TY - JOUR T1 - Targeted Chromosomal Barcoding Establishes Direct Genotype-Phenotype Associations for Antibiotic Resistance in Mycobacterium abscessus. JF - Microbiol Spectr Y1 - 2023 A1 - Juan Calvet-Seral A1 - Estefanía Crespo-Yuste A1 - Vanessa Mathys A1 - Rodriguez-Villalobos, Hector A1 - Pieter-Jan Ceyssens A1 - Martin, Anandi A1 - Jesús Gonzalo-Asensio AB -

A bedaquiline-resistant Mycobacterium abscessus isolate was sequenced, and a candidate mutation in the gene was identified as responsible for the antibiotic resistance phenotype. To establish a direct genotype-phenotype relationship of this mutation which results in a Asp-to-Ala change at position 29 (D29A), we developed a recombineering-based method consisting of the specific replacement of the desired mutation in the bacterial chromosome. As surrogate bacteria, we used two M. abscessus bedaquiline-susceptible strains: ATCC 19977 and the SL541 clinical isolate. The allelic exchange substrates used in recombineering carried either the sole D29A mutation or a genetic barcode of silent mutations in codons flanking the D29A mutation. After selection of bedaquiline-resistant M. abscessus colonies transformed with both substrates, we obtained equivalent numbers of recombinants. These resistant colonies were analyzed by allele-specific PCR and Sanger sequencing, and we demonstrated that the presence of the genetic barcode was linked to the targeted incorporation of the desired mutation in its chromosomal location. All recombinants displayed the same MIC to bedaquiline as the original isolate, from which the D29A mutation was identified. Finally, to demonstrate the broad applicability of this method, we confirmed the association of bedaquiline resistance with the A64P mutation in analysis performed in independent M. abscessus strains and by independent researchers. Antimicrobial resistance (AMR) threatens the effective prevention and treatment of an ever-increasing range of infections caused by microorganisms. On the other hand, infections caused by affect people with chronic lung diseases, and their incidence has grown alarmingly in recent years. Further, these bacteria are known to easily develop AMR to the few therapeutic options available, making their treatment long-lasting and challenging. The recent introduction of new antibiotics against , such as bedaquiline, makes us anticipate a future when a plethora of antibiotic-resistant strains will be isolated and sequenced. However, in the era of whole-genome sequencing, one of the challenges is to unequivocally assign a biological function to each identified polymorphism. Thus, in this study, we developed a fast, robust, and reliable method to assign genotype-phenotype associations for putative antibiotic-resistant polymorphisms in .

M3 - 10.1128/spectrum.05344-22 ER - TY - JOUR T1 - Telacebec Interferes with Virulence Lipid Biosynthesis Protein Expression and Sensitizes to Other Antibiotics JF - Microorganisms Y1 - 2023 A1 - Zhou, Zhiyu A1 - Wattiez, Ruddy A1 - Patricia Constant A1 - Hedia Marrakchi A1 - Karine Soetaert A1 - Vanessa Mathys A1 - Véronique Fontaine A1 - Zeng, Sheng VL - 11 CP - 10 M3 - 10.3390/microorganisms11102469 ER - TY - JOUR T1 - The 2021 WHO catalogue of complex mutations associated with drug resistance: A genotypic analysis. JF - Lancet Microbe Y1 - 2022 A1 - Timothy M Walker A1 - Paolo Miotto A1 - Claudio U Köser A1 - Philip W Fowler A1 - Jeff Knaggs A1 - Iqbal, Zamin A1 - Martin Hunt A1 - Leonid Chindelevitch A1 - Maha Farhat A1 - Daniela Maria Cirillo A1 - Iñaki Comas A1 - James Posey A1 - Shaheed V Omar A1 - Timothy EA Peto A1 - Anita Suresh A1 - Swapna Uplekar A1 - Sacha Laurent A1 - Rebecca E Colman A1 - Carl-Michael Nathanson A1 - Matteo Zignol A1 - Ann Sarah Walker A1 - Derrick W Crook A1 - Nazir Ismail A1 - Timothy C Rodwell A1 - Vanessa Mathys KW - Drug Resistance KW - mutations KW - Tuberculosis AB -

Background: Molecular diagnostics are considered the most promising route to achieving rapid, universal drug susceptibility testing for complex (MTBC). We aimed to generate a WHO endorsed catalogue of mutations to serve as a global standard for interpreting molecular information for drug resistance prediction.

Methods: A candidate gene approach was used to identify mutations as associated with resistance, or consistent with susceptibility, for 13 WHO endorsed anti-tuberculosis drugs. 38,215 MTBC isolates with paired whole-genome sequencing and phenotypic drug susceptibility testing data were amassed from 45 countries. For each mutation, a contingency table of binary phenotypes and presence or absence of the mutation computed positive predictive value, and Fisher's exact tests generated odds ratios and Benjamini-Hochberg corrected p-values. Mutations were graded as Associated with Resistance if present in at least 5 isolates, if the odds ratio was >1 with a statistically significant corrected p-value, and if the lower bound of the 95% confidence interval on the positive predictive value for phenotypic resistance was >25%. A series of expert rules were applied for final confidence grading of each mutation.

Findings: 15,667 associations were computed for 13,211 unique mutations linked to one or more drugs. 1,149/15,667 (7·3%) mutations were classified as associated with phenotypic resistance and 107/15,667 (0·7%) were deemed consistent with susceptibility. For rifampicin, isoniazid, ethambutol, fluoroquinolones, and streptomycin, the mutations' pooled sensitivity was >80%. Specificity was over 95% for all drugs except ethionamide (91·4%), moxifloxacin (91·6%) and ethambutol (93·3%). Only two resistance mutations were classified for bedaquiline, delamanid, clofazimine, and linezolid as prevalence of phenotypic resistance was low for these drugs.

Interpretation: This first WHO endorsed catalogue of molecular targets for MTBC drug susceptibility testing provides a global standard for resistance interpretation. Its existence should encourage the implementation of molecular diagnostics by National Tuberculosis Programmes.

Funding: UNITAID, Wellcome, MRC, BMGF.

VL - 3 CP - 4 M3 - 10.1016/S2666-5247(21)00301-3 ER - TY - JOUR T1 - The small-molecule SMARt751 reverses resistance to ethionamide in acute and chronic mouse models of tuberculosis. JF - Sci Transl Med Y1 - 2022 A1 - Flipo, Marion A1 - Frita, Rosangela A1 - Marilyne Bourotte A1 - Maria S Martínez-Martínez A1 - Markus Boesche A1 - Gary W Boyle A1 - Geo Derimanov A1 - Gerard Drewes A1 - Pablo Gamallo A1 - Sonja Ghidelli-Disse A1 - Stephanie Gresham A1 - Elena Jiménez A1 - Jaime de Mercado A1 - Esther Pérez-Herrán A1 - Esther Porras-De Francisco A1 - Joaquín Rullas A1 - Patricia Casado A1 - Florence Leroux A1 - Catherine Piveteau A1 - Kiass, Mehdi A1 - Vanessa Mathys A1 - Karine Soetaert A1 - Véronique Megalizzi A1 - Tanina, Abdalkarim A1 - Wintjens, René A1 - Antoine, Rudy A1 - Brodin, Priscille A1 - Delorme, Vincent A1 - Moune, Martin A1 - Djaout, Kamel A1 - Stéphanie Slupek A1 - Kemmer, Christian A1 - Gitzinger, Marc A1 - Lluis Ballell A1 - Alfonso Mendoza-Losana A1 - Sergio Lociuro A1 - Déprez, Benoit A1 - David Barros-Aguirre A1 - Modesto J Remuiñán A1 - Willand, Nicolas A1 - Baulard, Alain R KW - Animals KW - Antitubercular Agents KW - Ethionamide KW - mice KW - Mycobacterium tuberculosis KW - Prodrugs KW - Tuberculosis AB -

The sensitivity of , the pathogen that causes tuberculosis (TB), to antibiotic prodrugs is dependent on the efficacy of the activation process that transforms the prodrugs into their active antibacterial moieties. Various oxidases of have the potential to activate the prodrug ethionamide. Here, we used medicinal chemistry coupled with a phenotypic assay to select the N-acylated 4-phenylpiperidine compound series. The lead compound, SMARt751, interacted with the transcriptional regulator VirS of , which regulates the operon encoding a monooxygenase that activates ethionamide. SMARt751 boosted the efficacy of ethionamide in vitro and in mouse models of acute and chronic TB. SMARt751 also restored full efficacy of ethionamide in mice infected with strains carrying mutations in the gene, which cause ethionamide resistance in the clinic. SMARt751 was shown to be safe in tests conducted in vitro and in vivo. A model extrapolating animal pharmacokinetic and pharmacodynamic parameters to humans predicted that as little as 25 mg of SMARt751 daily would allow a fourfold reduction in the dose of ethionamide administered while retaining the same efficacy and reducing side effects.

