caa a22 4500 445318643 CHVBK 20180317142656.0 cr unu---uuuuu 170323e20110801xx s 000 0 eng 10.1007/s10126-010-9329-4 doi (NATIONALLICENCE)springer-10.1007/s10126-010-9329-4 A Reverse Genetics System for the Great Lakes Strain of Viral Hemorrhagic Septicemia Virus: the NV Gene is Required for Pathogenicity [Elektronische Daten] [Arun Ammayappan, Gael Kurath, Tarin Thompson, Vikram Vakharia] Viral hemorrhagic septicemia virus (VHSV), belonging to the genus Novirhabdovirus in the family of Rhabdoviridae, causes a highly contagious disease of fresh and saltwater fish worldwide. Recently, a novel genotype of VHSV, designated IVb, has invaded the Great Lakes in North America, causing large-scale epidemics in wild fish. An efficient reverse genetics system was developed to generate a recombinant VHSV of genotype IVb from cloned cDNA. The recombinant VHSV (rVHSV) was comparable to the parental wild-type strain both in vitro and in vivo, causing high mortality in yellow perch (Perca flavescens). A modified recombinant VHSV was generated in which the NV gene was substituted with an enhanced green fluorescent protein gene (rVHSV-ΔNV-EGFP), and another recombinant was made by inserting the EGFP gene into the full-length viral clone between the P and M genes (rVHSV-EGFP). The in vitro replication kinetics of rVHSV-EGFP was similar to rVHSV; however, the rVHSV-ΔNV-EGFP grew 2 logs lower. In yellow perch challenges, wtVHSV and rVHSV induced 82-100% cumulative per cent mortality (CPM), respectively, whereas rVHSV-EGFP produced 62% CPM and rVHSV-ΔNV-EGFP caused only 15% CPM. No reversion of mutation was detected in the recovered viruses and the recombinant viruses stably maintained the foreign gene after several passages. These results indicate that the NV gene of VHSV is not essential for viral replication in vitro and in vivo, but it plays an important role in viral replication efficiency and pathogenicity. This system will facilitate studies of VHSV replication, virulence, and production of viral vectored vaccines. Springer Science+Business Media, LLC, 2010 Viral hemorrhagic septicemia virus nationallicence Reverse genetics nationallicence Synthetic biology nationallicence Pathogenesis nationallicence Yellow perch nationallicence Non-virion gene nationallicence Ammayappan Arun Center of Marine Biotechnology, University of Maryland Biotechnology Institute, Baltimore, 701 East Pratt Street, 21202-3101, Baltimore, MD, USA aut Kurath Gael US Geological Survey, Western Fisheries Research Center, 98115, Seattle, WA, USA aut Thompson Tarin US Geological Survey, Western Fisheries Research Center, 98115, Seattle, WA, USA aut Vakharia Vikram Center of Marine Biotechnology, University of Maryland Biotechnology Institute, Baltimore, 701 East Pratt Street, 21202-3101, Baltimore, MD, USA aut Marine Biotechnology Springer-Verlag 13/4(2011-08-01), 672-683 1436-2228 13:4<672 2011 13 10126 https://doi.org/10.1007/s10126-010-9329-4 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s10126-010-9329-4 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Ammayappan Arun Center of Marine Biotechnology, University of Maryland Biotechnology Institute, Baltimore, 701 East Pratt Street, 21202-3101, Baltimore, MD, USA aut NATIONALLICENCE 700 1- Kurath Gael US Geological Survey, Western Fisheries Research Center, 98115, Seattle, WA, USA aut NATIONALLICENCE 700 1- Thompson Tarin US Geological Survey, Western Fisheries Research Center, 98115, Seattle, WA, USA aut NATIONALLICENCE 700 1- Vakharia Vikram Center of Marine Biotechnology, University of Maryland Biotechnology Institute, Baltimore, 701 East Pratt Street, 21202-3101, Baltimore, MD, USA aut NATIONALLICENCE 773 0- Marine Biotechnology Springer-Verlag 13/4(2011-08-01), 672-683 1436-2228 13:4<672 2011 13 10126 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer