Celebrating 10 years of innovation in TB research by the Nanopore Community

Today, tuberculosis (TB) remains one of the deadliest infectious diseases in the world. In 2022, 1.3 million people died from TB, and an estimated 10.6 million people fell ill with TB1 — it is present in all countries and age groups.

Drug resistance is a major threat to TB control and remains a public health crisis, therefore rapid and comprehensive detection methods directly from sputum (or decontaminated sputum) samples are vital to tackle this global challenge.

In February 2014, Oxford Nanopore Technologies launched the MinION Access Programme. Since then, members of the Nanopore Community have worked tirelessly to innovate and develop methods to provide faster and extensive data for the identification and characterisation of TB.

They found that the long sequencing reads generated by nanopore technology provided a more comprehensive view of Mycobacterium tuberculosis genomes, enabling the identification of a broad range of drug resistance mutations, including novel mutations. This, in addition to the fast turnaround time from sample to answer, meant that researchers quickly adopted nanopore sequencing as a better alternative to traditional M. tuberculosis culture and molecular methods.

Here we take a look at some of the TB research using nanopore sequencing from the last 10 years, and the future potential of nanopore sequencing to be used for the clinical identification of TB and its drug resistance*.

First steps to rapid and comprehensive identification of TB with nanopore sequencing

Just nine months after the MinION Access Programme was launched, the Nanopore Community had demonstrated the impact nanopore sequencing could have on TB research.

In their proof-of-concept study, Bradley et al. showed that they could rapidly identify and predict antibiotic resistance from M. tuberculosis isolates2. Furthermore, the team demonstrated it was feasible to generate a clinician-friendly report within three minutes using just a laptop and a MinION.

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A year later, in December 2016, Votintseva et al. showcased how whole-genome sequencing on a MinION device could potentially identify TB directly from respiratory samples in a single day, removing the need for culture-based methods that take several weeks3.

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Pioneering the use of portable, real-time nanopore sequencing in TB research

In 2018, scientists in Madagascar used a MinION device to perform DNA sequencing in-country for the first time to rapidly identify TB and its drug resistance profile. This achievement was only possible due to the portability and low cost of the MinION device.

Up until this point, DNA sequencing of TB samples was restricted to high-income countries due to the cost and complexity of alternative sequencing technologies — nanopore sequencing enables low- and middle-income countries access to DNA sequencing to research TB.

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Complete, high-quality assembly of an extensively drug-resistant TB strain using nanopore sequencing

Also in 2018, Bainomugisa et al. used nanopore sequencing to assemble a high-quality, complete genome of the modern Beijing lineage strain of TB responsible for drug resistance outbreaks in the Western Province of Papua New Guinea4.

Using a MinION device, the team resolved all the repetitive pe/ppe gene families, which are thought to contribute to virulence. Traditionally, these gene families have been inaccessible to short-read sequencing technologies because of their repetitive and GC-rich nature.

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Positive evaluation of nanopore sequencing-based targeted analysis of drug-resistant TB

On World TB Day in 2022, Oxford Nanopore, in collaboration with Quadram Institute Bioscience (QIB), announced that it had developed a new rapid workflow shown to identify drug-resistant TB in a Stage I study.

When validated, the sequencing workflow has the potential to help clinicians tackle TB globally in the future; especially in resource-limited settings where the MinION device can be used in distributed networks. The Oxford Nanopore sequencing solution is available today only as a life science research tool.

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Characterisation of drug resistance in TB with targeted nanopore sequencing

In 2023, Shannon Murphy and colleagues at the Wadsworth Center, New York State Department of Health, developed a rapid, culture-free, targeted sequencing assay that used MinION Flow Cells on both MinION and GridION devices to characterise drug resistance profiles of TB directly from respiratory samples5.

The team demonstrated that their targeted assay was just as accurate at identifying resistance mutations as whole-genome sequencing analysis. But importantly, the assay turnaround time showed a more than two-week improvement compared with culture and whole-genome sequencing workflows at a similar cost, and their method also offers additional utility for cultures that are too low quality for whole-genome sequencing analysis.

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World Health Organization (WHO) supports the use of targeted sequencing, including a test under development from Oxford Nanopore, to detect drug resistance in TB

After almost 10 years of research, in 2023, the WHO announced that a rapid sequencing solution being developed by Oxford Nanopore meets the class-based performance criteria to detect drug resistance after TB diagnosis, to guide clinical decision-making for drug-resistant TB treatment.

The Oxford Nanopore solution is among the first sequencing-based tests to be supported by the WHO in the field of TB — a significant milestone in the effort to combat one of the most persistent threats in public health globally.

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Looking to the future: nanopore sequencing as a powerful tool in the fight against TB

With ongoing advancements in technology and method development showcased over the past decade, including genomic assays directly from respiratory samples, nanopore sequencing is poised to transform comprehensive TB drug resistance testing.

WHO support of sequencing-based tests, including a rapid sequencing solution being developed by Oxford Nanopore, signifies a crucial step forward in leveraging nanopore technology for TB control efforts, offering the potential for more accurate, timely, and accessible genomic profiling of drug-resistant TB.

An ongoing strategic partnership between Oxford Nanopore and bioMérieux aiming to improve health outcomes globally by exploring selected opportunities to bring nanopore sequencing to the infectious disease diagnostics market holds promise for nanopore sequencing as a powerful tool in the fight against global TB drug resistance.

*Oxford Nanopore Diagnostics are developing a TB drug resistance (TB-DR) test for clinical use as described in the WHO rapid communication (July 2023). The end-to-end workflow, inclusive of reagents, GridION device, and software will be manufactured under ISO 13485 and will be submitted for registration with global regulatory bodies.

  1. World Health Organization. Tuberculosis Fact Sheet. https://www.who.int/news-room/fact-sheets/detail/tuberculosis (2023) [Accessed 12 March 2024]

  2. Bradley, P. et al. Rapid antibiotic-resistance predictions from genome sequence data for Staphylococcus aureus and Mycobacterium tuberculosis. Nat. Commun. 6:10063 (2015). DOI: https://doi.org/10.1038/ncomms10063

  3. Votintseva, A.A. et al. Same-day diagnostic and surveillance data for tuberculosis via whole-genome sequencing of direct respiratory samples. J. Clin. Microbiol. 55(5):1285–1298 (2017). DOI: https://doi.org/10.1128/jcm.02483-16

  4. Bainomugisa, A. et al. A complete high-quality MinION nanopore assembly of an extensively drug-resistant Mycobacterium tuberculosis Beijing lineage strain identifies novel variation in repetitive PE/PPE gene regions. Microb. Genom. 4(7):e000188 (2018). DOI: https://doi.org/10.1099/mgen.0.000188

  5. Murphy, S.G. et al. Direct detection of drug-resistant Mycobacterium tuberculosis using targeted next generation sequencing. Front. Public Health. 11:1206056 (2023). DOI: https://doi.org/10.3389/fpubh.2023.1206056