Design and development of a novel recombinant multi-epitope antigen: A promising candidate for melioidosis disease diagnosis.

Melioidosis, a deadly seasonal disease prevalent in the tropics and subtropics, is caused by exposure to the opportunistic pathogen Burkholderia pseudomallei. Diagnosing the disease is challenging due to its wide range of clinical symptoms and reliance on slow bacterial culture. Although molecular assays like real-time PCR offers high sensitivity and specificity, they are not suitable for field use and require trained personnel. Immunological tests often suffer from reduced accuracy in endemic regions due to the background seropositivity and the application of crude antigen preparations. To address these limitations, this study focused on developing a novel recombinant multi-epitope antigen that incorporates recurrent linear and conformational B-cell epitopes derived from six B. pseudomallei antigens with different sub-cellular localization. A 1071 bp synthetic gene encoding 17 epitopes was cloned in the pET-28a(+) prokaryotic expression system and recombinant protein was over-expressed under IPTG induction in E. coli BL21 cells. The potential of rBpMEP as a diagnostic antigen was further evaluated through immunoblotting using melioidosis-positive and healthy human serum. The rBpMEP-based iELISA exhibited a sensitivity of 83.3% and specificity of 94.1%. The findings demonstrated that rBpMEP holds significant potential as a serodiagnostic antigen for developing assays for melioidosis diagnosis and disease surveillance.