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Anna Toffan - One of the best experts on this subject based on the ideXlab platform.
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Lumpfish (Cyclopterus lumpus, Linnaeus) is susceptible to viral nervous necrosis: Result of an experimental infection with different genotypes of Betanodavirus.
Journal of fish diseases, 2019Co-Authors: Anna Toffan, Maria De Salvador, Felix Scholz, Tobia Pretto, Alessandra Buratin, Hamish D. Rodger, Marica Toson, Argelia Cuenca, Niccolò VendraminAbstract:In recent years, the use of cleaner fish for biological control of sea lice has increased considerably. Along with this, a number of infectious diseases have emerged. The aim of this study was to investigate the susceptibility of lumpfish (Cyclopterus lumpus) to Betanodavirus since it was detected in asymptomatic wild wrasses in Norway and Sweden. Three Betanodaviruses were used to challenge lumpfish: one RGNNV genotype and two BFNNV genotypes. Fish were injected and monitored for 4 weeks. Brain samples from clinically affected specimens, from weekly randomly selected fish and survivors were subjected to molecular testing, viral isolation, histopathology and immunohistochemistry. Reduced survival was observed but was attributed to tail-biting behaviour, since no nervous signs were observed throughout the study. Betanodavirus RNA was detected in all samples, additionally suggesting an active replication of the virus in the brain. Viral isolation confirmed molecular biology results and revealed a high viral titre in BFNNV-infected groups associated with typical lesions in brains and eyes of survivor fish. We concluded that lumpfish are susceptible to Betanodavirus, as proven by the high viral titre and brain lesions detected, but further studies are necessary to understand if Betanodavirus can cause clinical disease in this species.
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Vaccination and immune responses of European sea bass (Dicentrarchus labrax L.) against Betanodavirus.
Fish & Shellfish Immunology, 2019Co-Authors: Francesco Buonocore, Noelia Nuñez-ortiz, Simona Picchietti, Elisa Randelli, Valentina Stocchi, Laura Guerra, Anna Toffan, Francesco Pascoli, Anna Maria Fausto, Massimo MazziniAbstract:Abstract This review summarizes the available knowledge on the immune defences of European sea bass against antigenic preparations derived from the viral encephalopathy and retinopathy virus (Betanodavirus), which represents a major threat to the health of this fish species. The nodavirus is widely present and differentiates into several strains that infect invertebrates (in insects, alphanodavirus) and teleost fish, and thus may represent a great problem for farmed fish species. Many efforts have been directed to discovering new immunizations to induce protection in sea bass, especially at young stages, and these efforts have included employing diverse Betanodavirus strains, antigen preparation, vaccination routes, and the addition of adjuvants and/or immunostimulants. The obtained results showed that inactivated preparations of Betanodavirus that were administered intraperitoneally may induce both immune recognition and protection. Attempts at performing mucosal immunization by immersion and/or oral administration, which is a vaccination route that is highly preferred for sea bass, have shown intriguing results, and more studies are necessary for its improvement. Overall, the objective of identifying a reliable vaccine that also cross-protects against different genotypes or reassortant viruses for use in European sea bass against Betanodavirus appears to be an attainable goal in the near future.
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Lack of in vivo cross-protection of two different Betanodavirus species RGNNV and SJNNV in European sea bass Dicentrachus labrax.