VL - 14 CP - 643 M3 - 10.1126/scitranslmed.aaz6280 ER - TY - JOUR T1 - A Bioinformatics Whole-Genome Sequencing Workflow for Clinical Mycobacterium tuberculosis Complex Isolate Analysis, Validated Using a Reference Collection Extensively Characterized with Conventional Methods and In Silico Approaches JF - J Clin Microbiol Y1 - 2021 A1 - Bert Bogaerts A1 - Thomas Delcourt A1 - Karine Soetaert A1 - Samira Boarbi A1 - Pieter-Jan Ceyssens A1 - Raf Winand A1 - Julien Van Braekel A1 - Sigrid C.J. De Keersmaecker A1 - Nancy Roosens A1 - Marchal, Kathleen A1 - Vanessa Mathys A1 - Kevin Vanneste KW - Antimicrobial resistance KW - Mycobacterium tuberculosis KW - national reference center KW - public health KW - single nucleotide polymorphism KW - VALIDATION KW - whole genome sequencing AB -

The use of whole genome sequencing (WGS) for routine typing of bacterial isolates has increased substantially in recent years. For (MTB), in particular, WGS has the benefit of drastically reducing the time to generate results compared to most conventional phenotypic methods. Consequently, a multitude of solutions for analyzing WGS MTB data have been developed, but their successful integration in clinical and national reference laboratories is hindered by the requirement for their validation, for which a consensus framework is still largely absent. We developed a bioinformatics workflow for (Illumina) WGS-based routine typing of MTB Complex (MTBC) member isolates allowing complete characterization including (sub)species confirmation and identification (16S, /RD, , Single Nucleotide Polymorphism (SNP)-based antimicrobial resistance (AMR) prediction, and pathogen typing (spoligotyping, SNP barcoding, and core genome MultiLocus Sequence Typing). Workflow performance was validated on a per-assay basis using a collection of 238 in-house sequenced MTBC isolates, extensively characterized with conventional molecular biology-based approaches supplemented with public data. For SNP-based AMR prediction, results from molecular genotyping methods were supplemented with modified datasets allowing to greatly increase the set of evaluated mutations. The workflow demonstrated very high performance with performance metrics >99% for all assays, except for spoligotyping where sensitivity dropped to ∼90%. The validation framework for our WGS-based bioinformatics workflow can aid standardization of bioinformatics tools by the MTB community and other SNP-based applications regardless of the targeted pathogen(s). The bioinformatics workflow is available for academic and non-profit usage through the Galaxy instance of our institute at https://galaxy.sciensano.be.

M3 - 10.1128/JCM.00202-21 ER - TY - JOUR T1 - Integrative transnational analysis to dissect tuberculosis transmission events along the migratory route from Africa to Europe. JF - J Travel Med Y1 - 2021 A1 - Miguel Martínez-Lirola A1 - Rana Jajou A1 - Vanessa Mathys A1 - Martin, Anandi A1 - Andrea Maurizio Cabibbe A1 - Ana Valera A1 - Pedro J Sola-Campoy A1 - Estefanía Abascal A1 - Sandra Rodríguez-Maus A1 - Jose Antonio Garrido-Cárdenas A1 - Magdalena Bonillo A1 - Álvaro Chiner-Oms A1 - Begoña López A1 - Silvia Vallejo-Godoy A1 - Iñaki Comas A1 - Patricia Muñoz A1 - Daniela Maria Cirillo A1 - Dick van Soolingen A1 - Laura Pérez-Lago A1 - Darío García de Viedma KW - Africa KW - Cluster Analysis KW - Europe KW - Genotype KW - Humans KW - Mycobacterium tuberculosis KW - Tuberculosis AB -

BACKGROUND: Growing international migration has increased the complexity of tuberculosis transmission patterns. Italy's decision to close its borders in 2018 made of Spain the new European porte entrée for migration from the Horn of Africa (HA). In one of the first rescues of migrants from this region at the end of 2018, tuberculosis was diagnosed in eight subjects, mainly unaccompanied minors.

METHODS: Mycobacterium tuberculosis isolates from these recently arrived migrants were analysed by Mycobacterial Interspersed Repetitive-Unit/Variable-Number of Tandem Repeat (MIRU-VNTR) and subsequent whole genome sequencing (WGS) analysis. Data were compared with those from collections from other European countries receiving migrants from the HA and a strain-specific PCR was applied for a fast searching of common strains. Infections in a cellular model were performed to assess strain virulence.

RESULTS: MIRU-VNTR analysis allowed identifying an epidemiological cluster involving three of the eight cases from Somalia (0 single-nucleotide polymorphisms between isolates, HA cluster). Following detailed interviews revealed that two of these cases had shared the same migratory route in most of the trip and had spent a long time at a detention camp in Libya. To confirm potential en route transmission for the three cases, we searched the same strain in collections from other European countries receiving migrants from the HA. MIRU-VNTR, WGS and a strain-specific PCR for the HA strain were applied. The same strain was identified in 12 cases from Eritrea diagnosed soon after their arrival in 2018 to the Netherlands, Belgium and Italy. Intracellular replication rate of the strain did not reveal abnormal virulence.

CONCLUSIONS: Our study suggests a potential en route transmission of a pan-susceptible strain, which caused at least 15 tuberculosis cases in Somalian and Eritrean migrants diagnosed in four different European countries.

VL - 28 CP - 4 M3 - 10.1093/jtm/taab054 ER - TY - JOUR T1 - Retrospective evaluation of routine whole genome sequencing of Mycobacterium tuberculosis at the Belgian National Reference Center, 2019. JF - Acta Clin Belg Y1 - 2021 A1 - Karine Soetaert A1 - Pieter-Jan Ceyssens A1 - Samira Boarbi A1 - Bert Bogaerts A1 - Thomas Delcourt A1 - Kevin Vanneste A1 - Sigrid C.J. De Keersmaecker A1 - Nancy Roosens A1 - Alexandra Vodolazkaia A1 - Marina Mukovnikova A1 - Vanessa Mathys AB -

OBJECTIVES: Since January 2019, the Belgian National Reference Center for Mycobacteria (NRC) has switched from conventional typing to prospective whole-genome sequencing (WGS) of all submitted complex (MTB) isolates. The ISO17025 validated procedure starts with semi-automated extraction and purification of gDNA directly from the submitted MGIT tubes, without preceding subculturing. All samples are then sequenced on an Illumina MiSeq sequencer and analyzed using an in-house developed and validated bioinformatics workflow to determine the species and antimicrobial resistance. In this study, we retrospectively compare results obtained via WGS to conventional phenotypic and genotypic testing, for all Belgian MTB strains analyzed in 2019 (n = 306).

RESULTS: In all cases, the WGS-based procedure was able to identify correctly the MTB species. Compared to MGIT drug susceptibility testing (DST), the sensitivity and specificity of genetic prediction of resistance to first-line antibiotics were respectively 100 and 99% (rifampicin, RIF), 90.5 and 100% (isoniazid, INH), 100 and 98% (ethambutol, EMB) and 61.1 and 100% (pyrazinamide, PZA). The negative predictive value was above 95% for these four first-line drugs. A positive predictive value of 100% was calculated for INH and PZA, 80% for RIF and 45% for EMB.

CONCLUSIONS: Our study confirms the effectiveness of WGS for the rapid detection of complex and its drug resistance profiles for first-line drugs even when working directly on MGIT tubes, and supports the introduction of this test into the routine workflow of laboratories performing tuberculosis diagnosis.

