Publications

determination of the complete nucleotide sequences of rna1 and rna2 from greasy grouper (epinephelus tauvina) nervous necrosis virus, singapore strain.	the complete nucleotide sequences of rna1 and rna2 from greasy grouper (epinephelus tauvina) nervous necrosis virus (ggnnv), singapore strain, were determined. 5' race and rna ligation were used to obtain the complete nucleotide sequences of the 5' and 3' non-coding regions (ncrs). ggnnv rna1 was determined to be 3103 nt long, containing an orf of 982 aa, while ggnnv rna2 was determined to be 1433 nt long, containing an orf of 338 aa. both ggnnv rnas are longer than those of other published beta ...	2001	11172107
establishment of an infectious rna transcription system for striped jack nervous necrosis virus, the type species of the betanodaviruses.	a system has been established to produce infectious rna transcripts for striped jack nervous necrosis virus (sjnnv), the type species of the betanodaviruses, which infect fish. an enzymological analysis suggested that both rna1 and rna2 of sjnnv have a 5' cap. both rnas were largely resistant to 3' polyadenylation and ligation, suggesting the presence of an interfering 3' structure, while a small quantity of viral rnas were polyadenylated in vitro. the complete 5' and 3' non-coding sequences of ...	2001	11602777
intramolecular disulfide bonding is essential for betanodavirus coat protein conformation.	here we report on the conformational changes that are responsible for the appearance of the dicentrarchus labrax encephalitis virus (dlev) coat protein as a doublet in sds-page. wild-type and mutated forms of the coat protein cdna were expressed in e. coli. the study of the resulting recombinant molecules excluded the possibility of the involvement of a precursor autocatalysis mechanism or a ribosomal frameshifting event in the doublet formation. the appearance of the coat protein doublet was fo ...	2002	12185275
interference of the life cycle of fish nodavirus with fish retrovirus.	interference of the life cycle of grouper nervous necrosis virus (gnnv), a member of the nodaviridae, genus betanodavirus, by snakehead retrovirus (snrv) has been studied in vitro. sgf-1, a new fish cell line that is persistently infected with snrv, was induced by inoculating snrv into the grouper fin cell line gf-1. culture supernatants and cell pellets from both gnnv-infected sgf-1 and gf-1 cells were collected and employed for virus productivity analysis. the yields of gnnv rna and capsid pro ...	2002	12237429
betanodavirus as a novel transneuronal tracer for fish.	in order to obtain a potential new tool to analyze networks of the central nervous system of teleost fishes, we tested a fish-pathogenic betanodavirus, sevenband grouper nervous necrosis virus (sgnnv), as a transneuronal tracer using the freshwater angelfish pterophyllum scalare as a test animal. intravitreous injections of sgnnv into the right eye resulted first in the labelings of neuronal cell bodies in the ganglion cell layers of the retina and then those in the inner and outer nuclear layer ...	2002	12359322
involvement of the terminus of grouper betanodavirus capsid protein in virus-like particle assembly.	dragon grouper, epinephelus lanceolatus, nervous necrosis virus (dgnnv) consists of 180 copies of capsid protein that encapsulates a bipartite genome of single-stranded (+)rnas. expressing the open reading frame (orf) of rna2 in escherichia coli forms virus-like particles (vlps) that resembles native virus. deleting n- and c-termini revealed different impacts on vlp formation. deletion of 35 or 52 residues at the n-terminus completely ruined the vlp assembly, presumably due to removal of positiv ...	2003	12556997
identification of nucleolus localization signal of betanodavirus ggnnv protein alpha.	betanodavirus greasy grouper (epinephelus tauvina) nervous necrosis viruses (ggnnv) protein alpha, a virus capsid protein, was detected in both nucleolus and cytoplasm of infected cells of asian sea bass (sb) and transfected cells of sb and cos-7 with pcdna3.1/rna2. to study its subcellular localization, orf of protein alpha with 338 aa was fused with enhanced green fluorescent protein (egfp) gene and was detected in transfected cells in the absence of other viral proteins. in both sb and cos-7 ...	2003	12642096
susceptibility of cultured juveniles of several marine fish to the sevenband grouper nervous necrosis virus.	piscine nodaviruses (betanodaviruses) have been tentatively divided into four genotypes (sjnnv, rgnnv, tpnnv and bfnnv) and it is suggested that host specificity is different among these genotypes. in the present study, a betanodavirus [sevenband grouper nervous necrosis virus (sgnnv)] belonging to the redspotted grouper nervous necrosis virus (rgnnv) genotype, to which most betanodaviruses from warm water fish are identified, was evaluated for its pathogenicity to hatchery-reared juveniles of s ...	2003	12962219
serological relationships among genotypic variants of betanodavirus.	betanodaviruses, the causative agents of viral nervous necrosis or viral encephalopathy and retinopathy, are divided into 4 genotypes based on the coat protein gene (rna2). in the present study, serological relationships among betanodavirus genotypic variants were examined by virus neutralization tests using rabbit antisera raised against purified virions of strains representative of each genotype. all 20 isolates examined shared epitopes for neutralizing, but they fell into 3 major serotypes (a ...	2003	14735917
inoculation of balb/c mice with fish-pathogenic nodaviruses.	to investigate the pathogenicity of fish-pathogenic nodaviruses (betanodaviruses) in mammals, weanling mice were inoculated with striped jack nervous necrosis virus (sjnnv; the type species of the genus betanodavirus) or redspotted grouper nervous necrosis virus (rgnnv), which belong to different genotypes. after inoculation with 10(7.5) tcid50, either intramuscularly or intraperitoneally, mice remained clinically normal for the 14-day observation period. in a subsequent short-term (72 h) experi ...	2004	15003479
complete sequence of rna1 and subgenomic rna3 of atlantic halibut nodavirus (ahnv).	the nodaviridae are divided into the alphanodavirus genus, which infects insects, and the betanodavirus genus, which infects fishes. betanodaviruses are the causative agent of viral encephalopathy and retinopathy (ver) in a number of cultivated marine fish species. the nodaviridae are small non-enveloped rna viruses that contain a genome consisting of 2 single-stranded positivesense rna segments: rna1 (3.1 kb), which encodes the viral part of the rna-dependent rna polymerase (rdrp); and rna2 (1. ...	2004	15109133
