Objective:To develop the rapid and efficient quantitative detection tool for nervous necrosis virus isolated from sevenband grouper Hyporhodus septemfasciatus.Methods:The viral genes of the NNV(SGYeosu08) isolated fro...Objective:To develop the rapid and efficient quantitative detection tool for nervous necrosis virus isolated from sevenband grouper Hyporhodus septemfasciatus.Methods:The viral genes of the NNV(SGYeosu08) isolated from sevenband grouper were phylogenetically analyzed.In addition,novel quantitative PCR primers based on the genomic sequence of SGYeosu08 isolate were designed and compared it with the conventional bio-assay method(TCID_(50)) using in vitro and in vivo samples.Results:The phylogenetic analysis of viral genes demonstrated the relationship of SGYeosu08 with members of red-spotted grouper nervous necrosis virus(RGNNV).The qNNV_Rl primer set(R1_F and R1_R) and the qNNV_R2 primer set(R2_F and R2_R) revealed 93%primer efficiency(regression:y=-0.2861 x + 9.9401,R^2= 0.9976)and the revealed 108%primer efficiency(regression:y=-0.3172 x + 10.0611,R^2= 0.9982),respectively.Its comparison with viral infectivity calculated by TCID_(50) method showed similar kinetic pattern at in vitro and NNV challenged fish(in vivo) samples.Conclusions:Result show that this method is rapid and efficient to diagnose NNV infection compare to traditional bioassay method(TCID_(50)).展开更多
Viral nervous necrosis(VNN)disease,caused by the nervous necrosis virus(NNV),is a devastating disease,leading to high mortality rate and huge economical loss in aquaculture.In the past 30 years,many studies on the vir...Viral nervous necrosis(VNN)disease,caused by the nervous necrosis virus(NNV),is a devastating disease,leading to high mortality rate and huge economical loss in aquaculture.In the past 30 years,many studies on the virus,host responses to the virus infection and diagnostics have yielded a lot of knowledge on developing measures to prevent the VNN disease.Although genetic improvement of disease resistance through breeding is inheritable and has long-lasting positive effect on aquaculture,it is a very challenging task in conventional selective breeding for improving disease resistance.With the advances in mapping quantitative trait loci(QTL)and genome wide association studies(GWAS)for NNV resistance in aquaculture species,DNA markers and genes associated with NNV resistance have been identified,making the application of marker-assisted selection(MAS)and genomic selection(GS)for NNV resistance possible.A few genes for NNV resistance are identified and are being used for genome editing to rapidly improve NNV resistance.In this review,we present the current knowledge on the NNV,host responses to NNV infection,diagnostic methods and vaccines available for NNV disease.In addition,we summarize the current status of conventional and molecular breeding for NNV resistance and highlight future directions,including genome editing for improving NNV resistance in aquaculture.展开更多
[Objective] To optimize the prokaryotic expression of MCP gene of red-spotted grouper nervous necrosis virus. [Method] The MCP gene was amplified from red-spotted grouper nervous necrosis viral genome by RT-PCR. The r...[Objective] To optimize the prokaryotic expression of MCP gene of red-spotted grouper nervous necrosis virus. [Method] The MCP gene was amplified from red-spotted grouper nervous necrosis viral genome by RT-PCR. The recombinant expression vector pRSET A-MCP was constructed and transformed into BL21(DE3)plysS to express proteins with induction in different media, at different pH, or at different temperatures. [Result] The expression level of recombinant bacteria reached a peak with induction under the following condition: SOB or LB medium, pH 7.0, 37 ℃ while the fusion protein was about 44.5 kD in molecular weight. [Conclusion] This study provided a basis for the development of RGNNV-MCP vaccine.展开更多