M3 - 10.1080/17843286.2021.1999588 ER - TY - JOUR T1 - Whole-genome sequencing for TB source investigations: principles of ethical precision public health. JF - Int J Tuberc Lung Dis Y1 - 2021 A1 - A Van Rie A1 - D G de Viedma A1 - C Meehan A1 - I Comas A1 - T H Heupink A1 - E De Vos A1 - W A de Oñate A1 - Vanessa Mathys A1 - Pieter-Jan Ceyssens A1 - Groenen, G A1 - F González-Candelas A1 - A Forier A1 - E Juengst AB -

Whole-genome sequencing (WGS) of allows rapid, accurate inferences about the sources, location and timing of transmission. However, in an era of heightened concern for personal privacy and science distrust, such inferences could result in unintended harm and undermine the public´s trust. We held interdisciplinary stakeholder discussions and performed ethical analyses of real-world illustrative cases to identify principles that optimise benefit and mitigate harm of WGS-driven TB source investigations. The speed and precision with which real-time WGS can be used to associate strains with sensitive information has raised important concerns. While detailed understanding of transmission events could mitigate harm to vulnerable patients and communities when otherwise unfairly blamed for TB outbreaks, the precision of WGS can also identify transmission events resulting in social blame, fear, discrimination, individual or location stigma, and the use of defaming language by the public, politicians and scientists. Public health programmes should balance the need to safeguard privacy with public health goals, transparency and individual rights, including the right to know who infects whom or where. Ethical challenges raised by real-time WGS-driven TB source investigation requires public health authorities to move beyond their current legal mandate and embrace transparency, privacy and community engagement.

VL - 25 CP - 3 M3 - 10.5588/ijtld.20.0886 ER - TY - JOUR T1 - Deep amplicon sequencing for culture-free prediction of susceptibility or resistance to 13 anti-tuberculous drugs. JF - Eur Respir J Y1 - 2020 A1 - Agathe Jouet A1 - Cyril Gaudin A1 - Nelly Badalato A1 - Allix-Béguec, Caroline A1 - Stéphanie Duthoy A1 - Alice Ferré A1 - Maren Diels A1 - Yannick Laurent A1 - Sandy Contreras A1 - Silke Feuerriegel A1 - Stefan Niemann A1 - Emmanuel André A1 - Michel K Kaswa A1 - Elisa Tagliani A1 - Andrea Cabibbe A1 - Vanessa Mathys A1 - Daniela Cirillo A1 - Bouke C de Jong A1 - Rigouts, Leen A1 - Supply, Philip AB -

Conventional molecular tests for detecting complex (MTBC) drug resistance on clinical samples cover a limited set of mutations. Whole genome sequencing (WGS) typically requires culture. Here, we evaluated the Deeplex Myc-TB targeted deep sequencing assay for prediction of resistance to 13 anti-tuberculous drugs/drug classes, directly applicable on sputum. With MTBC DNA tests, the limit of detection was 100-1000 genome copies for fixed resistance mutations. Deeplex Myc-TB captured in silico 97.1-99.3% of resistance phenotypes correctly predicted by WGS from 3651 MTBC genomes. On 429 isolates, the assay predicted 92.2% of 2369 first- and second-line phenotypes, with a sensitivity of 95.3% and specificity of 97.4%. Fifty-six of 69 (81.2%) residual discrepancies with phenotypic results involved pyrazinamide, ethambutol, and ethionamide, and low-level rifampicin- or isoniazid-resistance mutations, all notoriously prone to phenotypic testing variability. Only 2 of 91 (2.2%) resistance phenotypes undetected by Deeplex Myc-TB had known resistance-associated mutations by WGS analysis outside Deeplex Myc-TB targets. Phenotype predictions from Deeplex Myc-TB analysis directly on 109 sputa from a Djibouti survey matched those of MTBSeq/PhyResSE/Mykrobe, fed with WGS data from subsequent cultures, with a sensitivity of 93.5/98.5/93.1% and specificity of 98.5/97.2/95.3%. Most residual discordances involved gene deletions/indels and 3-12% heteroresistant calls undetected by WGS analysis, or natural pyrazinamide resistance of globally rare " strains then unreported by Deeplex Myc-TB. On 1494 arduous sputa from a Democratic Republic of the Congo survey, 14 902 of 19 422 (76.7%) possible susceptible or resistance phenotypes could be predicted culture-free. Deeplex Myc-TB may enable fast, tailored tuberculosis treatment.

M3 - 10.1183/13993003.02338-2020 ER - TY - JOUR T1 - Recurrent tuberculosis in a young child JF - Pediatr Infect Dis J Y1 - 2020 A1 - Alexandra Dreesman A1 - Oriane Stévart A1 - Catherine Adler A1 - Vanessa Mathys A1 - Françoise Mouchet AB -

A young child, 19 months of age, presented with a second episode of tuberculosis after full recovery from initial tuberculosis disease 6 months earlier. Mycobacterium tuberculosis strains isolated from both episodes were genotyped and differed from one another. We present the first case of proven tuberculosis reinfection in a likely immunocompetent child, living in a high-risk environment favorable for exposition to M. tuberculosis but in a low-incidence country.

M3 - 10.1097/INF.0000000000002685 ER - TY - JOUR T1 - Antibiotic resistance prediction for Mycobacterium tuberculosis from genome sequence data with Mykrobe. JF - Wellcome Open Res Y1 - 2019 A1 - Martin Hunt A1 - Phelim Bradley A1 - Lapierre, Simon Grandjean A1 - Simon Heys A1 - Mark Thomsit A1 - Michael B Hall A1 - Kerri M Malone A1 - Penelope Wintringer A1 - Timothy M Walker A1 - Daniela M Cirillo A1 - Iñaki Comas A1 - Maha R Farhat A1 - Phillip Fowler A1 - Jennifer Gardy A1 - Nazir Ismail A1 - Thomas A Kohl A1 - Vanessa Mathys A1 - Matthias Merker A1 - Stefan Niemann A1 - Shaheed Vally Omar A1 - Vitali Sintchenko A1 - Grace Smith A1 - Dick van Soolingen A1 - Supply, Philip A1 - Sabira Tahseen A1 - Mark Wilcox A1 - Irena Arandjelovic A1 - Tim E A Peto A1 - Derrick W Crook A1 - Zamin Iqbal AB -

Two billion people are infected with , leading to 10 million new cases of active tuberculosis and 1.5 million deaths annually. Universal access to drug susceptibility testing (DST) has become a World Health Organization priority. We previously developed a software tool, , which provided offline species identification and drug resistance predictions for from whole genome sequencing (WGS) data. Performance was insufficient to support the use of WGS as an alternative to conventional phenotype-based DST, due to mutation catalogue limitations.  Here we present a new tool, , which provides the same functionality based on a new software implementation. Improvements include i) an updated mutation catalogue giving greater sensitivity to detect pyrazinamide resistance, ii) support for user-defined resistance catalogues, iii) improved identification of non-tuberculous mycobacterial species, and iv) an updated statistical model for Oxford Nanopore Technologies sequencing data. is released under MIT license at https://github.com/mykrobe-tools/mykrobe. We incorporate mutation catalogues from the CRyPTIC consortium et al. (2018) and from Walker et al. (2015), and make improvements based on performance on an initial set of 3206 and an independent set of 5845 Illumina sequences. To give estimates of error rates, we use a prospectively collected dataset of 4362 . Using culture based DST as the reference, we estimate to be 100%, 95%, 82%, 99% sensitive and 99%, 100%, 99%, 99% specific for rifampicin, isoniazid, pyrazinamide and ethambutol resistance prediction respectively. We benchmark against four other tools on 10207 (=5845+4362) samples, and also show that gives concordant results with nanopore data.  We measure the ability of -based DST to guide personalized therapeutic regimen design in the context of complex drug susceptibility profiles, showing 94% concordance of implied regimen with that driven by phenotypic DST, higher than all other benchmarked tools.

VL - 4 M3 - 10.12688/wellcomeopenres.15603.1 ER - TY - JOUR T1 - In vitro activity of bedaquiline against slow-growing nontuberculous mycobacteria. JF - J Med Microbiol Y1 - 2019 A1 - Martin, Anandi A1 - Godino, Isabel Torres A1 - Diana Angelica Aguilar-Ayala A1 - Vanessa Mathys A1 - Nacer Lounis A1 - Villalobos, Hector Rodriguez KW - Bedaquiline KW - Mycobacteria KW - nontuberculous AB -

Bedaquiline (BDQ) is a recently approved antibiotic for the treatment of multidrug-resistant tuberculosis, but its potential against slow-growing mycobacteria (SGM) is still unknown. The objective of this study was to determine the in vitro activity of BDQ on SGM by assessing their MIC and minimal bactericidal concentration (MBC). The MIC of BDQ against 17 clinical isolates including Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium chimaera, Mycobacterium kansasii and Mycobacterium simiae species was determined by the resazurin microtitre assay and the MBC by the c.f.u. determination on 7H10 agar plates. BDQ has a bacteriostatic activity on all SGM tested with a MIC range from 0.03 to 0.007 µg ml and surprisingly a good bactericidal activity on the majority of the isolates tested with an MBC of 1-2 µg ml . Based on these preliminary results BDQ seems to be very promising for treatment of diseases caused by SGM.

