Development and validation of indirect ELISAs based on multiepitope recombinant outer membrane protein fusion antigens for caprine brucellosis diagnosis

Abstract Background Brucella spp. is the causative agent of brucellosis, an important zoonotic disease of major public health concern that is responsible for economic losses in livestock production worldwide. Brucella melitensis , which is prevalent in goats, is the most virulent species for humans. Thus, rapid and accurate diagnostics are crucial for control. In this study, indirect ELISAs were developed and evaluated using multiepitope recombinant fusion proteins derived from Brucella outer membrane proteins (OMPs) for serodiagnosis of caprine brucellosis. Results Three recombinant fusion proteins (OMP31/28/25, OMP28/25, and OMP31) were used to develop indirect ELISAs. Diagnostic performance was evaluated using 400 goat serum samples and compared against (1) the combined Rose Bengal test (RBT) and microtiter plate agglutination test (MPAT) and (2) a commercial ELISA. The sensitivity and specificity of the OMP31/28/25 ELISA were 91.34% and 84.62%, respectively, compared with the RBT+MPAT, and the sensitivity and specificity were 91.39% and 91.97%, respectively, compared with the commercial ELISA. The sensitivity and specificity of the OMP28/25 assay were 95.28% and 82.78%, respectively, compared with the RBT+MPAT and 94.7% and 89.96%, respectively, compared with the commercial ELISA. Both fusion protein-based ELISAs had Cohen’s kappa coefficients of 0.71–0.83, indicating good-to-very good agreement with the reference methods. The OMP31 ELISA demonstrated inadequate diagnostic performance. Conclusions The developed indirect ELISAs using recombinant fusion antigens (OMP31/28/25 and OMP28/25) offered robust diagnostic performance, comparable to that of the reference methods. These assays demonstrate potential as cost-effective tools for caprine brucellosis diagnosis and surveillance, but broader field validation is needed.