membrane association of greasy grouper nervous necrosis virus protein a and characterization of its mitochondrial localization targeting signal.	localization of rna replication to intracellular membranes is a universal feature of positive-strand rna viruses. the betanodavirus greasy grouper (epinephelus tauvina) nervous necrosis virus (ggnnv) is a positive-rna virus with one of the smallest genomes among rna viruses replicating in fish cells. to understand the localization of ggnnv replication complexes, we generated polyclonal antisera against protein a, the ggnnv rna-dependent rna polymerase. protein a was detected at 5 h postinfection ...	2004	15163743
effect of temperature on betanodavirus infection in ssn-1 cell line.		2004	15372978
genomic classification of new betanodavirus isolates by phylogenetic analysis of the coat protein gene suggests a low host-fish species specificity.	viral encephalopathy and retinopathy is a devastating disease that causes neurological disorders and high mortality in a large number of cultivated marine fish species around the world. it is now established that several viral strains classified in the genus betanodavirus of the family nodaviridae are the aetiological agents of this disease. betanodaviruses can be classified into four genotypes based on the coat protein gene sequence. here, the coat protein genes of the three major strains isola ...	2004	15448371
pcr-based detection of betanodaviruses from cultured and wild marine fish with no clinical signs.	betanodaviruses are the causative agents of viral nervous necrosis (vnn) or viral encephalopathy and retinopathy (ver) in cultured marine fish. a total of 131 apparently healthy fish from 30 species were collected in two geographically remote aquaculture areas, yashima bay (kagawa prefecture) and tamanoura bay (nagasaki prefecture), in japan. the brains of fish were examined by reverse transcriptase-polymerase chain reaction (rt-pcr) and nested pcr to detect the coat protein gene of betanodaviru ...	2004	15482425
white tail disease of the giant freshwater prawn, macrobrachium rosenbergii: separation of the associated virions and characterization of mrnv as a new type of nodavirus.	white tail disease of the farmed freshwater prawn, macrobrachium rosenbergii, is the cause of mortalities in the french west indies, china and india. two different sized particles, both developing in the cytoplasm of target cells, are found associated with diseased animals. these two viruses were separated, purified and subsequently characterized. the larger one, called mrnv, is icosahedral in shape and 27 nm in diameter. its genome is composed of two fragments of linear single-stranded rna (ss- ...	2005	15660790
persistent infection of betanodavirus in a novel cell line derived from the brain tissue of barramundi lates calcarifer.	in order to obtain an in vitro system for studying the mechanism of persistent infection of fish nodavirus, a novel cell line (bb) was established from the brain tissue of a barramundi, lates calcarifer, which had survived viral nervous necrosis disease. the cell line has been subcultured > 100 times. the persistence of fish nodavirus designated as barramundi brain nervous necrosis virus (bbnnv) in the bb cells was demonstrated by: (1) the detection of the infectious virus in the culture superna ...	2005	16060261
early endocytosis pathways in ssn-1 cells infected by dragon grouper nervous necrosis virus.	many fish undergo betanodavirus infection. to study the infection process of dragon grouper nervous necrosis virus (dgnnv), native virus and virus-like particles (vlps) were used to analyse the binding and internalization in ssn-1 cells. the binding of dgnnv and vlps to ssn-1 cells was demonstrated using western blotting and immunofluorescence microscopy. as estimated by indirect elisa, the dgnnv particles bound ssn-1 cells in a dose-dependent manner up to 8 x 10(4) particles per cell. the bindi ...	2005	16099914
development of a sensitive and quantitative diagnostic assay for fish nervous necrosis virus based on two-target real-time pcr.	the aim of the present work was to develop two new independent sybr green i-based real-time pcr assays for both detection and quantification of betanodavirus, an rna virus that infects several species of marine teleost fish causing massive mortalities in larvae and juveniles. the assays utilized two pairs of primers targeting highly conserved regions of both the rna molecules forming the betanodavirus genome: rna1 encoding the rna-dependent rna polymerase (rdrp) and rna2 encoding the coat protei ...	2005	16146676
characterization of striped jack nervous necrosis virus subgenomic rna3 and biological activities of its encoded protein b2.	striped jack nervous necrosis virus (sjnnv), which infects fish, is the type species of the genus betanodavirus. this virus has a bipartite genome of positive-strand rnas, designated rnas 1 and 2. a small rna (ca. 0.4 kb) has been detected from sjnnv-infected cells, which was newly synthesized and corresponded to the 3'-terminal region of rna1. rapid amplification of cdna ends analysis showed that the 5' end of this small rna (designated rna3) initiated at nt 2730 of the corresponding rna1 seque ...	2005	16186236
genetic characterisation of coat protein gene of betanodavirus isolates from different fish species.		2005	16244964
betanodavirus b2 is an rna interference antagonist that facilitates intracellular viral rna accumulation.	betanodaviruses are small positive-sense bipartite rna viruses that infect a wide variety of fish species and are notorious for causing lethal outbreaks in juvenile fish hatcheries worldwide. the function of a small nonstructural protein, b2, encoded by the subgenomic rna3 of betanodaviruses, has remained obscure. greasy grouper nervous necrosis virus, a betanodavirus model, was used to develop a facile dna-based reverse genetics system that recapitulated the virus infection cycle, and we used t ...	2006	16352533
betanodavirus induces phosphatidylserine exposure and loss of mitochondrial membrane potential in secondary necrotic cells, both of which are blocked by bongkrekic acid.	in this study, we show how the red spotted grouper nervous necrosis virus (rgnnv) causes loss of mitochondrial membrane potential and promotes host secondary apoptotic necrosis. rgnnv viral proteins such as protein alpha (42 kda) and protein a (110 kda) were quickly expressed between 12 h and 24 h postinfection (p.i.) in gl-av cells. annexin v staining revealed that the nnv infection of gl-av cells induced phosphatidylserine (ps) externalization and development of bulb-like vesicles (bleb format ...	2006	16430940
temperature-dependency of betanodavirus infection in ssn-1 cell line.	this study examined the in vitro effects of temperature on betanodavirus infection in the ssn-1 cell line. a betanodavirus isolated from moribund sea bass fry dicentrarchus labrax farmed in the adriatic sea and characterised as a rgnnv (redspotted grouper nervous necrosis virus) genotype was used. virus-infected ssn-1 cells were incubated at temperatures between 10 and 30 degrees c and observed for cytopathic effects daily for 15 d. cell-free and cell-associated viral growth were evaluated by 50 ...	2006	16610592