M3 - 10.1099/jmm.0.001025 ER - TY - JOUR T1 - Isoniazid Bactericidal Activity Involves Electron Transport Chain Perturbation. JF - Antimicrob Agents Chemother Y1 - 2019 A1 - Zeng, Sheng A1 - Karine Soetaert A1 - Faustine Ravon A1 - Marie Vandeput A1 - Dirk Bald A1 - Kauffmann, Jean-Michel A1 - Vanessa Mathys A1 - Wattiez, Ruddy A1 - Véronique Fontaine AB -

Accumulating evidence suggests that the bactericidal activity of some antibiotics may not be directly initiated by target inhibition. The activity of isoniazid (INH), a key first-line bactericidal antituberculosis drug currently known to inhibit mycolic acid synthesis, becomes extremely poor under stress conditions, such as hypoxia and starvation. This suggests that the target inhibition may not fully explain the bactericidal activity of the drug. Here, we report that INH rapidly increased BCG cellular ATP levels and enhanced oxygen consumption. The INH-triggered ATP increase and bactericidal activity were strongly compromised by Q203 and bedaquiline, which inhibit mycobacterial cytochrome and FF ATP synthase, respectively. Moreover, the antioxidant -acetylcysteine (NAC) but not 4-hydroxy-2,2,6,6-tetramethylpiperidin-1-oxyl (TEMPOL) abrogated the INH-triggered ATP increase and killing. These results reveal a link between the energetic (ATP) perturbation and INH's killing. Furthermore, the INH-induced energetic perturbation and killing were also abrogated by chemical inhibition of NADH dehydrogenases (NDHs) and succinate dehydrogenases (SDHs), linking INH's bactericidal activity further to the electron transport chain (ETC) perturbation. This notion was also supported by the observation that INH dissipated mycobacterial membrane potential. Importantly, inhibition of cytochrome oxidase significantly reduced cell recovery during INH challenge in a culture settling model, suggesting that the respiratory reprogramming to the cytochrome oxidase contributes to the escape of INH killing. This study implicates mycobacterial ETC perturbation through NDHs, SDHs, cytochrome , and FF ATP synthase in INH's bactericidal activity and pinpoints the participation of the cytochrome oxidase in protection against this drug under stress conditions.

VL - 63 CP - 3 M3 - 10.1128/AAC.01841-18 ER - TY - Generic T1 - RNA-Based susceptibility testing of Mycobacterium tuberculosis Y1 - 2019 A1 - An Van den Bossche A1 - Roby Bhattacharyya A1 - Jean-Yves Coppee A1 - Alain Baulard A1 - Deborah Hung A1 - Vanessa Mathys A1 - Pieter-Jan Ceyssens JF - 29th ECCMID CY - Amsterdam, The Netherlands CP - ESCMID ER - TY - JOUR T1 - Strong increase of true and false positive mycobacterial cultures sent to the National Reference Centre in Belgium, 2007 to 2016. JF - Euro Surveill Y1 - 2019 A1 - Karine Soetaert A1 - Lorenzo Subissi A1 - Pieter-Jan Ceyssens A1 - Vanfleteren, Brigitte A1 - Marianne Chantrenne A1 - Tommi Asikainen A1 - Els Duysburgh A1 - Vanessa Mathys AB -

IntroductionIn 2007, a new federal legislation in Belgium prohibited non-biosafety level 3 laboratories to process culture tubes suspected of containing mycobacteria.AimTo present mycobacterial surveillance/diagnosis data from the Belgian National Reference Centre for mycobacteria (NRC) from 2007 to 2016.MethodsThis retrospective observational study investigated the numbers of analyses at the NRC and false positive cultures (interpreted as containing mycobacteria at referring clinical laboratories, but with no mycobacterial DNA detected by PCR in the NRC). We reviewed mycobacterial species identified and assessed trends over time of proportions of nontuberculous mycobacteria (NTM) vs complex (MTBc), and false positive cultures vs NTM.ResultsFrom 2007 to 2016, analyses requests to the NRC doubled from 12.6 to 25.3 per 100,000 inhabitants. A small but significant increase occurred in NTM vs MTBc proportions, from 57.9% (587/1,014) to 60.3% (867/1,437) (p < 0.001). Although NTM infection notification is not mandatory in Belgium, we annually received up to 8.6 NTM per 100,000 inhabitants. predominated (ca 20% of NTM cultures), but culture numbers rose significantly, from 13.0% (74/587) of NTM cultures in 2007 to 21.0% (178/867) in 2016 (RR: 1.05; 95% CI: 1.03-1.07). The number of false positive cultures also increased, reaching 43.3% (1,097/2,534) of all samples in 2016.ConclusionWe recommend inclusion of NTM in sentinel programmes. The large increase of false positive cultures is hypothesised to result from processing issues prior to arrival at the NRC, highlighting the importance of sample decontamination/transport and equipment calibration in peripheral laboratories.

VL - 24 CP - 11 M3 - 10.2807/1560-7917.ES.2019.24.11.1800205 ER - TY - JOUR T1 - Transcriptional profiling of a laboratory and clinical Mycobacterium tuberculosis strain suggests respiratory poisoning upon exposure to delamanid. JF - Tuberculosis (Edinb) Y1 - 2019 A1 - An Van den Bossche A1 - Hugo Varet A1 - Amandine Sury A1 - Odile Sismeiro A1 - Rachel Legendre A1 - Jean-Yves Coppee A1 - Vanessa Mathys A1 - Pieter-Jan Ceyssens KW - Mycobacterium tuberculosis KW - Nitroimidazoles KW - Transcriptome AB -

Tuberculosis (TB) is the most deadly infectious disease worldwide. To reduce TB incidence and counter the spread of multidrug resistant TB, the discovery and characterization of new drugs is essential. In this study, the transcriptional response of two Mycobacterium tuberculosis strains to a pressure of the recently approved delamanid is investigated. Total RNA sequencing revealed that the response to this bicyclic nitroimidazole shows many similarities with pretomanid, an anti-tuberculous drug from the same class. Although delamanid is found to inhibit cell wall synthesis, the expression of genes involved in this process were only mildly affected. In contrast, a clear parallel was found with components that affect aerobic respiration. This demonstrates that, besides the inhibition of cell wall synthesis, respiratory poisoning plays a fundamental role in the bactericidal effect of delamanid. Remarkably, the most highly induced genes comprise poorly characterized genes for which functional characterization might hint to the target molecule(s) of delamanid and its exact mode(s) of action.

VL - 117 M3 - 10.1016/j.tube.2019.05.002 ER - TY - JOUR T1 - Transcriptional profiling of a laboratory and clinical Mycobacterium tuberculosis strain suggests respiratory poisoning upon exposure to delamanid JF - Tuberculosis Y1 - 2019 A1 - An Van den Bossche A1 - Hugo Varet A1 - Amandine Sury A1 - Odile Sismeiro A1 - Rachel Legendre A1 - Jean-Yves Coppee A1 - Vanessa Mathys A1 - Pieter-Jan Ceyssens KW - Delamanid KW - Mycobacterium tuberculosis KW - RNA sequencing KW - Transciptomics AB -

Tuberculosis (TB) is the most deadly infectious disease worldwide. To reduce TB incidence and counter the

spread of multidrug resistant TB, the discovery and characterization of new drugs is essential. In this study, the

transcriptional response of two Mycobacterium tuberculosis strains to a pressure of the recently approved delamanid

is investigated. Total RNA sequencing revealed that the response to this bicyclic nitroimidazole shows

many similarities with pretomanid, an anti-tuberculous drug from the same class. Although delamanid is found

to inhibit cell wall synthesis, the expression of genes involved in this process were only mildly affected. In

contrast, a clear parallel was found with components that affect aerobic respiration. This demonstrates that,

besides the inhibition of cell wall synthesis, respiratory poisoning plays a fundamental role in the bactericidal

effect of delamanid. Remarkably, the most highly induced genes comprise poorly characterized genes for which

functional characterization might hint to the target molecule(s) of delamanid and its exact mode(s) of action.