Viral nervous necrosis (VNN) is a worldwide disease among teleost fish. In the mainland of China, VNN was first identified in 2 species of hatchery-reared groupers, Epinephelus akaara and E. coioides. In the present s...Viral nervous necrosis (VNN) is a worldwide disease among teleost fish. In the mainland of China, VNN was first identified in 2 species of hatchery-reared groupers, Epinephelus akaara and E. coioides. In the present study, samples were collected from larvae of E. akaara with signs of VNN in Dayawan bay which is located in southern China. The complete viral coat protein gene was amplified using extracted total RNA as template. The reverse transcription polymerase chain reaction (RT-PCR) amplification was performed using primers containing a heterologous restriction site for NotI. PCR products were cloned into pcDNA3 vector and sequenced. The results indicated that the coat protein gene of Dayawan isolate (RG-CN) was 1056 bases in length and contained a single open reading frame (ORF) of 1017 bases encoding a protein of 338 amino acids. The sequence similarities between the coat protein gene of RG-CN and other 8 isolates of NNV from Dayawan, Taiwan, Japan, Singapore and France were 79.1%-99.5% at the nucleotide level and 83.7%-100% at the amino acid level, respectively.The homology between RG-CN and the other 5 isolates from groupers was high and relatively lower between RG-CN and guppy nervous necrosis virus (GNNV), sea bass nervous necrosis virus (DIEV), but more divergences existed between RG-CN and striped jack nevous necrosis virus (SJNNV). Compared with SJNNV, RG-CN and the other 7 isolates all lacked 6 nucleotides and 2 amino acids in the same positions. Based on the result of molecular phylogenetic analysis, Dayawan isolate belongs to red-spotted grouper nervous necrosis virus (RGNNV) genotype.展开更多
As a highly important fish virus,nervous necrosis virus(NNV)has caused severe economic losses to the aquaculture industry worldwide.Autophagy,an evolutionarily conserved intracellular degradation process,is involved i...As a highly important fish virus,nervous necrosis virus(NNV)has caused severe economic losses to the aquaculture industry worldwide.Autophagy,an evolutionarily conserved intracellular degradation process,is involved in the pathogenesis of several viruses.Although NNV can induce autophagy to facilitate infection in grouper fish spleen cells,how it initiates and mediates autophagy pathways during the initial stage of infection is still unclear.Here,we found that red-spotted grouper NNV(RGNNV)induced autophagosome formation in two fish cell lines at 1.5 and 3 h post infection,indicating that autophagy is activated upon entry of RGNNV.Moreover,autophagic detection showed that RGNNV entry induced incomplete autophagy by impairing the fusion of autophagosomes with lysosomes.Further investigation revealed that binding of the RGNNV capsid protein(CP)to the Lateolabrax japonicus heat shock protein HSP90ab1(LjHSP90ab1),a cell surface receptor of RGNNV,contributed to RGNNV invasion-induced autophagy.Finally,we found that CP blocked the interaction of L.japonicus protein kinase B(AKT)with LjHSP90ab1 by competitively binding the NM domain of LjHSP90ab1 to inhibit the AKT-mechanistic target of the rapamycin(MTOR)pathway.This study provides novel insight into the relationship between NNV receptors and autophagy,which may help clarify the pathogenesis of NNV.展开更多
Using the extended homogenous balance method, we obtainabundant exact solution structures ofa (2+1)dimensional integrable model, the generalized Nizhnik-Novikov-Veselov equation. By means of the leading order termanal...Using the extended homogenous balance method, we obtainabundant exact solution structures ofa (2+1)dimensional integrable model, the generalized Nizhnik-Novikov-Veselov equation. By means of the leading order termanalysis, the nonlinear transformations of generalized Nizhnik-Novikov-Veselov equation are given first, and then somespecial types of single solitary wave solution and the multisoliton solutions are constructed.展开更多
