Publications by authors named "Sebastien Gagneux"

173 Publications

Genomic analyses of Mycobacterium tuberculosis from human lung resections reveal a high frequency of polyclonal infections.

Nat Commun 2021 May 11;12(1):2716. Epub 2021 May 11.

Instituto de Biomedicina de Valencia IBV-CSIC, Valencia, Spain.

Polyclonal infections occur when at least two unrelated strains of the same pathogen are detected in an individual. This has been linked to worse clinical outcomes in tuberculosis, as undetected strains with different antibiotic resistance profiles can lead to treatment failure. Here, we examine the amount of polyclonal infections in sputum and surgical resections from patients with tuberculosis in the country of Georgia. For this purpose, we sequence and analyse the genomes of Mycobacterium tuberculosis isolated from the samples, acquired through an observational clinical study (NCT02715271). Access to the lung enhanced the detection of multiple strains (40% of surgery cases) as opposed to just using a sputum sample (0-5% in the general population). We show that polyclonal infections often involve genetically distant strains and can be associated with reversion of the patient's drug susceptibility profile over time. In addition, we find different patterns of genetic diversity within lesions and across patients, including mutational signatures known to be associated with oxidative damage; this suggests that reactive oxygen species may be acting as a selective pressure in the granuloma environment. Our results support the idea that the magnitude of polyclonal infections in high-burden tuberculosis settings is underestimated when only testing sputum samples.
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http://dx.doi.org/10.1038/s41467-021-22705-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8113332PMC
May 2021

High burden of tuberculosis infection and disease among people receiving medication-assisted treatment for substance use disorder in Tanzania.

PLoS One 2021 29;16(4):e0250038. Epub 2021 Apr 29.

Swiss Tropical and Public Health Institute, Basel, Switzerland.

Objective: To determine the prevalence of tuberculosis (TB) disease and infection as well as incident TB disease among people who use drugs (PWUD) attending Medication Assisted Treatment (MAT) clinics in Dar-es-Salaam, Tanzania.

Methods: In this prospective cohort study, a total of 901 consenting participants were enrolled from November 2016 to February 2017 and a structured questionnaire administered to them through the open data kit application on android tablets. Twenty-two months later, we revisited the MAT clinics and reviewed 823 of the 901 enrolled participant's medical records in search for documentation on TB disease diagnosis and treatment. Medical records reviewed included those of participants whom at enrolment were asymptomatic, not on TB disease treatment, not on TB preventive therapy and those who had a documented tuberculin skin test (TST) result.

Results: Of the 823 medical records reviewed 22 months after enrolment, 42 had documentation of being diagnosed with TB disease and initiated on TB treatment. This is equivalent to a TB disease incidence rate of 2,925.2 patients per 100,000 person years with a total follow up time of 1,440 person-years. At enrolment the prevalence of TB disease and TB infection was 2.6% and 54% respectively and the HIV prevalence was 44% and 16% among females and males respectively.

Conclusion: PWUD attending MAT clinics bear an extremely high burden of TB and HIV and are known to have driven TB epidemics in a number of countries. Our reported TB disease incidence is 12 times that of the general Tanzanian incidence of 237 per 100,000 further emphasizing that this group should be prioritized for TB screening, testing and treatment. Gender specific approaches should also be developed as female PWUDs are markedly more affected with HIV and TB disease than male PWUDs.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0250038PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8084249PMC
April 2021

Melting the Eis: non-detection of kanamycin resistance markers by routine diagnostic tests and identification of new -promoter variants.

Antimicrob Agents Chemother 2021 Apr 26. Epub 2021 Apr 26.

DSI-NRF Centre of Excellence for Biomedical Tuberculosis Research; South African Medical Research Council Centre for Tuberculosis Research; Division of Molecular Biology and Human Genetics, Faculty of Medicine and Health Sciences, Stellenbosch University, Cape Town.

-promoter mutations can confer reduced kanamycin susceptibility. GenoType MTBDR a widely used assay evaluating this region, wrongly classified 17/410 isolates as -promoter wildtype. 6/17 isolates harbored mutations known to confer kanamycin resistance, and the remainder harbored either novel -promoter mutations (7/11) or disputed mutations (4/11). GenoType MTBDR can miss established and new variants that cause reduced susceptibility. These data highlight the importance of reflex phenotypic kanamycin testing.
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http://dx.doi.org/10.1128/AAC.02502-20DOI Listing
April 2021

Local adaptation in populations of endemic to the Indian Ocean Rim.

F1000Res 2021 1;10:60. Epub 2021 Feb 1.

Institut Pasteur de Madagascar, Antananarivo, Madagascar.

