A Multi-Epitope Fusion Protein-Based p-ELISA Method for Diagnosing Bovine and Goat Brucellosis

In recent years, the incidence of brucellosis has increased annually, causing tremendous economic losses to animal husbandry in a lot of countries. Therefore, developing rapid, sensitive, and specific diagnostic techniques is critical to control the spread of brucellosis. In this study, bioinformatics technology was used to predict the B cell epitopes of the main outer membrane proteins of Brucella, and the diagnostic efficacy of each epitope was verified by an indirect enzyme-linked immunosorbent assay (iELISA). Then, a fusion protein containing 22 verified epitopes was prokaryotically expressed and used as an antigen in paper-based ELISA (p-ELISA) for serodiagnosis of brucellosis. The multi-epitope-based p-ELISA was evaluated using a collection of brucellosis-positive and -negative sera collected from bovine and goat, respectively. Receiver operating characteristic (ROC) curve analysis showed that the sensitivity and specificity of detection-ELISA in diagnosing goat brucellosis were 98.85 and 98.51%. The positive and the negative predictive values were 99.29 and 98.15%, respectively. In diagnosing bovine brucellosis, the sensitivity and specificity of this method were 97.85 and 96.61%, with the positive and negative predictive values being identified as 98.28 and 97.33%, respectively. This study demonstrated that the B cell epitopes contained in major antigenic proteins of Brucella can be a very useful antigen source in developing a highly sensitive and specific method for serodiagnosis of brucellosis.