VL - 117 M3 - 10.1016/j.tube.2019.05.002 ER - TY - Generic T1 - Transcriptional profiling of Mycobacterium tuberculosis suggests respiratory poisoning upon exposure to delamanid Y1 - 2019 A1 - An Van den Bossche A1 - Hugo Varet A1 - Amandine Sury A1 - Odile Sismeiro A1 - Rachel Legendre A1 - Jean-Yves Coppee A1 - Vanessa Mathys A1 - Pieter-Jan Ceyssens AB -

Tuberculosis (TB) is the most deadly infectious disease worldwide. To reduce TB incidence and counter the spread of multidrug resistant TB, the discovery and characterization of new drugs is essential. In this study, the transcriptional response of two Mycobacterium tuberculosis strains to a pressure of the recently approved delamanid is investigated. Total RNA sequencing revealed that the response to this bicyclic nitroimidazole shows many similarities with pretomanid, an anti-tuberculous drug from the same class. Although delamanid is found to inhibit cell wall synthesis, the expression of genes involved in this process were only mildly affected. In contrast, a clear parallel was found with components that affect aerobic respiration. This demonstrates that, besides the inhibition of cell wall synthesis, respiratory poisoning plays a fundamental role in the bactericidal effect of delamanid. Remarkably, the most highly induced genes comprise poorly characterized genes for which functional characterization might hint to the target molecule(s) of delamanid and its exact mode(s) of action.

JF - 40th ESM conference CY - Valencia, Spain CP - European Society of Mycobacteria ER - TY - JOUR T1 - Aloe Emodin Reduces Phthiodiolone Dimycocerosate Potentiating Vancomycin Susceptibility on Mycobacteria JF - Indian Journal of Microbiology Y1 - 2018 A1 - Céline Rens A1 - Pieter-Jan Ceyssens A1 - Françoise Laval A1 - Lefèvre, Philippe A1 - Vanessa Mathys A1 - Daffé, Mamadou A1 - Véronique Fontaine M3 - 10.1007/s12088-018-0734-0 ER - TY - JOUR T1 - A cluster of multidrug-resistant Mycobacterium tuberculosis among patients arriving in Europe from the Horn of Africa: a molecular epidemiological study. JF - Lancet Infect Dis Y1 - 2018 A1 - Timothy M Walker A1 - Matthias Merker A1 - Astrid M Knoblauch A1 - Peter Helbling A1 - Otto D Schoch A1 - J van der Werf A1 - Katharina Kranzer A1 - Lena Fiebig A1 - Stefan Kröger A1 - Walter Haas A1 - Harald Hoffmann A1 - Indra, Alexander A1 - Adrian Egli A1 - Daniela M Cirillo A1 - Jérôme Robert A1 - Thomas R Rogers A1 - Ramona Groenheit A1 - Anne T Mengshoel A1 - Vanessa Mathys A1 - Marjo Haanperä A1 - Dick van Soolingen A1 - Stefan Niemann A1 - Erik C Böttger A1 - Peter M Keller KW - Cluster KW - Tuberculosis AB -

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).

M3 - 10.1016/S1473-3099(18)30004-5 ER - TY - JOUR T1 - Meropenem-clavulanate for drug-resistant tuberculosis: a follow-up of relapse-free cases. JF - Int J Tuberc Lung Dis Y1 - 2018 A1 - M C Payen A1 - I Muylle A1 - Vandenberg, O A1 - Vanessa Mathys A1 - M Delforge A1 - S Van den Wijngaert A1 - N Clumeck A1 - De Wit, S KW - Carbapenems KW - repurposed drugs KW - XDR-TB AB -

BACKGROUND: Extensively drug-resistant tuberculosis (XDR-TB), defined as TB caused by a Mycobacterium strain resistant to at least rifampicin, isoniazid, any fluoroquinolone and one of the injectable anti-tuberculosis drugs, remains a worldwide public health threat. Among repurposed drugs empirically used for XDR-TB cases, carbapenems have been studied in vitro and in animal models, with encouraging results. However, only short-term follow-up data from clinical studies are currently available.

OBJECTIVES: To study the long-term follow-up of XDR-TB cases treated with a regimen containing meropenem-clavulanate (M/Clav).

DESIGN: Retrospective observational case series study at a single hospital.

METHODS: All hospitalised drug-resistant TB patients who received M/Clav as part of their treatment from 2009 to 2016 were included. Demographic and clinical data were extracted from medical records.

RESULTS: Eighteen XDR-TB patients were included in the analysis. The successful outcome and mortality rates were respectively 83.3% and 11.1%. No relapses were observed in cured patients after a median follow-up of 4 years. No specific adverse events were attributed to treatment with M/Clav.

CONCLUSION: The rate of sustained successful treatment outcome observed here is far higher than the 26% observed in the 2014 World Health Organization XDR-TB cohort, suggesting that carbapenems may be beneficial for the treatment of difficult-to-treat TB cases.

VL - 22 CP - 1 M3 - 10.5588/ijtld.17.0352 ER - TY - JOUR T1 - The potential use of rifabutin for treatment of patients diagnosed with rifampicin-resistant tuberculosis. JF - J Antimicrob Chemother Y1 - 2018 A1 - Michael G Whitfield A1 - Robin M Warren A1 - Vanessa Mathys A1 - Lesley Scott A1 - Elise De Vos A1 - Wendy Stevens A1 - Elizabeth M Streicher A1 - Groenen, Guido A1 - Frederick A Sirgel A1 - Annelies Van Rie KW - rifabutin KW - Tuberculosis AB -

Background: Use of the Xpert MTB/RIF assay has increased the number of people diagnosed with rifampicin-resistant tuberculosis (RR-TB), especially in South Africa where Xpert is now the initial diagnostic for individuals with TB symptoms. We hypothesized that a proportion of RR-TB patients determined by Xpert can be treated with a rifabutin-containing regimen.

Methods: Rifabutin susceptibility by rpoB mutation was assessed in 349 individuals from South Africa and 172 from Belgium. rpoB polymorphisms were identified by Sanger sequencing. Rifampicin and rifabutin susceptibility was assessed phenotypically. A systematic review was performed to comprehensively collate information on rifabutin susceptibility by rpoB polymorphism. Rifabutin susceptibility was assigned to rpoB polymorphisms based on their positive likelihood ratios and ORs.

Results: One hundred and twelve rpoB polymorphisms (67.9% from literature) were identified from all 2045 RR-TB patients, of which 17 polymorphisms could be classified as susceptible/resistant to rifabutin. Eleven polymorphisms were associated with rifabutin susceptibility. The 516GTC mutation was the most common, representing 70% (South Africa) and 76% (Belgium) of all rifabutin-susceptible isolates. At a population level, the 11 polymorphisms associated with rifabutin susceptibility occurred in 33.2% and 16.6% of all South African and Belgian patients diagnosed with RR-TB, respectively.

Conclusions: Identification of the exact rpoB polymorphism leading to the diagnosis of RR-TB has the potential to allow inclusion of rifabutin in the treatment regimen of a substantial proportion of RR-TB patients. A randomized controlled trial evaluating the efficacy of a rifabutin-containing TB treatment regimen in these selected patients is needed to provide the evidence required for a change in policy.

M3 - 10.1093/jac/dky248 ER - TY - JOUR T1 - Prediction of Susceptibility to First-Line Tuberculosis Drugs by DNA Sequencing JF - New England Journal of MedicineNew England Journal of MedicineN Engl J Med Y1 - 2018 A1 - The CRyPTIC Consortium and the 100000 Genomes Project A1 - Allix-Béguec, Caroline A1 - Irena Arandjelovic A1 - Lijun Bi A1 - Patrick Beckert A1 - Maryline Bonnet A1 - Phelim Bradley A1 - Andrea Cabibbe A1 - Irving Cancino-Muñoz A1 - Mark J. Caulfield A1 - Angkana Chaiprasert A1 - Daniela Cirillo A1 - David Clifton A1 - Iñaki Comas A1 - Derrick W Crook A1 - Maria R De Filippo A1 - Han De Neeling A1 - Roland Diel A1 - Francis A. Drobniewski A1 - Kiatichai Faksri A1 - Maha R. Farhat A1 - Joy Fleming A1 - Philip Fowler A1 - Tom A. Fowler A1 - Gao, Qian A1 - Jennifer Gardy A1 - Deborah Gascoyne-Binzi A1 - Ana-Luiza Gibertoni-Cruz A1 - Ana Gil-Brusola A1 - Tanya Golubchik A1 - Ximena Gonzalo A1 - Louis Grandjean A1 - Guangxue, He A1 - Jennifer L. Guthrie A1 - Sarah Hoosdally A1 - Martin Hunt A1 - Zamin Iqbal A1 - Nazir Ismail A1 - James Johnston A1 - Faisal Khanzada A1 - Chiea Khor A1 - Thomas A. Kohl A1 - Clare Kong A1 - Sam Lipworth A1 - Liu, Qingyun A1 - Gugu Maphalala A1 - Elena Martinez A1 - Vanessa Mathys A1 - Matthias Merker A1 - Paolo Miotto A1 - Mistry, Nerges A1 - David A.J. Moore A1 - Megan Murray A1 - Stefan Niemann A1 - Rick T.-H. Ong A1 - Tim E.A. Peto A1 - James E. Posey A1 - Prammananan, Therdsak A1 - Alexander Pym A1 - Camilla Rodrigues A1 - Mabel Rodrigues A1 - Timothy Rodwell A1 - Gian M. Rossolini A1 - Elisabeth Sánchez Padilla A1 - Marco Schito A1 - Xin Shen A1 - Jay Shendure A1 - Vitali Sintchenko A1 - Alex Sloutsky A1 - Grace E. Smith A1 - Matthew Snyder A1 - Karine Soetaert A1 - Angela M. Starks A1 - Supply, Philip A1 - Suriyapol, Prapat A1 - Sabira Tahseen A1 - Patrick Tang A1 - Teo, Yik-Ying A1 - Thuong N.T. Thuong A1 - Guy Thwaites A1 - Tortoli, Enrico A1 - Shaheed V. Omar A1 - Dick van Soolingen A1 - Sarah A. Walker A1 - Timothy M. Walker A1 - Mark Wilcox A1 - Daniel J. Wilson A1 - David Wyllie A1 - Yang, Yang A1 - Hongtai Zhang A1 - Zhao, Yanlin A1 - Zhu, Baoli AB -

