Virulence Comparison of Three Buhl-Subtype Isolates of Infectious Pancreatic Necrosis Virus in Brook Trout Fry

Abstract

Infectious pancreatic necrosis virus (IPNV) is an important aquatic pathogen that can cause high mortality in populations of young salmonids. To determine the molecular basis of virulence, the fry of brook trout Salvelinus fontinalis were experimentally infected with three different A1-serotype, Buhl-subtype isolates of IPNV. The three isolates were selected on the basis of results of previously completed virulence assays, which indicated that the isolates had substantially differing virulence levels. To confirm this, mortalities from each treatment were recorded for the duration of the experiment (62 d), along with observation of any clinical disease signs. Mortalities began on day 5 postexposure, peaked on day 7, and then rapidly decreased for all three isolates tested. Diseased fry exhibited whirling, ascites, abdominal hemorrhaging, and prostration on the bottom of the tank. Daily virus titers from live fish were determined for 10 d postexposure (dpe), as well as at 28 and 62 dpe. Viral titers were correlated with fish weight to determine statistical significance. Fish weight was found to correlate negatively to virus titer for the two least-virulent isolates. Initial sequencing results demonstrated sequence homology for the viral capsid protein VP3, whereas slight differences among isolates were observed for the viral capsid protein VP2. The VP2 region was sequenced for each isolate at three times—before being introduced into the fish, during the epizootic, and 2 months after exposure—to determine whether major changes existed in the VP2 region that might account for the differences in virulence. Data indicate that two amino acid differences in the VP2 region exist, at residues 217 and 286, distinguishing the least-virulent isolates and the most-virulent isolate. These amino acid differences might account for the disparity in expressed virulence.