Lineage 1 (L1) and 3 (L3) are two lineages of the complex (MTBC) causing tuberculosis (TB) in humans. L1 and L3 are prevalent around the rim of the Indian Ocean, the region that accounts for most of the world's new TB cases. Despite their relevance for this region, L1 and L3 remain understudied. We analyzed 2,938 L1 and 2,030 L3 whole genome sequences originating from 69 countries. We reconstructed the evolutionary history of these two lineages and identified genes under positive selection. We found a strongly asymmetric pattern of migration from South Asia toward neighboring regions, highlighting the historical role of South Asia in the dispersion of L1 and L3. Moreover, we found that several genes were under positive selection, including genes involved in virulence and resistance to antibiotics. For L1 we identified signatures of local adaptation at the locus, a gene coding for a secreted effector that targets the human endosomal sorting complex, and is included in several vaccine candidates. Our study highlights the importance of genetic diversity in the MTBC, and sheds new light on two of the most important MTBC lineages affecting humans.
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http://dx.doi.org/10.12688/f1000research.28318.2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7921886.2PMC
February 2021

Genomic epidemiological analysis identifies high relapse among individuals with recurring tuberculosis and provides evidence of recent household-related transmission of tuberculosis in Ghana.

Int J Infect Dis 2021 Mar 2;106:13-22. Epub 2021 Mar 2.

Noguchi Memorial Institute for Medical Research, College of Health Sciences, University of Ghana (UG), Ghana; West African Centre for Cell Biology of Infectious Pathogens, UG, Ghana.

Objective: To retrospectively investigate the cause of recurring tuberculosis (rcTB) among participants with pulmonary TB recruited from a prospective population-based study conducted between July 2012 and December 2015.

Methods: Mycobacterium tuberculosis complex isolates obtained from rcTB cases were characterized by standard mycobacterial genotyping tools, whole-genome sequencing, and phylogenetic analysis carried out to assess strain relatedness.

Results: The majority (58.3%, 21/36) of study participants with rcTB episodes had TB recurrence within 12 months post treatment. TB strains with isoniazid (INH) resistance were found in 19.4% (7/36) of participants at the primary episode, of which 29% (2/7) were also rifampicin-resistant. On TB recurrence, an INH-resistant strain was found in a larger proportion of participants, 27.8% (10/36), of which 40% (4/10) were MDR-TB strains. rcTB was attributed to relapse (same strain) in 75.0% (27/36) of participants and 25.0% (9/36) to re-infection.

Conclusion: Our findings indicate that previous unresolved infectiondue to inadequate treatment, may be the major cause of rcTB.
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http://dx.doi.org/10.1016/j.ijid.2021.02.110DOI Listing
March 2021

Phylogenomics of reveals a new lineage and a complex evolutionary history.

Microb Genom 2021 02;7(2)

University of Basel, Basel, Switzerland.

Human tuberculosis (TB) is caused by members of the complex (MTBC). The MTBC comprises several human-adapted lineages known as , as well as two lineages (L5 and L6) traditionally referred to as . Strains of L5 and L6 are largely limited to West Africa for reasons unknown, and little is known of their genomic diversity, phylogeography and evolution. Here, we analysed the genomes of 350 L5 and 320 L6 strains, isolated from patients from 21 African countries, plus 5 related genomes that had not been classified into any of the known MTBC lineages. Our population genomic and phylogeographical analyses showed that the unclassified genomes belonged to a new group that we propose to name MTBC lineage 9 (L9). While the most likely ancestral distribution of L9 was predicted to be East Africa, the most likely ancestral distribution for both L5 and L6 was the Eastern part of West Africa. Moreover, we found important differences between L5 and L6 strains with respect to their phylogeographical substructure and genetic diversity. Finally, we could not confirm the previous association of drug-resistance markers with lineage and sublineages. Instead, our results indicate that the association of drug resistance with lineage is most likely driven by sample bias or geography. In conclusion, our study sheds new light onto the genomic diversity and evolutionary history of , and highlights the need to consider the particularities of each MTBC lineage for understanding the ecology and epidemiology of TB in Africa and globally.
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http://dx.doi.org/10.1099/mgen.0.000477DOI Listing
February 2021

Evaluation of drug susceptibility profile of Mycobacterium tuberculosis Lineage 1 from Brazil based on whole genome sequencing and phenotypic methods.

Mem Inst Oswaldo Cruz 2021 29;115:e200520. Epub 2021 Jan 29.

Fundação Oswaldo Cruz-Fiocruz, Instituto Oswaldo Cruz, Laboratório de Biologia Molecular Aplicada a Micobactérias, Rio de Janeiro, RJ, Brasil.

Background: The evaluation of procedures for drug susceptibility prediction of Mycobacterium tuberculosis based on genomic data against the conventional reference method test based on culture is realistic considering the scenario of growing number of tools proposals based on whole-genome sequences (WGS).

Objectives: This study aimed to evaluate drug susceptibility testing (DST) outcome based on WGS tools and the phenotypic methods performed on isolates of M. tuberculosis Lineage 1 from the state of Pará, Brazil, generally associated with low levels of drug resistance.

Methodology: Culture based DST was performed using the Proportion Method in Löwenstein-Jensen medium on 71 isolates that had been submitted to WGS. We analysed the seven main genome sequence-based tools for resistance and lineage prediction applied to M. tuberculosis and for comparison evaluation we have used the Kappa concordance test.

Findings: When comparing the WGS-based tools against the DST, we observed the highest level of agreement using TB-profiler. Among the tools, TB-profiler, KvarQ and Mykrobe were those which identified the largest number of TB-MDR cases. Comparing the four most sensitive tools regarding resistance prediction, agreement was observed for 43 genomes.