sequence analysis of coat protein gene of wuhan nodavirus isolated from insect.	wuhan nodavirus (whnv) particles are isometric, non-enveloped, and about 29 nm in diameter. in the previous study, we determine its physiochemical characterization and the nucleotide sequence of the larger genomic segment, rna1 and identify it a nodavirus. whnv rna1 is 3,149 nt in length, encoding protein a, catalytic subunit of rna-dependent rna polymerase (rdrp). in this report, we complete the sequence determination of the smaller genomic segment, rna2 of whnv. whnv rna2 is determined to be 1 ...	2006	16644053
persistence of betanodavirus in barramundi brain (bb) cell line involves the induction of interferon response.	the bb cell line derived from the brain tissue of a barramundi (lates calcarifer) that survived nervous necrosis virus (nnv) infection is persistently infected with nnv. to elucidate whether interferon (ifn) plays a role in the mechanism of nnv-persistent infection in bb cell line, a virus-negative control cell line was obtained by treating bb cells with nnv-specific rabbit antiserum for 5 subcultures. after the treatment, nnv titer or rna or capsid protein was no longer detected in the cured bb ...	2006	16698284
sequestration and protection of double-stranded rna by the betanodavirus b2 protein.	betanodavirus b2 belongs to a group of functionally related proteins from the sense-strand rna virus family nodaviridae that suppress cellular rna interference. the b2 proteins of insect alphanodaviruses block rna interference by binding to double-stranded rna (dsrna), thus preventing dicer-mediated cleavage and the subsequent generation of short interfering rnas. we show here that the fish betanodavirus b2 protein also binds dsrna. binding is sequence independent, and maximal binding occurs wit ...	2006	16809288
betanodavirus infection in the freshwater model fish medaka (oryzias latipes).	betanodaviruses, the causal agents of viral nervous necrosis in marine fish, have bipartite, positive-sense rna genomes. as their genomes are the smallest and simplest among viruses, betanodaviruses have been studied in detail as model viruses by using a genetic-engineering system, as has occurred with the insect alphanodaviruses, the other members of the family nodaviridae. however, studies of virus-host interactions have been limited, as betanodaviruses basically infect marine fish at early de ...	2006	16847129
detection of betanodavirus in juvenile barramundi, lates calcarifer (bloch), by antigen capture elisa.	betanodavirus infection of fish has been responsible for mass mortalities in aquaculture hatcheries worldwide. betanodaviruses possess a bipartite single-stranded rna genome consisting of the 3.1 kb rna1 encoding an rna-dependent rna polymerase and the b2 protein, while the 1.4 kb rna2 encodes the viral nucleocapsid protein, alpha. a panel of six monoclonal antibodies against the alpha protein of greasy grouper nervous necrosis virus (ggnnv) was developed for use in diagnostics. all antibodies r ...	2006	16866926
phylogeny of betanodaviruses and molecular evolution of their rna polymerase and coat proteins.	the betanodaviruses are the causative agent of the disease viral nervous necrosis in fishes. betanodavirus genome consists of two single-stranded positive-sense rna molecules (rna1 and rna2). rna1 gene encodes the rna polymerase, named also protein a, while rna2 encodes the coat protein precursor, the cpp protein. we investigated the evolutionary relationships among betanodaviruses working on partial sequences of both rna1 and rna2. phylogenetic analyses were performed by applying a maximum like ...	2007	16990016
induction of a protective immune response against viral nervous necrosis in the european sea bass dicentrarchus labrax by using betanodavirus virus-like particles.	betanodaviruses are causative agents of viral nervous necrosis (vnn), a devastating disease of cultured marine fish worldwide. virus particles contain a single type of coat protein that spontaneously assembles into virus-like particles (vlps) when expressed in a baculovirus expression system. in the present study, the immunogenicity of betanodavirus vlps and the protection they confer against vnn in the european sea bass dicentrarchus labrax were investigated. enzyme-linked immunosorbent assay a ...	2006	17005697
susceptibility of the fish cell line saf-1 to betanodavirus.		2006	17026673
detection of betanodaviruses in apparently healthy aquarium fishes and invertebrates.	betanodaviruses are the causative agents of viral nervous necrosis (vnn) in cultured marine fish. a total of 237 apparently healthy aquarium fish, marine (65 species) and freshwater (12 species) fishes and marine invertebrates (4 species), which were stocked in a commercial aquarium in seoul, south korea, were collected from november 2005 to february 2006. the brains of the fish and other tissues of the invertebrates were examined by reverse transcriptase-polymerase chain reaction (rt-pcr) and n ...	2006	17106229
detection of dnas homologous to betanodavirus genome rnas in barfin flounder verasper moseri and japanese flounder paralichthys olivaceus.	i investigated the presence of dna homologous to genome rna1 and rna2 (rna1 dna and rna2 dna) of betanodaviruses - the causative agent of viral nervous necrosis (vnn) --in eggs, sperm, ovarian cavity fluid, larvae, and juveniles of barfin flounder verasper moseri and larvae and juveniles of japanese flounder paralichthys olivaceus collected at 6 sites in hokkaido, japan, from 1994 to 2001. rna1 dna and rna2 dna were detected by pcr in 13 and 33 % of barfin flounder samples and 0 and 69% of japan ...	2006	17190201
anti-bcl-2 family members, zfbcl-x(l) and zfmcl-1a, prevent cytochrome c release from cells undergoing betanodavirus-induced secondary necrotic cell death.	nervous necrosis virus (nnv)-induced, host cell apoptosis mediates secondary necrosis by an ill-understood process. in this study, redspotted grouper nervous necrosis virus (rgnnv) is shown to induce mitochondria-mediated necrotic cell death in gl-av cells (fish cells) via cytochrome c release, and anti-apoptotic proteins are shown to protect these cells from death. western blots revealed that cytochrome c release coincided with disruption of mitochondrial ultrastructure and preceded necrosis, b ...	2007	17245642
dissection of double-stranded rna binding protein b2 from betanodavirus.	betanodaviruses are small rna viruses that infect teleost fish and pose a considerable threat to marine aquaculture production. these viruses possess a small protein, termed b2, which binds to and protects double-stranded rna. this prevents cleavage of virus-derived double-stranded rnas (dsrnas) by dicer and subsequent production of small interfering rna (sirna), which would otherwise induce an rna-silencing response against the virus. in this work, we have performed charged-to-alanine scanning ...	2007	17376906