报道了一种可以快速、灵敏地检测七带石斑鱼神经坏死病毒(NNV)的逆转录环介导等温扩增方法(RT-LAMP)。该方法参考赤点石斑鱼NNV病毒的主衣壳蛋白(MCP)基因的保守序列,设计1对引物克隆出七带石斑鱼NNV的基因序列,利用Primer Explorer V4...报道了一种可以快速、灵敏地检测七带石斑鱼神经坏死病毒(NNV)的逆转录环介导等温扩增方法(RT-LAMP)。该方法参考赤点石斑鱼NNV病毒的主衣壳蛋白(MCP)基因的保守序列,设计1对引物克隆出七带石斑鱼NNV的基因序列,利用Primer Explorer V4软件设计了3对针对所克隆NNV基因序列的特异性引物,建立了RT-LAMP的反应体系,并分别对反应系统的引物浓度、反应温度和时间进行了优化,形成了七带石斑鱼NNV病毒RT-LAMP检测技术。实践应用结果表明,在63℃的最佳反应温度下,采用RT-LAMP检测技术经过45 min就能完成一次检测,准确率达到100%。本研究建立的RT-LAMP检测NNV技术设备要求简单,为七带石斑鱼等石斑鱼无NNV受精卵和仔稚鱼的生产现场检测和筛选提供了一种方便、灵敏的检测方法。展开更多
With the help of an improved mapping approach and a linear-variable-separation approach, a new family of exact solutions with arbitrary functions of the （2＋1）-dimensional Nizhnik-Novikov-Veselov system （NNV） is d...With the help of an improved mapping approach and a linear-variable-separation approach, a new family of exact solutions with arbitrary functions of the （2＋1）-dimensional Nizhnik-Novikov-Veselov system （NNV） is derived. Based on the derived solutions and using some multi-valued functions, we find a few new folded solitary wave excitations for the （2＋1）-dimensional NNV system.展开更多
Exact periodic-wave solutions to the generalized Nizhnik-Novikov-Veselov (NNV) equation are obtained by using the extended Jacobi elliptic-function method, and in the limit case, the solitary wave solution to NNV equa...Exact periodic-wave solutions to the generalized Nizhnik-Novikov-Veselov (NNV) equation are obtained by using the extended Jacobi elliptic-function method, and in the limit case, the solitary wave solution to NNV equation are also obtained.展开更多
Some new structures and interactions of solitons for the （2＋1）-dimensional Nizhnik-Novikov-Veselov equation are revealed with the help of the idea of the bilinear method and variable separation approach. The soluti...Some new structures and interactions of solitons for the （2＋1）-dimensional Nizhnik-Novikov-Veselov equation are revealed with the help of the idea of the bilinear method and variable separation approach. The solutions to describe the interactions between two dromions, between a line soliton and a y-periodic soliton, and between two y-periodic solitons are included in our results. Detailed behaviors of interaction are illustrated both analytically and in graphically. Our analysis shows that the interaction properties between two solitons are related to the form of interaction constant. The form of interaction constant and the dispersion relationship are related to the form of the seed solution （u0, v0, w0 ） in Backlund transformation.展开更多
基金a part of the project titled'Production of diagnostic antibodies for viral diseases in aquatic animals'(Project No.20150259)funded by the Ministry of Oceans and Fisheries,Korea
文摘Objective:To develop the rapid and efficient quantitative detection tool for nervous necrosis virus isolated from sevenband grouper Hyporhodus septemfasciatus.Methods:The viral genes of the NNV(SGYeosu08) isolated from sevenband grouper were phylogenetically analyzed.In addition,novel quantitative PCR primers based on the genomic sequence of SGYeosu08 isolate were designed and compared it with the conventional bio-assay method(TCID_(50)) using in vitro and in vivo samples.Results:The phylogenetic analysis of viral genes demonstrated the relationship of SGYeosu08 with members of red-spotted grouper nervous necrosis virus(RGNNV).The qNNV_Rl primer set(R1_F and R1_R) and the qNNV_R2 primer set(R2_F and R2_R) revealed 93%primer efficiency(regression:y=-0.2861 x + 9.9401,R^2= 0.9976)and the revealed 108%primer efficiency(regression:y=-0.3172 x + 10.0611,R^2= 0.9982),respectively.Its comparison with viral infectivity calculated by TCID_(50) method showed similar kinetic pattern at in vitro and NNV challenged fish(in vivo) samples.Conclusions:Result show that this method is rapid and efficient to diagnose NNV infection compare to traditional bioassay method(TCID_(50)).
基金This study was supported by the internal fund of the Temasek Life Sciences Laboratory,Singapore and a graduate research scholarship from Tropical Marine Science Institute,National University of Singapore.
