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Genotyping of the fish rhabdovirus, viral hemorrhagic septicemia, by restriction fragment length polymorphisms

The aim of this study was to develop a standardized molecular assay   that used limited resources and equipment for routine genotyping of   isolates of the fish rhabdovirus, viral haemorrhagic septicaemia   virus (VHSV). Computer generated restriction maps, based on 62   unique full-length (1524 nt) sequences of the VHSV glycoprotein (G)   gene, were used to predict restriction fragment length polymorphism   (RFLP) patterns that were subsequently grouped and compared with a   phylogenetic analysis of the G-gene sequences of the same set of   isolates. Digestion of PCR amplicons from the full-length G-gene by   a set of three restriction enzymes was predicted to accurately   enable the assignment of the VHSV isolates into the four major   genotypes discovered to date. Further sub-typing of the isolates   into the recently described sub-lineages of genotype I was possible   by applying three additional enzymes. Experimental evaluation of the   method consisted of three steps: (i) RT-PCR amplification of the G-  gene of VHSV isolates using purified viral RNA as template, (ii)   digestion of the PCR products with a panel of restriction   endonucleases and (iii) interpretation of the resulting RFLP   profiles. The RFLP analysis was shown to approximate the level of   genetic discrimination obtained by other, more labour-intensive,   molecular techniques such as the ribonuclease protection assay or   sequence analysis. In addition, 37 previously uncharacterised   isolates from diverse sources were assigned to specific genotypes.   While the assay was able to distinguish between marine and   continental isolates of VHSV, the differences did not correlate with   the pathogenicity of the isolates.    (VHS); (VHSv); (viral hemorrhagic septicemia); (viral haemorrhagic   septicaemia); (viral hemmorrhagic septicemia)

In progress
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Scope of Study
Laboratory Investigation
Scale of Phenomena
Impact of Pollutants
Exotic Species
Resource Management
Biology And Life History
Parasites and Diseases

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