BACKGROUND: The World Health Organization recommends drug-susceptibility testing of Mycobacterium tuberculosis complex for all patients with tuberculosis to guide treatment decisions and improve outcomes. Whether DNA sequencing can be used to accurately predict profiles of susceptibility to first-line antituberculosis drugs has not been clear.METHODS: We obtained whole-genome sequences and associated phenotypes of resistance or susceptibility to the first-line antituberculosis drugs isoniazid, rifampin, ethambutol, and pyrazinamide for isolates from 16 countries across six continents. For each isolate, mutations associated with drug resistance and drug susceptibility were identified across nine genes, and individual phenotypes were predicted unless mutations of unknown association were also present. To identify how whole-genome sequencing might direct first-line drug therapy, complete susceptibility profiles were predicted. These profiles were predicted to be susceptible to all four drugs (i.e., pansusceptible) if they were predicted to be susceptible to isoniazid and to the other drugs or if they contained mutations of unknown association in genes that affect susceptibility to the other drugs. We simulated the way in which the negative predictive value changed with the prevalence of drug resistance.RESULTS: A total of 10,209 isolates were analyzed. The largest proportion of phenotypes was predicted for rifampin (9660 [95.4%] of 10,130) and the smallest was predicted for ethambutol (8794 [89.8%] of 9794). Resistance to isoniazid, rifampin, ethambutol, and pyrazinamide was correctly predicted with 97.1%, 97.5%, 94.6%, and 91.3% sensitivity, respectively, and susceptibility to these drugs was correctly predicted with 99.0%, 98.8%, 93.6%, and 96.8% specificity. Of the 7516 isolates with complete phenotypic drug-susceptibility profiles, 5865 (78.0%) had complete genotypic predictions, among which 5250 profiles (89.5%) were correctly predicted. Among the 4037 phenotypic profiles that were predicted to be pansusceptible, 3952 (97.9%) were correctly predicted.CONCLUSIONS: Genotypic predictions of the susceptibility of M. tuberculosis to first-line drugs were found to be correlated with phenotypic susceptibility to these drugs. (Funded by the Bill and Melinda Gates Foundation and others.).

SN - 0028-4793 M3 - doi: 10.1056/NEJMoa1800474 ER - TY - Generic T1 - RNA-based drug susceptibility testing of Mycobacterium tuberculosis Y1 - 2018 A1 - An Van den Bossche A1 - Roby Bhattacharyya A1 - Jean-Yves Coppee A1 - Leen Rigouts A1 - Alain Baulard A1 - Alexandra Vodolazkaia A1 - Deborah Hung A1 - Vanessa Mathys A1 - Pieter-Jan Ceyssens AB -

Background

Multidrug resistance of Tuberculosis strains (MDR-TB) are one of the major WHO health concerns. One of the challenges that hampers the effective response to MDR-TB is the long turnaround time of phenotypic Drug Susceptibility Testing (DST). To counter this, new fast and sensitive DNA-based methods were successfully introduced over the last years. However, these (a) are based on the knowledge on resistance mutations, (b) do not distinguish living from dead cells, (c) ignore all intrinsic resistance mechanisms, and (d) ignore the influence of compensatory mutations.

Objectives

We introduce a next-generation diagnostic test based on quantification of drug-specific RNA biomarkers. The basic principle is that a brief antibiotic exposure triggers specific transcriptional responses in susceptible, but not in resistant, microbes within a few hours. This has the advantage that long culture-dependent steps are avoided, yet the resistance phenotype is detected independent of the specific cause of resistance.

Materials & Methods

First, the global transcriptional response of two TB strains to 10 anti-TB drugs was determined using RNAtaq-Seq. A set of highly responsive genes was selected for each drug and RNA-targeting probes were designed.

Next, the RNA-based DST was developed in 96 well format. In short, 200 µl of a positively flagged MGITTM (BD) culture is spiked with a drug, while a replicate is incubated in absence of the drug. Multiplex mRNA quantification is performed directly on crude cell lysates using a combination of the bead-based MagPixTM (Luminex) and QuantigeneTM Plex (Thermo Fisher) technology. The normalized expression levels are combined to one numeric value which determines the drug susceptibility of the investigated strain.

Results

We successfully developed 8 primary sets of RNA biomarkers for ten 1st-line, 2nd-line and new drugs. Taking isoniazid as proof of principle, we present a biomarker set of 5 responsive genes and 3 normalizing genes, which enables to distinguish susceptible, low- and high resistant TB strains after 6 hours incubation. Next, preliminary results demonstrate that the biomarker sets can successfully discriminate between susceptible and resistance strains for the selected drugs.

Conclusion

We present a robust, RNA-based DST without the need for RNA extraction. The assay was proven to be efficient for isoniazid. With a total of 8 biomarker sets under optimization, the drug resistance profile of up to 14 drugs can be determined.

JF - 2nd St. Petersburg Symposium on Tuberculosis and Mycobacteria: Molecular Approach CY - Sint Petersburg CP - Institut Pasteur Sint Petersburg ER - TY - Generic T1 - RNA-based drug susceptibility testing of Mycobacterium tuberculosis Y1 - 2018 A1 - An Van den Bossche A1 - Leen Rigouts A1 - Jean-Yves Coppee A1 - Vanessa Mathys A1 - Pieter-Jan Ceyssens KW - Mycobacterium tuberculosis; antimicrobial resistance; diagnostics AB -

To counter the long turnaround time of standard phenotypic Drug Susceptibility Testing (DST) of M. tuberculosis (Mtb), multiple DNA-based methods were successfully introduced over the last years. Although these are fast and sensitive, they (a) are based on the knowledge on resistance mutations which is limited for especially 2nd-line and new drugs, (b) do not distinguish living from death cells, (c) ignore all intrinsic resistance mechanisms like efflux pump overexpression, and (d) ignore the multifactorial influence of compensatory mutations. Here, we introduce a next-generation diagnostic test based on quantification of antibiotic-specific RNA biomarkers. The basic principle is that a brief antibiotic exposure triggers specific transcriptional responses in susceptible, but not in resistant, microbes within minutes to a few hours. A major advantage of this method is that it avoids a long culture-dependent step, yet detects the resistance phenotype, independent of the specific cause of resistance.

First, the global transcriptional response of H37Rv and clinical Mtb strains on ten anti-TB drugs including Bedaquiline, Pyrazinamide and Delamanid was determined, to identify RNA Biomarkers. In a next phase, the RNA-based DST was developed in 96 well format. In short, 200 µl of a positively flagged MGIT culture is spiked with a specific concentration of a drug, while a control sample is incubated in absence of the drug. Multiplex mRNA quantification is performed directly on crude cell lysate using a combination of the bead-based MagPixTM (Luminex) and QuantigeneTM Plex (Thermo Fischer) technology. Normalized, relative genes expression levels (control vs drug) are combined to one numeric value which determines the drug susceptibility of the investigated strain.