Main Conclusions: Drug resistance profiling using next-generation sequencing offers rapid assessment of resistance-associated mutations, therefore facilitating rapid access to effective treatment.
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http://dx.doi.org/10.1590/0074-02760200520DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7849176PMC
February 2021

The within-host evolution of antimicrobial resistance in Mycobacterium tuberculosis.

FEMS Microbiol Rev 2020 Dec 15. Epub 2020 Dec 15.

Swiss Tropical and Public Health Institute, Basel, Switzerland.

Tuberculosis (TB) has been responsible for the greatest number of human deaths due to an infectious disease in general, and due to antimicrobial resistance (AMR) in particular. The etiological agents of human TB are a closely-related group of human-adapted bacteria that belong to the Mycobacterium tuberculosis complex (MTBC). Understanding how MTBC populations evolve within-host may allow for improved TB treatment and control strategies. In this Review, we highlight recent works that have shed light on how AMR evolves in MTBC populations within individual patients. We discuss the role of heteroresistance in AMR evolution, and review the bacterial, patient, and environmental factors that likely modulate the magnitude of heteroresistance within-host. We further highlight recent works on the dynamics of MTBC genetic diversity within-host, and discuss how spatial substructures in patients' lungs, spatiotemporal heterogeneity in antimicrobial concentrations, and phenotypic drug tolerance likely modulates the dynamics of MTBC genetic diversity in patients during treatment. We note the general characteristics that are shared between how the MTBC and other bacterial pathogens evolve in humans, and highlight the characteristics unique to the MTBC.
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http://dx.doi.org/10.1093/femsre/fuaa071DOI Listing
December 2020

Natural Polymorphisms in Mycobacterium tuberculosis Conferring Resistance to Delamanid in Drug-Naive Patients.

Antimicrob Agents Chemother 2020 10 20;64(11). Epub 2020 Oct 20.

Institute of Medical Microbiology, University of Zurich, Zurich, Switzerland

Mutations in the genes of the F signaling pathway of complex, including , , , , , and , can lead to delamanid resistance. We searched for such mutations among 129 strains from Asia, South America, and Africa using whole-genome sequencing; 70 (54%) strains had at least one mutation in one of the genes. For 10 strains with mutations, we determined the MIC of delamanid. We found one strain from a delamanid-naive patient carrying the natural polymorphism Tyr29del () that was associated with a critical delamanid MIC.
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http://dx.doi.org/10.1128/AAC.00513-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7577131PMC
October 2020

HIV Coinfection Is Associated with Low-Fitness Variants in Rifampicin-Resistant Mycobacterium tuberculosis.

Antimicrob Agents Chemother 2020 09 21;64(10). Epub 2020 Sep 21.

Swiss Tropical and Public Health Institute, Basel, Switzerland

We analyzed 312 drug-resistant genomes of isolates collected from HIV-coinfected and HIV-negative TB patients from nine countries with a high tuberculosis burden. We found that rifampicin-resistant strains isolated from HIV-coinfected patients carried disproportionally more resistance-conferring mutations in that are associated with a low fitness in the absence of the drug, suggesting these low-fitness variants can thrive in the context of reduced host immunity.
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http://dx.doi.org/10.1128/AAC.00782-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7508592PMC
September 2020

Multiple Merger Genealogies in Outbreaks of Mycobacterium tuberculosis.

Mol Biol Evol 2021 01;38(1):290-306

Department of Plant Biodiversity and Breeding Informatics, Institute of Plant Breeding, Seed Science and Population Genetics, University of Hohenheim, Stuttgart, Germany.

The Kingman coalescent and its developments are often considered among the most important advances in population genetics of the last decades. Demographic inference based on coalescent theory has been used to reconstruct the population dynamics and evolutionary history of several species, including Mycobacterium tuberculosis (MTB), an important human pathogen causing tuberculosis. One key assumption of the Kingman coalescent is that the number of descendants of different individuals does not vary strongly, and violating this assumption could lead to severe biases caused by model misspecification. Individual lineages of MTB are expected to vary strongly in reproductive success because 1) MTB is potentially under constant selection due to the pressure of the host immune system and of antibiotic treatment, 2) MTB undergoes repeated population bottlenecks when it transmits from one host to the next, and 3) some hosts show much higher transmission rates compared with the average (superspreaders). Here, we used an approximate Bayesian computation approach to test whether multiple-merger coalescents (MMC), a class of models that allow for large variation in reproductive success among lineages, are more appropriate models to study MTB populations. We considered 11 publicly available whole-genome sequence data sets sampled from local MTB populations and outbreaks and found that MMC had a better fit compared with the Kingman coalescent for 10 of the 11 data sets. These results indicate that the null model for analyzing MTB outbreaks should be reassessed and that past findings based on the Kingman coalescent need to be revisited.
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http://dx.doi.org/10.1093/molbev/msaa179DOI Listing
January 2021

A sister lineage of the Mycobacterium tuberculosis complex discovered in the African Great Lakes region.

Nat Commun 2020 06 9;11(1):2917. Epub 2020 Jun 9.

