Seroprevalence of Enzootic Teschen Disease in the Wild Boar Population in Ukraine.

Teschen disease is an acute fatal enterovirus encephalomyelitis of pigs, characterized by a range of central nervous system disorders. The cause of porcine enterovirus encephalomyelitis is the picornavirus porcine teschovirus-1 (PTV-1). There are at least 12 disctinct serotypes of PTVs, where PTV-2 to PTV-12 serogroups are associated with other forms of disease (Talfan disease or poliomyelitis suum) or benign enzootic paresis. Combined, PTVs have been found to have a high seroprevalence, up to 65%, in healthy pig populations in Europe. PTVs have also been detected in wild boar, including the divergent PTV-13 serogroup; wild suids may represent a sylvatic reservoir capable of carrying the virus long distances. In Ukraine, Teschen disease is widespread and causes lethal disease in domestic pigs. To understand temporal and geographical distribution of Teschen disease virus (PTV-1) in wild boar in Ukraine (2001-2013), we analyzed seroprevalence of 6840 blood serum samples from hunted suids using a virus microneutralization assay. A total of 1364 samples (19.9%) were seropositive, with average antibody titer ratios 5.89 ± 0.03 log2 (range 5-12 log2). Teschen seroprevalence was temporally and geographically concentrated in the northern and western regions of Ukraine, corresponding to forested regions (polissya) and overlapping with wild boar populations and habitats, suggesting endemicity in wild boar. The virus sporadically emerged in central, southern, and eastern forested regions, suggesting long-distance movement of infected wild suids. Thus, wild boar should be monitored for potential transboundary spread in forested and mountain regions and spillover of PTVs to domestic swine populations.

Porcine Astrovirus Type 3 in Central Nervous System of Swine with Polioencephalomyelitis.

Using next-generation sequencing, we identified and genetically characterized a porcine astrovirus type 3 strain found in tissues from the central nervous system of 1 piglet and 3 sows with neurologic signs and nonsuppurative polioencephalomyelitis. Further studies are needed to understand the potential for cross-species transmission and clinical impact.

Detection of a novel sapelovirus in central nervous tissue of pigs with polioencephalomyelitis in the USA.

An approximately 3,000 finishing swine operation in the United States experienced an outbreak of an atypical neurologic disease in 11-weeks-old pigs with an overall morbidity of 20% and case fatality rate of 30%. The clinical onset and progression of signs in affected pigs varied but included inappetence, compromised ambulation, ataxia, incoordination, mental dullness, paresis, paralysis and decreased response to environmental stimuli. Tissues from affected pigs were submitted for diagnostic investigation. Histopathologic examination of the cerebrum, cerebellum and spinal cord revealed severe lymphoplasmacytic and necrotizing polioencephalomyelitis with multifocal areas of gliosis and neuron satellitosis, suggestive of a neurotropic viral infection. Bacterial pathogens were not isolated by culture of neurologic tissue from affected pigs. Samples tested by polymerase chain reaction (PCR) were negative for pseudorabies virus and atypical porcine pestivirus. Immunohistochemistry for porcine reproductive and respiratory syndrome virus, porcine circovirus and Listeria was negative. Porcine sapelovirus (PSV) was identified in spinal cord by a nested PCR used to detect porcine enterovirus, porcine teschovirus and PSV. Next-generation sequencing of brainstem and spinal cord samples identified PSV and the absence of other or novel pathogens. In addition, Sapelovirus A mRNA was detected in neurons and nerve roots of the spinal cord by in situ hybridization. The PSV is genetically novel with an overall 94% amino acid identity and 86% nucleotide identity to a recently reported sapelovirus from Korea. This is the first case report in the United States associating sapelovirus with severe polioencephalomyelitis in pigs.

Experimental teschovirus encephalomyelitis in gnotobiotic pigs.

