Faster TB tracking from sputum using nanopore sequencing

Tuberculosis is caused by Mycobacterium tuberculosis (MTB) and tracking how it spreads requires genomic data. Because MTB grows slowly in the lab, whole genome sequencing (WGS) results are often delayed, slowing public health responses. Only a few teams have tried to speed this up by sequencing directly from respiratory specimens rather than waiting for cultures, and most of those efforts have focused on predicting drug resistance. In the study led by Laura Pérez‐Lago, researchers took a different tack: they aimed to accelerate precise delineation of transmission by coupling culture-free WGS to a surveillance programme and, for the first time in this line of work, they used flexible nanopore sequencing to do it. The team selected 71 sputa and applied a simple laboratory step to deplete human DNA, deliberately avoiding costly and cumbersome capture-bait alternatives. By sequencing directly from sputum and integrating results with ongoing surveillance, the study tests whether genomic information useful for mapping who infected whom can be produced close to the time of diagnosis rather than waiting for culture-based data.

The study used direct nanopore sequencing on 71 sputa after removing human DNA, without performing capture-bait enrichment. The authors report that optimal results—defined as >90% genome covered, mean coverage >45× and >70% genome covered >20×—were obtained from 33.8% of cases. Those high-quality results allowed assignment of every new case with such data to transmission clusters close to diagnosis. Another 12.6% of samples produced suboptimal data, with coverage ranging from 15.5%-90.92% at >10× depth. To make use of those weaker datasets, the team developed a rescue pipeline based on identifying informative SNPs acting as markers for relevant transmission clusters in their population. Using those marker SNPs, the pipeline enabled pre-allocation of new cases to pre-existing clusters and, in some instances, allowed the researchers to determine precise genomic relationships with preceding cases in a cluster. Overall, the approach yielded epidemiologically valuable information directly from sputum in approximately half the samples analysed.

This work represents a step forward in producing comparative genomic information at diagnosis rather than after culture, demonstrating that a substantial fraction of cases can be placed into transmission context without lengthy lab growth. By showing that nanopore sequencing combined with a human DNA depletion step and a targeted rescue pipeline can generate cluster assignments or at least pre-assignments, the study points toward faster, more flexible genomic surveillance. Importantly, the method avoids capture-bait enrichment, simplifying the lab workflow and reducing cost and complexity. While not every sputum yielded complete genomes, the ability to exploit informative SNPs from suboptimal data increases the number of cases that can contribute to real-time transmission mapping. Coupling culture-free WGS to routine surveillance programmes, as tested here, offers a practical route to bring genomic epidemiology closer to the point of diagnosis and to make genomic insights available earlier in the public health response.