文摘Viral nervous necrosis(VNN)disease,caused by the nervous necrosis virus(NNV),is a devastating disease,leading to high mortality rate and huge economical loss in aquaculture.In the past 30 years,many studies on the virus,host responses to the virus infection and diagnostics have yielded a lot of knowledge on developing measures to prevent the VNN disease.Although genetic improvement of disease resistance through breeding is inheritable and has long-lasting positive effect on aquaculture,it is a very challenging task in conventional selective breeding for improving disease resistance.With the advances in mapping quantitative trait loci(QTL)and genome wide association studies(GWAS)for NNV resistance in aquaculture species,DNA markers and genes associated with NNV resistance have been identified,making the application of marker-assisted selection(MAS)and genomic selection(GS)for NNV resistance possible.A few genes for NNV resistance are identified and are being used for genome editing to rapidly improve NNV resistance.In this review,we present the current knowledge on the NNV,host responses to NNV infection,diagnostic methods and vaccines available for NNV disease.In addition,we summarize the current status of conventional and molecular breeding for NNV resistance and highlight future directions,including genome editing for improving NNV resistance in aquaculture.
基金Supported by Science and Technology Planning Project of Guangdong Province,China(2003B21502,2005B20301016)~~
文摘[Objective] To optimize the prokaryotic expression of MCP gene of red-spotted grouper nervous necrosis virus. [Method] The MCP gene was amplified from red-spotted grouper nervous necrosis viral genome by RT-PCR. The recombinant expression vector pRSET A-MCP was constructed and transformed into BL21(DE3)plysS to express proteins with induction in different media, at different pH, or at different temperatures. [Result] The expression level of recombinant bacteria reached a peak with induction under the following condition: SOB or LB medium, pH 7.0, 37 ℃ while the fusion protein was about 44.5 kD in molecular weight. [Conclusion] This study provided a basis for the development of RGNNV-MCP vaccine.
文摘Viral nervous necrosis (VNN) is a worldwide disease among teleost fish. In the mainland of China, VNN was first identified in 2 species of hatchery-reared groupers, Epinephelus akaara and E. coioides. In the present study, samples were collected from larvae of E. akaara with signs of VNN in Dayawan bay which is located in southern China. The complete viral coat protein gene was amplified using extracted total RNA as template. The reverse transcription polymerase chain reaction (RT-PCR) amplification was performed using primers containing a heterologous restriction site for NotI. PCR products were cloned into pcDNA3 vector and sequenced. The results indicated that the coat protein gene of Dayawan isolate (RG-CN) was 1056 bases in length and contained a single open reading frame (ORF) of 1017 bases encoding a protein of 338 amino acids. The sequence similarities between the coat protein gene of RG-CN and other 8 isolates of NNV from Dayawan, Taiwan, Japan, Singapore and France were 79.1%-99.5% at the nucleotide level and 83.7%-100% at the amino acid level, respectively.The homology between RG-CN and the other 5 isolates from groupers was high and relatively lower between RG-CN and guppy nervous necrosis virus (GNNV), sea bass nervous necrosis virus (DIEV), but more divergences existed between RG-CN and striped jack nevous necrosis virus (SJNNV). Compared with SJNNV, RG-CN and the other 7 isolates all lacked 6 nucleotides and 2 amino acids in the same positions. Based on the result of molecular phylogenetic analysis, Dayawan isolate belongs to red-spotted grouper nervous necrosis virus (RGNNV) genotype.
