Detection of Viral Hemorrhagic Septicemia Virus by Quantitative Reverse Transcription Polymerase Chain Reaction from Two Fish Species at Two Sites in Lake Superior

Abstract

Viral hemorrhagic septicemia virus (VHSV) was first detected in the Laurentian Great Lakes in 2005 during a mortality event in the Bay of Quinte, Lake Ontario. Subsequent analysis of archived samples determined that the first known isolation of VHSV in the Laurentian Great Lakes was from a muskellunge Esox masquinongy collected in Lake St. Clair in 2003. By the end of 2008, mortality events and viral isolations had occurred in all of the Laurentian Great Lakes except Lake Superior. In 2009, a focused disease surveillance program was designed to determine whether VHSV was also present in Lake Superior. In this survey, 874 fish from 7 sites along the U.S. shoreline of Lake Superior were collected during June 2009. Collections were focused on nearshore species known to be susceptible to VHSV. All fish were dissected individually by using aseptic techniques and were tested for the presence of VHSV genetic material by use of a quantitative reverse transcription (qRT) polymerase chain reaction (PCR) targeting the viral nucleoprotein gene. Seventeen fish from two host species at two different sites tested positive at low levels for VHSV. All attempts to isolate virus in cell culture were unsuccessful. However, the presence of viral RNA was confirmed independently in five fish by using a nested PCR that targeted the glycoprotein (G) gene. Partial G gene sequences obtained from three fish were identical to the corresponding sequence from the original 2003 VHSV isolate (MI03) from muskellunge. These detections represent the earliest evidence for the presence of VHSV in Lake Superior and illustrate the utility of the highly sensitive qRT-PCR assay for disease surveillance in aquatic animals.

Role of Viral Hemorrhagic Septicemia Virus Matrix (M) Protein in Suppressing Host Transcription

“…VHSV-IVb caused massive die-offs among many freshwater species during the next decade and continues to pose a potential threat to both fish farming and the sport fishing industry in the Great Lakes watershed (28)(29)(30). VHSV-IVb has been isolated from at least 31 fish species, including muskellunge, yellow perch, and walleye (28), and has been detected in all five of the Laurentian Great Lakes (31,32). However, unlike the European Ia sublineage, IVb has low pathogenicity for rainbow trout (33).…”

Detection and surveillance of viral hemorrhagic septicemia virus using real-time RT-PCR. I. Initial comparison of four protocols

“…Clearly the Hope assay is not suitable for surveillance or diagnostic purposes when attempting to detect all genotypes, as it is not likely that this assay will detect VHSV genotypes I and II and was not considered further. However, this assay has clearly demonstrated to be a useful tool for surveillance of VHSV IVb in the Great Lakes (Bain et al 2010, Cornwell et al 2011. Phase I data established that while the Garver assay had efficiency estimates closer to 100% than the Phelps assay, the Phelps assay performed better in detecting all genotype representatives.…”

“…VHSV can become endemic in fish populations, with some individual fish serving as carriers of the virus and periodically shedding the virus, which causes repeat episodes of disease in naïve fish populations (Hershberger et al 2010. Since its emergence in the Laurentian Great Lakes of North America, the novel sublineage of VHSV genotype IV has caused several large-scale die-offs of wild fish and spread into the 5 Great Lakes, a number of inland lakes, and several streams (Elsayed et al 2006, Gagné et al 2007, Groocock et al 2007, Lumsden et al 2007, Cornwell et al 2011. VHSV is now endemic in the Great Lakes and represents a serious risk to farmed fish populations within the Great Lakes watershed as well as a risk for dissemination of the virus to other watersheds within or outside the USA by movement of fish (reviewed by Faisal et al 2012).…”

Low prevalence of VHSV detected in round goby collected in offshore regions of Lake Ontario

“…The first reports of VHSV type IVb in the Great Lakes occurred in muskellunge (Esox masquinongy) in Lake St. Clair during 2003 and 2005 and in freshwater drum (Aplodinotus grunniens) in the Bay of Quinte during 2005 (Elsayed et al, 2006;Lumsden et al, 2007). Since these first detections, VHSV has been detected throughout the Great Lakes (Bain et al, 2010;Cornwell et al, 2011;Frattini et al, 2011).…”

“…During that period, VHSV was only detected in regions where it was previously known to occur. Additional surveillance efforts have detected VHSV throughout the Great Lakes in the absence of concurrent mortality events and led to the earliest evidence for VHSV in Lake Superior by molecular methods (Bain et al, 2010;Cornwell et al, 2011. These surveillance efforts have expanded the known geographic and host range of VHSV as well as identified risk factors for VHSV infection in near-shore populations.…”