Distant Strains of the Fish Rhabdovirus VHSV Maintain a Sixth Functional Cistron Which Codes for a Nonstructural Protein of Unknown Function

AbstractWe used direct RNA sequencing to determine the genomic organization of the region downstream from the G gene of viral hemorrhagic septicemia virus (VHSV), a fish rhabdovirus. This region contains a gene coding for a protein, identified as nonvirion protein (NV), and the gene coding for the RNA polymerase (L), Thus, VHSV genome organization was confirmed to be 3′-N-P-M-G-NV-L-5′ In both a virulent European (07-71) and an avirulent North American (Makah) strain, the NV gene is transcribed into a small mRNA that codes for a protein of 122 amino acids. It has no significant sequence similarity with the infectious hematopoietic necrosis virus NV protein nor with any other known protein. We expressed the NV protein as a fusion protein with the glutathione S-transferase of Schistosoma japonicum and used the purified fusion protein to immunize rabbits. The rabbit antiserum precipitated from infected cell extracts — and not from noninfected cells or purified virions — a protein of 14 kDa, well in accordance with the expected NV gene product size. The prediction that the NV protein is a nonstructural protein is supported by its absence from mature virions although it is present in infected cells

Fish responses to viruses : From interferon to T-cells

Viruses affecting farmed fish species, such as the rainbow trout, have been studied extensively because they cause significant economic losses. Studies on vaccines developed against Viral Hemorrhagic Septicemia (VHS) have provided evidence of an effective and specific response based on neutralizing antibodies, as well as of an immune memory. Various techniques of differential transcript analysis were used to investigate the non-specific leukocyte response to the VHS virus in trout. As in mammals, this response was dependent on interferon-responsive genes. A VDJ junction spectratyping approach of transcripts of the specific T-cell antigen receptor (TCR) was also developed to examine the specific cellular response in rainbow trout. This approach was used to show the existence of complex antiviral T-cell responses in fish.Les virus des poissons d'intérêt agronomique, comme la truite arc-en-ciel, ont été bien étudiés parce qu'ils causent des dommages significatifs dans les élevages. Ainsi, des vaccins ont été développés contre la Septicémie Hémorragique Virale (SHV), montrant l'existence d'une réponse efficace et spécifique basée sur les anticorps neutralisants et celle d'une mémoire immunitaire. La réponse non spécifique des leucocytes de truite induite par le virus de la SHV a pu être explorée par différentes techniques d'analyse différentielle des transcrits. Il a ainsi été démontré que les gènes induits par l'interféron orchestrent cette réponse, comme c'est aussi le cas chez les mammifères. Enfin, une stratégie de spectratypage des longueurs de jonctions VDJ des transcrits du récepteur spécifique de l'antigène des lymphocytes T (TCR) a été développée pour l'étude de la réponse cellulaire spécifique chez la truite arc-en-ciel. Cette approche a permis de démontrer l'existence de réponses T antivirales complexes chez les poissons

Bacterial diseases of farmed fish

An overview of bacterial diseases in farmed fish is presented, including bacterial species involved, predisposing factors, routes of infection, modes of transmission, and control. INRA studies on the main bacterial pathogen of salmonid fish worldwide, Flavobacterium psychrophilum, are also presented. Once techniques of culture, identification, and molecular typing of isolates were defined, a reproducible model of experimental infection was developed using the rainbow trout. Molecular studies improved considerably the taxonomy and phylogeny of the whole Flavobacteriaceae family. The full genome of a F. psychrophilum strain has recently been sequenced. Its analysis has already identified many genes involved in the pathogen’s lifestyle, especially in its virulence towards the host.Un aperçu général des infections bactériennes chez les poissons d'élevage est d'abord présenté: espèces bactériennes en cause, facteurs favorisants, types d'infection, modes de transmission, et méthodes de lutte. Les études menées par l'INRA sur le principal pathogène bactérien des salmonidés dans le monde, Flavobacterium psychrophilum, sont ensuite passées en revue. Après les techniques de culture, d'identification et de typage moléculaire des souches, un modèle expérimental reproductible de la maladie sur la truite arc-en-ciel a été mis au point. Les études moléculaires ont considérablement affiné la taxonomie et la phylogénie de l'ensemble de la famille des Flavobacteriaceae. Le génome complet d'une souche de F. psychrophilum vient d'être séquencé. Son analyse a déjà révélé de nombreux gènes impliqués dans le mode de vie de la bactérie, en particulier dans sa virulence pour l'hôte