Univ. Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019 - UMR 9017, CIIL - Center for Infection and Immunity of Lille, 59000, Lille, France.

The human- and animal-adapted lineages of the Mycobacterium tuberculosis complex (MTBC) are thought to have expanded from a common progenitor in Africa. However, the molecular events that accompanied this emergence remain largely unknown. Here, we describe two MTBC strains isolated from patients with multidrug resistant tuberculosis, representing an as-yet-unknown lineage, named Lineage 8 (L8), seemingly restricted to the African Great Lakes region. Using genome-based phylogenetic reconstruction, we show that L8 is a sister clade to the known MTBC lineages. Comparison with other complete mycobacterial genomes indicate that the divergence of L8 preceded the loss of the cobF genome region - involved in the cobalamin/vitamin B12 synthesis - and gene interruptions in a subsequent common ancestor shared by all other known MTBC lineages. This discovery further supports an East African origin for the MTBC and provides additional molecular clues on the ancestral genome reduction associated with adaptation to a pathogenic lifestyle.
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http://dx.doi.org/10.1038/s41467-020-16626-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7283319PMC
June 2020

Whole Genome Sequencing and Spatial Analysis Identifies Recent Tuberculosis Transmission Hotspots in Ghana.

Front Med (Lausanne) 2020 19;7:161. Epub 2020 May 19.

Noguchi Memorial Institute for Medical Research, College of Health Sciences, University of Ghana, Accra, Ghana.

Whole genome sequencing (WGS) is progressively being used to investigate the transmission dynamics of complex (MTBC). We used WGS analysis to resolve traditional genotype clusters and explored the spatial distribution of confirmed recent transmission clusters. Bacterial genomes from a total of 452 MTBC isolates belonging to large traditional clusters from a population-based study spanning July 2012 and December 2015 were obtained through short read next-generation sequencing using the illumina HiSeq2500 platform. We performed clustering and spatial analysis using specified R packages and ArcGIS. Of the 452 traditional genotype clustered genomes, 314 (69.5%) were confirmed clusters with a median cluster size of 7.5 genomes and an interquartile range of 4-12. Recent tuberculosis (TB) transmission was estimated as 24.7%. We confirmed the wide spread of a Cameroon sub-lineage clone with a cluster size of 78 genomes predominantly from the Ablekuma sub-district of Accra metropolis. More importantly, we identified a recent transmission cluster associated with isoniazid resistance belonging to the Ghana sub-lineage of lineage 4. WGS was useful in detecting unsuspected outbreaks; hence, we recommend its use not only as a research tool but as a surveillance tool to aid in providing the necessary guided steps to track, monitor, and control TB.
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http://dx.doi.org/10.3389/fmed.2020.00161DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7248928PMC
May 2020

Effect of history of tuberculosis on specificity of Xpert MTB/RIF.

Eur Respir J 2020 09 24;56(3). Epub 2020 Sep 24.

Foundation for Innovative New Diagnostics, Geneva, Switzerland.

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http://dx.doi.org/10.1183/13993003.00343-2020DOI Listing
September 2020

Interaction between host genes and Mycobacterium tuberculosis lineage can affect tuberculosis severity: Evidence for coevolution?

PLoS Genet 2020 04 30;16(4):e1008728. Epub 2020 Apr 30.

Department of Population and Quantitative Health Sciences, Case Western Reserve University School of Medicine, Cleveland, Ohio, United States of America.

Genetic studies of both the human host and Mycobacterium tuberculosis (MTB) demonstrate independent association with tuberculosis (TB) risk. However, neither explains a large portion of disease risk or severity. Based on studies in other infectious diseases and animal models of TB, we hypothesized that the genomes of the two interact to modulate risk of developing active TB or increasing the severity of disease, when present. We examined this hypothesis in our TB household contact study in Kampala, Uganda, in which there were 3 MTB lineages of which L4-Ugandan (L4.6) is the most recent. TB severity, measured using the Bandim TBscore, was modeled as a function of host SNP genotype, MTB lineage, and their interaction, within two independent cohorts of TB cases, N = 113 and 121. No association was found between lineage and severity, but association between multiple polymorphisms in IL12B and TBscore was replicated in two independent cohorts (most significant rs3212227, combined p = 0.0006), supporting previous associations of IL12B with TB susceptibility. We also observed significant interaction between a single nucleotide polymorphism (SNP) in SLC11A1 and the L4-Ugandan lineage in both cohorts (rs17235409, meta p = 0.0002). Interestingly, the presence of the L4-Uganda lineage in the presence of the ancestral human allele associated with more severe disease. These findings demonstrate that IL12B is associated with severity of TB in addition to susceptibility, and that the association between TB severity and human genetics can be due to an interaction between genes in the two species, consistent with host-pathogen coevolution in TB.
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http://dx.doi.org/10.1371/journal.pgen.1008728DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7217476PMC
April 2020

Mycobacterium tuberculosis associated with severe tuberculosis evades cytosolic surveillance systems and modulates IL-1β production.

