Construction, Expression and Evaluation of Recombinant VP2 Protein for serotype-independent Detection of FMDV Seropositive Animals in Egypt

摘要

Foot-and-mouth disease virus (FMDV) is one of the most devastating viral pathogens of cloven-hoofed animals. The detection of antibodies (Ab) against FMDV structural proteins (SP) using virus neutralization test (VNT) and liquid-phase blocking ELISA (LPBE) is the standard procedure in use for monitoring seroconversion in animals post vaccination, the prevalence of infection-surveillance, proving clinical cases and seronegative status of FMDV-free/na?ve-animals prior transportation. However, due to variations within SP of FMDV serotypes, each serotype-specific Ab should be detected separately which is laborious and time-consuming. Accordingly, it is crucial to develop a sensitive, rapid, and accurate test capable of detecting FMDV-specific Ab, regardless its serotype. This study describes the heterologous expression of VP2 protein in E. coli, and its evaluation as a capture antigen in a simple indirect ELISA for serotype-independent detection of anti-FMDV Ab. Sequence analysis revealed that the VP2-coding sequence is considerably conserved among FMDV serotypes. The recombinant VP2 (rVP2), a 22?kDa polypeptide, was purified to near homogeneity by affinity chromatography under native conditions. Immunoreactivity of the rVP2 was confirmed by using a panel of positive sera including sera from animals vaccinated with the local trivalent vaccine and guinea pig FMDV antiserum, which is routinely used as tracing/detecting Ab in LPBE testing. The results obtained from the VP2-based ELISA were comparable to those determined by VNT and LPBE standard diagnostic assays. Specificity and sensitivity of rVP2 in capturing anti-FMDV Ab were 98.3% and 100%, respectively. The developed VP2-ELISA is proved reliable and time-efficient assay for detection of FMDV seropositive animals, regardless the FMDV serotype that can be implemented in a combination with VNT and/or LPBE for rapid diagnosis of an ongoing FMDV infection.