The assay was optimized for parameters as cell density, incubation time and lysis method. I will present the results, showing that following a biomarker set of 5 responsive genes and 3 normalizing genes enables to distinguish low- and high resistant Mtb strains after an incubation step of 6 hours. With a total of 8 biomarker sets under optimization, the phenotypic drug resistance profile of up to 14 drugs can be determined for any combination of antibiotics in 96 well format.

 

JF - Combating resistance : microbes and vectors CY - Paris, France CP - Institute Pasteur ER - TY - Generic T1 - RNA-based drug susceptibility testing of Mycobacterium tuberculosis Y1 - 2018 A1 - An Van den Bossche A1 - Roby Bhattacharyya A1 - Jean-Yves Coppee A1 - Leen Rigouts A1 - Alain Baulard A1 - Deborah Hung A1 - Vanessa Mathys A1 - Pieter-Jan Ceyssens KW - drug susceptibility testing KW - Mycobacterium tuberculosis JF - 39th Annual Congress of the European Society of Mycobacteriology CY - Dresden, Germany CP - ESM ER - TY - JOUR T1 - Time-and-motion tool for the assessment of working time in tuberculosis laboratories: a multicentre study. JF - Int J Tuberc Lung Dis Y1 - 2018 A1 - Vanessa Mathys A1 - E Roycroft A1 - P Raftery A1 - R Groenheit A1 - B D Folkvardsen A1 - D Homorodean A1 - E Vasiliauskiene A1 - L Vasiliauskaite A1 - C Kodmon A1 - J van der Werf A1 - F Drobniewski A1 - V Nikolayevskyy KW - hands-on time KW - laboratory diagnosis KW - Workload AB -

SETTING: Implementation of novel diagnostic assays in tuberculosis (TB) laboratory diagnosis requires effective management of time and resources.

OBJECTIVE: To further develop and assess at multiple centres a time-and-motion (T&M) tool as an objective means for recording the actual time spent on running laboratory assays.

DESIGN: Multicentre prospective study conducted in six European Union (EU) reference TB laboratories.

RESULTS: A total of 1060 specimens were tested using four laboratory assays. The number of specimens per batch varied from one to 60; a total of 64 recordings were performed. Theoretical hands-on times per specimen (TTPS) in h:min:s for Xpert® MTB/RIF, mycobacterial interspersed repetitive unit-variable number of tandem repeats genotyping, Ziehl-Neelsen staining and manual fluorescence microscopy were respectively 00:33:02 ± 00:12:32, 00:13:34 ± 00:03:11, 00:09:54 ± 00:00:53 and 00:06:23 ± 00:01:36. Variations between laboratories were predominantly linked to the time spent on reporting and administrative procedures. Processing specimens in batches could help save time in highly automated assays (e.g., line-probe) (TTPS 00:14:00 vs. 00:09:45 for batches comprising 7 and 31 specimens, respectively).

CONCLUSIONS: The T&M tool can be considered a universal and objective methodology contributing to workload assessment in TB diagnostic laboratories. Comparison of workload between laboratories could help laboratory managers justify their resource and personnel needs for the implementation of novel, time-saving, cost-effective technologies, as well as identify areas for improvement.

VL - 22 CP - 4 M3 - 10.5588/ijtld.17.0564 ER - TY - JOUR T1 - Case report of a false positive result of the Xpert(®) MTB/RIF assay for rifampicin resistance in Mycobacterium tuberculosis complex. JF - Acta Clin Belg Y1 - 2017 A1 - Claessens, Jolien A1 - Vanessa Mathys A1 - Derdelinckx, Inge A1 - Saegeman, Veroniek AB -

In the present case, we report a false positive result for the detection of rifampicin (RIF) resistance by the Xpert(®) MTB/RIF assay, version G4.Miliary Mycobacterium tuberculosis infection (miliary TB) was suspected in a 50-year old Angolan woman. Imaging of the thorax and abdomen displayed diffuse lesions. The Xpert(®) MTB/RIF assay conducted on the broncho-alveolar lavage (BAL) fluid was positive for TB and positive for RIF resistance. Confirmatory molecular tests and the phenotypic drug susceptibility determination supported the diagnosis of TB but not RIF resistance. The patient was treated successfully with a conventional therapeutic scheme. Because, the Xpert(®) MTB/RIF assay allows the simultaneous detection of TB and RIF resistance, the World Health Organisation (WHO) recommends its use as initial diagnostic test, over microscopy, culture and phenotypic drug susceptibility testing. Even though specificity of the Xpert(®) MTB/RIF assay version G4 is high, false positive test results remain possible and have to be considered for the interpretation of the RIF resistance detection by Xpert(®) MTB/RIF assay.

VL - 72 CP - 3 U1 - http://www.ncbi.nlm.nih.gov/pubmed/26400761?dopt=Abstract M3 - 10.1179/2295333715Y.0000000072 ER - TY - JOUR T1 - Consensus numbering system for the rifampicin resistance-associated rpoB gene mutations in pathogenic mycobacteria. JF - Clin Microbiol Infect Y1 - 2017 A1 - E André A1 - Goeminne, L A1 - Cabibbe, A A1 - Beckert, P A1 - B Kabamba Mukadi A1 - Vanessa Mathys A1 - Gagneux, S A1 - Niemann, S A1 - Van Ingen, J A1 - Cambau, E KW - antibiotics KW - Antitubercular KW - Bacterial Proteins KW - Consensus KW - DNA-Directed RNA Polymerases KW - Escherichia coli KW - Escherichia coli Proteins KW - Genotype KW - Genotyping Techniques KW - Humans KW - Microbial Sensitivity Tests KW - Mutant Proteins KW - Mutation KW - Mycobacterium KW - Mycobacterium tuberculosis KW - Rifampin KW - Terminology as Topic AB -

The rpoB gene codes for the RNA polymerase β subunit, which is the target of rifampicin, an essential drug in the treatment of tuberculosis and other mycobacterial infections. This gene is present in all bacteria, but its length and nucleotide sequence vary between bacterial species, including mycobacteria. Mutations in the rpoB gene alter the structure of this protein and cause drug resistance. To describe the resistance-associated mutations, the scientific and medical communities have been using, since 1993, a numbering system based on the Escherichia coli sequence annotation. Using E. coli reference for describing mutations in mycobacteria leads to misunderstandings, particularly with the increasing use of whole genome sequencing, which brought an alternative numbering system based on the Mycobacterium tuberculosis rpoB sequence. We propose using a consensus numbering system for the reporting of resistance mutations based on the reference genomes from the species interrogated (such as strain H37Rv for M. tuberculosis). This manuscript provides the necessary figures and tables allowing researchers, microbiologists and clinicians to easily convert other annotation systems into one common language.

VL - 23 CP - 3 U1 - http://www.ncbi.nlm.nih.gov/pubmed/27664776?dopt=Abstract M3 - 10.1016/j.cmi.2016.09.006 ER - TY - Generic T1 - Global transcriptional analyses of Mycobacterium tuberculosis using old and new drugs Y1 - 2017 A1 - An Van den Bossche A1 - Pieter-Jan Ceyssens A1 - Roby Bhattacharyya A1 - Deborah Hung A1 - Vanessa Mathys KW - antibiotics KW - Mycobacterium tuberculosis KW - Transcriptomics AB -

Background: With 1.5 million deaths in 2014, the most recent report of the WHO ranks tuberculosis (TB) alongside HIV as a leading cause of death by infectious diseases. In combination, the spread of (multi-)drug resistant Mycobacterium tuberculosis is rising (480 000 new cases), impeding the treatment of infected patients.

Current TB drug resistance research is mainly focused on the genomic level, using WGS to search for resistance-causing mutations. However, transcriptional analyses can be a powerful tool as well. Analyses on susceptible strains provide information on the response to the stress a drug is causing, showing which mechanisms are activated to counter the effect of the drug. This might lead to new strategies for the development of new/complimentary drugs. On the other hand, this response gives an view on the working mechanism of the drug. Moreover, comparing the response of sensitive and resistant strains can yield additional information, like the activation of specific efflux pumps.

However, due to the high cost of RNAseq analyses, genome wide transcriptional analyses of drug influences on M. tuberculosis remain rather limited. Here we present a global study of the response of two pansensitive M. tuberculosis strains to eight TB-drugs. By using the approach called RNAtag-Seq, multiple samples were combined in one run, reducing time and cost.