Nat Commun 2020 04 23;11(1):1949. Epub 2020 Apr 23.

i3S-Instituto de Investigação e Inovação em Saúde, University of Porto, Porto, Portugal.

Genetic diversity of Mycobacterium tuberculosis affects immune responses and clinical outcomes of tuberculosis (TB). However, how bacterial diversity orchestrates immune responses to direct distinct TB severities is unknown. Here we study 681 patients with pulmonary TB and show that M. tuberculosis isolates from cases with mild disease consistently induce robust cytokine responses in macrophages across multiple donors. By contrast, bacteria from patients with severe TB do not do so. Secretion of IL-1β is a good surrogate of the differences observed, and thus to classify strains as probable drivers of different TB severities. Furthermore, we demonstrate that M. tuberculosis isolates that induce low levels of IL-1β production can evade macrophage cytosolic surveillance systems, including cGAS and the inflammasome. Isolates exhibiting this evasion strategy carry candidate mutations, generating sigA recognition boxes or affecting components of the ESX-1 secretion system. Therefore, we provide evidence that M. tuberculosis strains manipulate host-pathogen interactions to drive variable TB severities.
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http://dx.doi.org/10.1038/s41467-020-15832-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7181847PMC
April 2020

Model-based integration of genomics and metabolomics reveals SNP functionality in .

Proc Natl Acad Sci U S A 2020 04 30;117(15):8494-8502. Epub 2020 Mar 30.

Institute of Molecular Systems Biology, ETH Zurich, 8093 Zurich, Switzerland

Human tuberculosis is caused by members of the complex (MTBC) that vary in virulence and transmissibility. While genome-wide association studies have uncovered several mutations conferring drug resistance, much less is known about the factors underlying other bacterial phenotypes. Variation in the outcome of tuberculosis infection and diseases has been attributed primarily to patient and environmental factors, but recent evidence indicates an additional role for the genetic diversity among MTBC clinical strains. Here, we used metabolomics to unravel the effect of genetic variation on the strain-specific metabolic adaptive capacity and vulnerability. To define the functionality of single-nucleotide polymorphisms (SNPs) systematically, we developed a constraint-based approach that integrates metabolomic and genomic data. Our model-based predictions correctly classify SNP effects in pyruvate kinase and suggest a genetic basis for strain-specific inherent baseline susceptibility to the antibiotic -aminosalicylic acid. Our method is broadly applicable across microbial life, opening possibilities for the development of more selective treatment strategies.
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http://dx.doi.org/10.1073/pnas.1915551117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7165461PMC
April 2020

An African origin for .

Evol Med Public Health 2020 31;2020(1):49-59. Epub 2020 Jan 31.

Molecular Parasitology and Infection Biology, Swiss Tropical and Public Health Institute, Basel, Switzerland.

Background And Objectives: and are two of the most important agents of tuberculosis in livestock and the most important causes of zoonotic tuberculosis in humans. However, little is known about the global population structure, phylogeography and evolutionary history of these pathogens.

Methodology: We compiled a global collection of 3364 whole-genome sequences from and originating from 35 countries and inferred their phylogenetic relationships, geographic origins and age.

Results: Our results resolved the phylogenetic relationship among the four previously defined clonal complexes of , and another eight newly described here. Our phylogeographic analysis showed that likely originated in East Africa. While some groups remained restricted to East and West Africa, others have subsequently dispersed to different parts of the world.

Conclusions And Implications: Our results allow a better understanding of the global population structure of and its evolutionary history. This knowledge can be used to define better molecular markers for epidemiological investigations of in settings where whole-genome sequencing cannot easily be implemented.

Lay Summary: During the last few years, analyses of large globally representative collections of whole-genome sequences (WGS) from the human-adapted Mycobacterium tuberculosis complex (MTBC) lineages have enhanced our understanding of the global population structure, phylogeography and evolutionary history of these pathogens. In contrast, little corresponding data exists for M. bovis, the most important agent of tuberculosis in livestock. Using whole-genome sequences of globally distributed M. bovis isolates, we inferred the genetic relationships among different M. bovis genotypes distributed around the world. The most likely origin of M. bovis is East Africa according to our inferences. While some M. bovis groups remained restricted to East and West Africa, others have subsequently dispersed to different parts of the world driven by cattle movements.
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http://dx.doi.org/10.1093/emph/eoaa005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7081938PMC
January 2020

Dispersal of Driven by Historical European Trade in the South Pacific.

Front Microbiol 2019 4;10:2778. Epub 2019 Dec 4.

School of Science, University of Waikato, Hamilton, New Zealand.

() is a globally distributed bacterial pathogen whose population structure has largely been shaped by the activities of its obligate human host. Oceania was the last major global region to be reached by Europeans and is the last region for which the dispersal and evolution of remains largely unexplored. Here, we investigated the evolutionary history of the Euro-American L4.4 sublineage and its dispersal to the South Pacific. Using a phylodynamics approach and a dataset of 236 global L4.4 genomes we have traced the origins and dispersal of L4.4 strains to New Zealand. These strains are predominantly found in indigenous Māori and Pacific people and we identify a clade of European, likely French, origin that is prevalent in indigenous populations in both New Zealand and Canada. Molecular dating suggests the expansion of European trade networks in the early 19th century drove the dispersal of this clade to the South Pacific. We also identify historical and social factors within the region that have contributed to the local spread and expansion of these strains, including recent Pacific migrations to New Zealand and the rapid urbanization of Māori in the 20th century. Our results offer new insight into the expansion and dispersal of in the South Pacific and provide a striking example of the role of historical European migrations in the global dispersal of .
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http://dx.doi.org/10.3389/fmicb.2019.02778DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6915100PMC
December 2019

Classifying recurrent Mycobacterium tuberculosis cases in Georgia using MIRU-VNTR typing.