Fish & shellfish immunology, 2017Co-Authors: Francesco Pascoli, Francesco Buonocore, Alessandra Buratin, Marica Toson, Giuseppe Scapigliati, Andrea Guazzo, Anna ToffanAbstract:Viral encephalopathy and retinopathy (VER) is a severe infective disease characterized by neuropathological changes in several fish species associated with high mortality. The etiological agent is a virus belonging to the Nodaviridae family, genus Betanodavirus. To date, four different Betanodavirus species have been officially recognized by International Committee on Taxonomy of Viruses (ICTV), namely the red-spotted grouper- (RGNNV), the striped jack- (SJNNV), the barfin flounder- (BFNNV) and the tiger puffer nervous necrosis virus (TPNNV). Moreover, two reassortants RGNNV/SJNNV and SJNNV/RGNNV have been described. Betanodaviruses can be classified into three different serotypes (A, B and C) that are antigenically different, so none (between serotype A and C) or partial (between serotype B and C) cross-immunoreactivity has been detected in vitro. In this study we investigated the in vivo cross-protection of the two main Betanodavirus species (RGNNV and SJNNV), which belong to distinct serotype, by immunizing intraperitoneally (IP) juvenile sea bass with formalin inactivated RGNNV and SJNNV vaccines, followed by a challenge with RGNNV. Fish IP vaccinated with inactivated RGNNV showed a high protection value (85%). Serological analyses highlighted a great specific anti-NNV immunoglobulin M (IgM) production against the homologous virus, while a good seroconversion with low neutralization property was highlighted against the heterologous virus. In fish IP vaccinated with inactivated SJNNV the protection recorded was equal to 25%, significantly lower respect to the one provided by RGNNV IP vaccine. ELISA test detected good IgM production against the homologous virus, and a lower, but still detectable IgM production against the heterologous one. By contrast, serum neutralization test highlighted a poorly detectable antibody production unable to neutralize either the homologous or the heterologous virus. These results confirm that the two serotypes are not cross-protective in vivo. According to these findings, the production of multivalent formulation, or at least the provision of different types of vaccines based on both fish and virus species requirement, should be recommended in order to broaden the range of protection.
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viral nervous necrosis in gilthead sea bream sparus aurata caused by reassortant Betanodavirus rgnnv sjnnv an emerging threat for mediterranean aquaculture
Scientific Reports, 2017Co-Authors: Anna Toffan, Francesco Pascoli, Tobia Pretto, Alessandra Buratin, Valentina Panzarin, Miriam Abbadi, Rosita Quartesan, Daniel Gijon, Francesc PadrosAbstract:: Viral nervous necrosis (VNN) certainly represents the biggest challenge for the sustainability and the development of aquaculture. A large number of economically relevant fish species have proven to be susceptible to the disease. Conversely, gilthead sea bream has generally been considered resistant to VNN, although it has been possible to isolate the virus from apparently healthy sea bream and sporadically from affected larvae and postlarvae. Unexpectedly, in 2014-2016 an increasing number of hatcheries in Europe have experienced mass mortalities in sea bream larvae. Two clinical outbreaks were monitored over this time span and findings are reported in this paper. Despite showing no specific clinical signs, the affected fish displayed high mortality and histological lesions typical of VNN. Fish tested positive for Betanodavirus by different laboratory techniques. The isolates were all genetically characterized as being reassortant strains RGNNV/SJNNV. A genetic characterization of all sea bream Betanodaviruses which had been isolated in the past had revealed that the majority of the strains infecting sea bream are actually RGNNV/SJNNV. Taken together, this information strongly suggests that RGNNV/SJNNV Betanodavirus possesses a particular tropism to sea bream, which can pose a new and unexpected threat to the Mediterranean aquaculture.
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Molecular Basis for Antigenic Diversity of Genus Betanodavirus.
PloS one, 2016Co-Authors: Valentina Panzarin, Anna Toffan, Alessandra Buratin, Miriam Abbadi, Marzia Mancin, Stine Braaen, Christel Moræus Olsen, Luca Bargelloni, Espen Rimstad, Giovanni CattoliAbstract:Betanodaviruses are the causative agents of viral nervous necrosis (VNN), a devastating disease for the Mediterranean mariculture. Four different Betanodavirus species are recognized, Striped jack-, Redspotted grouper-, Tiger puffer-, and Barfin flounder nervous necrosis virus (SJNNV, RGNNV, TPNNV and BFNNV), but there is little knowledge on their antigenic properties. In order to describe the serological relationships among different Betanodavirus genotypes, serum neutralization assays were performed using rabbit polyclonal antisera against eight fish nodaviruses that cover a wide species-, temporal-, spatial- and genetic range. The results indicate that the SJNNV and RGNNV are antigenically distinct, constituting serotypes A and C, respectively. The TPNNV and BFNNV, the latter representing cold-water Betanodaviruses, are antigenically related and cluster within serotype B. The reassortant viruses RGNNV/SJNNV and SJNNV/RGNNV group within serotypes A and C, respectively, indicating that the coat protein encoded by RNA2 acts as major immunoreactivity determinant. Immunostaining of in vitro expressed wild type and chimeric capsid proteins between the RGNNV and the SJNNV species indicated that the C-terminal part of the capsid protein retains the immunoreactive portion. The amino acid (aa) residues determining RGNNV and SJNNV antigenic diversity were mapped to aa residues 217-256 and aa 257-341, respectively. Neutralization of reverse genetics derived chimeric viruses indicated that these areas determine the neutralizing epitopes. The data obtained are crucial for the development of targeted serological tests for the diagnosis of VNN, and informative for development of cross-protective vaccines against various Betanodavirus genotypes.
