The purpose of this study was to determine the effect of formalin fixation on the immunohistochemical detection of porcine reproductive and respiratory syndrome (PRRS) viral antigen in lungs of experimentally and naturally infected pigs. In separate trials, five 24-day-old pigs and six 10-day-old pigs were housed as separate groups in isolation and inoculated intranasally with 10(5.5) TCID50 of an isolate of PRRS virus (PRRSV; P129). The older and younger pigs were euthanatized at 7 and 10 days post inoculation (dpi), respectively. At necropsy, all pigs had gross and microscopic lung lesions typical of PRRS, and PRRSV was isolated from all pigs. To insure uniform fixation, lungs from each pig were cut into 1-cm-thick slices and immersed into 10% neutral-buffered formalin. After fixation in formalin for 8 hours or 1, 2, 3, 5, 6, 8, 10, and 15 days, 3 lung sections from some or all pigs were processed for histological examination using routine methods. Immunohistochemical staining for PRRSV antigen was positive at the following times (days unless otherwise stated) after fixation (percentage of pigs staining positive for PRRSV in parentheses): 8 hours (100); 1 (100); 2 (100); 3 (80); 5 (33); and 6, 8, 10, and 15 (0-all negative). To further evaluate the effects of formalin fixation on PRRSV immunodetection, 31 field cases of PRRS were selected for immunohistochemistry (IHC). Over a 3-month period, submitted cases were selected from the Purdue University Animal Disease Diagnostic Laboratory, W. Lafayette, Indiana, for IHC if 1) the clinical history included respiratory disease, 2) PRRSV was isolated from lung and/or serum from the submitted pigs or tissues, 3) at least 1 section of lung fixed in 10% neutral-buffered formalin was submitted for IHC, and 4) the duration of fixation could be accurately determined from the case history. Of the 31 PRRSV-infected pig cases meeting the selection criteria, 23 were fixed in formalin for 4 days or less. Twenty-one of these 23 (91%) were positive by IHC. Two of 8 cases fixed for greater than 4 days (25%) were positive by IHC. In practical terms, 1-day shipping of fixed samples to a laboratory followed by routine tissue processing within a laboratory should not adversely affect immunohistochemical detection of PRRS viral antigen. But a delay in shipping or processing of more than 2 days could reduce or prevent the detection of PRRS viral antigen by IHC.

In order to accurately diagnose bacterial kidney disease caused by Renibacterium salmoninarum in steelhead trout, kidney tissue from experimentally infected fish was evaluated using a commercially available enzyme-linked immunosorbent assay (ELISA) test kit, fluorescent antibody (FA) testing, bacteriologic culture, and histopathology. Seventy-five steelhead trout were randomly assigned to 1 of 4 groups and intraperitoneally inoculated with 0.15 ml saline ( n = 20), 1 × 1010 organisms/ml ( n = 18), 1 × 108 organisms/ml ( n = 18), or 1 × 106 organisms/ml ( n = 19) of R. salmoninarum. ELISA, FA, and bacteriologic culture were positive for R. salmoninarum from the kidney tissue of the 2 groups infected with the highest doses. Although the ELISA and FA tests were accurate when compared to the bacteriologic culture from the 2 groups infected with the higher doses of the organism, they were less sensitive at the lowest level of inoculum. Histopathology was not specific for this disease; however, all infected fish had a marked proliferative histiocytic interstitial nephritis, characterized by marked expansion of the renal hematopoietic tissue by histiocytes without tissue necrosis. Other microscopic findings included splenitis and myositis (at the injection site) of some fish.

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