Development and validation of recombinant A27L based indirect-ELISA for Sheeppox and Goatpox disease in India

Sheeppox and Goatpox are highly contagious transboundary viral diseases of sheep and goats caused by Capripoxvirus, respectively. This study describes the development of indirect ELISA and its serodiagnostic potential as an alternative to the gold standard serum neutralization test (SNT). The homologue of vaccinia virus, ORF 117 (A27L) gene of the Romanian Fenner (RF) strain of Sheeppox virus (SPPV) was used for producing the full-length recombinant A27L (rA27L) protein (∼22 kDa) in a prokaryotic expression system. The immunogenic potential of purified rA27L was confirmed by detecting its specific reactivity with SPPV hyperimmune sera in Western Blot analysis. An indirect ELISA based on rA27L was developed and optimized. In comparison to SNT, the rA27L-ELISA assay exhibited a diagnostic specificity of 96% and 98%, while the sensitivity was 98% and 97%, with sheep and goat sera, respectively. No cross-reactivity was noted with other transboundary diseases such as Orf, Foot-and-Mouth disease, Peste des Petits Ruminants, Haemorrhagic septicemia and Brucella. A total of 508 serum samples were tested in total for the preliminary analysis. Among these, seropositivity rates for sheep (n = 270) and goats (n = 238) were 31.85% and 47.47%, respectively. Further, to determine accuracy of developed test a total of 1731 serum samples were analyzed from the states with high small ruminant populations in India which included Andhra Pradesh, Telangana, Karnataka, West Bengal, and Gujarat. Overall seropositivity recorded were 24% in sheep and 19.12% in goats. Among the population of sheep and goats examined, the females and animals in the age group of 1-2 years showed high Capripoxvirus antibodies. This study highlighted the potential utility of rA27L protein as a safe and alternative serodiagnostic reagent, unlike the SNT, for serosurveillance and seromonitoring of Capripoxvirus infection in sheep and goats.