Paul Hick - One of the best experts on this subject based on the ideXlab platform.
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Recurrent outbreaks of viral nervous necrosis in intensively cultured barramundi (Lates calcarifer) due to horizontal transmission of Betanodavirus and recommendations for disease control
Aquaculture, 2011Co-Authors: Paul Hick, Glenn Schipp, Jérôme Bosmans, John Humphrey, Richard WhittingtonAbstract:Abstract Betanodavirus is a significant constraint to marine finfish aquaculture worldwide and the development of control strategies will depend on accurate data about mechanisms of infection. A repeated cross-sectional survey for Betanodavirus infection using a real-time reverse transcriptase-polymerase chain reaction assay and virus isolation was conducted in barramundi ( Lates calcarifer ) at a hatchery in Australia to determine whether transmission occurred vertically from the broodstock or horizontally from the environment. Six consecutive production batches were studied in 2007–2008. Subclinical infection was detected in Batch 2007-1 with onset of infection after 26 d of age, reaching a true prevalence > 90.9% (lower 95% probability limit) by 40 d. Two days later an outbreak of VNN with 100% mortality occurred in 12 d old larvae in Batch 2007-2, due to infection with an identical Betanodavirus, which was not detected in this batch 7 d previously. A point source epidemic initiated by horizontal transmission from the environment (probably seawater) to Batch 1 then Batch 2 was confirmed. Betanodavirus was not endemic in the hatchery and was probably not vertically transmitted because: (i) infection was not detected in the eggs or larvae of any batch; (ii) infection was not detected in 4 out of 6 batches; (iii) infection was not identified in 20 broodstock; iv) freedom from infection of life history stages was determined with a very high degree of confidence; v) similar data were derived from archival samples from a batch in 2005. The data suggested that outbreaks were initiated because of vulnerabilities in the water supply and spread because biosecurity measures failed. The age of fish appeared to be a major risk factor for susceptibility to disease.
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Preparation of fish tissues for optimal detection of Betanodavirus
Aquaculture, 2010Co-Authors: Paul Hick, Alison Tweedie, Richard J. WhittingtonAbstract:Abstract Betanodaviruses, the causative agents of viral nervous necrosis (VNN) disease, cause significant production losses in aquaculture worldwide. To improve the detection of Betanodavirus, methods of extraction of the virus and its RNA from the tissues of sub-clinically infected barramundi (Lates calcarifer) and Australian bass (Macquaria novemaculeata) were evaluated. Compared to crude manual processing, the yield of Betanodavirus was increased by 3.5-fold when tissues were disrupted by micro-bead beating and 2-fold using an optimised manual pestle grinding method (p
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optimisation and validation of a real time reverse transcriptase polymerase chain reaction assay for detection of Betanodavirus
Journal of Virological Methods, 2010Co-Authors: Paul Hick, Richard WhittingtonAbstract: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 TCID50 units per reaction. The assay was highly repeatable (standard deviation of estimated log10 (viral copies) 0.10 ± 0.08) and reproducible (standard deviation of estimated log10 (viral copies) 0.08 ± 0.06). Diagnostic accuracy was assessed in 2193 samples comprising tissue homogenates from Australian bass (Macquaria novemaculeata) and barramundi (Lates calcarifer), and also in SSN-1 tissue culture supernatants, using virus isolation in striped snake head (SSN-1) cell culture as the gold standard. Diagnostic sensitivity and specificity were 100% when the assay was applied to Australian bass tissue and SSN-1 tissue culture supernatants, but for barramundi tissue were 99.1% and 92.8%, respectively. The apparent imperfect specificity was shown by specific amplification of alternate regions of the Betanodavirus genome to be due to the lower sensitivity of virus isolation. This is the first study to report the diagnostic performance of a RT-qPCR assay for detection of Betanodavirus.
