Research Projects

Background While the current laboratory algorithms being followed in most TB laboratory settings in India involve phenotypic drug susceptibility tests (DST) and/or targeted genotypic methods such as line probe assays, these methods have their disadvantages. Next generation sequencing has revolutionized the tracking and genomic epidemiology of many pathogens of public health importance including M.tuberculosis in many developed country settings. However, the deployment of this vital tool in the public health laboratories involved in tuberculosis detection and susceptibility testing in Indian settings has lagged behind. Novelty Our proposal seeks to optimize and standardize next generation sequencing protocols for detection of anti-tuberculosis drug resistance directly from sputum specimens. Using long-read next generation sequencing platform (Oxford Nanopore MK1C), we aim to evaluate the feasibility of incorporating NGS protocols into the current algorithms for anti-tuberculosis drug susceptibility testing and the potential of deploying them in field settings. Objectives 1. To optimize next generation sequencing protocols using long-read portable sequencing chemistry for detecting anti-tuberculosis drug resistance. 2. To evaluate the performance of portable next generation sequencing technology in detecting and predicting drug susceptibility profiles of patients/samples positive for M.tuberculosis as compared to the current gold standards in use. 3. Compare next generation sequencing protocol in terms of sensitivity to detect drug resistance targets and to calculate concordance with current methods such as line probe assays and culture-based methods. 4. Analyze the turn-around time for profiling M.tuberculosis DST using currently deployed phenotypic and genotypic methods as compared to portable next generation sequencing assays directly from sputum samples. 5. To explore the feasibility of optimizing and simplifying work flows for portable next generation sequencing to be deployed in intermediate reference laboratory settings. Methods We will standardize protocols for detection of anti-tuberculosis drug resistance using long-read portable next generation sequencing platforms. The concordance of the results from NGS methods with the current laboratory algorithms will be calculated as well as the turn-around-times. In addition, capacity building workshops for NGS methods at IRL and district levels will be enabled. Expected Outcomes The proposed project is an endeavor in capacity building in public health laboratories at different levels for detection of anti-tuberculosis drug mutations using next generation sequencing and introducing NGS at the programmatic level in the country for MDR TB diagnosis.