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

Dehui Yin,Qiongqiong Bai,Han Li,Xiling Wu,Mingjun Sun,Jihong Shao,Jinpeng Zhang

doi:10.3389/fvets.2021.708008

Abstract

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.

Highlights

Brucellosis, as a re-emerging zoonosis, puts human health at risk and causes tremendous losses in animal husbandry around the world, especially in developing countries [1]
The current most commonly used antigens for diagnosing brucellosis mainly depend on Brucella whole cell and lipopolysaccharide (LPS), which can cross-react with the antibodies aroused by other bacteria, such as Yersinia enterocolitica serotype O:9 and Escherichia coli O:157
The results showed that the fusion protein did not cross-react with other bacteria according to an S/N (OD450, sample/negative) > 2.1, paper-based ELISA (p-ELISA) Method for Diagnosing Brucellosis

Summary

Introduction

Brucellosis, as a re-emerging zoonosis, puts human health at risk and causes tremendous losses in animal husbandry around the world, especially in developing countries [1]. Due to the lack of specific clinical manifestations, brucellosis is misdiagnosed as other febrile diseases, such as dengue fever, malaria, or viral bleeding diseases [3, 4]. In animals, this disease is often neglected as there are no symptoms at the early stage of infection. Among the many techniques currently used for diagnosing brucellosis, serological diagnosis methods are the most widely used. It is still meaningful to develop new diagnostic antigens to improve the specificity and sensitivity of serological diagnostic methods for brucellosis [6]

Methods

Results

Conclusion