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Optimisation and validation of a real-time reverse transcriptase-polymerase chain reaction assay for detection of Betanodavirus.
Journal of virological methods, 2009Co-Authors: Paul Hick, Richard WhittingtonAbstract: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
Richard Whittington - One of the best experts on this subject based on the ideXlab platform.
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Recurrent outbreaks of viral nervous necrosis in intensively cultured barramundi (Lates calcarifer) due to horizontal transmission of Betanodavirus and recommendations for disease control
Aquaculture, 2011Co-Authors: Paul Hick, Glenn Schipp, Jérôme Bosmans, John Humphrey, Richard WhittingtonAbstract:Abstract Betanodavirus is a significant constraint to marine finfish aquaculture worldwide and the development of control strategies will depend on accurate data about mechanisms of infection. A repeated cross-sectional survey for Betanodavirus infection using a real-time reverse transcriptase-polymerase chain reaction assay and virus isolation was conducted in barramundi ( Lates calcarifer ) at a hatchery in Australia to determine whether transmission occurred vertically from the broodstock or horizontally from the environment. Six consecutive production batches were studied in 2007–2008. Subclinical infection was detected in Batch 2007-1 with onset of infection after 26 d of age, reaching a true prevalence > 90.9% (lower 95% probability limit) by 40 d. Two days later an outbreak of VNN with 100% mortality occurred in 12 d old larvae in Batch 2007-2, due to infection with an identical Betanodavirus, which was not detected in this batch 7 d previously. A point source epidemic initiated by horizontal transmission from the environment (probably seawater) to Batch 1 then Batch 2 was confirmed. Betanodavirus was not endemic in the hatchery and was probably not vertically transmitted because: (i) infection was not detected in the eggs or larvae of any batch; (ii) infection was not detected in 4 out of 6 batches; (iii) infection was not identified in 20 broodstock; iv) freedom from infection of life history stages was determined with a very high degree of confidence; v) similar data were derived from archival samples from a batch in 2005. The data suggested that outbreaks were initiated because of vulnerabilities in the water supply and spread because biosecurity measures failed. The age of fish appeared to be a major risk factor for susceptibility to disease.
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optimisation and validation of a real time reverse transcriptase polymerase chain reaction assay for detection of Betanodavirus
Journal of Virological Methods, 2010Co-Authors: Paul Hick, Richard WhittingtonAbstract: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 TCID50 units per reaction. The assay was highly repeatable (standard deviation of estimated log10 (viral copies) 0.10 ± 0.08) and reproducible (standard deviation of estimated log10 (viral copies) 0.08 ± 0.06). Diagnostic accuracy was assessed in 2193 samples comprising tissue homogenates from Australian bass (Macquaria novemaculeata) and barramundi (Lates calcarifer), and also in SSN-1 tissue culture supernatants, using virus isolation in striped snake head (SSN-1) cell culture as the gold standard. Diagnostic sensitivity and specificity were 100% when the assay was applied to Australian bass tissue and SSN-1 tissue culture supernatants, but for barramundi tissue were 99.1% and 92.8%, respectively. The apparent imperfect specificity was shown by specific amplification of alternate regions of the Betanodavirus genome to be due to the lower sensitivity of virus isolation. This is the first study to report the diagnostic performance of a RT-qPCR assay for detection of Betanodavirus.
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Optimisation and validation of a real-time reverse transcriptase-polymerase chain reaction assay for detection of Betanodavirus.
Journal of virological methods, 2009Co-Authors: Paul Hick, Richard WhittingtonAbstract: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
S Hammami - One of the best experts on this subject based on the ideXlab platform.