A central nervous system (CNS) disorder characterized by non-suppurative encephalomyelitis with neurological signs was induced experimentally in gnotobiotic pigs by intravenous and oral or intranasal inoculation of the porcine teschovirus (PTV) Toyama 2002 strain isolated from breeding pigs in Japan. Lesions consisting of perivascular cuffing of mononuclear cells, focal gliosis, neuronal necrosis and neuronophagia were observed in the brainstem, cerebellum and spinal cord. Non-suppurative ganglionitis in the spinal ganglion and neuritis in the spinal root were also observed. Regardless of the route of inoculation, all pigs infected experimentally with PTV showed a similar distribution of CNS lesions. Histological lesions in the CNS caused by oral or intranasal inoculation of the virus were mild compared with those induced by intravenous infection. Immunohistochemically, the distribution of PTV antigens corresponded closely with the distribution of brain lesions. PTV particles were detected via electron microscopy in the cytoplasm of nerve cells and the endothelial cells of blood vessels in the spinal cord of inoculated pigs. Polymerase chain reaction analysis demonstrated the presence of PTV RNA in the CNS, tonsils and large intestines of 21 of the 22 pigs inoculated. Direct CNS invasion via the blood vessels appears to be a major route of infection for PTV. The gnotobiotic pig provides a useful model for further study of PTV pathogenesis.

Porcine teschovirus polioencephalomyelitis in western Canada.

Beginning in 2002, a small number of pig farms in western Canada began reporting 4-7-week-old pigs with bilateral hind-end paresis or paralysis. Low numbers of pigs were affected, some died, most had to be euthanized, and those that survived had reduced weight gains and neurological deficits. Necropsies revealed no gross lesions, but microscopic lesions consisted of a nonsuppurative polioencephalomyelitis, most severe in the brain stem and spinal cord. The lesions were most consistent with a viral infection. Tests for circovirus, Porcine reproductive and respiratory syndrome virus, coronavirus, Rabies virus, and Pseudorabies virus were negative. Using immunohistochemistry, virus neutralization, fluorescent antibody test, and nested reverse transcription polymerase chain reaction, Porcine teschovirus was identified in tissues from affected individuals. To the authors' knowledge, this is the first report of teschovirus encephalitis in western Canada and the first reported case of polioencephalomyelitis in pigs in Canada, where teschovirus was confirmed as the cause.

Paralytic rabies in swine

Rabies transmitted by vampire bats was diagnosed in pigs with paralysis of the pelvic limbs. Diffuse nonsuppurative encephalomyelitis, affecting mainly the spinal cord, was observed histologically. Despite the various diagnosis of rabies in pigs this is the first report of clinical signs and pathology of rabies transmitted by vampire bats.

Genomic analysis of two porcine encephalomyocarditis virus strains isolated in China.

Two strains of encephalomyocarditis virus (EMCV), designated BJC3 and HB1, were isolated from an aborted fetus and the heart tissue of a dead piglet that had pericardial fluid, respectively. The complete genomic sequences of the two viruses were determined and analyzed. The size of the genomes of BJC3 and HB1 were 7746 and 7735 nucleotides, respectively, including poly(A) tails. Comparative analysis with the genomic sequences of other EMCV strains showed that BJC3 and HB1 shared higher identity (92.5-99.6%) with BEL-2887A/91, EMCV-R and PV21, but lower identity (83.3-84.6%) with EMC-B, EMC-D and D variants, and only 81.0% with Mengo virus. Two amino acid mutations in the leader protein of the two viruses and one amino acid substitution in VP1 of BJC3 were found in comparison to other EMCV strains Phylogenetic analysis based on the amino acid sequences of the entire ORF revealed that the two Chinese isolates BJC3 and HB1 clustered together with the strains BEL-2887/91, EMCV-R and PV21, which belong to the same genetic subgroup as EMCV-30. Our results provide genomic information for EMCV isolated in China.

[Express method for diagnostics of enzootic encephalomyelitis (Teschen disease) in pigs].

