Progress 08/01/07 to 07/31/09

Outputs
OUTPUTS: PROJECT OBJECTIVES: 1. Develop a laboratory model of infection for channel catfish and rainbow trout. 2. Determine the sensitivity of detection of VHSV with qRT-PCR. 3. Provide reference laboratory services for the fish health community. BENEFITS: Viral Hemorrhagic Septicemia Virus (VHSV) is considered to be one of the most important viral pathogens of finfish worldwide and is listed as a Reportable Animal Pathogen by the OIE (World Animal Health Organization). The first reported isolation of VHSV in the Great Lakes Basin occurred in 2005 and was associated with a massive fish kill among freshwater drum and other species that occurred in the Canadian portion of Lake Ontario. During 2006, the known host and geographic range of the virus expanded explosively, and VHSV was isolated from fish in Lake Huron, Lake St. Clair, Lake Erie, Lake Ontario and the Saint Lawrence River where it caused significant mortality in a variety of important sport fish. This emerging strain of VHSV appears to have an exceptionally broad host range as the virus has infected several additional species of freshwater fish in the region; however, the magnitude of mortality in some of the wild populations has been difficult to assess or has not been reported. Fisheries managers and representatives of the aquaculture industry in both the US and Canada are extremely concerned about the spread of this exceptionally virulent virus into new populations of native freshwater fish or the introduction of VHSV into the private sector aquaculture industry. The emergence of VHSV in the freshwater environment of North America has brought with it a number of high priority research and technical assistance needs to help protect US aquaculture from the potential impacts of this virus. We addressed several of the most immediate research needs under this project. Those needs included determining the relative susceptibility of channel catfish and rainbow trout to infection with VHSV Genotype IVb. These two species represent two of the most important aquacultured fish species in the United States. We also developed a quantitative RT-PCR for the detection of VHSV IVb. This assay constitutes an invaluable tool for surveillance efforts that can provide accurate and rapid results regarding the absence of VSHV IVb. We also developed a laboratory infrastructure in which isolates of VHSV IVb will be characterized in a consistent manner. Such information will be invaluable in following the epidemiology of this emerging pathogen of fish. PARTICIPANTS: Dr. Paul R. Bowser, Professor of Aquatic Animal Medicine Department of Microbiology and Immunology College of Veterinary Medicine, Cornell University, Ithaca, New York 14853-6401 Phone: 607-253-4029 ; FAX: 607-253-3384; e-mail: prb4@cornell.edu, Dr. James W. Casey, Associate Professor of Virology Department of Microbiology and Immunology College of Veterinary Medicine, Cornell University, Ithaca, New York 14853-6401 Phone: 607-253-3579; FAX: 607-253-3384; e-mail: jwc3@cornell.edu, Dr. James R. Winton, Chief, Fish Health Section Western Fisheries Research Center 6505 NE 65th Street Seattle, WA 98115-5016 Phone: 1-206-526-6587; FAX: 1-206-526-6654; e-mail: jim_winton@usgs.gov, Dr. Gael Kurath Fish Health Section Western Fisheries Research Center 6505 NE 65th St. Seattle, WA 98115 USA Phone: 206-526-6583; FAX: 206-526-6654; e-mail: gael_kurath@usgs.gov TARGET AUDIENCES: 30 presentations were given to various stock holder groups. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
PROGRESS AND PRINCIPAL ACCOMPLISHMENTS: 1. Develop a laboratory model of infection for channel catfish and rainbow trout. Infection trials were conducted with channel catfish and rainbow trout at both 15C and 10C. The fish were injected with 106 plaque forming units (PFU) of VHSV Genotype IVb (isolate MI03). The trials were conducted for 28 days. Fish were sampled at scheduled collection times over the 28-day trial and liver, kidneys, spleen and heart were prepared for inoculation onto EPC cells as well as prepared for assay by the qRT-PCR. In all cases, little to no signs of disease were observed. However, virus was detected by cell culture and qRT-PCR in fish at multiple time points during the trials. Under the conditions of our infection trials, channel catfish and rainbow trout appear to have a relatively low susceptibility to infection with VHSV IVb. 2. Determine the sensitivity of detection of VHSV with qRT-PCR. A quantitative RT-PCR (qRT-PCR) was developed for VHSV IVb, based on the N gene of the virus. To date this assay has been applied to over 10,000 samples generated through this study as well as other efforts focused on VHSV. We consider this test to be capable of detecting the virus when it is present in low quantities. In our hands we feel confident that we can detect the virus when it is present at levels between 10 and 100 genome copies. This level of detection can be compared to that of cell culture requiring approximately 10,000 genome copies to generate a VHSV-positive cell culture isolation. We have submitted a publication to the peer reviewed literature that describes the assay. 3. Provide reference laboratory services for the fish health community. A database of isolates of VHSV IVb has been established at the USGS Western Fisheries Research Center. Fish health laboratories, including ours, have forwarded isolates to the USGS WFRC for partial sequencing. To date the Aquatic Animal Health Program at Cornell has forwarded 44 VHSV IVb isolates to the WFRC for this effort. The total collection currently includes over 80 isolates of the Great Lakes VHSV IVb. 4. There were 30 Paper Presentations given regarding this project.

Publications

• Bowser, P. R. 2009. Fish Diseases: Viral Hemorrhagic Septicemia (VHS). Northeast Regional Aquaculture Center, USDA. Fact Sheet 201-2009.