PLoS One 2019 18;14(10):e0223610. Epub 2019 Oct 18.

Swiss Tropical and Public Health Institute, Basel, Switzerland.

Introduction: Recurrent tuberculosis (TB) is one of the main challenges in TB control. Genotyping based on Mycobacterial Interspersed Repetitive Units-Variable Tandem Repeats (MIRU-VNTR) has been widely used to differentiate between relapse and reinfection, which are the two main causes of recurrent TB. There is a lack of data regarding the causes of TB recurrence in Georgia, and while differentiating between relapse and reinfection plays a key role in defining appropriate interventions, the required genotyping methodologies have not been implemented. The objective of this study was to implement MIRU-VNTR genotyping at the National Center for Tuberculosis and Lung Diseases (NCTBLD) and differentiate between relapse and reinfection in multidrug resistant (MDR-) TB patients from Tbilisi, Georgia.

Methods: Recurrent MDR tuberculosis cases from 2014-2016 diagnosed at NCTLD were included in the study when bacterial samples from both episodes were available. Genotyping based on the MIRU-VNTR 24 loci was implemented and used for differentiating between relapse and reinfection. Paired samples showing the same MIRU-VNTR pattern or one locus difference were classified as relapse, while two and more loci differences were treated as reinfection. Exact logistic regression was used to identify predictors of recurrence.

Results: Thirty two MDR-TB patients (64 samples) were included and MIRU-VNTR 24 typing was performed on the corresponding paired samples. Of the 32 patients, 25 (83.3%) were identified as relapse while 5 (16.7%) were due to re-infection. Patients with a history of incarceration were significantly associated with TB reinfection (p< 0.05).

Conclusion: Recurrent TB in MDR patients in Georgia are mainly caused by relapse, raising concerns on the efficacy of the TB control program. An association between incarceration and reinfection likely reflects high levels of ongoing TB transmission in prisons, indicating the need for better TB infection control measures in these settings. Our results add to the rationale for implementing genotypic surveillance of TB more broadly to support TB control in Georgia.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0223610PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6799914PMC
March 2020

The Genetic Background Modulates the Evolution of Fluoroquinolone-Resistance in Mycobacterium tuberculosis.

Mol Biol Evol 2020 Jan;37(1):195-207

Swiss Tropical and Public Health Institute, Basel, Switzerland.

Fluoroquinolones (FQ) form the backbone in experimental treatment regimens against drug-susceptible tuberculosis. However, little is known on whether the genetic variation present in natural populations of Mycobacterium tuberculosis (Mtb) affects the evolution of FQ-resistance (FQ-R). To investigate this question, we used nine genetically distinct drug-susceptible clinical isolates of Mtb and measured their frequency of resistance to the FQ ofloxacin (OFX) in vitro. We found that the Mtb genetic background led to differences in the frequency of OFX-resistance (OFX-R) that spanned two orders of magnitude and substantially modulated the observed mutational profiles for OFX-R. Further, in vitro assays showed that the genetic background also influenced the minimum inhibitory concentration and the fitness effect conferred by a given OFX-R mutation. To test the clinical relevance of our in vitro work, we surveyed the mutational profile for FQ-R in publicly available genomic sequences from clinical Mtb isolates, and found substantial Mtb lineage-dependent variability. Comparison of the clinical and the in vitro mutational profiles for FQ-R showed that 51% and 39% of the variability in the clinical frequency of FQ-R gyrA mutation events in Lineage 2 and Lineage 4 strains, respectively, can be attributed to how Mtb evolves FQ-R in vitro. As the Mtb genetic background strongly influenced the evolution of FQ-R in vitro, we conclude that the genetic background of Mtb also impacts the evolution of FQ-R in the clinic.
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http://dx.doi.org/10.1093/molbev/msz214DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6984360PMC
January 2020

The molecular clock of Mycobacterium tuberculosis.

PLoS Pathog 2019 09 12;15(9):e1008067. Epub 2019 Sep 12.

Department of Medical Parasitology and Infection Biology, Swiss Tropical and Public Health Institute, Basel, Switzerland.

