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Whole genome sequencing of Mycobacterium tuberculosis: current standards and open issues

Abstract

Whole genome sequencing (WGS) of Mycobacterium tuberculosis has rapidly progressed from a research tool to a clinical application for the diagnosis and management of tuberculosis and in public health surveillance. This development has been facilitated by drastic drops in cost, advances in technology and concerted efforts to translate sequencing data into actionable information. There is, however, a risk that, in the absence of a consensus and international standards, the widespread use of WGS technology may result in data and processes that lack harmonization, comparability and validation. In this Review, we outline the current landscape of WGS pipelines and applications, and set out best practices for M. tuberculosis WGS, including standards for bioinformatics pipelines, curated repositories of resistance-causing variants, phylogenetic analyses, quality control and standardized reporting.

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Fig. 1: Whole genome sequencing of Mycobacterium tuberculosis.
Fig. 2: Standard workflow for whole genome sequencing of Mycobacterium tuberculosis complex isolates.
Fig. 3: Current and potential approaches for determining resistance-related polymorphisms.
Fig. 4: Epidemiological and within-host applications of SNP-based comparisons between Mycobacterium tuberculosis complex isolates.

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Acknowledgements

C.J.M. and L.R. are also affiliated with BCCM/ITM Mycobacterial Culture Collection, Institute of Tropical Medicine, Antwerp, Belgium. J.G. is also affiliated with the BC Centre for Disease Control, Vancouver, Canada. B.O.-A. is also affiliated with the Center for Global Health Security and Diplomacy, Ottawa, Canada. M.S. is also affiliated with the University of Arizona, Tucson, AZ, USA. I.C. is also affiliated with the CIBER in Epidemiology and Public Health, Spain. C.J.M., B.O.-A., L.R. and B.C.d.J. are supported by a European Research Council grant (INTERRUPTB; no. 311725). I.C. and G.A.G. are supported by a European Research Council grant (TB-ACCELERATE; no. 638553). T.C.R. receives salary support from the not-for-profit organization Foundation for Innovative New Diagnostics (the terms of this arrangement have been reviewed and approved by the University of California, San Diego). T.M.W. is an NIHR Academic Clinical Lecturer. J.L.G. and J.G. receive funding from the University of British Columbia, Vancouver, Canada. T.A.K., C.U., V.D. and S.N. receive funding from the German Center for Infection Research (DZIF) and are funded by the Deutsche Forschungsgemeinschaft (German Research Foundation) under Germany’s Excellence Strategy (EXC 22167–390884018). L.V., T.H.H. and A.V.R. are funded by FWO Odysseus G0F8316N. M.R.F. is supported by the US National Institutes of Health BD2K K01 (MRF ES026835). P.S. is supported by the Agence Nationale de la Recherche (ANR-16-CE35-0009).

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Nature Reviews Microbiology thanks T. McHugh, V. Sintchenko, and other anonymous reviewer(s), for their contribution to the peer review of this work.

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C.J.M., G.A.G., T.A.K., L.V., A.D., M.E., M.R.F., J.L.G., K.L., P.M., B.O.-A., V.D., P.S., A.S., C.U., D.v.S., Y.Z., M.S., J.G., D.M.C., S.N., I.C. and A.V.R. researched the data for the article. C.J.M., G.A.G., T.A.K., L.V., A.D., M.E., M.R.F., J.L.G., K.L., P.M., B.O.-A., V.D., P.S., A.S., C.U., D.v.S. Y.Z., M.d.V., S.G., T.H.H., L.R., E.T., T.M.W., R.M.W., M.S., J.G., D.M.C., S.N., I.C. and A.V.R. substantially contributed to the discussion of the content. C.J.M., G.A.G., T.A.K., L.V., A.D., M.E., M.R.F., J.L.G., K.L., P.M., B.O.-A., V.D., P.S., A.S., C.U., D.v.S., Y.Z., M.S., J.G., D.M.C., S.N., I.C. and A.V.R. wrote the article. All authors reviewed and edited the manuscript before submission.

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Correspondence to Inaki Comas or Annelies Van Rie.

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P.S. was a consultant for Genoscreen. All other authors declare no competing interests.

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Related links

Human, Heredity and Health in Africa Consortium: https://h3abionet.org

ReSeqTB: http://www.reseqtb.org

TORCH consortium: https://torch-consortium.com/vliruos

ERLTB-Net: https://ecdc.europa.eu/en/about-us/partnerships-and-networks/disease-and-laboratory-networks/erltb-net

Supplementary information

Glossary

Mycobacterium tuberculosis complex

(MTBC). The genetically related group of organisms within the genus Mycobacterium that cause tuberculosis in humans or animals.

Drug susceptibility testing

(DST). A procedure to determine if clinical isolates are resistant to antibiotics either by testing the inhibition in culture (phenotypic DST) or by identifying drug resistance-associated mutations (genotypic DST).

Source investigation

The first case in a group of related individuals that transmitted the disease. Usually, identified during the development of an epidemiological investigation.

Löwenstein–Jensen

A selective culture solid medium commonly used to isolate Mycobacterium tuberculosis complex strains.

Mycobacteria Growth Indicator Tube

A tube that contains mycobacteria-selective culture liquid medium and is usually coupled to an automated instrument to read the results.

PE and PPE gene families

Families of genes that encode virulence factors in Mycobacterium tuberculosis complex strains. They have signature (proline)–proline–glutamate ((P)PE) motifs at their amino terminus.

Core genome MLST

A scheme that converts genome-wide SNP data into an allele-numbering system using a preselected set of core genes.

Whole genome MLST

A scheme that converts genome-wide SNP data into an allele-numbering system using a preselected set of core genes and additional accessory genes.

Contact tracing

The identification of possible contacts that interacted with an infected person (index case), often through questionnaires and interviews.

Mycobacterial interspersed repetitive unit variable-number tandem repeat

Mycobacterium tuberculosis complex (MTBC)-specific variable tandem repeat locus used to genotype MTBC strains.

WGS pipelines

The bioinformatics section of the whole genome sequencing workflow, starting from raw sequencing files through to SNP calling and analyses.

WGS workflows

All steps involved (from culturing to SNP calling and analyses) for whole genome sequencing of an isolate.

BioCompute Object

(BCO). A framework for standardized reporting of computational parameters for a whole genome sequencing pipeline.

Spoligotyping

A PCR-based approach based on the amplification of spacers in the CRISPR region of the Mycobacterium tuberculosis complex (MTBC). It is used for genotyping MTBC strains.

Effective reproduction number

The average number of secondary cases per infectious case.

Microevolution

Genetic changes within a population, resulting in separate subpopulations.

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Meehan, C.J., Goig, G.A., Kohl, T.A. et al. Whole genome sequencing of Mycobacterium tuberculosis: current standards and open issues. Nat Rev Microbiol 17, 533–545 (2019). https://doi.org/10.1038/s41579-019-0214-5

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