fish sexual maturity, sex control breeding and conservation of the rare and endangered fish, which has been studied and applied in a variety of teleost, including freshwater and marine fish. In this review, we systematically summarized the progress of germ cell transplantation in fish, pointed out the key points for the implementation of this technique, and discussed its promising application.