From immunoinformatics insights to assay development: establishment of a peptide-based indirect elisa for cynomolgus tuberculosis detection.

Tuberculosis (TB) is a severe zoonotic disease that represents a considerable threat to the health of captive non-human primates (NHPs). Conventional serological diagnostic methods often suffer from insufficient sensitivity and specificity, making them difficult to meet the requirements of large-scale screening in monkey colonies. In this study, we predicted and screened Mycobacterium tuberculosis antigenic epitopes using immunoinformatic approaches, and identified a novel antigenic peptide MP71 from the MPT64 protein. Based on the MP71-KLH conjugate, we further developed an indirect ELISA assay. In both the learning cohort (n = 19) and independent validation cohort (n = 21), the assay exhibited favorable and stable diagnostic performance: in the validation cohort, the AUC was >0.94 (95% CI: 0.8471 to 1.000), with a sensitivity of >92% and a specificity of 100%, together with favorable repeatability. The established ELISA represents a rapid, simple, and reliable serological tool that can be used for routine surveillance and large-scale preliminary screening of tuberculosis in monkeys.