A large new subset of TRIM genes highly diversified by duplication and positive selection in teleost fish

<p>Abstract</p> <p>Background</p> <p>In mammals, the members of the tripartite motif (TRIM) protein family are involved in various cellular processes including innate immunity against viral infection. Viruses exert strong selective pressures on the defense system. Accordingly, antiviral TRIMs have diversified highly through gene expansion, positive selection and alternative splicing. Characterizing immune TRIMs in other vertebrates may enlighten their complex evolution.</p> <p>Results</p> <p>We describe here a large new subfamily of TRIMs in teleosts, called finTRIMs, identified in rainbow trout as virus-induced transcripts. FinTRIMs are formed of nearly identical RING/B-box regions and C-termini of variable length; the long variants include a B30.2 domain. The zebrafish genome harbors a striking diversity of finTRIMs, with 84 genes distributed in clusters on different chromosomes. A phylogenetic analysis revealed different subsets suggesting lineage-specific diversification events. Accordingly, the number of <it>fintrim </it>genes varies greatly among fish species. Conserved syntenies were observed only for the oldest <it>fintrims</it>. The closest mammalian relatives are <it>trim16 </it>and <it>trim25</it>, but they are not true orthologs. The B30.2 domain of zebrafish finTRIMs evolved under strong positive selection. The positions under positive selection are remarkably congruent in finTRIMs and in mammalian antiviral TRIM5α, concentrated within a viral recognition motif in mammals. The B30.2 domains most closely related to finTRIM are found among NOD-like receptors (NLR), indicating that the evolution of TRIMs and NLRs was intertwined by exon shuffling.</p> <p>Conclusion</p> <p>The diversity, evolution, and features of finTRIMs suggest an important role in fish innate immunity; this would make them the first TRIMs involved in immunity identified outside mammals.</p

Genetic Resistance to Rhabdovirus Infection in Teleost Fish Is Paralleled to the Derived Cell Resistance Status

Genetic factors of resistance and predisposition to viral diseases explain a significant part of the clinical variability observed within host populations. Predisposition to viral diseases has been associated to MHC haplotypes and T cell immunity, but a growing repertoire of innate/intrinsic factors are implicated in the genetic determinism of the host susceptibility to viruses. In a long-term study of the genetics of host resistance to fish rhabdoviruses, we produced a collection of double-haploid rainbow trout clones showing a wide range of susceptibility to Viral Hemorrhagic Septicemia Virus (VHSV) waterborne infection. The susceptibility of fibroblastic cell lines derived from these clonal fish was fully consistent with the susceptibility of the parental fish clones. The mechanisms determining the host resistance therefore did not associate with specific host immunity, but rather with innate or intrinsic factors. One cell line was resistant to rhabdovirus infection due to the combination of an early interferon IFN induction - that was not observed in the susceptible cells - and of yet unknown factors that hamper the first steps of the viral cycle. The implication of IFN was well consistent with the wide range of resistance of this genetic background to VSHV and IHNV, to the birnavirus IPNV and the orthomyxovirus ISAV. Another cell line was even more refractory to the VHSV infection through different antiviral mechanisms. This collection of clonal fish and isogenic cell lines provides an interesting model to analyze the relative contribution of antiviral pathways to the resistance to different viruses

Origin and Evolution of TRIM Proteins: New Insights from the Complete TRIM Repertoire of Zebrafish and Pufferfish