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complete sequencing of tunisian redspotted grouper nervous necrosis virus Betanodavirus capsid gene and rna dependent rna polymerase gene
Journal of Fish Diseases, 2010Co-Authors: N Chérif, Nellie Gagné, David B. Groman, Frederick S. B. Kibenge, Tokinori Iwamoto, Carmencita V. Yason, S HammamiAbstract: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 study reports on the characterization of the full RNA2 sequence of a Tunisian Betanodavirus with a length of 1433 nt, containing a 339 amino acid open-reading frame encoding the CP, and typing to the redspotted grouper nervous necrosis virus Ia genotype following phylogenetic analysis. The homology of the capsid protein to other Betanodaviruses or alphanodaviruses was compared. In addition, a full length RNA1 sequence of 3104 nt encoding a 982 amino acid RNA-dependent RNA polymerase was obtained.
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Complete sequencing of Tunisian redspotted grouper nervous necrosis virus Betanodavirus capsid gene and RNA‐dependent RNA polymerase gene
Journal of fish diseases, 2009Co-Authors: N Chérif, Nellie Gagné, David B. Groman, Frederick S. B. Kibenge, Tokinori Iwamoto, Carmencita V. Yason, S HammamiAbstract: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 study reports on the characterization of the full RNA2 sequence of a Tunisian Betanodavirus with a length of 1433 nt, containing a 339 amino acid open-reading frame encoding the CP, and typing to the redspotted grouper nervous necrosis virus Ia genotype following phylogenetic analysis. The homology of the capsid protein to other Betanodaviruses or alphanodaviruses was compared. In addition, a full length RNA1 sequence of 3104 nt encoding a 982 amino acid RNA-dependent RNA polymerase was obtained.
Isabel Bandín - One of the best experts on this subject based on the ideXlab platform.
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Betanodavirus infection in primary neuron cultures from sole
Veterinary Research, 2018Co-Authors: Sandra Souto, J G Olveira, Carlos P. Dopazo, Lucía Vázquez-salgado, Isabel BandínAbstract:Nervous necrosis virus (NNV), G. Betanodavirus, is the causative agent of viral encephalopathy and retinopathy, a disease that causes mass mortalities in a wide range of fish species. Betanodaviruses are neurotropic viruses and their replication in the susceptible fish species seems to be almost entirely restricted to nerve tissue. However, none of the cell lines used for NNV propagation has a nervous origin. In this study, first we established a protocol for the primary culture of neurons from Senegalese sole, which made it possible to further study virus-host cell interactions. Then, we compared the replication of three NNV strains with different genotypes (SJNNV, RGNNV and a RGNNV/SJNNV reassortant strain) in sole neuron primary cultures and E-11 cells. In addition, to study how two amino acid substitutions at the c-terminal of the capsid protein (positions 247 and 270) affect the binding to cell receptors, a recombinant strain was also tested. The results show that sole neural cells enabled replication of all the tested NNV strains. However, the recombinant strain shows a clearly delayed replication when compared with the wt strain. This delay was not observed in virus replicating in E-11 cells, suggesting a viral interaction with different cell receptors. The establishment of a sole primary neuronal culture protocol provides an important tool for research into Betanodavirus infection in sole.
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Betanodavirus infection in bath-challenged Solea senegalensis juveniles: A comparative analysis of RGNNV, SJNNV and reassortant strains.
Journal of fish diseases, 2018Co-Authors: Sandra Souto, J G Olveira, M. Carmen Alonso, Carlos P. Dopazo, Isabel BandínAbstract:Senegalese sole has been shown to be highly susceptible to Betanodavirus infection, although virulence differences were observed between strains. To study the mechanisms involved in these differences, we have analysed the replication in brain tissue of three strains with different genotypes during 15 days after bath infection. In addition, possible portals of entry for Betanodavirus into sole were investigated. The reassortant RGNNV/SJNNV and the SJNNV strain reached the brain after 1 and 2 days postinfection, respectively. Although no RGNNV replication was detected until day 3-4 postinfection, at the end of the experiment this strain yielded the highest viral load; this is in accordance with previous studies in which sole infected with the reassortant showed more acute signs and earlier mortality than the RGNNV and SJNNV strains. Differences between strains were also observed in the possible portals of entry. Thus, whereas the reassortant strain could infect sole mainly through the skin or the oral route, and, to a minor extent, through the gills, the SJNNV strain seems to enter fish only through the gills and the RGNNV strain could use all tissues indistinctly. Taken together, all these results support the hypothesis that reassortment has improved Betanodavirus infectivity for sole.
