Characterization of mucosal immunity in rainbow trout Oncorhynchus mykiss challenged with infectious hematopoietic necrosis virus: Identification of antiviral activity

Cutaneous and lower gastrointestinal (GI) tract mucus of rainbow trout Oncorhynchus mykiss were evaluated for immunological responses following exposure to infectious hematopoietic necrosis virus (IHNV). Juvenile rainbow trout (mean weight 5.4 g) were infected with IHNV (isolate 220-90) by waterborne exposure to 10(4) plaque forming units (pfu) ml(-1) or by an injection challenge of 10(4) pfu fish(-1). Cutaneous mucus and lower GI tract washings were analyzed for virus neutralizing activity (antiviral activity) using a complement-dependent neutralization procedure. Antiviral activity was primarily detected in the GI tract and was highest in mock infected fish (not exposed to virus). This activity decreased rapidly over time in fish challenged by the injection route. Specific antibodies were not detected in cutaneous mucus or GI tract washings by ELISA. Virus was isolated in external and GI tract mucus from both waterborne and injection challenged fish, and showed significantly higher titers in fish challenged by the injection route. Histological examination of fish exposed to IHNV showed a moderate inflammatory response in the mucosal epithelial layer of the skin with minor focal areas of cellular necrosis in severely infected fish. When the study was repeated, in addition to cutaneous mucus and GI tract washings, serum was monitored for antibody production in juvenile rainbow trout (mean weight 2.4 g) out to 28 days post-exposure (dpe). Antiviral activity was confirmed in the GI mucus, and virus neutralizing antibodies were detected in the serum at 21 and 28 dpe in IHNV challenged fish. Antibodies in skin and GI mucus were not detected by ELISA. Virus antigen was visualized by immunohistochemistry in the kidney, spleen, liver, heart, pancreatic tissue, and smooth muscle of the stomach wall. Results from these experiments reveal innate mechanisms of viral resistance which may be important as a first line of defense against IHNV.