RESUMO
An ovine testis cell line (OA3.Ts) was evaluated and compared with primary lamb kidney (LK) cells for its utility in capripoxvirus propagation and titration. A comparison of OA3.Ts cell growth kinetics and morphology at low (<33) and high (34-36) passage levels indicated a difference in both characteristics. However, viral titers determined in low and high passage OA3.Ts cells were comparable with those obtained using LK cells. Capripoxvirus infection of OA3.Ts and LK cells resulted in a similar cytopathic effect, which allowed for the detection of discrete viral plaques following immunostaining with capripoxvirus-specific antiserum.
Assuntos
Capripoxvirus/fisiologia , Testículo/citologia , Ensaio de Placa Viral/veterinária , Cultura de Vírus/veterinária , Animais , Linhagem Celular , Rim/citologia , Masculino , Ovinos , Coloração e Rotulagem , Ensaio de Placa Viral/métodos , Cultura de Vírus/métodosRESUMO
Sheeppox and goatpox are caused by viruses that are members of the genus Capripoxvirus, and globally result in significant production losses. To improve the understanding of disease pathogenesis and evaluate host species preferences, sheep and goats were inoculated either with a capripoxvirus isolate from Yemen or from a recent outbreak in Vietnam. Blood, swabs and tissues were collected at various time points following experimental challenge and assessed for viral DNA content using real-time PCR and infectivity using virus isolation. The Yemen isolate was considerably more pathogenic in goats with 100 % mortality and morbidity compared with sheep with 0 % mortality and 100 % morbidity. The Vietnam isolate was also more pathogenic in goats with 100 % morbidity and an estimated 33 % mortality rate compared with mild morbidity and a 0 % mortality rate in sheep. Higher viral titres were observed in nasal, oral and conjunctival swabs from goats inoculated with either the Yemen or Vietnam isolate compared with those collected from sheep. Although the highest viral titres were detected in primary and secondary skin lesions in sheep and goats, the severity of clinical disease observed in each species varied according to the inoculum used. Whereas both the Yemen and Vietnam isolates clearly caused more severe disease in goats, the Yemen isolate was also moderately pathogenic in sheep. The Vietnam isolate, in contrast, caused only very mild disease in sheep. Limited DNA sequencing revealed ORF 074 of the Vietnam isolate to be identical to that of several goatpox virus isolates from China, suggesting a possible Chinese origin.
Assuntos
Capripoxvirus/isolamento & purificação , Capripoxvirus/patogenicidade , Doenças das Cabras/virologia , Infecções por Poxviridae/veterinária , Doenças dos Ovinos/virologia , Estruturas Animais/virologia , Animais , Sangue/virologia , Capripoxvirus/genética , DNA Viral/química , DNA Viral/genética , DNA Viral/isolamento & purificação , Cabras , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/métodos , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Ovinos , Análise de Sobrevida , Vietnã , IêmenRESUMO
Viruses in the genus Capripoxvirus, family Poxviridae, cause sheeppox, goatpox and lumpy skin disease, which are the most serious poxvirus diseases of production animals. Despite the considerable threat that these viruses pose to livestock production and global trade in sheep, goats, cattle and their products, convenient and effective serodiagnostic tools are not readily available. To develop a more effective antibody detection capability, selected open reading frames from capripoxvirus DNA were amplified and expressed in Escherichia coli as His-tagged fusion proteins. By screening 42 candidate antigens, two sheeppox virus virion core proteins that were expressed efficiently, purified readily using affinity chromatography and reactive against capripoxvirus immune sera in an indirect enzyme-linked immunosorbent assay (ELISA) were identified. The ELISA performed favourably when sera from sheep and goats infected experimentally with virulent capripoxvirus isolates were tested, with sensitivity and diagnostic specificity ranging between 95 and 97%, but it was unable to detect antibodies reliably in vaccinated sheep or goats. Furthermore, no cross-reactivity with antibodies against orf virus was detected. This assay offers the prospect of a convenient and standardised ELISA-based serodiagnostic test, with no requirement for infectious reagents, that is well suited to high-throughput capripoxvirus surveillance on a flock or herd basis.
Assuntos
Anticorpos Antivirais/sangue , Antígenos Virais , Capripoxvirus/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Doenças das Cabras/diagnóstico , Infecções por Poxviridae/veterinária , Doenças dos Ovinos/diagnóstico , Animais , Antígenos Virais/genética , Capripoxvirus/genética , Clonagem Molecular , Escherichia coli/genética , Doenças das Cabras/imunologia , Cabras , Infecções por Poxviridae/diagnóstico , Proteínas Recombinantes/genética , Sensibilidade e Especificidade , Ovinos , Doenças dos Ovinos/imunologiaRESUMO
Sheeppox virus and goatpox virus cause systemic disease in sheep and goats that is often associated with high morbidity and high mortality. To increase understanding of the pathogenesis of these diseases, we undertook quantitative time-course studies in sheep and goats following intradermal inoculation of Nigerian sheeppox virus or Indian goatpox virus in their respective homologous hosts. Viremia, determined by virus isolation and real-time PCR, cleared within 2 to 3 weeks post inoculation. Peak shedding of viral DNA and infectious virus in nasal, conjunctival and oral secretions occurred between 10 and 14 days post inoculation, and persisted at low levels for up to an additional 3 to 6 weeks. Although gross lesions developed in multiple organ systems, highest viral titers were detected in skin and in discrete sites within oronasal tissues and gastrointestinal tract. The temporal distribution of infectious virus and viral DNA in tissues suggests an underlying pathogenesis that is similar to smallpox and monkeypox where greatest viral replication occurs in the skin. Our data demonstrate that capripoxvirus infections in sheep and goats provide additional and convenient models which are suitable not only for evaluation of poxvirus-specific vaccine concepts and therapeutics, but also study of poxvirus-host interactions.
Assuntos
Capripoxvirus/patogenicidade , Doenças das Cabras/patologia , Infecções por Poxviridae/veterinária , Doenças dos Ovinos/patologia , Eliminação de Partículas Virais , Animais , Capripoxvirus/isolamento & purificação , DNA Viral/análise , DNA Viral/sangue , DNA Viral/isolamento & purificação , Doenças das Cabras/virologia , Cabras/virologia , Especificidade de Órgãos , Reação em Cadeia da Polimerase , Infecções por Poxviridae/patologia , Infecções por Poxviridae/virologia , Doenças dos Ovinos/virologia , Viremia/veterinária , Viremia/virologiaRESUMO
We evaluated the performance of three commercial measles immunoglobulin M enzyme immunoassays from Meddens, Denka Seiken, and Behring. The sensitivities were determined to be 96.7% for the Meddens and Denka Seiken assays and 87.9% for the Behring assay. The specificities of the assays were determined to be 94.6% for Meddens, 98.2% for Denka Seiken, and 98.7% for Behring.