Results of creating the home ELISA test-system for diagnostics of enzootic encephalomyelitis (Teschen disease) has been presented. Biological components of diagnosticums has been created and described, reaction stages have been optimized. Sensitivity of the test-system for identification of antibodies to PTV-1 averaged 93%, specificity--100%. In the "sandwich"-variant the strains may be classified as serotype 1.

[Purification of viral drugs for development of diagnostic aids of Teschen disease]. / Ochyshchennia virusnykh preparativ dlia rozrobky diahnostykumiv khvoroby Teshena.

Optimal methods of purification and concentration of teschevirus strain of the pigs of the first serotype "Chernigivsky-2372" have been chosen for development of the set of Teschen pig discase diagnosticums. It has been established that differential centrifugation with following ultracentrifugation in the gradient of saccharose density permits to deprive viruses from host cell proteins, empty virus capsides and defect virus particles. That is the method fit for purification of virus preparations for the solid-phase immunoenzyme analysis.

A wild-type porcine encephalomyocarditis virus containing a short poly(C) tract is pathogenic to mice, pigs, and cynomolgus macaques.

Previous studies using wild-type Encephalomyocarditis virus (EMCV) and Mengo virus, which have long poly(C) tracts (61 to 146 C's) at the 5' nontranslated region of the genome, and variants of these viruses genetically engineered to truncate or substitute the poly(C) tracts have produced conflicting data on the role of the poly(C) tract in the virulence of these viruses. Analysis of the nucleotide sequence of an EMCV strain isolated from an aborted swine fetus (EMCV 30/87) revealed that the virus had a poly(C) tract that was 7- to 10-fold shorter than the poly(C) tracts of other EMCV strains and 4-fold shorter than that of Mengo virus. Subsequently, we investigated the virulence and pathogenesis of this naturally occurring short-poly(C)-tract-containing virus in rodents, pigs, and nonhuman primates. Infection of C57BL/6 mice, pigs, and cynomolgus macaques resulted in similar EMCV 30/87 pathogenesis, with the heart and brain as the primary sites of infections in all three animals, but with different disease phenotypes. Sixteen percent of EMCV 30/87-infected pigs developed acute fatal cardiac failure, whereas the rest of the pigs were overtly asymptomatic for as long as 90 days postinfection (p.i.), despite extensive myocardial and central nervous system (CNS) pathological changes. In contrast, mice infected with >/==" BORDER="0">4 PFU of EMCV 30/87 developed acute encephalitis that resulted in the death of all animals (n = 25) between days 2 and 7 p.i. EMCV 30/87-infected macaques remained overtly asymptomatic for 45 days, despite extensive myocardial and CNS pathological changes and viral persistence in more than 50% of the animals. The short poly(C) tract in EMCV 30/87 (CUC(5)UC(8)) was comparable to that of strain 2887A/91 (C(10)UCUC(3)UC(10)), another recent porcine isolate.

Porcine encephalomyocarditis virus persists in pig myocardium and infects human myocardial cells.

Recent advances toward using pig tissues in human transplantation have made it necessary to determine the risk of transmitting zoonotic viruses from pigs to humans or vice versa. We investigated the suitability of the porcine encephalomyocarditis virus (EMCV) model for such studies by determining its ability to persist in pigs, escape detection by routine serological methods, and infect human cells. Intraperitoneal inoculation of 5-week-old pigs with EMCV-30, a strain isolated from commercial pigs, resulted in acute cellular degeneration, infiltration of lymphocytes, and apoptosis in myocardium in 13 of 15 (86.7%) pigs during the acute phase of disease (3 to 21 days postinfection), followed by less-severe lymphocytic infiltration and apoptosis in 5 of 10 (50%) pigs during the chronic phase of the disease (day 45 to 90 postinfection). In the brain, lymphocytic infiltration, neuronal degeneration, and gliosis were observed in 26 to 33% of pigs in the acute phase of disease whereas perivascular cuffing was the predominant feature during chronic disease. EMCV antigens and RNA were demonstrated in the myocardium and brain during the chronic phase of disease. Analysis of 100 commercial pigs that were negative for EMCV antibodies identified two pig hearts positive for EMCV RNA. Porcine EMCV productively infected primary human cardiomyocytes as demonstrated by immunostaining using a monoclonal antibody specific for EMCV RNA polymerase, which is expressed only in productively infected cells, and by a one-step growth curve that showed production of 100 to 1,000 PFU of virus per cell within 6 h. The findings that porcine EMCV can persist in pig myocardium and can infect human myocardial cells make it an important infectious agent to screen for in pig-to-human cardiac transplants and a good model for xenozoonosis.

