Hybrid recombinant Omp 22, 25, and 31 immunodominant epitopes can be used for serodiagnosis of brucellosis

• Published in: Journal of immunological methods 2021-10, Vol.497, p.113123-113123, Article 113123
• Main Authors: Dehghani, Sima, Sabzehei, Faezeh, Taromchi, Amir Hossein, Mobaien, Ahmad Reza, Arsang-Jang, Shahram
• Format: Article
• Language: English
• Subjects: B-cell epitopes; Brucellosis; ELISA; Outer membrane protein; Recombinant protein
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/j.jim.2021.113123

Brucellosis is a well-known infectious disease in most parts of the world, especially in developing countries, common between humans and animals. Brucellosis is diagnosed by serological tests based on lipopolysaccharides (LPSs), which are bacterial cell wall antigens, and due to the similarities between LPSs antigens of some gram-negative bacterias, false-positive responses are inevitable. Alternatively, Outer membrane proteins (Omps), as antigenic conserved membrane proteins, can be used to diagnose brucellosis instead of LPS antigens. In this study, by using bioinformatics tools, linear B-cell epitopes were selected from Omp22, Omp25, and Omp31 antigens and fused with the rigid KP linker (K = Lysine, P=Proline). Designed gene cassette was cloned into pET-28a (+) vector and expressed recombinant protein was purified using Ni-NTA chromatography column and was confirmed with Poly-Histidine-HRP antibody. Finally, recombinant protein's seroreactivity with serum samples from 37 patients and 27 healthy individuals was evaluated by western blotting and enzyme-linked immunosorbent assay (ELISA) methods. Western blotting results showed high reactivity of the recombinant protein with serum samples of Brucella infected patients. ELISA results were analyzed using the receiver operating curve (ROC). Optical density cut-off point, accuracy, sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and Youden index J for recombinant protein were > 0.809, 84.37%,83.78%,88.89%,88.57%, 79.31% and 0.72 respectively. Western blotting and ELISA results showed that our recombinant protein has good sensitivity and specificity for the diagnosis of brucellosis. [Display omitted] •To accurately detection of brucellosis, alternative antigens with conserved sequences are needed for diagnosis.•By using bioinformatics tools, six linear B-cell epitopes were identified from three antigenic outer membrane proteins.•Each epitope was connected to the other epitope with an appropriate linker.•Candidate recombinant fused epitopes were able to react with antibodies at the serological assays.