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Reassortant Betanodavirus infection in turbot (Scophthalmus maximus).
Journal of fish diseases, 2016Co-Authors: Sandra Souto, J G Olveira, Carlos P. Dopazo, Isabel BandínAbstract:In this study, the susceptibility of turbot juveniles to two Betanodavirus strains was assessed, a RGNNV/SJNNV reassortant (Ss160.03) and a SJNNV strain. The reassortant isolate exhibits a slightly modified SJNNV CP, with two amino acid substitutions in the C-terminal domain (positions 247 and 270). To analyse the role of these residues as virulence and host determinants in turbot, three recombinant strains (rSs160.03247 , rSs160.03270 , rSs160.03247+270 ) harbouring site-specific mutations in the CP sequence were also tested in experimental trials. Moderate mortalities (up to 50%) were recorded at 18 °C in the fish challenged with the Ss160.03 strain, whereas low mortalities (17%) were observed in the group challenged with the SJNNV strain. A slight decrease (around 10%) was observed in the mortalities caused by the mutants rSs160.03247 and rSs160.03270 , whilst the mutation of both positions reduced mortality by more than half of that observed in fish challenged with the wild strain. These results are confirmed by the replication in brain tissues, because whereas the wild strain was detected from 5 to 30 dpi and reached the highest viral load, the recombinant virus harbouring both mutations was not detected in the brain until 20 dpi and with a moderate viral load.
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Influence of temperature on Betanodavirus infection in Senegalese sole (Solea senegalensis).
Veterinary microbiology, 2015Co-Authors: Sandra Souto, J G Olveira, Isabel BandínAbstract:In this study Senegalese sole juveniles were experimentally infected with a reassortant Betanodavirus strain at three different temperatures: 22 °C, 18 °C and 16 °C by bath challenge and cohabitation. The results obtained showed that virus virulence decreased by reducing the water temperature. At 22 °C mortalities reached 100%, at 18 °C they ranged from 75 to 80% and at 16 °C only 8% of the fish died. In addition, horizontal transmission was demonstrated regardless of the rearing temperature. At 16 °C active viral replication was detected up to 66 days post-infection, but no signs of the disease were observed and only a very low level of mortality was recorded. The increase in water temperature from 16 to 22 °C caused a quick rise in the viral load and a subsequent outbreak of mortalities. These findings demonstrate that this reassortant Betanodavirus strain can cause a persistent infection in Senegalese sole at low temperatures (16 °C) for long periods of time, and when temperature increases the virus is able to trigger an acute infection and provoke high mortalities.
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In vitro and in vivo characterization of molecular determinants of virulence in reassortant Betanodavirus
Journal of General Virology, 2015Co-Authors: Sandra Souto, Emilie Mérour, Stephane Biacchesi, Michel Bremont, José G. Olveira, Isabel BandínAbstract:We previously reported that Betanodavirus reassortant strains [redspotted grouper nervous necrosis virus/striped jack nervous necrosis virus (SJNNV)] isolated from Senegalese sole (Solea senegalensis) exhibited a modified SJNNV capsid amino acid sequence, with changes at aa 247 and 270. In the current study, we investigated the possible role of both residues as putative virulence determinants. Three recombinant viruses harbouring site-specific mutations in the capsid protein sequence, rSs160.03(247) (S247A), rSs160.03(270) (S270N) and rSs160.03(247+270) (S247A/S270N), were generated using a reverse genetics system. These recombinant viruses were studied in cell culture and in vivo in the natural fish host. The three mutant viruses were shown to be infectious and able to replicate in E-11 cells, reaching final titres similar to the WT virus, although with a somewhat slower kinetics of replication. When the effect of the amino acid substitutions on virus pathogenicity was evaluated in Senegalese sole, typical clinical signs of Betanodavirus infection were observed in all groups. However, fish mortality induced by all three mutant viruses was clearly affected. Roughly 40% of the fish survived in these three groups in contrast with the WT virus which killed 100% of the fish. These data demonstrated that aa 247 and 270 play a major role in Betanodavirus virulence although when both mutated aa 247 and 270 are present, corresponding recombinant virus was not further attenuated.