Whole-genome sequencing of M. tuberculosis directly from clinical samples for high-resolution genomic epidemiology and drug resistance surveillance: an observational study.

Background Direct whole-genome sequencing (dWGS) of M. tuberculosis (MTB) from clinical specimens will be a major breakthrough in tuberculosis diagnosis and control. To date, dWGS has never been used in genomic epidemiology and its accuracy in transmission inference remains unknown. Here we address the technical challenges imposed by dWGS and use it for the accurate inference of transmission clusters and prediction of drug susceptibility. Methods An end-to-end workflow for dWGS was optimized and implemented, and 37 clinical specimens from tuberculosis patients were sequenced. Additionally, 780 clinical isolates covering 90% of all 2014-2016 TB cases in the Comunidad Valenciana (Spain) were whole-genome sequenced. The complete tuberculosis transmission network in the region was built including the clinical specimens and drug susceptibility profiles were predicted. Findings We were able to provide a complete genomic analysis within a week of specimen receipt, at a cost of €217 per sample. After sequencing 37 clinical specimens, 22 smear-positive (85%), and six smear-negative (55%) ones met the quality criteria for downstream analysis. All clinical specimens clustered with their matching culture isolates (median distance = 0 SNPs). 16 specimens were accurately assigned to 10 transmission clusters in the region while 12 were unique cases. Transmission inferences and drug-susceptibility predictions from dWGS data were concordant with sequences from corresponding sputum and phenotypic drug-susceptibility testing. Interpretation dWGS can be used to accurately delineate transmission clusters of tuberculosis and conduct culture-independent surveillance. As compared to conventional approaches, dWGS allows researchers to perform genomic epidemiology in real-time as well as drug resistance surveillance in settings where culture and drug susceptibility testing are not available.