Tripartite motif proteins (TRIM) constitute a large family of proteins containing a RING-Bbox-Coiled Coil motif followed by different C-terminal domains. Involved in ubiquitination, TRIM proteins participate in many cellular processes including antiviral immunity. The TRIM family is ancient and has been greatly diversified in vertebrates and especially in fish. We analyzed the complete sets of trim genes of the large zebrafish genome and of the compact pufferfish genome. Both contain three large multigene subsets - adding the hsl5/trim35-like genes (hltr) to the ftr and the btr that we previously described - all containing a B30.2 domain that evolved under positive selection. These subsets are conserved among teleosts. By contrast, most human trim genes of the other classes have only one or two orthologues in fish. Loss or gain of C-terminal exons generated proteins with different domain organizations; either by the deletion of the ancestral domain or, remarkably, by the acquisition of a new C-terminal domain. Our survey of fish trim genes in fish identifies subsets with different evolutionary dynamics. trims encoding RBCC-B30.2 proteins show the same evolutionary trends in fish and tetrapods: they evolve fast, often under positive selection, and they duplicate to create multigenic families. We could identify new combinations of domains, which epitomize how new trim classes appear by domain insertion or exon shuffling. Notably, we found that a cyclophilin-A domain replaces the B30.2 domain of a zebrafish fintrim gene, as reported in the macaque and owl monkey antiretroviral TRIM5α. Finally, trim genes encoding RBCC-B30.2 proteins are preferentially located in the vicinity of MHC or MHC gene paralogues, which suggests that such trim genes may have been part of the ancestral MHC

La glycoproteine du virus rabique. Antigenicite, variabilite de la sequence et heterogeneite

SIGLEAvailable from INIST (FR), Document Supply Service, under shelf-number : T 80568 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Réponses antivirales des poissons : de l'interféron aux lymphocytes T

National audienceViruses affecting farmed fish species, such as the rainbow trout, have been studied extensively because they cause significant economic losses. Studies on vaccines developed against Viral Hemorrhagic Septicemia (VHS) have provided evidence of an effective and specific response based on neutralizing antibodies, as well as of an immune memory. Various techniques of differential transcript analysis were used to investigate the non-specific leukocyte response to the VHS virus in trout. As in mammals, this response was dependent on interferon-responsive genes. A VDJ junction spectratyping approach of transcripts of the specific T-cell antigen receptor (TCR) was also developed to examine the specific cellular response in rainbow trout. This approach was used to show the existence of complex antiviral T-cell responses in fish.Les virus des poissons d'intérêt agronomique, comme la truite arc-en-ciel, ont été bien étudiés parce qu'ils causent des dommages significatifs dans les élevages. Ainsi, des vaccins ont été développés contre la Septicémie Hémorragique Virale (SHV), montrant l'existence d'une réponse efficace et spécifique basée sur les anticorps neutralisants et celle d'une mémoire immunitaire. La réponse non spécifique des leucocytes de truite induite par le virus de la SHV a pu être explorée par différentes techniques d'analyse différentielle des transcrits. Il a ainsi été démontré que les gènes induits par l'interféron orchestrent cette réponse, comme c'est aussi le cas chez les mammifères. Enfin, une stratégie de spectratypage des longueurs de jonctions VDJ des transcrits du récepteur spécifique de l'antigène des lymphocytes T (TCR) a été développée pour l'étude de la réponse cellulaire spécifique chez la truite arcen- ciel. Cette approche a permis de démontrer l'existence de réponses T antivirales complexes chez les poissons

Infections et immunité des poissons : apport du modèle vSHV/truite arc-en-ciel

National audienceUne nouvelle approche de l'étude de l'immunité des poissons est apparue avec le développement de la génomique, les poissons ayant joué un rôle important dans la réalisation du séquençage des génomes eucaryotes. L'étude de leurs pathologies s'intègre ainsi de plus en plus à la recherche sur le contrôle des agents infectieux en vue d'une compréhension globale du système immunitaire des vertébrés et de ses mécanismes adaptatifs et régulateurs complexes