• Role and value of whole genome sequencing in studying tuberculosis transmission.

      Nikolayevskyy, Vlad; Niemann, Stefan; Anthony, Richard; Van Soolingen, Dick; Tagliani, Elisa; Ködmön, Csaba; van der Werf, Marieke J; Cirillo, Daniela Maria (2019-04-10)
      Tuberculosis (TB) remains a serious public health threat worldwide. Theoretically ultimate resolution of whole genome sequencing (WGS) for Mycobacterium tuberculosis complex (MTBC) strain classification makes this technology very attractive for epidemiological investigations. To summarise the evidence available in peer-reviewed publications on the role and place of WGS in detection of TB transmission. 69 peer-reviewed publications identified in Pubmed database. Evidence from >30 publications suggests that a cut-off value of <6 single nucleotide polymorphisms (SNPs) between strains efficiently excludes cases that are not the result of recent transmission and could be used for the identification of drug-sensitive isolates involved in direct human-to-human TB transmission. Sensitivity of WGS to identify epidemiologically linked isolates is high reaching 100% in eight studies with specificity (17 - 95%) highly dependent on the settings. Drug resistance and specific phylogenetic lineages may be associated with accelerated mutation rates affecting genetic distances. WGS can be potentially used to distinguish between true relapses and re-infections but in high-incidence low-diversity settings this would require consideration of epidemiologic links and minority alleles. Data from four studies looking into within host diversity highlight a need for developing criteria for acceptance or rejection of WGS relatedness results depending on the proportion of minority alleles.
    • Towards standardisation: comparison of five whole genome sequencing (WGS) analysis pipelines for detection of epidemiologically linked tuberculosis cases.

      Jajou, Rana; Kohl, Thomas A; Walker, Timothy; Norman, Anders; Cirillo, Daniela Maria; Tagliani, Elisa; Niemann, Stefan; de Neeling, Albert; Lillebaek, Troels; Anthony, Richard M; et al. (2019-12-01)
      BackgroundWhole genome sequencing (WGS) is a reliable tool for studying tuberculosis (TB) transmission. WGS data are usually processed by custom-built analysis pipelines with little standardisation between them.AimTo compare the impact of variability of several WGS analysis pipelines used internationally to detect epidemiologically linked TB cases.MethodsFrom the Netherlands, 535 Mycobacterium tuberculosis complex (MTBC) strains from 2016 were included. Epidemiological information obtained from municipal health services was available for all mycobacterial interspersed repeat unit-variable number of tandem repeat (MIRU-VNTR) clustered cases. WGS data was analysed using five different pipelines: one core genome multilocus sequence typing (cgMLST) approach and four single nucleotide polymorphism (SNP)-based pipelines developed in Oxford, United Kingdom; Borstel, Germany; Bilthoven, the Netherlands and Copenhagen, Denmark. WGS clusters were defined using a maximum pairwise distance of 12 SNPs/alleles.ResultsThe cgMLST approach and Oxford pipeline clustered all epidemiologically linked cases, however, in the other three SNP-based pipelines one epidemiological link was missed due to insufficient coverage. In general, the genetic distances varied between pipelines, reflecting different clustering rates: the cgMLST approach clustered 92 cases, followed by 84, 83, 83 and 82 cases in the SNP-based pipelines from Copenhagen, Oxford, Borstel and Bilthoven respectively.ConclusionConcordance in ruling out epidemiological links was high between pipelines, which is an important step in the international validation of WGS data analysis. To increase accuracy in identifying TB transmission clusters, standardisation of crucial WGS criteria and creation of a reference database of representative MTBC sequences would be advisable.
    • Use of a Whole Genome Sequencing-based approach for Mycobacterium tuberculosis surveillance in Europe in 2017-2019: an ECDC pilot study.

      Tagliani, Elisa; Anthony, Richard; Kohl, Thomas A; de Neeling, Albert; Nikolayevskyy, Vlad; Ködmön, Csaba; Maurer, Florian P; Niemann, Stefan; van Soolingen, Dick; van der Werf, Marieke J; et al. (2020-07-30)
      Whole genome sequencing (WGS) can be used for molecular typing and characterisation of Mycobacterium tuberculosis complex (MTBC) strains. We evaluated the systematic use of a WGS-based approach for MTBC surveillance involving all European Union/European Economic Area (EU/EEA) countries and highlight the challenges and lessons learned to be considered for the future development of a WGS-based surveillance system.WGS and epidemiological data of patients with rifampicin (RR) and multi-drug resistant (MDR)-tuberculosis (TB) were collected from EU/EEA countries between January 2017 and December 2019. WGS-based genetic relatedness analysis was performed using a standardised approach including both core genome multilocus sequence typing (cgMLST), and single nucleotide polymorphism (SNP)-based calculation of distances on all WGS data that fulfilled minimum quality criteria to ensure data comparability.From 25 countries, 2218 RR/MDR-MTBC isolates were collected. Fifty-six cross-border clusters with increased likelihood of recent transmission (≤5 SNPs distance) comprising a total of 316 RR/MDR-MTBC isolates were identified. The cross-border clusters included between two and thirty resistant isolates from two to six countries unravelling different RR/MDR-TB transmission patterns in Western and Eastern EU.This pilot study shows that a WGS-based surveillance system is not only feasible but can efficiently elucidate the dynamics of in-country and cross-border RR/MDR-TB transmission across EU/EEA countries. Lessons learned from this study highlight how the establishment of an EU/EEA centralised WGS-based surveillance system for TB will require strengthening of national integrated systems performing prospective WGS surveillance and the development of clear procedures to facilitate international collaboration for the investigation of cross-border clusters.
    • Whole genome sequencing of Mycobacterium tuberculosis: current standards and open issues.

      Meehan, Conor J; Goig, Galo A; Kohl, Thomas A; Verboven, Lennert; Dippenaar, Anzaan; Ezewudo, Matthew; Farhat, Maha R; Guthrie, Jennifer L; Laukens, Kris; Miotto, Paolo; et al. (2019-09-01)