Negligible risk associated with the movement of processed rainbow trout, Oncorhynchus mykiss (Walbaum), from an infectious haematopoietic necrosis virus (IHNV) endemic area

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Abstract
To assess the risk of transmission of infectious haematopoietic necrosis virus (IHNV) associated with the movement of processed rainbow trout, Oncorhynchus mykiss, from an area where the virus is endemic, 240 freshly eviscerated fish (225–500 g) exhibiting spinal curvature or spinal compression types of deformities were tested for IHNV by virus isolation and polymerase chain reaction (PCR) techniques. Commercially produced rainbow trout, approximately 1‐year‐old, that exhibited spinal deformities were considered to have had a high likelihood of having survived an outbreak of IHN. Serological analysis of fish exhibiting spinal curvature or spinal compression types of deformities for anti‐IHNV antibodies resulted in 71 and 50% of the serum samples, respectively, with detectable neutralization activity suggesting previous infection with IHNV. A portion of the skin and muscle in the area of the deformity was collected, as well as brain tissue from each commercially processed fish. Tissue homogenates were tested for IHNV using the epithelioma papulosum cyprini (EPC) cell line pretreated with polyethylene glycol and the chinook salmon embryo (CHSE‐214) cell line using standard methods. Nested, reverse transcriptase (RT)‐PCR for the detection of IHNV used the central 1231 bp portion of the glycoprotein (G) gene. All brain and skin‐muscle homogenates were negative for IHNV by virus isolation and by nested RT‐PCR. To assess virus clearance experimentally, groups of 100 specific‐pathogen‐free (SPF) rainbow trout (mean weight, 100 g) were either intraperitoneally injected with IHNV or sham infected and held separately in 385 L aquaria in SPF water at 15 °C. In the first study, a total of 33% of the infected fish died. Ten days after the last mortality or 34 days after post‐infection, six fish from each treatment were killed at weekly intervals for 6 weeks. Kidney and brain homogenates from individual fish were tested by virus isolation and nested RT‐PCR using the nucleoprotein (N) gene. All tissue homogenates were negative for IHNV. When this study was repeated, 54% of the infected fish died and beginning 34 days post‐infection, six fish from each treatment were killed at biweekly intervals for 8 weeks, and kidney and brain homogenates from individual fish were tested by virus isolation and quantitative real‐time PCR utilizing primers and fluorescent labelled probes generated for the specific identification of the N and G genes of IHNV. One brain specimen from an IHN survivor obtained 34 days post‐infection was positive by both virus isolation and quantitative PCR; however, all remaining tissue specimens were negative over the 8‐week period. Intraperitoneal injection of supernatants from pooled kidney or brain tissue obtained from control or IHN survivor fish into SPF rainbow trout (mean weight, 10 g) also failed to establish virus infection. A strong humoral response was detected in the IHN survivors in both challenge studies and is suggested as a mechanism responsible for virus clearance. These results provide scientific information that can be used to assess the risk associated with the movement of processed rainbow trout from an IHNV endemic area.