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Paralichthys olivaceus rhabdovirus (PORV), which is associated with high mortality rates in flounder, was isolated in China in 2005. Here, we provide an annotated sequence record of PORV, the genome of which comprises 11,182 nucleotides and contains six genes in the order 3′-N-P-M-G-NV-L-5′. Phylogenetic analysis based on glycoprotein sequences of PORV and other rhabdoviruses showed that PORV clusters with viral haemorrhagic septicemia virus (VHSV), genus Novirhabdovirus, family Rhabdoviridae. Further phylogenetic analysis of the combined amino acid sequences of six proteins of PORV and VHSV strains showed that PORV clusters with Korean strains and is closely related to Asian strains, all of which were isolated from flounder. In a comparison in which the sequences of the six proteins were combined, PORV shared the highest identity (98.3 %) with VHSV strain KJ2008 from Korea. [ABSTRACT FROM AUTHOR]

Abstract: Plasmid DNAs containing Siniperca chuatsi rhabdovirus (SCRV) glycoprotein gene (pcDNA-G) and nucleoprotein gene (pcDNA-N) were constructed, and used to determine the antiviral immune response elicited by DNA vaccination in mandarin fish. In vitro and in vivo expression of the plasmid constructs was confirmed in transfected cells and muscle tissues of vaccinated fish by Western blot, indirect immunofluorescence or RT-PCR analysis. Fish injected with pcDNA-G exhibited protective effect against SCRV challenge with a relative percent survival (RPS) of 77.5%, but no significant protection (RPS of 2.5%) was observed in fish vaccinated with pcDNA-N. Immunohistochemical analysis showed that vaccination with pcDNA-G decreased histological lesions and suppressed the virus replication in fish target organs, e.g. kidney, liver, spleen, gill and heart. Transcriptional analysis further revealed that the expression levels of type I IFN system genes including interferon regulation factor-7 (IRF-7) gene, myxovirus resistance (Mx) gene and virus inhibitory protein (Viperin) gene were strongly up-regulated after injection with pcDNA-G, whereas the level of transcription of immunoglobulin M (IgM) gene did not show a statistically significant change. These results reveal that type I IFN antiviral immune response is rapidly triggered by the plasmid DNA containing rhabdovirus glycoprotein gene in fish, which offers an explanation of molecular mechanisms for DNA vaccination inducing mandarin fish resist to SCRV disease. [Copyright &y& Elsevier]

Abstract: Co-infection of two viruses has been observed in mandarin fish (Siniperca chuatsi), but the two viruses have not been characterized. In this study, a rhabdovirus has been isolated from the co-infected two viruses extracted from the diseased mandarin fish, and its morphological structure and partial biochemical and biophysical characteristics have been observed and analyzed. The isolated rhabdovirus has a typical bullet shape, and is therefore called S. chuatsi rhabdovirus (SCRV). And, the isolated rhabdovirus produced a higher titer (108.5 TCID50 ml−1) than did the co-infecting viruses (106.5 TCID50 ml−1). Subsequently, the viral genome RNA was extracted, and used as template to clone the complete nucleoprotein (N) gene by RT-PCR amplification. Cloning and sequencing of the SCRV N protein revealed 42%–31% amino acid identities to that of trout rhabdovirus 903/87 and the rhabdoviruses in genus Vesiculovirus. SDS-PAGE separation of the isolated SCRV and other two rhabdoviruses also revealed obvious polypeptide profile difference. Moreover, the anti-SCRV N protein antibody was prepared, and the anti-SCRV N protein antibody only could recognize the SCRV N protein, whereas no antigenicity was detected in other two rhabdoviruses. The data suggested that the SCRV should be a rhabdovirus member related to the genus Vesiculovirus in the Rhabdoviridae. [Copyright &y& Elsevier]

Abstract: Genomic sequence of Scophthalmus maximus rhabdovirus (SMRV) isolated from diseased turbot has been characterized. The complete genome of SMRV comprises 11,492 nucleotides and encodes five typical rhabdovirus genes N, P, M, G and L. In addition, two open reading frames (ORF) are predicted overlapping with P gene, one upstream of P and smaller than P (temporarily called Ps), and another in P gene which may encodes a protein similar to the vesicular stomatitis virus C protein. The C ORF is contained within the P ORF. The five typical proteins share the highest sequence identities (48.9%) with the corresponding proteins of rhabdoviruses in genus Vesiculovirus. Phylogenetic analysis of partial L protein sequence indicates that SMRV is close to genus Vesiculovirus. The first 13 nucleotides at the ends of the SMRV genome are absolutely inverse complementarity. The gene junctions between the five genes show conserved polyadenylation signal (CATGA7) and intergenic dinucleotide (CT) followed by putative transcription initiation sequence A(A/G)(C/G)A(A/G/T), which are different from known rhabdoviruses. The entire Ps ORF was cloned and expressed, and used to generate polyclonal antibody in mice. One obvious band could be detected in SMRV-infected carp leucocyte cells (CLCs) by anti-Ps/C serum via Western blot, and the subcellular localization of Ps-GFP fusion protein exhibited cytoplasm distribution as multiple punctuate or doughnut shaped foci of uneven size. [ABSTRACT FROM AUTHOR]

Abstract: Two MAbs (3C7 and 3C9) against flounder Paralichthys olivaceus rhabdovirus (PORV) were generated with hybridoma cell fusion technology and characterized by an indirect enzyme-linked immunosorbent assay, isotype test, Western blot and immunodot analysis and immunofluorescence assay. Isotyping tests demonstrated that both of the two MAbs belonged to IgM subclass. Western blot analysis showed the MAbs reacted with 42, 30, and 22kDa viral proteins, which were localized within the cytoplasm of PORV-infected grass carp ovary (GCO) cells analyzed by indirect immunofluorescences tests. The MAb 3C7 was also selected at random for detecting virus antigens in the inoculated grass carp tissues by immunohistochemistry assay. Flow cytometry tests showed that at the 36h postinfection (0.25PFU/cell), the 23% PORV-infected GCO cells could be distinguished from the uninfected cells with the MAb 3C7. Such MAbs could be useful for diagnosis and potential treatment of viral infection. [Copyright &y& Elsevier]