Tissue distribution and genetic typing of porcine circoviruses in pigs with naturally occurring congenital tremors.

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 PCV-infected 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.

Viral replication and lesions in BALB/c mice experimentally inoculated with porcine circovirus isolated from a pig with postweaning multisystemic wasting disease.

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.

Infection of pigs by Aujeszky's disease virus via the breath of intranasally inoculated pigs.

Aujeszky's disease is a worldwide problem in the pig industry. In this experiment, four pigs chosen to act as shedder pigs were intranasally infected with Aujeszky's disease virus. Next, on three consecutive days, eight recipient pigs were exposed to the breath of a pair of shedder pigs via a mask-to-mask module. Except for the virtual absence of CNS signs, shedder pigs expressed clinical signs that were similar to pigs infected naturally or experimentally. Only mild respiratory signs occurred in recipient pigs, but all were infected by aerosols of Aujeszky's disease virus as evidenced by seroconversion. The pig is a much more sensitive indicator of airborne virions than our aerosol collection methods. We conclude that the mild respiratory disease acquired by the aerogenous route in recipient pigs is an easily managed model for studying the transmission of airborne respiratory infections and the immune responses to this type of infection.

Ultrastructure of porcine circovirus in persistently infected PK-15 cells.

The ultrastructure of porcine circovirus was examined in persistently infected porcine kidney (PK)-15 cells. Virus-infected PK-15 cells had large numbers of intracytoplasmic inclusions, and a few cells had intranuclear inclusions. Intracytoplasmic inclusions were dispersed throughout the cytoplasm but were most numerous in the perinuclear cytoplasm. Inclusion were of various sizes, round to oval, and electron dense and were of two general types. Inclusions of the first type were small (0.1-0.5 microm diameter), not surrounded by trilaminar membranes, and granular with indistinct margins that blended with surrounding cytoplasm. Some contained 12+/-2-nm-diameter icosahedral virions in loose aggregates or rarely forming paracrystalline arrays. Small inclusions could be sites of viral assembly or maturation. Intracytoplasmic inclusions of the second type were larger (0.5-5.0 microm diameter) and more numerous and had abrupt margins surrounded by trilaminar membranes. They were more electron dense than small inclusions and were heterogeneous, containing various proportions of aggregated virions, electron-dense crystalline lamellae of 5 nm periodicity, and/or whorls of myelinoid membranes. Virions usually formed paracrystalline arrays and occasionally were loosely aggregated. Larger inclusions were typical of autophagolysosomes. Intranuclear inclusions were not membrane bound and were often associated with reticulated nucleoli or aggregates of heterochromatin. Some inclusions were irregularly shaped aggregates of indistinct, circular 10-12-nm-diameter viruslike particles. Others were 0.1-1.0 microm in diameter, round or ring shaped, dense, and finely granular, with sharply demarcated margins.

Infection of pigs by aerosols of Aujeszky's disease virus and their shedding of the virus.

On three consecutive days, six pigs were exposed for 15 minutes to aerosols of Aujeszky's disease virus. The total estimated dose was 4.5 log 10 TCID50. Within each isolation room, a sentinel pig was placed on a deck two feet away from the infected pig. The breath of the pigs that had inhaled the aerosols was collected on days 3, 7 and 13. The respiratory and other clinical signs of the infected pigs resembled those in field cases of Aujeszky's disease. All the pigs infected with Aujeszky's disease virus seroconverted within seven to 10 days after infection. Among the sentinel pigs, clinical signs were minimal and only three seroconverted.

Porcine reproductive and respiratory syndrome virus does not exacerbate Mycoplasma hyopneumoniae infection in young pigs.

