In-silico designing of epitope-based vaccine against the seven banded grouper nervous necrosis virus affecting fish species

Abstract European sea bass (Dicentrarchus labrax) is severely affected by nervous necrosis disease, caused by nervous necrosis virus (NNV). Two out of the four genotypes of this virus (red-spotted grouper nervous necrosis virus, RGNNV; and striped jack nervous necrosis virus, SJNNV) have been detected in sea bass, although showing different levels of virulence to this fish species. Thus, sea bass is highly susceptible to RGNNV, whereas outbreaks caused by SJNNV have not been reported in this fish species. The role of the capsid protein (Cp) amino acids 247 and 270 in the virulence of a RGNNV isolate to sea bass has been evaluated by the generation of recombinant RGNNV viruses harbouring SJNNV-type amino acids in the above mentioned positions (Mut247Dl965, Mut270Dl965 and Mut247 + 270Dl965). Viral in vitro and in vivo replication, virus virulence and fish immune response triggered by these viruses have been analysed. Mutated viruses replicated on E-11 cells, although showing some differences compared to the wild type virus, suggesting that the mutations can affect the viral cell recognition and entry. In vivo, fish mortality caused by mutated viruses was 75% lower, and viral replication in sea bass brain was altered compared to non-mutated virus. Regarding sea bass immune response, mutated viruses triggered a lower induction of IFN I system and inflammatory response-related genes. Furthermore, mutations caused changes in viral serological properties (especially the mutation in amino acid 270), inducing higher seroconversion and changing antigen recognition.

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Betanodavirus infection in the freshwater model fish medaka (Oryzias latipes)

Journal of General Virology ◽

10.1099/vir.0.81761-0 ◽

2006 ◽

Vol 87 (8) ◽

pp. 2333-2339 ◽

Cited By ~ 31

Author(s):

Ryo Furusawa ◽

Yasushi Okinaka ◽

Toshihiro Nakai

Keyword(s):

Marine Fish ◽

Fish Species ◽

Reverse Genetics ◽

Developmental Stages ◽

Rna Virus ◽

Oryzias Latipes ◽

Engineering System ◽

Nervous Necrosis Virus ◽

Necrosis Virus ◽

Model Fish

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 developmental stages (larval and juvenile). These fish are only available for a few months of the year and are not suitable for the construction of a reverse-genetics system. To overcome these problems, several freshwater fish species were tested for their susceptibility to betanodaviruses. It was found that adult medaka (Oryzias latipes), a well-known model fish, was susceptible to both Striped jack nervous necrosis virus (the type species of the genus Betanodavirus) and Redspotted grouper nervous necrosis virus (RGNNV), which have different host specificities in marine fish species. Infected medaka exhibited erratic swimming and the viruses were localized specifically in the brain, spinal cord and retina of the infected fish, similar to the pattern of infection in naturally infected marine fish. Moreover, medaka were susceptible to RGNNV at the larval stage. This is the first report of a model virus–model host infection system in fish. This system should facilitate elucidation of the mechanisms underlying RNA virus infections in fish.

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Indigenous versus Lessepsian Hosts: Nervous Necrosis Virus (NNV) in Eastern Mediterranean Sea Fish

Viruses ◽

10.3390/v12040430 ◽

2020 ◽

Vol 12 (4) ◽

pp. 430 ◽

Cited By ~ 1

Author(s):

Yael Lampert ◽

Ran Berzak ◽

Nadav Davidovich ◽

Arik Diamant ◽

Nir Stern ◽

...

Keyword(s):

Invasive Species ◽

Mediterranean Sea ◽

Marine Environment ◽

Fish Species ◽

Eastern Mediterranean ◽

Nervous Necrosis Virus ◽

Indigenous Species ◽

Necrosis Virus ◽

Marine Aquaculture ◽

The Mediterranean

Viruses are among the most abundant and diverse biological components in the marine environment. In finfish, viruses are key drivers of host diversity and population dynamics, and therefore, their effect on the marine environment is far-reaching. Viral encephalopathy and retinopathy (VER) is a disease caused by the marine nervous necrosis virus (NNV), which is recognized as one of the main infectious threats for marine aquaculture worldwide. For over 140 years, the Suez Canal has acted as a conduit for the invasion of Red Sea marine species into the Mediterranean Sea. In 2016–2017, we evaluated the prevalence of NNV in two indigenous Mediterranean species, the round sardinella (Sardinella aurita) and the white steenbras (Lithognathus mormyrus) versus two Lessepsian species, the Randall’s threadfin bream (Nemipterus randalli) and the Lessepsian lizardfish (Saurida lessepsianus). A molecular method was used to detect NNV in all four fish species tested. In N. randalli, a relatively newly established invasive species in the Mediterranean Sea, the prevalence was significantly higher than in both indigenous species. In S. lessepsianus, prevalence varied considerably between years. While the factors that influence the effective establishment of invasive species are poorly understood, we suggest that the susceptibility of a given invasive fish species to locally acquired viral pathogens such as NVV may be important, in terms of both its successful establishment in its newly adopted environment and its role as a reservoir ‘host’ in the new area.