[Typing of 17 porcine enterovirus isolates from polio encephalomyelitis cases during the years 1983-1991]. / Typisierung von 17 porzinen Enterovirusisolaten aus Polioenzephalomyelitisfällen der Jahre 1983-1991.

Seventeen strains of porcine enteroviruses (PEV) were isolated from organs of 119 pigs with symptoms of polioencephalomyelitis submitted to the State Veterinary Institute Arnsberg/Westphalia between 1983 and 1991. 15 isolates originated from the central nervous system and 2 from organ suspensions made up of a brain-spleen-pool and a lung-spleen-lymph node-pool, respectively. Isolates were assigned to 7 PEV types which were present at the following frequencies: PEV1: 2x; PEV2: 6x; PEV4: 3x; PEV5: 1x; PEV6: 2x; PEV 12: 1x; PEV 13: 2x. Mortality rates of affected groups exhibited an age-dependent curvilinear relationship suggesting that the PEV involved possessed a rather similar low to medium grade neurovirulence, irrespective of type. Exceptions were 1 herd with 100% mortality at the age of 10-18 weeks from which PEV2 strain 6793/83 was isolated (described earlier) and a second herd with 18% mortality at the age of 14-18 weeks from which PEV types 1, 2 and 4 were recovered. Sensitivity of 5 cell lines for the isolation of PEV was compared. Rates of isolation from organ suspensions which had proved positive in any of the cell lines tested were as follows: PS-EK: 77%; IB-RS-2: 63%; ST: 56%; PK-15: 47%; BHK21 (CT): 10%.

Virus isolation studies in an outbreak of porcine encephalomyelitis.

An outbreak of central nervous system disease affecting young pigs occurred in the fall of 1981 in eastern Ontario. A diagnosis of viral encephalomyelitis was made on pathological grounds and virus isolation studies were subsequently initiated to determine the causative agent. Cultural isolation procedures using several biological systems failed to detect virus in nervous tissues from affected animals. Direct extraction of similar tissues by combined biochemical and biophysical procedures yielded nonenveloped , spherical particles with a diameter of 30 nm and a buoyant density of 1.34 g/mL in CsCl. A tentative diagnosis of enterovirus infection was made on this basis.

[Method of preparation of the specimens for combined immunocytological examinations by light and electron microscopy]. / Sposob prigotovleniia praparatov dlia sovmeshchennogo immunotsitologicheskogo issledovaniia v svetovom i elektronnom mikroskopakh.

A method for making preparations for combined immunocytological examinations in light and electron microscopes is described. An infected cell culture grown on slides (a piglet kidney cell culture infected with Teschen virus) was fixed with 2.5% glutaraldehyde in phosphate buffer, pH 7.2-7.4 for 10-15 min and incubated with a conjugate (peroxidase-labeled immune serum to the causative agent of the infection under study). After postfixation, cytochemical reaction for peroxidase was performed, then the material was fixed with 2% OsO4 solution in phosphate buffer, pH 7,2-7,4, dehydrated in an ascending series of alcohols and gradually impregnated with polymerising resins (epon, araldit, etc.). The cells were embedded in foil baths, and after polymerization and separation of the slide and foil preparations with good light microscopy properties. In the light microscope, the desired parts of cells or a cell were selected, removed, cut in an ultratome and examined in electron microscope.