Research Projects

Worldwide, impact of tuberculosis remains considerable, with around 10.8 million cases and 1.25 million deaths reported in 2023. India represented 27% of global TB cases, with extrapulmonary TB (EPTB) accounting for 24% of these instances. EPTB affects organs outside the lungs, such as lymph nodes, intestines, pleura, and skin, posing significant diagnostic challenges because of its paucibacillary nature and the complexity of the sites involved. Existing diagnostic methods, including smear microscopy, culture, histopathology, and nucleic acid amplification tests, have limitations regarding sensitivity, specificity, cost, and the need for trained professionals. Therefore, there is a critical need for the development of a rapid and precise diagnostic method for EPTB. Inspired by this necessity, proposed study intends to establish a new, rapid, efficient, and low-cost nanobiosensing platform for point-of-care diagnosis of EPTB, focusing on M.tb tb-specific biomarkers. Studies suggest that immunodominant proteins such as ESAT-6, CFP-10, Ag85B, MPT-51, and MPT-64 can be used as the diagnostic biomarkers, as they are commonly found in cerebrospinal fluid, urine, and pleural fluid of individuals infected with M. tb. Notably, deletion of RD1 locus of ESAT-6 and CFP-10 in BCG vaccine can reduce cross-reactivity with other Mycobacterium species. MPT-64 is often employed in culture methods to differentiate M. tb from non-tuberculous mycobacteria, while Ag85B is essential for transfer of mycolic acid during cell wall synthesis. MPT-51 significantly contributes to the host immune response. Collectively, these antigens present promising targets for developing rapid and specific nanobiosensing platforms for the diagnosis of EPTB. Initially, a panel of M.tb specific biomarkers (ESAT-6, CFP-10, Ag85B, MPT-51, MPT-64) will be expressed, purified, and characterized. Subsequently, aptamers will be generated and validated against these biomarkers. These aptamers will then be conjugated with engineered nanomaterials (gold and silver nanoparticles, graphene oxide, and quantum dots) to create functionalized nanocomposites for improved signal transduction. The next phase will involve integrating these nanocomposites into an easy-to-use electrochemical biosensing platform. The sensor will be evaluated for its binding efficiency, specificity, and detection limits, with its diagnostic performance confirmed through testing clinical samples, including cerebrospinal fluid, pleural fluid, and urine (n=100). Additionally, an exploration of an aptamer cocktail targeting multiple antigens will be aimed to enhance the sensitivity and specificity of the assay. This platform aims to deliver rapid (30 min), cost-effective (₹200/test), and equipment-free diagnostics, making it appropriate for low-resource environments. Additionally, the strategy could significantly enhance EPTB diagnosis and will contribute to National TB Elimination Program of India.