Initial experiences withMycobacterium tuberculosisDNA extraction for downstream Deeplex Myc-TB targeted deep sequencing in a high burden setting

The propensity for M. tuberculosis to develop resistance and the lack of clinical tools for the rapid determination of such resistance has long significantly complicated tuberculosis (TB) therapeutics. Targeted next-generation sequencing (NGS) has improved our understanding of the genetic basis and identification of drug-resistant TB. However, to achieve accurate results reliable enough for clinical implementation, high-quality M. tuberculosis DNA must be extracted from patient-derived samples within high burden routine laboratory workflows. In advance of a large cluster RCT in the Western Cape of South Africa evaluating the Deeplex Myc-TB targeted NGS assay (GenoScreen; Lille, France), we sought to compare DNA extraction methods for both early MGIT culture-positive samples and processed patient sputum. Given the lack of reference standard method, we assessed a representative set of DNA extraction protocols, including the GenoScreen-recommended method, in parallel in South Africa and at UC San Francisco. Our findings provide preliminary insights into an optimal DNA extraction method for the utilization of Deeplex Myc-TB in routine laboratory settings and can inform future experiments evaluating newer generation assays.