Abstract. Eight-week-old BALB/c mice were either sham inoculated (control mice) or were inoculated intraperitoneally (IP) and intranasally (IN) with a single (sPCV mice) or multiple (mPCV mice) doses of porcine circovirus 2 (PCV2). Four control mice and 4 sPCV mice were sacrificed 7, 14, 28, and 42 days postinoculation (PI). All 4 mPCV mice were sacrificed 42 days PI. In addition, 7-day and 14-day pregnant BALB/c mice were either sham inoculated (control mice) or were inoculated IP and IN with a single dose of PCV2. Newborn mice were euthanatized 1, 8, and 15 days after birth. Necropsies were performed on all euthanatized mice and tissues were collected for histopathology, electron microscopy, in situ hybridization, and polymerase chain reaction (PCR). PCV2 replicated in 8-week-old BALB/c mice that were inoculated with PCV2 and caused fetal infection when inoculated into pregnant BALB/c mice at 7 days and 14 days of gestation. PCV was detected by in situ hybridization and PCR in sPCV mice on days 7, 14, 28, and 42 PI; in mPCV mice on day 42 PI; and in newborn mice from mothers inoculated with PCV at 7 days and 14 days of gestation at 1, 8, and 15 days after birth, but not in control mice. No clinical signs or gross lesions were found in sPCV or mPCV mice during the study. Microscopic lesions in sPCV mice and mPCV mice were characterized by expansion of germinal centers in lymphoid organs with large numbers of histiocytic cells and lymphoblasts, apoptosis of histiocytic cells in germinal centers, and mild lymphoid depletion of the paracortex. PCV nucleic acid was detected in the nuclei and cytoplasm of histiocytes and apoptotic cells in germinal centers in lymphoid tissues as well as in the nuclei of hepatocytes in the liver, in the nuclei of renal tubular epithelial cells, and in the cytoplasm of single lymphocytes in the thymus. Congenitally infected mice only had PCV nucleic acid detected in putative Kupffer cells in livers.
Abstract. Congenital tremors (CT) type A2 is associated with porcine circovirus (PCV) and deficient and abnormal myelin. The aim of this study was to determine the tissue distribution and genetic type of PCV in 1-2-day-old pigs with naturally occurring CT type A2 using in situ hybridization, polymerase chain reaction (PCR), and indirect fluorescent antibody tests on frozen tissue sections. CT-affected and clinically normal pigs were selected from 4 farms in the midwestern USA that were undergoing outbreaks of CT type A2. All CT and most normal pigs were infected with PCV. PCV was widely distributed in tissues of infected pigs and was most common in tissues of the central nervous system and liver. In all infected pigs, there were more PCVinfected cells in brain and spinal cord than in nonneural tissues. CT pigs had many more PCV-infected cells in the brain and spinal cord than did clinically normal pigs because of a more diffuse distribution and a larger proportion of infected cells. The cells most commonly infected with PCV in brain and spinal cord were large neurons. In nonneural tissues, macrophages were the most frequent cell type infected. PCR analysis demonstrated only PCV type 2 and not PCV type 1 in all PCV-infected pigs on all 4 farms.
Neonatal diarrhea induced by bovine group A rotavirus causes significant economic loss in the dairy and beef industry due to increased morbidity and mortality, treatment costs, and reduced growth rates. The objective of this study was to evaluate a human group A rotavirus assay (ImmunoCardSTAT Rotavirus [ICS-RV]) as an on-site diagnostic test for bovine rotavirus. When used with a collection of bovine diarrhea samples submitted to the Virology Section of the Diagnostic Center for Population and Animal Health at Michigan State University and compared to a bovine group A rotavirus-specific reverse transcription-PCR (RT-PCR), the ICS-RV assay had a sensitivity and specificity of 87.0 and 93.6%, respectively. A commercially available group A rotavirus enzyme-linked immunosorbent assay (ELISA) (Pathfinder; Sanofi Diagnostics, Redmond, Wash.), when used with the same fecal sample collection and compared to the same RT-PCR, had a sensitivity and specificity of 78.3 and 67.7%, respectively. Subsequently, the ICS-RV assay, RT-PCR, and a different commercially available group A rotavirus ELISA (Rotaclone; Meridian Diagnostics, Cincinnati, Ohio) were used to evaluate fecal samples collected from neonatal calves experimentally infected with bovine rotavirus. When diarrheic fecal samples that were positive for bovine rotavirus by RT-PCR were evaluated, the ICS-RV assay and the Rotaclone assay detected bovine rotavirus 85 and 95% of the time, respectively. Based on these studies, the ICS-RV assay appears to be an excellent test for detecting group A bovine rotaviruses. This assay may be useful as an on-site diagnostic test for veterinarians as an aid in the management of bovine neonatal diarrhea.
Porcine reproductive and respiratory syndrome (PRRS), formerly called swine infertility and respiratory syndrome and mystery swine disease, is an important viral disease of swine causing late-term reproductive failure in sows and gilts and interstitial pneumonia in pigs. 3,6-9 PRRS virus is an enveloped RNA virus approximately 62 nm in diameter, which is relatively sensitive to environmental conditions. The virus is sensitive to lipid solvents and is completely inactivated in 45 minutes at 56 C. Virus infectivity titers can be reduced by 90% at a pH < 5 or > 7. 1 Currently, definitive diagnosis of this disease depends on isolation of PRRS virus from clinically ill animals. Harsh environmental conditions during shipment of samples to a diagnostic laboratory could result in inactivation of the virus and preclude virus isolation.The purpose of this study was to evaluate the stability of PRRS virus in serum and tissues under the typical time and temperature conditions of transportation of samples from the field to a laboratory.Five 7-day-old piglets were placed on nursery decks in an animal isolation room at Purdue University School of Veterinary Medicine and fed a liquid milk replacer a for the duration of the study. Prior to inoculation, all piglets were seronegative by an indirect fluorescent antibody test and were negative for PRRS virus by virus isolation from serum. 4 At 8 days of age, all pigs were intranasally inoculated with 10 5 TCID 50 of an Indiana isolate of PRRS virus (IND-5).Seven days postinoculation (DPI), pigs were euthanized, and serum and tissues were collected. From each piglet, the right lung, spleen, and thymus were removed and diced into approximately 100-mm 3 cubes. Cubes of each organ sample were thoroughly mixed and separated into equivalent subsamples. These subsamples were individually placed into wells of 6-well cell culture cluster plates, and plates were wrapped in plastic to prevent desiccation during storage. Serum from each pig was aliquoted into 2-ml samples in plastic serum tubes. Tissues and serum samples were stored at -20, 4, or 25 C. At necropsy (time 0) and at 24, 48, and 72 hours following necropsy, tissues and serum from each pig were removed from each storage temperature for virus isolation. At time 0 only, virus was titrated in each tissue.Tissue samples were processed for inoculation by grinding them to a 10% suspension in phosphate-buffered saline solution supplemented with penicillin, streptomycin, and gentamycin using Tenbroeck tissue grinders. At time 0, serial lo-fold dilutions of serum and each tissue suspension were processed for virus isolation to determine initial virus titers. At other sample times, only the original 10% tissue suspension or undiluted serum were processed for virus isolation. After cold centrifugation at 3,000 rpm for 15 minutes, supernatant fluids were filtered through a 0.45-µm filter, and From the Animal Disease Diagnostic Laboratory, Purdue University, 1175 ADDL, West Lafayette, IN 47907-1175.
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