The purpose of this study was to determine if Porcine Reproductive and Respiratory Syndrome (PRRS) virus infection altered the severity of acute Mycoplasma hyopneumoniae (MH) infection in young pigs. Twenty five, 3-week-old male pigs were randomly assigned by litter and weight to one of 3 groups. Groups 1 (PRRS only, n = 5) and 2 (PRRS + MH, n = 10) were inoculated intranasally with PRRS virus (IN-5 isolate, 10(5) TCID50) and viremia in all pigs was confirmed by virus isolation from serum 3 days later. Group 3 (MH only, n = 10) was inoculated at the same time with virus free culture media. Seven days after virus inoculation, Groups 2 and 3 were inoculated intratracheally with MH (strain P-5722-3, 10(7) CCU). All pigs were euthanized and necropsied 28 days later, when maximum lesions of mycoplasmosis occurs. Pigs in group 1 did not cough and had no gross lung lesions, but were still viremic at necropsy. MH was isolated from all pigs in groups 2 (avg. log 5.2 +/- 1.3) and 3 (avg. log 5.1 +/- 1.5), but differences were not significant (P = 0.87). Similarly, there were no differences in average days coughing (8.9 +/- 2.8 v 11.2 +/- 4.5, P = 0.17), grossly pneumonic lung (16.5% v 17%, P = 0.91), or microscopic lung lesion scores (10.1 +/- 2.6 v 11.1 +/- 1.9, P = 0.35) between pigs in groups 2 and 3. Under these experimental conditions, PRRS virus infection did not increase the severity of experimental Mycoplasma hyopneumoniae infection in young pigs.

Characterization of infection with endemic porcine reproductive and respiratory syndrome virus in a swine herd.

Infection of 8-week-old pigs with endemic porcine reproductive and respiratory syndrome (PRRS) virus was detected on a farm that had an epidemic of PRRS in 1989. During the 2.5 years since the original epidemic, reproductive performance of the breeding herd had been within acceptable limits, but mortality had periodically exceeded one-fourth of the pigs in the nursery (195 died of 761 weaned, 25.6%). Investigators attempted to determine the age and humoral immune status of pigs infected with endemic PRRS virus on the farm. Serum obtained from 9 groups of 1- to 18-week-old pigs (10 pigs/group) was examined for PRRS virus by virus isolation. Serum was obtained from 8 sows that had farrowed within the preceding 24 hours. Serum from the sows was obtained weekly until litters were weaned at 3 weeks of age. Serum was obtained from 27 newborn pigs (3 to 4 newborn pigs from each of the 8 sows) prior to intake of colostrum and at weekly or biweekly intervals until the pigs were 20 to 21 weeks of age. Isolation of PRRS virus and indirect fluorescent antibody serologic testing were performed on these serum samples. In another study, serum was obtained for serologic testing from 10 sows in each of 6 parity groups. The PRRS virus was isolated from serum of only 3- to 12-week-old pigs. The 8 sows and their 27 pigs were seronegative for PRRS virus during the 3-week lactation period. By 10 weeks of age, 18 of the 27 suckling pigs were still alive and had seroconverted (titers > or = 1:20).(ABSTRACT TRUNCATED AT 250 WORDS)

Mice and rats (laboratory and feral) are not a reservoir for PRRS virus.

Porcine reproductive and respiratory syndrome (PRRS) is caused by an unclassified arterivirus. The syndrome was first reported in the USA in 1987 as epizootics of reproductive failure in sows and respiratory disease in nursery, growing, and fattening pigs. An enzootic form of the disease has now emerged, characterized by interstitial pneumonia and an increased incidence of secondary infections. Because the disease has now become enzootic on many farms, rodents were investigated as a possible reservoir for the infection. Wild rodents from an endemically infected farm were trapped, and virus isolation for PRRS virus (PRRSV) was attempted using porcine primary alveolar macrophage cultures. PRRSV was not isolated from serum and selected pooled tissues (thymus, lung, and spleen) of 14 feral mice and 2 feral rats. Also, transmission experiments were carried out on 3-week-old Balb/c mice and 12-week-old Fischer-344 rats to determine if these species were susceptible to infection. The rodents were inoculated intranasally, orally, and intraperitoneally with a virus proven to transmit PRRS to pigs. Virus isolations from selected pooled tissues (lung, spleen, thymus, and kidney) and from serum were negative, and there were neither gross nor microscopic lesions. Weight gains and white blood cell counts were not significantly different between treated and control groups. These results indicate that rodents are not susceptible to infection with PRRSV and therefore are probably not a reservoir for the disease.

Shedding patterns in swine of virulent and attenuated pseudorabies virus.

