Development of a Novel Reverse Transcription Loop-Mediated Isothermal Amplification Assay for the Rapid Detection of African Horse Sickness Virus.

African horse sickness (AHS) is a disease of equids caused by African Horse Sickness Virus (AHSV) and is transmitted by Culicoides midges. AHS is endemic in sub-Saharan Africa, but during the past century, outbreaks of significant economic importance and elevated mortality have been recorded in Northern African countries, the Iberian and Arabian Peninsula, the Middle East and the Indian subcontinent. Effective control combines the application of early warning systems, accurate laboratory diagnosis and reporting, animal movement restrictions, suitable vaccination and surveillance programs, and the coordination of all these measures by efficient veterinary services. Conventional reverse-transcriptase (RT) PCR (RT-PCR) and real-time RT-PCR (rRT-PCR) assays have improved the sensitivity and rapidity of diagnosing AHS, resulting in the adoption of these methods as recommended tests by the World Organisation for Animal Health (OIE). However, currently these assays are only performed within laboratory settings; therefore, the development of field diagnostics for AHS would improve the fast implementation of control policies. Loop-mediated isothermal amplification (LAMP) is an isothermal, autocycling, strand-displacement nucleic acid amplification technique which can be performed in the field. LAMP assays are attractive molecular assays because they are simple to use, rapid, portable and have sensitivity and specificity within the range of rRT-PCR. This study describes the development of a novel RT-LAMP assay for the detection of AHSV. The AHSV RT-LAMP assay has an analytical sensitivity of 96.1% when considering an rRT-PCR cut-off value of CT  > 36, or 91.3% when no rRT-PCR cut-off is applied. Diagnostic sensitivity and specificity were 100%. This assay provides for a rapid and low cost AHS diagnostic for use in the field.

Experimental infection of common warthogs (Phacochoerus africanus) and bushpigs (Potamochoerus larvatus) with classical swine fever virus II: A comparative histopathological study.

Wild African Suidae, the common warthog (Phacochoerus africanus) and bushpig (Potamochoerus larvatus), were experimentally infected with classical swine fever (CSF) virus following the diagnosis of CSF subtype 2.1 in domestic pigs in South Africa in 2005. No data regarding the susceptibility or potential lesions of these African wild suids are available. Seven subadult warthogs and six bushpigs were captured and infected intranasally with the South African isolate. Two in-contact control animals of the same species in each experiment verified intra-species transmission. Surviving animals were euthanized after 44 days. Formalin-fixed tissue samples collected from them as well as animals euthanized during the trial were evaluated for histological lesions. The warthogs, which were clinically normal throughout the study, developed histological lesions that were inconsistently present and sometimes subtle. Three individuals, including one in-contact control, developed distinct lympho-plasmacytic cuffing in their brains. Subtle lesions included scant lympho-plasmacytic infiltration of various organs, occasionally accompanied by perivascular cuffing. In contrast, the bushpigs developed overt clinical signs similar to CSF in domestic pigs. Four of six animals, including two in-contact controls, died or were euthanized during the trial. On postmortem examination, intestinal necrosis and ulceration, purulent rhinitis and pneumonia were present. Affected animals developed lymphoid necrosis and depletion whilst surviving individuals showed perivascular cuffing in multiple organs. From the present work, we conclude that these wild Suidae are susceptible to CSF virus and intra-species transmission under experimental conditions can occur.

Experimental infection of common warthogs (Phacochoerus africanus) and bushpigs (Potamochoerus larvatus) with classical swine fever virus. I: Susceptibility and transmission.

An incursion of classical swine fever virus (CSFV) into the domestic pig population in South Africa, identified in 2005, raised the concern that infection might spread to wildlife species and be maintained in these hosts. This study sought to determine whether two wildlife Suidae species present in South Africa, the bushpig (Potamochoerus larvatus) and the common warthog (Phacochoerus africanus), could support productive CSFV infection. Both species could be infected with CSFV and transmitted infection to in-contact animals of the same species. Viral antigen and RNA genome were detected in blood/serum and animals that survived initial infection seroconverted approximately 10-14 days post-inoculation. Viral RNA remained detectable in nasal and saliva secretions for prolonged periods until monitoring ended at 42-44 days after initial challenge. These data suggest that both Suidae species could serve to spread circulating CSFV within wild populations, with implications for disease control.