identification of b-cell epitopes on the betanodavirus capsid protein.	the pepscan procedure was used to identify betanodavirus b-cell epitopes recognized by neutralizing mouse monoclonal antibodies (mabs) and serum samples obtained from sea bass, dicentrarchus labrax, naturally infected with betanodavirus. pepscan was performed with a panel of thirty-four 12-mer synthetic peptides that mimicked the entire betanodavirus capsid protein. sea bass serum samples reacted strongly with three regions of the capsid protein comprising amino acid residues 1-32, 91-162 and 18 ...	2007	17584439
rna-dependent rna polymerase from atlantic halibut nodavirus contains two signals for localization to the mitochondria.	nodaviruses encode an rna-dependent rna polymerase called protein a that is responsible for replication of the viral rna segments. the intracellular localization of protein a from a betanodavirus isolated from atlantic halibut (ahnv) was studied in infected fish cells and in transfected mammalian cells expressing myc-tagged wild type protein a and mutants. in infected cells protein a localized to cytoplasmic structures resembling mitochondria and in transfected mammalian cells the ahnv protein a ...	2007	17602779
identification of critical residues in nervous necrosis virus b2 for dsrna-binding and rnai-inhibiting activity through by bioinformatic analysis and mutagenesis.	it is known that the non-structural b2 protein of nervous necrosis virus (nnv) plays an important role in viral replication and can inhibit the rna interference system of the host cell. moreover, the mechanism of nnv b2 protein to inhibit rnai is by sequestration and protection of double strand (ds) rna. in the flock house virus (fhv), a model alphanodavirus, the structural and mutational analysis of b2 identified that the positively charged arg54 of the alpha2 helix mediated the dsrna-binding a ...	2007	17669362
isolation and genetic characterization of betanodavirus from wild marine fish from the adriatic sea.		2007	17682880
the interferon response is involved in nervous necrosis virus acute and persistent infection in zebrafish infection model.	betanodavirus, a small positive-sense bipartite rna virus notoriously affecting marine aquaculture worldwide has been extensively studied in vitro. however, impending studies in elucidating virus-host interactions have been limiting due to the lack of appropriate animal disease models. therefore, in this study, we have attempted to successfully establish nnv infection in zebrafish (danio rerio) showing typical nnv symptoms and which could potentially serve as an in vivo model for studying virus ...	2008	17727953
inhibition of betanodavirus infection by inhibitors of endosomal acidification.	betanodaviruses, members of the family nodaviridae, have small positive-stranded bipartite rna genomes and are the causal agent of viral nervous necrosis (vnn) in many species of marine farmed fish. in the aquaculture industry, outbreaks of betanodavirus infection and spread in larval and juvenile fish result in devastating damage and heavy economic loss. although an urgent need exists to develop drugs that inhibit betanodavirus infection, there have been no reports about anti-betanodavirus drug ...	2007	17891330
potential for the replication of the betanodavirus redspotted grouper nervous necrosis virus in human cell lines.	the determination of the host ranges of viruses is important because of the possible emergence of infectious agents, which may result from the zoonotic transmission of animal viruses to humans. the family nodaviridae, whose members are non-enveloped, positive-stranded bipartite rna viruses, is comprised of the genera alphanodavirus and betanodavirus, whose members predominantly infect insects and fish, respectively. the alphanodaviruses can also infect suckling mice and suckling hamsters, result ...	2008	17906832
screening of freshwater fish species for their susceptibility to a betanodavirus.	betanodaviruses, the causative agents of viral nervous necrosis in marine fish, have bipartite positive-sense rna genomes. because the genomes are the smallest and simplest among viruses, betanodaviruses have been well studied using a reversed genetics system as model viruses. however, studies of virus-host interactions have progressed slowly because permissive hosts for betanodaviruses (basically larvae and juveniles of marine fish) are only available for limited periods of the year and are not ...	2007	17972753
molecular detection of betanodaviruses from apparently healthy wild marine invertebrates.	one hundred eighteen samples (21 species) of wild marine invertebrates were collected from western and southern coastal area of korean peninsula. four of 78 (18 species) samples collected at namhae (south) area were positive for nodavirus in nested pcr test. of the 40 samples (5 species) collected at hwanghae (west) areas, all samples were negative for nodavirus in both rt-pcr and nested pcr tests. positive nested pcr results were obtained from the following species: charybdis bimaculata charybd ...	2008	18076900
new clade of betanodaviruses detected in wild and farmed cod (gadus morhua) in norway.	betanodaviruses have been isolated and detected in both farmed and wild fish species worldwide. they are classified in five clusters, and all are connected to mortalities in farmed fish. the clusters do not represent specific geographical areas or host species, but one cluster, barfin flounder nervous necrosis virus (bfnnv), is mainly associated with cold water fish species. this study presents the first species-specific clade within the bfnnv cluster. this clade consists of six isolates from wi ...	2008	18175044
molecular detection of betanodavirus in wild marine fish populations in korea.	viral nervous necrosis (vnn) is a worldwide disease affecting several species of cultured marine fish. in korea, vnn has been identified in several species of cultured marine fish. in this study, the authors present data of the amplified nested polymerase chain reaction product (420 bp) of 21 nodavirus strains from different species of apparently healthy wild marine fish on the southern coast of korea. phylogenetic analysis based on the partial nucleotide sequence (177 bases) of the rna2 coat pr ...	2008	18182506
efficacies of inactivated vaccines against betanodavirus in grouper larvae (epinephelus coioides) by bath immunization.	betanodavirus is the pathogen of viral nervous necrosis (vnn) disease that has caused mass mortality among many species of marine fish at larval stage. in this study, the efficacy of inactivated betanodavirus was evaluated by bath-immunization and bath-challenge of orange-spotted grouper (epinephelus coioides) at early larval stage. two kinds of chemicals were used for inactivation of the virus, and the relative percent survival (rps) values of 0.4mm binary ethylenimine (bei)-inactivated vaccine ...	2008	18276044
