Recombinant CA Protein-Based ELISA for Serological Detection of Small Ruminant Lentiviruses Antibodies

Maedi-visna (MV) and caprine arthritis encephalitis (CAE) are important viral diseases of sheep and goats. The diseases are caused by a group of genetically closely related lentiviruses known as small ruminant lentiviruses (SRLV), and are collectively referred to as SRLV infections. As the majority of sheep and goats infected with SRLV are asymptomatic, the disease is often overlooked. However, SRLV infection can significantly reduce animal productivity and impede animal trade. Currently, SRLV infection is widespread worldwide, but knowledge of its prevalence in China is limited due to a lack of cost-effective testing. In this study, we successfully developed an indirect enzyme-linked immunosorbent assay (iELISA) based on the SRLV capsid protein (CA) (p28) for the specific detection of anti-SRLV antibodies in serum. Using the application of checkerboard titration to optimize the iELISA assay conditions, the cut-off value was determined to be 0.09 by analyzing S/P values of 181 negative sera against SRLV that were confirmed with western blotting (WB). The method showed good reproducibility, with intra- and inter-assay coefficients of variation (CV) values less than 6.60%. A specificity test showed that the iELISA had no serological cross-reaction with six common small ruminant pathogens. Moreover, it was found to be 400-1600 times more sensitive than the available AGID test. The 93 clinical serum samples that tested SRLV-positive using the iELISA were all confirmed as positive using WB, indicating that the method has a low false positive rate. The iELISA was then applied to assess 4786 clinical serum samples from 13 cities in six provinces in China. The results show that the SRLV positivity rate of sera ranged between 0.85% and 40.00%, with the overall seroprevalence of SRLV being 10.64%. Our results indicate that the developed iELISA will serve as a valuable and efficient screening tool for the large-scale preliminary surveillance and monitoring of SRLV infections in both sheep and goats.