The molecular clock and its phylogenetic applications to genomic data have changed how we study and understand one of the major human pathogens, Mycobacterium tuberculosis (MTB), the etiologic agent of tuberculosis. Genome sequences of MTB strains sampled at different times are increasingly used to infer when a particular outbreak begun, when a drug-resistant clone appeared and expanded, or when a strain was introduced into a specific region. Despite the growing importance of the molecular clock in tuberculosis research, there is a lack of consensus as to whether MTB displays a clocklike behavior and about its rate of evolution. Here we performed a systematic study of the molecular clock of MTB on a large genomic data set (6,285 strains), covering different epidemiological settings and most of the known global diversity. We found that sampling times below 15-20 years were often insufficient to calibrate the clock of MTB. For data sets where such calibration was possible, we obtained a clock rate between 1x10-8 and 5x10-7 nucleotide changes per-site-per-year (0.04-2.2 SNPs per-genome-per-year), with substantial differences between clades. These estimates were not strongly dependent on the time of the calibration points as they changed only marginally when we used epidemiological isolates (sampled in the last 40 years) or three ancient DNA samples (about 1,000 years old) to calibrate the tree. Additionally, the uncertainty and the discrepancies in the results of different methods were sometimes large, highlighting the importance of using different methods, and of considering carefully their assumptions and limitations.
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http://dx.doi.org/10.1371/journal.ppat.1008067DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6759198PMC
September 2019

Genetic variability and consequence of Mycobacterium tuberculosis lineage 3 in Kampala-Uganda.

PLoS One 2019 9;14(9):e0221644. Epub 2019 Sep 9.

Department of Medical Microbiology, College of Health Sciences, Makerere University, Kampala, Uganda.

Background: Limited data existed exclusively describing Mycobacterium tuberculosis lineage 3 (MTB-L3), sub-lineages, and clinical manifestations in Kampala, Uganda. This study sought to elucidate the circulating MTB-L3 sub-lineages and their corresponding clinical phenotypes.

Method: A total of 141 M. tuberculosis isolates were identified as M. tuberculosis lineage 3 using Single nucleotide polymorphism (SNP) marker analysis method. To ascertain the sub-lineages/sub-strains within the M. tuberculosis lineage 3, the direct repeat (DR) loci for all the isolates was examined for sub-lineage specific signatures as described in the SITVIT2 database. The infecting sub-strains were matched with patients' clinical and demographic characteristics to identify any possible association.

Result: The data showed 3 sub-lineages circulating with CAS 1 Delhi accounting for 55% (77/141), followed by CAS 1-Kili 16% (22/141) and CAS 2/CAS 8% (12/141). Remaining isolates 21% (30/141) were unclassifiable. To explore whether the sub-lineages differ in their ability to cause increased severe disease, we used extent of lung involvement as a proxy for severe disease. Multivariable analysis showed no association between M. tuberculosis lineage 3 sub-lineages with severe disease. The risk factors associated with severe disease include having a positive smear (OR = 9.384; CI 95% = 2.603-33.835), HIV (OR = 0.316; CI 95% = 0.114-0.876), lymphadenitis (OR = 0. 171; CI 95% = 0.034-0.856) and a BCG scar (OR = 0.295; CI 95% = 0.102-0.854).

Conclusion: In Kampala, Uganda, there are three sub-lineages of M. tuberculosis lineage 3 that cause disease of comparable severity with CAS-Dehli as the most prevalent. Having HIV, lymphadenitis, a BCG scar and a smear negative status is associated with reduced severe disease.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0221644PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6733460PMC
March 2020

Genome-wide mutational biases fuel transcriptional diversity in the Mycobacterium tuberculosis complex.

Nat Commun 2019 09 5;10(1):3994. Epub 2019 Sep 5.

Instituto de Biomedicina de Valencia, IBV-CSIC, Valencia, Spain.

The Mycobacterium tuberculosis complex (MTBC) members display different host-specificities and virulence phenotypes. Here, we have performed a comprehensive RNAseq and methylome analysis of the main clades of the MTBC and discovered unique transcriptional profiles. The majority of genes differentially expressed between the clades encode proteins involved in host interaction and metabolic functions. A significant fraction of changes in gene expression can be explained by positive selection on single mutations that either create or disrupt transcriptional start sites (TSS). Furthermore, we show that clinical strains have different methyltransferases inactivated and thus different methylation patterns. Under the tested conditions, differential methylation has a minor direct role on transcriptomic differences between strains. However, disruption of a methyltransferase in one clinical strain revealed important expression differences suggesting indirect mechanisms of expression regulation. Our study demonstrates that variation in transcriptional profiles are mainly due to TSS mutations and have likely evolved due to differences in host characteristics.
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http://dx.doi.org/10.1038/s41467-019-11948-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6728331PMC
September 2019

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

Nat Rev Microbiol 2019 09;17(9):533-545

Foundation for Innovative New Diagnostics, Geneva, Switzerland.

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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http://dx.doi.org/10.1038/s41579-019-0214-5DOI Listing
September 2019

Mycobacterium tuberculosis lineage 1 genetic diversity in Pará, Brazil, suggests common ancestry with east-African isolates potentially linked to historical slave trade.

Infect Genet Evol 2019 09 3;73:337-341. Epub 2019 Jun 3.

