Identification and evaluation of T cell epitopes of Rv0585c from Mycobacterium tuberculosis

Abstract

Objective: To investigate the human T cell epitopes of Mycobacterium (M.) tuberculosis Rv0585c protein antigen and their immunogenicity and provide evidence for the development of specific tuberculosis immune diagnostic techniques and tuberculosis vaccine. Methods: We synthesized peptides from M. tuberculosis Rv0585c protein antigen predicted by TE-predict and IEDB human T cell epitope prediction tool. The cellular immunoreactivity of the predicted peptides was evaluated through ELISpot assay with the peripheral blood monouclear cells (PBMC) of clinical tuberculosis patients. In animal experiments, BALB/c mice were respectively immunized with high dose (100 μg/mice) and low dose (50 μg/mice) of the peptides of Rv0585c, at the same time, high dose (50 μg/mice) and low dose (20 μg/mice) of Ag85B protein were used in positive control group. The levels of IFN-γ, IL-2, IL-4 and IL-10 were tested with ELISA kit respectively. Results: By means of bioinformatics technique, 66 human T cell epitopes of Rv0585c were predicted, from which9 peptides concentrated epitopes were synthesized for the animal immune experiments. Peptides P10110, P10112 and P10117 were confirmed to be antigenic. The sensitivity and specificity of P10110, P10112 and P10117 were 14.00%, 12.00%, 6.00% and 100.00%, 100.00%, 97.96% respectively when they were used as diagnostic reagents of tuberculosis. The sensitivity and specificity were 22.00% and 97.96% when the epitopes were combined together. The results of animal immunity test showed that high levels of cytokines IFN-γ, IL-2, IL-4 and IL-10 were induced by high and low dose of P10110, and high levels of IFN-γ、IL-2 and IL-10 were induced by high and low dose of P10112, which were much higher than that in negative controls, respectively (P<0.001). Conclusion: Rv0585c, including its human T cell epitopes, has good immunogenicity and immunoreactivity, stimulating the body to produce a stronger cellular immune response and has better potential application value in cellular diagnosis of tuberculosis and the development of new type of tuberculosis vaccine.