Targeted and whole-genome sequencing for drug resistance and genetic relatedness inference of rifampicin-resistant Mycobacterium tuberculosis: an in-depth comparison.

BACKGROUND

Challenges in tuberculosis (TB) control have fuelled interest in routine next generation sequencing (NGS). However, the comparative performance of targeted NGS (tNGS) and whole genome sequencing (WGS) for drug resistance and genetic relatedness inference remains unclear.

METHODS

In this cross-sectional study, we compared WGS (MICK-MAGMA platform) and the Deeplex MycTB tNGS assay in 90 patients with rifampicin-resistant TB in South Africa. A pairwise analysis was conducted for a total of 60 isolates, for which tNGS was conducted directly on DNA from sputum, and WGS on cultured isolates from the same samples.

RESULTS

Drug resistance inference was highly concordant (≥92%) for most drugs, but lower for isoniazid (82%) and ethionamide (78%). Mixed infections were more commonly detected in tNGS (6.7%) than WGS (1.7%), though likely due to analytical errors. tNGS detected more minor variants (allelic frequency <25%) than WGS (76 versus 32), with minimal overlap. Most minor variants were of unknown significance; some likely stemmed from contamination or sequencing errors. Heteroresistance involving minor variants was rare (4.7% in tNGS, 0% in WGS). WGS provided lineage and sublineage information for all samples, while tNGS reported lineage for 67% and sublineage for 20%. WGS classified more samples as genetically unrelated (76%) than tNGS (40%).

CONCLUSION

In this cohort, drug resistance inference was largely concordant between tNGS and WGS. WGS offered higher resolution for genetic relatedness, while tNGS showed greater sensitivity for minor variants. Further research is needed to clarify the clinical relevance of minor variants and assess the utility of WGS for transmission control.