Institut de Biologie Intégrative de la Cellule, I2BC, UMR9198, CEA, CNRS, Univ. Paris-Sud, Univ. Paris-Saclay, 91198 Gif-sur-Yvette cedex, France; Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto-SP, Brazil. Electronic address:

Lineage 1 (L1) is one of seven Mycobacterium tuberculosis complex (MTBC) lineages. The objective of this study was to improve the complex taxonomy of L1 using phylogenetic SNPs, and to look for the origin of the main L1 sublineage prevalent in Para, Brazil. We developed a high-throughput SNPs-typing assay based on 12-L1-specific SNPs. This assay allowed us to experimentally retrieve SNP patterns on nine of these twelve SNPs in 277 isolates previously tentatively assigned to L1 spoligotyping-based sublineages. Three collections were used: Pará-Brazil (71); RIVM, the Netherlands (102), Madagascar (104). One-hundred more results were generated in Silico using the PolyTB database. Based on the final SNPs combination, the samples were classified into 11 clusters (C1-C11). Most isolates within a SNP-based cluster shared a mutual spoligotyping-defined lineage. However, L1/EAI1-SOM (SIT48) and L1/EAI6-BGD1 (SIT591) showed a poor correlation with SNP data and are not monophyletic. L1/EAI8-MDG and L1/EAI3-IND belonged to C5; this result suggests that they share a common ancestor. L1.1.3/SIT129, a spoligotype pattern found in SNPs-cluster C6, was found to be shared between Pará/Brazil and Malawi. SIT129 was independently found to be highly prevalent in Mozambique, which suggests a migration history from East-Africa to Brazil during the 16th-18th slave trade period to Northern Brazil.
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http://dx.doi.org/10.1016/j.meegid.2019.06.001DOI Listing
September 2019

Transition bias influences the evolution of antibiotic resistance in Mycobacterium tuberculosis.

PLoS Biol 2019 05 13;17(5):e3000265. Epub 2019 May 13.

Institute of Integrative Biology, ETH Zurich, Switzerland.

Transition bias, an overabundance of transitions relative to transversions, has been widely reported among studies of the rates and spectra of spontaneous mutations. However, demonstrating the role of transition bias in adaptive evolution remains challenging. In particular, it is unclear whether such biases direct the evolution of bacterial pathogens adapting to treatment. We addressed this challenge by analyzing adaptive antibiotic-resistance mutations in the major human pathogen Mycobacterium tuberculosis (MTB). We found strong evidence for transition bias in two independently curated data sets comprising 152 and 208 antibiotic-resistance mutations. This was true at the level of mutational paths (distinct adaptive DNA sequence changes) and events (individual instances of the adaptive DNA sequence changes) and across different genes and gene promoters conferring resistance to a diversity of antibiotics. It was also true for mutations that do not code for amino acid changes (in gene promoters and the 16S ribosomal RNA gene rrs) and for mutations that are synonymous to each other and are therefore likely to have similar fitness effects, suggesting that transition bias can be caused by a bias in mutation supply. These results point to a central role for transition bias in determining which mutations drive adaptive antibiotic resistance evolution in a key pathogen.
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http://dx.doi.org/10.1371/journal.pbio.3000265DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6532934PMC
May 2019

Pathways and associated costs of care in patients with confirmed and presumptive tuberculosis in Tanzania: A cross-sectional study.

BMJ Open 2019 04 20;9(4):e025079. Epub 2019 Apr 20.

Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland.

Objective: To assess pathways and associated costs of seeking care from the onset of symptoms to diagnosis in patients with confirmed and presumptive tuberculosis (TB).

Design: Cross-sectional study.

Setting: District hospital in Dar es Salaam, Tanzania.

Participants: Bacteriologically confirmed TB and presumptive TB patients.

Primary And Secondary Outcome Measures: We calculated distance in metres and visualised pathways to healthcare up to five visits for the current episode of sickness. Costs were described by medians and IQRs, with comparisons by gender and poverty status.

Results: Of 100 confirmed and 100 presumptive TB patients, 44% of confirmed patients sought care first at pharmacies after the onset of symptoms, and 42% of presumptive patients did so at hospitals. The median visits made by confirmed patients was 2 (range 1-5) and 2 (range 1-3) by presumptive patients. Patients spent a median of 31% of their monthly household income on health expenditures for all visits. The median total direct costs were higher in confirmed compared with presumptive patients (USD 27.4 [IQR 18.7-48.4] vs USD 19.8 [IQR 13.8-34.0], p=0.02), as were the indirect costs (USD 66.9 [IQR 35.5-150.0] vs USD 46.8 [IQR 20.1-115.3], p<0.001). The indirect costs were higher in men compared with women (USD 64.6 [IQR 31.8-159.1] vs USD 55.6 [IQR 25.1-141.1], p<0.001). The median total distance from patients' household to healthcare facilities for patients with confirmed and presumptive TB was 2338 m (IQR 1373-4122) and 2009 m (IQR 986-2976) respectively.

Conclusions: Patients with confirmed TB have complex pathways and higher costs of care compared with patients with presumptive TB, but the costs of the latter are also substantial. Improving access to healthcare and ensuring integration of different healthcare providers including private, public health practitioners and patients themselves could help in reducing the complex pathways during healthcare seeking and optimal healthcare utilisation.
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http://dx.doi.org/10.1136/bmjopen-2018-025079DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6528007PMC
April 2019