zebrafish anti-apoptotic protein zfbcl-xl can block betanodavirus protein alpha-induced mitochondria-mediated secondary necrosis cell death.	betanodavirus protein alpha induces cell apoptosis or secondary necrosis by a poorly understood process. in the present work, red spotted grouper nervous necrosis virus (rgnnv) rna 2 was cloned and transfected into tissue culture cells (gf-1) which then underwent apoptosis or post-apoptotic necrosis. in the early apoptotic stage, progressive phosphatidylserine externalization was evident at 24h post-transfection (p.t.) by annexin v-fluos staining. tunel assay revealed apoptotic cells at 24-72 h ...	2008	18276161
genetic analysis of betanodaviruses in subclinically infected aquarium fish and invertebrates.	betanodaviruses causing viral nervous necrosis (vnn) have been detected and isolated from several species of cultured marine fish worldwide. in korea, vnn was identified in several species of cultured marine fish. this study presents data on the amplified nested pcr product (420 bp) of 11 nodavirus strains from different species of apparently healthy aquarium fish and invertebrates collected in one private commercial aquarium in korea. phylogenetic analyses based on the partial nucleotide sequen ...	2008	18293033
antemortem versus postmortem methods for detection of betanodavirus in senegalese sole (solea senegalensis).	the suitability of nested reverse transcription polymerase chain reaction (nrt-pcr) to detect betanodavirus in blood samples from naturally infected senegalese sole (solea senegalensis) was evaluated in comparison with other diagnostic methods. results indicated that histologic examination of brain lesions could be regarded as the most consistent indicator of nodavirus infection in this species. the nrt-pcr showed low to moderate levels of detection; the best values were obtained in brain sample ...	2008	18319436
macrobrachium rosenbergii nodavirus infection in m. rosenbergii (de man) with white tail disease cultured in taiwan.	white tail disease (wtd) is a serious problem in macrobrachium rosenbergii hatcheries and nursery ponds in asia. the causative agents have been identified as m. rosenbergii nodavirus (mrnv) and its associated extra small virus. this is the first report demonstrating mrnv virus in m. rosenbergii displaying wtd signs in taiwan by reverse transcriptase-polymerase chain reaction (rt-pcr). amplified fragments of 850 and 425 bp for rna-1 and rna-2 of mrnv, respectively, were obtained by rt-pcr. rt-pcr ...	2008	18471097
establishment of reverse genetics system of betanodavirus for the efficient recovery of infectious particles.	betanodaviruses, a member of the family nodaviridae, have small positive-stranded bipartite rna genomes and are the causal agent of viral nervous necrosis in marine-farmed fish. to facilitate the study of betanodavirus, infectious cdna clones of its two genomic rnas were generated. the full-length cdna of the new redspotted grouper nervous necrosis virus strain (sg2001nag) rna1 and rna2 were co-transcribed by t7 rna polymerase in baby hamster kidney cells expressing t7 rna polymerase. the transc ...	2008	18508134
efficient propagation of betanodavirus in a murine astrocytoma cell line.	betanodavirus, a bipartite rna virus of fishes and a member of nodaviridae family, targets nervous tissues and is the causative agent of viral nervous necrosis in marine farmed fish. betanodavirus is thought to be propagated only in fish cells because betanodavirus has only been isolated in fish and it is not well propagated in mammalian culture cells. however, the host specificity of betanodavirus has not yet been well analyzed. to analyze the host specificity of betanodavirus, various mammalia ...	2008	18556083
variable region of betanodavirus rna2 is sufficient to determine host specificity.	betanodaviruses, the causative agents of viral nervous necrosis in marine fish, have bipartite positive-sense rna genomes. the viruses have been classified into 4 distinct types based on nucleotide sequence similarities in the variable region (the so-called t4 region) of the smaller genomic segment rna2 (1.4 kb). betanodaviruses have marked host specificity, although the primary structures of the viral rnas and encoded proteins are similar among the viruses. we have previously demonstrated, usin ...	2008	18589996
role of the dxxdxd motif in the assembly and stability of betanodavirus particles.	piscine betanodavirus possesses a bipartite genome of single-stranded (+)rnas. rna2 cdna of dragon grouper nervous necrosis virus (dgnnv) has been expressed previously to form virus-like particles (vlps), which are highly similar to the native virion. experiments with calcium-chelating or reducing/oxidizing reagents showed that the dgnnv vlps required only calcium for particle assembly. with the recombinant vlps, site-directed mutagenesis can be employed to investigate the roles of calcium-bindi ...	2008	18626568
comparisons among the complete genomes of four betanodavirus genotypes.	betanodaviruses, the causative agents of viral nervous necrosis in marine fish, have bipartite positive-sense rna genomes and have been classified (based on analysis of rna2 sequences) into 4 genotypes: tiger puffer nervous necrosis virus (tpnnv), barfin flounder nervous necrosis virus (bfnnv), striped jack nervous necrosis virus (sjnnv), and redspotted grouper nervous necrosis virus (rgnnv). full-length genomes of tpnnv and bfnnv were sequenced for the first time in this study. their sequence d ...	2008	18717064
viral encephalopathy and retinopathy of dicentrarchus labrax and sparus aurata farmed in tunisia.	viruses belonging to the nodaviridae family cause disease worldwide among a large number of species of marine fish, and have been described in all continents. in the present study, a total of 69 farmed tunisian sea bass (dicentrarchus labrax) and 24 sea bream (sparus aurata) samples were tested monthly for the detection of betanodavirus. the virus was identified in both species using indirect immunofluorescence assays (ifat) and rt-pcr. in addition sequence analysis of part of the coat protein g ...	2009	18982421
betanodavirus non-structural protein b2: a novel necrotic death factor that induces mitochondria-mediated cell death in fish cells.	the betanodavirus non-structural protein b2 plays a role in silencing rna interference (rnai), which mediated regulation of animal and plant innate immune responses, but little is known regarding the role of b2 in cell death. the present study examined the effects of b2 on mitochondria-mediated necrotic cell death in grouper liver (gl-av) cells. b2 was expressed at 12 h post-infection (pi), with increased expression between 24 and 72 h pi by western blot. b2 was transiently expressed to investig ...	2009	19116179
