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Demonstration of nanopore sequencing for the detection of tuberculosis and other infectious diseases in low-and-middle-income countries

A A Slyzkyi(1) E Tiemersma(1) B Myrzaliev(1) L Mtei(1) H Nguyen(1) A van den Bout(4) L Manrho(4) N Hermans(4) K Stoffers(4) E Ubbelohde(1,3) P Lempens(1) C Mulder(1,2) R Anthony(3) J de Beer(4) A van der Zanden(4) K Kremer(1)

1:KNCV Tuberculosis Foundation; 2:Amsterdam Institute for Global Health and Development (AIGHD); 3:National Institute for Public Health and the Environment; 4:Laboratory for Microbiology Twente Achterhoek (Labmicta)

Genome sequencing has been used mainly in large institutes for research and surveillance purposes, but recent innovations, such as the portable and low-cost MinION devices (Oxford Nanopore Technologies), enable genome sequencing close to point-of-care. We aim to demonstrate the use of nanopore sequencing with MinION devices for the diagnosis of tuberculosis (TB), COVID-19, other infectious diseases, and AMR in Kyrgyzstan, Tanzania, and Vietnam.

Training of project country staff has taken place, study protocols were developed, and procurement and importation procedures are ongoing. Nanopore sequencing studies at national level laboratories in the project countries will start in Q2, 2023. Subsequent implementation at decentralized settings follows later.

In the Netherlands, assay development and validation experiments were done. An optimized approach for nanopore sequencing of SARS-CoV-2 was developed. It achieved sufficient and more equally distributed coverage of the SARS-CoV-2 genome. An in-house TB assay was designed and compared by using MinION sequencing with two commercial assays developed for Illumina sequencing. The assays performed well on strains, but showed variable coverage of certain resistance genes in sputum samples. In addition, nanopore sequencing of mixtures of DNA of a multidrug resistant TB strain and a susceptible strain by using the Deeplex Myc-TB (Genoscreen) assay showed that mixed infections could be detected down to 10% abundance.

We aim to generate evidence on the effectiveness, feasibility, acceptability, and costs of using nanopore sequencing for the diagnosis of TB, other infectious diseases, and AMR. Challenges and critical issues will be highlighted guiding future development and scale up.

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