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Targeted genome sequencing of drug resistant tuberculosis: a laboratory perspective

T Niemann     V Mohr     C Utpatel     L Araujo     V Dreyer    I Barilar     T A Kohl     S Niemann(1)

1: Research Center Borstel, Leibniz Lung Center

Drug resistant (DR), multidrug resistant (MDR), pre-extensively (pre-XDR), and extensively DR (XDR) Mycobacterium tuberculosis complex (MTBC) strains represent a serious challenge for tuberculosis (TB) control. Key to control is the fast resistance prediction, which is still a great challenge and not widely available in high incidence settings. Here, targeted genome sequencing has become a promising tool for rapid DR/MDR/Pre-XDR/XDR TB detection from clinical specimens such as sputum. However, implementation of sequencing technologies in low- and middle-income countries is still a challenge. To tackle this question, we developed and tested a comprehensive toolbox for the implementation of targeted next generation sequencing based on the Deeplex® Myc-TB assay (Genoscreen, France). The assay is capable of detecting resistance associated mutations to 13 TB drugs  The toolbox includes infrastructure checklists, standardized methods and laboratory workflows, standard operating procedures (SOPs) and extensive training materials. Critical components are infrastructure assessment tools, optimized workflows for DNA extraction, library preparation, targeted sequencing, and standardized tools for quality control for all steps involved in sequencing, all of which linked to respective training materials. The developed workflows have been tested and optimized by in-house evaluations and in pilot projects in Namibia, Eswatini, and Mozambique. While implementation of genome sequencing in low- and middle-income countries is still a challenge, it can be successfully performed using our toolbox including optimized laboratory workflows, detailed SOPs, and comprehensive trainings. Challenges remain that e.g. are related to supply chains, constant laboratory support and further method development.

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