genetic analysis and pathogenicity of betanodavirus isolated from wild redspotted grouper epinephelus akaara with clinical signs.	diseased wild redspotted grouper epinephelus akaara were collected from seto inland sea, ehime prefecture, in august 2002. fish showed erratic swimming behavior and inflation of the swim bladder. the fish brains were positive for nodavirus in both rt-pcr and nested pcr. the sequence of the nested pcr product (177 nt) was closely related to that of a known betanodavirus, redspotted grouper nervous necrosis virus. when juvenile sevenband grouper e. septemfasciatus were challenged intravitreously w ...	2009	19130166
betanodavirus non-structural protein b1: a novel anti-necrotic death factor that modulates cell death in early replication cycle in fish cells.	the functions of the betanodavirus non-structural protein b1 is still unknown. we examined b1 expression patterns and investigated novel cell death regulatory functions for this viral protein following rgnnv infection in fish cells. the b1 gene (336 nt) was cloned from the redspotted grouper nervous necrosis virus (rgnnv) genome. b1 mrna was rapidly expressed in the fish cells from viral rna3 at 12 h post-infection (p.i.). at the protein level, expression was low at 12 h p.i., and then increased ...	2009	19136133
protection conferred against viral nervous necrosis by simultaneous inoculation of aquabirnavirus and inactivated betanodavirus in the sevenband grouper, epinephelus septemfasciatus (thunberg).	an aquabirnavirus (abv) and a formalin-inactivated betanodavirus [redspotted grouper nervous necrosis virus (rgnnv)] were investigated for their potential to prevent rgnnv-induced viral nervous necrosis (vnn) in the sevenband grouper, epinephelus septemfasciatus (thunberg). three groups of fish were injected intramuscularly with abv, intraperitoneally with inactivated rgnnv (irgnnv) or with both abv and irgnnv. at 3, 7, 14, 21 and 28 days post-injection (p.i.), fish were challenged by intramuscu ...	2009	19261047
b2 protein from betanodavirus is expressed in recently infected but not in chronically infected fish.	betanodavirus infects both larvae and juvenile fish and can cause the disease viral encephalopathy and retinopathy (ver). during an acute outbreak of ver, infected individuals display several clinical signs of infection, i.e. abnormal swimming pattern and loss of appetite. betanodaviruses can also cause chronic or persistent infection where the infected individuals show no clinical signs of infection. during infection the viral sub-genomic rna3 and the rna3-encoded b2 protein are expressed. anti ...	2009	19326790
immune responses of asian sea bass, lates calcarifer bloch, against an inactivated betanodavirus vaccine.	asian sea bass, lates calcarifer (bloch), exhibited strong immune responses against a single injection of the formalin-inactivated red-spotted grouper nervous necrosis virus (rgnnv), a betanodavirus originally isolated in japan. fish produced neutralizing antibodies at high titre levels from days 10 (mean titre 1:480) to 116 (1:1280), with the highest titre at day 60 post-vaccination (1:4480). when fish were challenged with the homologous rgnnv at day 54 post-vaccination, there were no mortaliti ...	2009	19364387
neutralizing antibody levels for protection against betanodavirus infection in sevenband grouper, epinephelus septemfasciatus (thunberg), immunized with an inactivated virus vaccine.	abstract an inactivated betanodavirus, red-spotted grouper nervous necrosis virus (rgnnv), is a vaccine candidate for viral nervous necrosis (vnn). the present study was conducted to examine inoculation doses of the vaccine and neutralizing antibody titre levels to protect fish against vnn. young sevenband grouper, epinephelus septemfasciatus, averaging 25.4 g, were immunized at 25 degrees c water temperature by a single intraperitoneal injection of formalin-inactivated rgnnv. fish immunized at ...	2009	19490392
outbreak of betanodavirus infection in tilapia, oreochromis niloticus (l.), in fresh water.	a betanodavirus associated with a massive mortality was isolated from larvae of tilapia, oreochromis niloticus, maintained in fresh water at 30 degrees c. histopathology revealed vacuolation of the nervous system, suggesting an infection by a betanodavirus. the virus was identified by indirect fluorescent antibody test in the ssn1 cell line and further characterized by sequencing of a pcr product. sequencing of the t4 region of the coat protein gene indicated a phylogenetic clustering of this is ...	2009	19500206
genetic heterogeneity of betanodaviruses in juvenile production trials of pacific bluefin tuna, thunnus orientalis (temminck & schlegel).	pacific bluefin tuna, thunnus orientalis (temminck & schlegel), is one of the most important commercially exploited fish species in the world, and juvenile production techniques have been developed for its culture and stock enhancement in japan. however, recent juvenile production has often failed because of the occurrence of viral nervous necrosis caused by betanodaviruses. in this study, we examined the genetic variability of betanodaviruses detected in the diseased juveniles to understand the ...	2009	19538459
comparative analysis of both genomic segments of betanodaviruses isolated from epizootic outbreaks in farmed fish species provides evidence for genetic reassortment.	sequencing of the full coding region of both genomic segments of seven betanodavirus strains isolated from different farmed species in spain and portugal revealed that six were reassortants, exhibiting a red-spotted grouper nervous necrosis virus (rgnnv)-type rna1 and a striped jack nervous necrosis virus (sjnnv)-type rna2. analysis of sequences of reassortant strains at both the genomic and protein levels revealed the existence of differences compared with type strains of both genotypes. these ...	2009	19710256
inactivation of nervous necrosis virus infecting grouper (epinephelus coioides) by epinecidin-1 and hepcidin 1-5 antimicrobial peptides, and downregulation of mx2 and mx3 gene expressions.	betanodaviruses are one of the serious pathogens in nervous necrosis viral (nnv) disease that brings about mortality in the larval stage of grouper (epinephelus coioides). in this study, the efficacy of pretreatment, co-treatment, and posttreatment with the antimicrobial epinecidin-1 and hepcidin 1-5 peptides against a betanodavirus was evaluated by intraperitoneal inoculation in grouper. the results showed that co-treatment of epinecidin-1 or hepcidin 1-5 with the virus was effective in promoti ...	2010	19819335
optimisation and validation of a real-time reverse transcriptase-polymerase chain reaction assay for detection of betanodavirus.	a rt-qpcr assay that was developed and optimised for detection of betanodaviruses was validated for use as a diagnostic test for viral nervous necrosis disease of fish. four betanodavirus genotypes were detected but the sensitivity was greatest for redspotted grouper nervous necrosis virus (rgnnv). the analytical sensitivity was 10-1000-fold greater than that of a nested rt-pcr assay and the limit of detection was <0.4 tcid(50) units per reaction. the assay was highly repeatable (standard deviat ...	2010	19891987
