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Background: Retrospective and descriptive molecular epidemiology studies have shown that Mycobacterium tuberculosis whole genome sequencing can identify outbreaks and disease transmission events with higher resolution than conventional epidemiological investigations. Those studies have strengthened our understanding of genomic polymorphisms correlating with person-to-person transmission and helped resolve putative transmission clusters. To date, systematic genomic surveillance programs implemented for M. tuberculosis were only implemented in low-incidence settings. The purpose of this study is to determine whether there is an impact of routine M. tuberculosis whole genome sequencing on tuberculosis case detection in a high-incidence setting., Methods: A cluster randomized controlled trial will be performed. Forty-eight rural village groups (or Fokontany) in the Vohibato district of Madagascar will be randomized to one of three interventions arms. Arm 1 (standard of care) involves healthcare facility-based passive case detection with smear microscopy testing. Arm 2 (best practice) consists of active case finding and Xpert MTB/RIF Ultra PCR testing followed by household contact investigations. Arm 3 (novel intervention) includes the same interventions as arm 2, with addition of sputum culture and M. tuberculosis whole genome sequencing for all newly diagnosed cases. In arm 3, molecular suggested putative outbreaks are investigated, and additional TB suspects are appropriately tested. The intervention observational period will be 2 years. The primary outcome will be the number of detected cases/100,000/year in each arm after 1 year of intervention., Discussion: This study is designed to determine whether there is an impact of prospective whole genome sequencing-based molecular typing on tuberculosis case detection in high-incidence settings. Investigating potential outbreaks and focusing active case finding in spatiotemporal settings where disease transmission is suggested by genomic typing is hypothesized to improve case detection in rural communities., Trial Registration: ClinicalTrials.gov NCT05406453 . Retrospectively registered on June 6, 2022., (© 2024. The Author(s).)

Targeted Next Generation Sequencing (tNGS) and Whole Genome Sequencing (WGS) are increasingly used for genotypic drug susceptibility testing (gDST) of Mycobacterium tuberculosis. Thirty-two multi-drugs resistant and 40 drug susceptible isolates from Madagascar were tested with Deeplex® Myc-TB and WGS using the Mykrobe analysis pipeline. Sixty-four of 72 (89 %) yielded concordant categorical gDST results for drugs tested by both assays. Mykrobe didn't detect pncA K96T, pncA Q141P, pncA H51P, pncA H82R, rrs C517T and rpsL K43R mutations, which were identified as minority variants in corresponding isolates by tNGS. One discrepancy (rrs C517T) was associated with insufficient sequencing depth on WGS. Deeplex® Myc-TB didn't detect inhA G-154A which isn't covered by the assay's amplification targets. Despite those targets being included in the Deeplex® Myc-TB assay, a pncA T47A and a deletion in gid were not identified in one isolate respectively. The evaluated WGS and tNGS gDST assays show high but imperfect concordance., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Philip Supply reports a relationship with GenoScreen that includes: consulting or advisory. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Tuberculosis rapid molecular assays, including GeneXpert MTB/RIF® and Loopamp MTBC Detection Kit®, are highly sensitive and specific. Such performance does not automatically translate in improved disease control and highly depends on their use, local epidemiology and the diagnostic algorithms they're implemented within. We evaluate the performance of both assays and assess their impact on additional cases notification when implemented within WHO recommended tuberculosis diagnostic algorithms in Madagascar., Methods: Five hundred forty eight presumptive pulmonary tuberculosis patients were prospectively recruited between November 2013 and December 2014 in Antananarivo, Madagascar, a high TB incidence sub-Saharan African urban setting. Both molecular assays were evaluated as first line or add-on testing following negative smear microscopy. Based on locally defined assay performance characteristics we measure the impact of both assays and WHO-recommended diagnostic algorithms on additional tuberculosis case notifications., Results: High sensitivity and specificity was confirmed for both GeneXpert MTB/RIF® (86.6% (95% CI 81.1-90.7%) and 97.4% (95% CI 94.9-98.8%)) and Loopamp MTBC Detection Kit® (84.6% (95% CI 78.9-89.0%) and 98.4% (95% CI 96.2-99.4%)). Implementation of GeneXpert MTB/RIF® and Loopamp MTBC Detection Kit® increased tuberculosis diagnostic algorithms sensitivity from 73.6% (95% CI 67.1-79.3%) up to 88.1% (95% CI 82.8-91.9%). This increase was highest when molecular assays were used as add-on testing following negative smear microscopy. As add-on testing, GeneXpert MTB/RIF® and Loopamp MTBC Detection Kit® respectively improved case detection by 23.8 and 21.2% (p < 0.05)., Conclusion: Including GeneXpert MTB/RIF® or Loopamp MTBC Detection Kit® molecular assays for TB detection on sputum samples from presumptive TB cases can significantly increase case notification in TB diagnostic centers. The TB case detection rate is further increased when those tests are use as second-line follow-on testing following negative smear microscopy results. A country wide scale-up and digital integration of molecular-based TB diagnosis assays shows promises for TB control in Madagascar.

Background: The risk of tuberculosis outbreaks among people fleeing hardship for refuge in Europe is heightened. We describe the cross-border European response to an outbreak of multidrug-resistant tuberculosis among patients from the Horn of Africa and Sudan., Methods: On April 29 and May 30, 2016, the Swiss and German National Mycobacterial Reference Laboratories independently triggered an outbreak investigation after four patients were diagnosed with multidrug-resistant tuberculosis. In this molecular epidemiological study, we prospectively defined outbreak cases with 24-locus mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) profiles; phenotypic resistance to isoniazid, rifampicin, ethambutol, pyrazinamide, and capreomycin; and corresponding drug resistance mutations. We whole-genome sequenced all Mycobacterium tuberculosis isolates and clustered them using a threshold of five single nucleotide polymorphisms (SNPs). We collated epidemiological data from host countries from the European Centre for Disease Prevention and Control., Findings: Between Feb 12, 2016, and April 19, 2017, 29 patients were diagnosed with multidrug-resistant tuberculosis in seven European countries. All originated from the Horn of Africa or Sudan, with all isolates two SNPs or fewer apart. 22 (76%) patients reported their travel routes, with clear spatiotemporal overlap between routes. We identified a further 29 MIRU-VNTR-linked cases from the Horn of Africa that predated the outbreak, but all were more than five SNPs from the outbreak. However all 58 isolates shared a capreomycin resistance-associated tlyA mutation., Interpretation: Our data suggest that source cases are linked to an M tuberculosis clone circulating in northern Somalia or Djibouti and that transmission probably occurred en route before arrival in Europe. We hypothesise that the shared mutation of tlyA is a drug resistance mutation and phylogenetic marker, the first of its kind in M tuberculosis sensu stricto., Funding: The Swiss Federal Office of Public Health, the University of Zurich, the Wellcome Trust, National Institute for Health Research (NIHR) Oxford Biomedical Research Centre (BRC), the Medical Research Council, BELTA-TBnet, the European Union, the German Center for Infection Research, and Leibniz Science Campus Evolutionary Medicine of the Lung (EvoLUNG)., (Copyright © 2018 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)