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Genomic classification of new betanodavirus isolates by phylogenetic analysis of the coat protein gene suggests a low host-fish species specificity

Journal of General Virology ◽

10.1099/vir.0.80264-0 ◽

2004 ◽

Vol 85 (10) ◽

pp. 3079-3087 ◽

Cited By ~ 77

Author(s):

Richard Thiéry ◽

Joëlle Cozien ◽

Claire de Boisséson ◽

Soasig Kerbart-Boscher ◽

Laurent Névarez

Keyword(s):

Coat Protein ◽

Virus Type ◽

Fish Species ◽

Coat Protein Gene ◽

Protein Gene ◽

Sea Bass ◽

Nervous Necrosis Virus ◽

Marine Fish Species ◽

Necrosis Virus ◽

Host Fish

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 isolated from sea bass (Dicentrarchus labrax) in France were found to be different. In addition, 21 novel strains of betanodavirus from several fish species from France, Spain, Tunisia and Tahiti were classified by using phylogenetic analysis of a partial sequence (383 nt) of the coat protein gene. Most of the isolates were grouped in the red-spotted grouper nervous necrosis virus type, which was subdivided into two subtypes, one of them containing only French isolates. Furthermore, an isolate obtained from sea bass during an outbreak at low temperature (15 °C) was classified as the barfin flounder nervous necrosis virus type. This is the first reported isolation from sea bass of such a strain, which is known to infect several cold-water marine fish species. In addition, a betanodavirus belonging to the striped jack nervous necrosis virus type was detected in Senagalese sole (Solea senegalensis) farmed in Spain, which is the first indication of the presence of this genotype outside Japan. These findings suggest that the different genotypes can infect a variety of fish species and thus have a low host-fish species specificity.

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GHSC70 Is Involved in the Cellular Entry of Nervous Necrosis Virus

Journal of Virology ◽

10.1128/jvi.02523-14 ◽

2014 ◽

Vol 89 (1) ◽

pp. 61-70 ◽

Cited By ~ 31

Author(s):

Jui-Shin Chang ◽

Shau-Chi Chi

Keyword(s):

Capsid Protein ◽

Fish Species ◽

Receptor Protein ◽

Economic Losses ◽

Nervous Necrosis Virus ◽

Specific Receptor ◽

Necrosis Virus ◽

Content Type ◽

Voltage Dependent ◽

Selective Channel

ABSTRACTNervous necrosis virus (NNV) is a devastating pathogen of cultured marine fish and has affected more than 40 fish species. NNV belongs to the betanodaviruses ofNodaviridaeand is a nonenveloped icosahedral particle with 2 single-stranded positive-sense RNAs. To date, knowledge regarding NNV entry into the host cell remains limited, and no NNV-specific receptor protein has been published. Using grouper fin cell line GF-1 and purified NNV capsid protein in a virus overlay protein binding assay (VOPBA), grouper heat shock cognate protein 70 (GHSC70) and grouper voltage-dependent anion selective channel protein 2 (GVDAC2) were investigated as NNV receptor protein candidates. We cloned and sequenced the genes for GHSC70 and GVDAC2 and expressed them inEscherichia colifor antiserum preparation. Knockdown of the expression of GHSC70 and GVDAC2 genes with specific short interfering RNAs (siRNAs) significantly downregulated viral RNA expression in NNV-infected GF-1 cells. By performing an immunoprecipitation assay, we confirmed that GHSC70 interacted with NNV capsid protein, while VDAC2 did not. Immunofluorescence staining and flow cytometry analysis revealed the presence of the GHSC70 protein on the cell surface. After a blocking assay, we detected the NNV RNA2 levels after 1 h of adsorption to GF-1 cells; the level was significantly lower in the cells pretreated with the GHSC70 antiserum than in nontreated cells. Therefore, we suggest that GHSC70 participates in the NNV entry of GF-1 cells, likely functioning as an NNV receptor or coreceptor protein.IMPORTANCEFish nodavirus has caused mass mortality of more than 40 fish species worldwide and resulted in huge economic losses in the past 20 years. Among the four genotypes of fish nodaviruses, the red-spotted grouper nervous necrosis virus (RGNNV) genotype exhibits the widest host range. In our previous study, we developed monoclonal antibodies with high neutralizing efficiency against grouper NNV in GF-1 cells, indicating that NNV-specific receptor(s) may exist on the GF-1 cell membrane. However, no NNV receptor protein has been published. In this study, we found GHSC70 to be an NNV receptor (or coreceptor) candidate through VOBPA and provided several lines of evidence demonstrating that GHSC70 protein has a role in the NNV entry step of GF-1 cells. To the best of our knowledge, this is the first report identifying grouper HSC70 and its role in NNV entry into GF-1 cells.

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