complete sequencing of tunisian redspotted grouper nervous necrosis virus betanodavirus capsid gene and rna-dependent rna polymerase gene.	finfish nodaviruses (betanodaviruses) can cause highly destructive infections in numerous species of farmed marine fish larvae and juveniles worldwide. the betanodavirus genome consists of two single-stranded positive-sense rna molecules (rna1 and rna2). the virus can be classified into four genotypes based on the partial sequences of the coat protein (cp) gene (t2 and t4 regions). currently, genomic sequence information for rna1 regions of rna2 outside of t2 and t4 is less well documented. this ...	2010	19912455
cellular and molecular immune responses of the sea bass (dicentrarchus labrax) experimentally infected with betanodavirus.	naïve sea bass juveniles (38.4 + or - 4.5 g) were intramuscularly infected with a sublethal dose of betanodavirus isolate 378/i03, followed after 43 days by a similar boosting. this infection resulted in an overall mortality of 7.6%. at various intervals, sampling of fish tissues was performed to investigate: i) b and t lymphocyte content in organs and tissues; ii), proliferation of leucocytes re-stimulated in vitro with inactivated virus; iii) presence of serum antibody specific for betanodavir ...	2010	19925869
antimicrobial peptides (amp) with antiviral activity against fish nodavirus.	nervous necrosis virus (nnv) is classified as betanodavirus of nodaviridae, and has caused mass mortality of numerous marine fish species at larval stage. antimicrobial peptides (amps) play an important role of innate immunity either against bacterial pathogens or viruses. up to date, little is known if any amp could effectively inhibit fish nodaviruses and its mechanism. in this study, the antiviral activities of three antimicrobial peptides (amps) against grouper nnv (gnnv) were screened in th ...	2010	20004246
protective immunity against viral nervous necrosis (vnn) in brown-marbled grouper (epinephelus fuscogutattus) following vaccination with inactivated betanodavirus.	viral nervous necrosis (vnn) caused by betanodaviruses has been recently implicated in serious mortalities of groupers in the grow-out culture system. a safe and effective vaccine against this disease is urgently needed. this study demonstrates that a single intramuscular vaccination with formalin-inactivated philippine strain of piscine betanodavirus (genotype: redspotted grouper nervous necrosis virus; rgnnv) induces potent immune responses and substantial protective immunity against an intram ...	2010	20018247
anti-viral mechanism of barramundi mx against betanodavirus involves the inhibition of viral rna synthesis through the interference of rdrp.	nervous necrosis virus (nnv) belongs to the betanodavirus of the nodaviridae family. it is the causative agent of viral nervous necrosis (vnn) disease, and has inflicted devastating damage on the world-wide aquaculture industry. the fish that survived after the outbreak of vnn become persistently nnv-infected carriers. nnv-persistent infection has been demonstrated in a barramundi brain (bb) cell line, and it involves the type i interferon (ifn) response with the expression of mx gene. however, ...	2010	20034570
phylogenetic analysis of betanodavirus isolates from australian finfish.	in australia, disease caused by betanodavirus has been reported in an increasing number of cultured finfish since the first report of mortalities in 1990. partial coat protein gene sequences from the t2 or t4 regions of 8 betanodaviruses from barramundi lates calcarifer, sleepy cod oxyeleotris lineolata, striped trumpeter latris lineata, barramundi cod cromileptes altivelis, australian bass macquaria novemaculata and gold-spotted rockcod epinephelus coioides from several australian states were d ...	2009	20099409
molecular detection of betanodavirus from the farmed fish, platax orbicularis (forsskal) (ephippidae), in french polynesia.		2010	20158581
roles of cysteines cys115 and cys201 in the assembly and thermostability of grouper betanodavirus particles.	the virus-like particle (vlp) assembled from capsid subunits of the dragon grouper nervous necrosis virus (dgnnv) is very similar to its native t = 3 virion. in order to investigate the effects of four cysteine residues in the capsid polypeptide on the assembly/dissociation pathways of dgnnv virions, we recombinantly cloned mutant vlps by mutating each cysteine to destroy the specific disulfide linkage as compared with thiol reduction to destroy all s-s bonds. the mutant vlps of c187a and c331a ...	2010	20446029
a combined rt-pcr and dot-blot hybridization method reveals the coexistence of sjnnv and rgnnv betanodavirus genotypes in wild meagre (argyrosomus regius).	to detect the possible coexistence of striped jack nervous necrosis virus (sjnnv) and red-spotted grouper nervous necrosis virus (rgnnv) genotypes in a single fish, a methodology based on the combination of pcr amplification and blot hybridization has been developed and applied in this study.	2010	20497277
development and validation of a real-time taqman pcr assay for the detection of betanodavirus in clinical specimens.	betanodaviruses are the causal agents of viral encephalo-retinopathy, an infectious disease affecting more than 40 marine fish species, characterized by high morbidity and mortality. because of its severe impact, robust diagnostic tools are required. the aim of this work was to develop and validate a real-time taqman pcr assay to detect betanodaviruses in clinical specimens by amplifying a conserved region of the rna2 strand. the method proved to be specific and sensitive, being capable of detec ...	2010	20532929
susceptibilities of medaka (oryzias latipes) cell lines to a betanodavirus.	betanodaviruses, members of the family nodaviridae, have bipartite, positive-sense rna genomes and are the causal agents of viral nervous necrosis in many marine fish species. recently, the viruses were shown to infect a few freshwater fish species including a model fish medaka (oryzias latipes). although virological study using cultured medaka cells would provide a lot of insight into virus-fish interactions in molecular aspects, no such cells have yet been tested for virus susceptibility.	2010	20624282
betanodavirus b2 causes atp depletion-induced cell death via mitochondrial targeting and complex ii inhibition in vitro and in vivo.	the betanodavirus non-structural protein b2 is a newly discovered necrotic death factor with a still unknown role in regulation of mitochondrial function. in the present study, we examined protein b2-mediated inhibition of mitochondrial complex ii activity, which results in atp depletion and thereby in a bioenergetic crisis in vitro and in vivo. expression of protein b2 was detected early at 24 h postinfection with red-spotted grouper nervous necrosis virus in the cytoplasm. later b2 was found i ...	2010	20870718