基金supported by the Pearl River S&T Nova Program of Guangzhou(201806010047)National Natural Science Foundation of China(32173001,3210284,31771587)+1 种基金China Postdoctoral Science Foundation Funded Project(2021M693678)Natural Science Foundation of Guangxi Province(2021GXNSFDA075015)。
文摘As a highly important fish virus,nervous necrosis virus(NNV)has caused severe economic losses to the aquaculture industry worldwide.Autophagy,an evolutionarily conserved intracellular degradation process,is involved in the pathogenesis of several viruses.Although NNV can induce autophagy to facilitate infection in grouper fish spleen cells,how it initiates and mediates autophagy pathways during the initial stage of infection is still unclear.Here,we found that red-spotted grouper NNV(RGNNV)induced autophagosome formation in two fish cell lines at 1.5 and 3 h post infection,indicating that autophagy is activated upon entry of RGNNV.Moreover,autophagic detection showed that RGNNV entry induced incomplete autophagy by impairing the fusion of autophagosomes with lysosomes.Further investigation revealed that binding of the RGNNV capsid protein(CP)to the Lateolabrax japonicus heat shock protein HSP90ab1(LjHSP90ab1),a cell surface receptor of RGNNV,contributed to RGNNV invasion-induced autophagy.Finally,we found that CP blocked the interaction of L.japonicus protein kinase B(AKT)with LjHSP90ab1 by competitively binding the NM domain of LjHSP90ab1 to inhibit the AKT-mechanistic target of the rapamycin(MTOR)pathway.This study provides novel insight into the relationship between NNV receptors and autophagy,which may help clarify the pathogenesis of NNV.
文摘Using the extended homogenous balance method, we obtainabundant exact solution structures ofa (2+1)dimensional integrable model, the generalized Nizhnik-Novikov-Veselov equation. By means of the leading order termanalysis, the nonlinear transformations of generalized Nizhnik-Novikov-Veselov equation are given first, and then somespecial types of single solitary wave solution and the multisoliton solutions are constructed.
文摘报道了一种可以快速、灵敏地检测七带石斑鱼神经坏死病毒(NNV)的逆转录环介导等温扩增方法(RT-LAMP)。该方法参考赤点石斑鱼NNV病毒的主衣壳蛋白(MCP)基因的保守序列,设计1对引物克隆出七带石斑鱼NNV的基因序列,利用Primer Explorer V4软件设计了3对针对所克隆NNV基因序列的特异性引物,建立了RT-LAMP的反应体系,并分别对反应系统的引物浓度、反应温度和时间进行了优化,形成了七带石斑鱼NNV病毒RT-LAMP检测技术。实践应用结果表明,在63℃的最佳反应温度下,采用RT-LAMP检测技术经过45 min就能完成一次检测,准确率达到100%。本研究建立的RT-LAMP检测NNV技术设备要求简单,为七带石斑鱼等石斑鱼无NNV受精卵和仔稚鱼的生产现场检测和筛选提供了一种方便、灵敏的检测方法。
基金supported by the Natural Science Foundation of Zhejiang Province under Grant No.Y604106the Scientific Research Foundation of Zhejiang Provincial Education Department under Grant No.20070568the Natural Science Foundation of Zhejiang Lishui University under Grant No.KZ08001
文摘With the help of an improved mapping approach and a linear-variable-separation approach, a new family of exact solutions with arbitrary functions of the （2＋1）-dimensional Nizhnik-Novikov-Veselov system （NNV） is derived. Based on the derived solutions and using some multi-valued functions, we find a few new folded solitary wave excitations for the （2＋1）-dimensional NNV system.
文摘Exact periodic-wave solutions to the generalized Nizhnik-Novikov-Veselov (NNV) equation are obtained by using the extended Jacobi elliptic-function method, and in the limit case, the solitary wave solution to NNV equation are also obtained.
基金The project supported by the State Key Laboratory of 0il/Gas Reservoir Geology and Exploitation ＂PLN0402＂The authors would like to thank Prof.Sen-Yue Lou for helpful discussions.
文摘Some new structures and interactions of solitons for the （2＋1）-dimensional Nizhnik-Novikov-Veselov equation are revealed with the help of the idea of the bilinear method and variable separation approach. The solutions to describe the interactions between two dromions, between a line soliton and a y-periodic soliton, and between two y-periodic solitons are included in our results. Detailed behaviors of interaction are illustrated both analytically and in graphically. Our analysis shows that the interaction properties between two solitons are related to the form of interaction constant. The form of interaction constant and the dispersion relationship are related to the form of the seed solution （u0, v0, w0 ） in Backlund transformation.