Shedding patterns of 2 virulent (P-2208 and KC-152-D) and 1 attenuated (BUK) strains of pseudorabies virus (PRV) were determined in groups of intranasally inoculated feeder pigs. Clinical signs observed following inoculation with the P-2208 and KC-152-D strains included increase in rectal temperatures up to 42.2 C, anorexia, severe respiratory disturbance, and fatal CNS signs in 2 cases. Clinical signs in pigs inoculated with 7.2 X 10(7) median tissue culture infective dose (TCID50) of the BUK strain were limited to depression and a rise in rectal temperatures to 40.5 C for 3 to 4 days. Evaluation of the efficacy of the virus isolation method used showed that the presence on swabs of only 12.5 TCID50 of the P-2208 strain or 8.4 TCID50 of the BUK strain resulted in a 50% chance of virus recovery. Intranasal inoculations with 500 TCID50 of the P-2208 or KC-152-D strain did not result in synchronous infection of the whole group. Intranasal inoculations with 5,000 TCID50 of the KC-152-D strain or 50,000 TCID50 of the P-2208 strain resulted in continuous virus shedding in all pigs between postinoculation days (PID) 4 and 13 (KC-152-D strain) or 14 (P-2208 strain). Some of the pigs in these 2 groups further shed the P-2208 or KC-152-D strain in a continuous or discontinuous pattern up to PID 19 (P-2208 strain) or 20 (KC-152-D strain). The time of onset or the level of virus neutralizing antibody production in individual pigs was not found to have an influence on their shedding patterns.(ABSTRACT TRUNCATED AT 250 WORDS)

Relationship of toxoplasma infections to other diseases in dogs.

Toxoplasma infection as detected by the immunofluorescent antibody (IFA) test was found in 13.2% of 448 dogs examined at the Purdue University Small Animal Clinic. Only titres of 1/64 and above were considered positive, but lesser titers were encountered more frequently. The highest titer observed was 1/1024 in 2 dogs one of which had idiopathic epilepsy and the other had dirofilariasis as well as hip dysplasia. The incidence of positive sera from male and female dogs were 11.7 and 14.5%, respectively. The highest frequency of infection occurred in young dogs up to 4 years of age and the lowest was in older animals. The presence of the high toxoplasmosis IFA titers in the sera of dogs seems to be associated with other disease conditions. Stress such as joint dislocations, fractures, helminth parasitism, and concomitant bacterial and viral infections, etc., was observed in 62.7% of the positive cases. Presence of tumors and renal disease conditions were associated with 28.8 and 5.2%, respectively, of the positive sera.

Possible role of wild mammals in transmission of pseudorabies to swine.

Of 73 wild and domestic mammals tested from an area endemic for pseudorabies in swine, 16 showed natural pseudorabies virus infection, 8 from farms with no pseudorabies history. In transmission experiments with swine and raccoons (Procyon lotor), pseudorabies was not transmitted between raccoons but was transmitted reciprocally between raccoons and swine by contact and when either consumed infected carrion of the other. The fluorescent antibody tissue section test proved valuable in diagnosis of pseudorabies, especially when employed with the virus isolation test.

Detection of pseudorabies virus antibodies: time of appearance by two serotests.

This report describes a time-course comparison of detection of pseudorabies virus antibodies in experimentally infected swine by the virus-neutralization (VN) and indirect hemagglutination tests. Specific antibody titers were observed by the IHA test at 5 days after swine were inoculated, but not until 12 days by the VN test. The predominant immunoglobulin (Ig) class present in the serums of the swine at 5 and 7 days after inoculation was IgM, as determined by sulfhydryl reductions. The VN test lacked sensitivity to early Ig levels (IgM) in these experimentally infected swine, while the indirect hemagglutination test was highly sensitive to these same levels. On the basis of these results, it is possible that the VN test may read early infections as pseudorabies virus negative, due to the low presence of IgG in these samples.

Indirect hemagglutination test for pseudorabies antibody detection in swine.

An indirect hemagglutination test for the detection of antibodies in swine serum specific for pseudorabies virus is described. The indirect hemagglutination test was less time consuming than the standardized virus nertralization test while being highly sensitive and accurate. Serum samples that were toxic to virus neutralization indicator cells were readily tested in the indirect hemagglutination test. The indirect hemagglutination test may also be more sensitive than the virus neutralization test for determining early titers in pseudorabies virus infections. Complete methodology is described.

Fluorescent antibody test in diagnosis of ulcerative enteritis.

A direct fluorescent antibody (FA) test to demonstrate the presence of Clostridium colinum in cryostat sections of liver and intestine from chickens with ulcerative enteritis ("quail disease") was investigated. Both field cases and experimentally infected birds were studied by FA, bacterial isolation of the clostridium, and histopathology. The FA test proved highly specific for Cl. colinum. It was concluded that the described direct FA procedure offers simplicity and speed over bacterial isolation procedures for etiologic diagnosis of ulcerative enteritis.