development and partial characterization of new marine cell line from brain of asian sea bass lates calcarifer for virus isolation.	a new cell line, asian sea bass brain (asbb), was derived from the brain tissue of asian sea bass lates calcarifer. this cell line was maintained in leibovitz l-15 media supplemented with 10% fetal bovine serum (fbs). the asbb cell line was subcultured more than 60 times over a period of 15 mo. the asbb cell line consists predominantly of fibroblastic-like cells and was able to grow at temperatures between 20°c and 30°c with an optimum temperature of 25°c. the growth rate of these cells increase ...	2010	21082288
betanodavirus up-regulates chaperone grp78 via er stress: roles of grp78 in viral replication and host mitochondria-mediated cell death.	whether viral pathogens that induce er stress responses benefit the host or the virus remains controversial. in this study we show that betanodavirus induced er stress responses up-regulate grp78, which regulates the viral replication and host cellular mitochondrial-mediated cell death. betanodavirus (redspotted grouper nervous necrosis virus, rgnnv) infection resulted in the following increased er stress responses in fish gf-1 grouper fin cells: (1) ire-1 and atf-6 sensors at 48 h post-infectio ...	2011	21170590
real-time rt-pcr detection of betanodavirus in naturally and experimentally infected fish from spain.	infections with betanodavirus affect a wide range of wild and farmed fish species throughout the world, mostly from the marine environment. the aim of this work was to develop and validate real-time rt-pcr assays for sensitive and specific detection of nodavirus in diseased or carrier fish. the new detection assay was used to study the transmission and development of nodavirus infection in juvenile sea bass, dicentrarchus labrax (l.), challenged by different routes, and also to screen for nodavi ...	2011	21306586
the reverse genetics applied to fish rna viruses.	abstract: aquaculture has expanded rapidly to become a major economic and food-producing sector worldwide these last 30 years. in parallel, viral diseases have emerged and rapidly spread from farm to farm causing enormous economic losses. the most problematic viruses encountered in the field are mainly, but not exclusively, rna viruses belonging to the novirhabdovirus, aquabirnavirus, alphavirus and betanodavirus genera. the recent establishment of reverse genetics systems to recover infectious ...	2011	21314978
host range, host specificity and hypothesized host shift events among viruses of lower vertebrates.	abstract: the successful replication of a viral agent in a host is a complex process that often leads to a species specificity of the virus and can make interspecies transmission difficult. despite this difficulty, natural host switch seems to have been frequent among viruses of lower vertebrates, especially fish viruses, since there are several viruses known to be able to infect a wide range of species. in the present review we will focus on well documented reports of broad host range, variatio ...	2011	21592358
optimization of betanodavirus culture and enumeration in striped snakehead fish cells.	an optimized culture method for detection of infection of fish with the red spotted grouper nervous necrosis virus (rgnnv) genotype of betanodavirus in striped snakehead (ssn-1, channa striatus) cells is described. inoculation of fish tissue homogenates at the same time or within 4 hr of seeding the ssn-1 cells was as sensitive as the method recommended by the world organization for animal health, where homogenates were adsorbed onto an established cell monolayer. such modification halved the ti ...	2011	21908274
Betanodavirus induces oxidative stress-mediated cell death that prevented by anti-oxidants and zfcatalase in fish cells.	The role of oxidative stress in the pathogenesis of RNA nervous necrosis virus infection is still unknown. Red-spotted grouper nervous necrosis virus (RGNNV) induced free radical species (ROS) production at 12-24 h post-infection (pi; early replication stage) in fish GF-1 cells, and then at middle replication stage (24-48 h pi), this ROS signal may upregulate some expressions of the anti-oxidant enzymes Cu/Zn SOD and catalase, and eventually expression of the transcription factor Nrf2. Furthermo ...	2011	21991373
viral encephalopathy and retinopathy outbreak in freshwater fish farmed in italy.	viral encephalopathy and retinopathy (ver), otherwise known as viral nervous necrosis (vnn), is a neuropathological condition affecting > 40 species of fish. although ver affects mainly marine fish, the disease has also been detected in certain species reared in freshwater environments. there are relatively few reports concerning the disease in freshwater species, and there is not much information on clinical signs. nevertheless, the most common clinical findings reported from affected freshwate ...	2011	21991664
Molecular epidemiology and evolutionary dynamics of betanodavirus in southern Europe.	Viral encephalopathy and retinopathy (VER) is one of the most devastating diseases for marine aquaculture, and similarly represents a threat to wild fish populations because of its high infectivity and broad host range. Betanodavirus, the causative agent of VER, is a small non-enveloped virus with a bipartite RNA genome comprising the RNA1 and RNA2 segments. We partially sequenced both RNA1 and RNA2 from 120 viral strains isolated from 2000 to 2009 in six different countries in Southern Europe. ...	2012	22036789
genome analysis of betanodavirus from cultured marine fish species in malaysia.	betanodavirus is the causative agent of the viral nervous necrosis (vnn) or viral encephalopathy and retinopathy disease in marine fish. this disease is responsible for most of the mass mortalities that occurred in marine fish hatcheries in malaysia. the genome of this virus consists of two positive-sense rna molecules which are the rna1 and rna2. the rna1 molecule contains the rdrp gene which encodes for the rna-dependent rna polymerase and the rna2 molecule contains the cp gene which encodes f ...	2011	22051446
transcriptome analysis of orange-spotted grouper (epinephelus coioides) spleen in response to singapore grouper iridovirus.	orange-spotted grouper (epinephelus coioides) is an economically important marine fish cultured in china and southeast asian countries. the emergence of infectious viral diseases, including iridovirus and betanodavirus, have severely affected food products based on this species, causing heavy economic losses. limited available information on the genomics of e. coioides has hampered the understanding of the molecular mechanisms that underlie host-virus interactions. in this study, we used a 454 p ...	2011	22078027