 

Material/methods: For each drug (isoniazid, rifampicin, ethambutol, capreomycin, amikacin, linezolid, moxifloxacin and bedaquilin), twice the critical concentration was added to the cultures of two pansensitive M. tuberculosis strains. Samples were taken 0h, 2h, 6h and 24h after the administration of the drug.  RNA was extracted combining bead-beating in TRIzol and the Direct-Zol purification kit. After quality control, the extracts were fragmented, barcoded and pooled, cDNA libraries were constructed and sequenced using the HiSeq technology (Illumina®). Reads were mapped to the reference genome and normalized read counts were calculated per gene.

 

Results: For each drug a specific transcriptional response was mapped, revealing lists of up- and down-regulated genes. As an example and in accordance with previous studies, the highest induced genes in the presence of isoniazid belong to a cluster of genes that encodes components of the FAS-II (fatty acid synthase II) complex, which is targeted by isoniazid. On the other hand, a remarkable down-regulation of the NADH-dehydrogenase (ndh) cluster genes was noted. This can be assigned to the dependence of isoniazid target inhA on NADH. Lowering the ndh-activity is also seen in resistant strains.

 

Conclusions: By using RNAtag-seq, a global study of the transcriptional response of M. tuberculosis to several drugs could be made. These analyses can reveal the working mechanisms of existing and new drugs and provide new insights on the mechanism of resistance of strains.

 

JF - ECCMID 2017 PB - ECCMID CY - Vienna, Austria CP - ESCMID ER - TY - Generic T1 - Global transcriptional analyses of Mycobacterium tuberculosis using old and new drugs Y1 - 2017 A1 - An Van den Bossche A1 - Pieter-Jan Ceyssens A1 - Roby Bhattacharyya A1 - Deborah Hung A1 - Vanessa Mathys KW - antibiotics KW - Mycobacterium tuberculosis KW - Transcriptomics AB -

Background: With 1.5 million deaths in 2014, the most recent report of the WHO ranks tuberculosis (TB) alongside HIV as a leading cause of death by infectious diseases. In combination, the spread of (multi-)drug resistant Mycobacterium tuberculosis is rising (480 000 new cases), impeding the treatment of infected patients.

Current TB drug resistance research is mainly focused on the genomic level, using WGS to search for resistance-causing mutations. However, transcriptional analyses can be a powerful tool as well. Analyses on susceptible strains provide information on the response to the stress a drug is causing, showing which mechanisms are activated to counter the effect of the drug. This might lead to new strategies for the development of new/complimentary drugs. On the other hand, this response gives an view on the working mechanism of the drug. Moreover, comparing the response of sensitive and resistant strains can yield additional information, like the activation of specific efflux pumps.

However, due to the high cost of RNAseq analyses, genome wide transcriptional analyses of drug influences on M. tuberculosis remain rather limited. Here we present a global study of the response of two pansensitive M. tuberculosis strains to eight TB-drugs. By using the approach called RNAtag-Seq, multiple samples were combined in one run, reducing time and cost.

 

Material/methods: For each drug (isoniazid, rifampicin, ethambutol, capreomycin, amikacin, linezolid, moxifloxacin and bedaquilin), twice the critical concentration was added to the cultures of two pansensitive M. tuberculosis strains. Samples were taken 0h, 2h, 6h and 24h after the administration of the drug.  RNA was extracted combining bead-beating in TRIzol and the Direct-Zol purification kit. After quality control, the extracts were fragmented, barcoded and pooled, cDNA libraries were constructed and sequenced using the HiSeq technology (Illumina®). Reads were mapped to the reference genome and normalized read counts were calculated per gene.

 

Results: For each drug a specific transcriptional response was mapped, revealing lists of up- and down-regulated genes. As an example and in accordance with previous studies, the highest induced genes in the presence of isoniazid belong to a cluster of genes that encodes components of the FAS-II (fatty acid synthase II) complex, which is targeted by isoniazid. On the other hand, a remarkable down-regulation of the NADH-dehydrogenase (ndh) cluster genes was noted. This can be assigned to the dependence of isoniazid target inhA on NADH. Lowering the ndh-activity is also seen in resistant strains.

 

Conclusions: By using RNAtag-seq, a global study of the transcriptional response of M. tuberculosis to several drugs could be made. These analyses can reveal the working mechanisms of existing and new drugs and provide new insights on the mechanism of resistance of strains.

 

JF - 38th Annual Congress of the European Society of Mycobacteriology PB - ESM CY - Sibenik, Craotia CP - ESM ER - TY - JOUR T1 - Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry for Combined Species Identification and Drug Sensitivity Testing in Mycobacteria. JF - J Clin Microbiol Y1 - 2017 A1 - Pieter-Jan Ceyssens A1 - Karine Soetaert A1 - Timke, Markus A1 - An Van den Bossche A1 - Sparbier, Katrin A1 - Koen De Cremer A1 - Kostrzewa, Markus A1 - Marijke Hendrickx A1 - Vanessa Mathys KW - Antitubercular Agents KW - Bacteriological Techniques KW - Humans KW - Mycobacterium tuberculosis KW - Nontuberculous Mycobacteria KW - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization AB -

Species identification and drug susceptibility testing (DST) of mycobacteria are important yet complex processes traditionally reserved for reference laboratories. Recent technical improvements in matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has started to facilitate routine mycobacterial identifications in clinical laboratories. In this paper, we investigate the possibility of performing phenotypic MALDI-based DST in mycobacteriology using the recently described MALDI Biotyper antibiotic susceptibility test rapid assay (MBT-ASTRA). We randomly selected 72 clinical Mycobacterium tuberculosis and nontuberculous mycobacterial (NTM) strains, subjected them to MBT-ASTRA methodology, and compared its results to current gold-standard methods. Drug susceptibility was tested for rifampin, isoniazid, linezolid, and ethambutol (M. tuberculosis, n = 39), and clarithromycin and rifabutin (NTM, n = 33). Combined species identification was performed using the Biotyper Mycobacteria Library 4.0. Mycobacterium-specific MBT-ASTRA parameters were derived (calculation window, m/z 5,000 to 13,000, area under the curve [AUC] of >0.015, relative growth [RG] of <0.5; see the text for details). Using these settings, MBT-ASTRA analyses returned 175/177 M. tuberculosis and 65/66 NTM drug resistance profiles which corresponded to standard testing results. Turnaround times were not significantly different in M. tuberculosis testing, but the MBT-ASTRA method delivered on average a week faster than routine DST in NTM. Databases searches returned 90.4% correct species-level identifications, which increased to 98.6% when score thresholds were lowered to 1.65. In conclusion, the MBT-ASTRA technology holds promise to facilitate and fasten mycobacterial DST and to combine it directly with high-confidence species-level identifications. Given the ease of interpretation, its application in NTM typing might be the first in finding its way to current diagnostic workflows. However, further validations and automation are required before routine implementation can be envisioned.

VL - 55 CP - 2 U1 - http://www.ncbi.nlm.nih.gov/pubmed/28003422?dopt=Abstract M3 - 10.1128/JCM.02089-16 ER - TY - JOUR T1 - Molecular epidemiology of Mycobacterium tuberculosis complex in Brussels, 2010-2013. JF - PLoS One Y1 - 2017 A1 - Vluggen, Christelle A1 - Soetaert, Karine A1 - Groenen, Guido A1 - Wanlin, Maryse A1 - Spitaels, Martine A1 - Arrazola de Oñate, Wouter A1 - Fauville-Dufaux, Maryse A1 - Saegerman, Claude A1 - Vanessa Mathys KW - ADOLESCENT KW - Adult KW - Bacterial Typing Techniques KW - Belgium KW - Child KW - Cluster Analysis KW - Contact Tracing KW - Drug Resistance, Multiple, Bacterial KW - Emigrants and Immigrants KW - Family Health KW - Female KW - Genetic Variation KW - Genotype KW - Hospitals, Urban KW - Humans KW - incidence KW - Male KW - middle aged KW - Mycobacterium tuberculosis KW - Phylogeny KW - Population Surveillance KW - Tuberculosis KW - Tuberculosis, Multidrug-Resistant KW - Urban Health KW - Young adult AB -

The tuberculosis (TB) incidence rate in Brussels-Capital Region is 3-fold higher than in Belgium as a whole. Eight years after the realization of initial prospective population-based molecular epidemiology investigations in this Region, a similar study over the period 2010-2013 was conducted. TB strains isolated from 945 patients were submitted to genotyping by standardized 24-locus-MIRU-VNTR typing and spoligotyping. The phylogenetic analysis showed that the LAM (16.7%) and Haarlem (15.7%) branches are the two most prevalent TB lineages circulating in Brussels. Analysis of the MDR subgroup showed an association with Beijing strains (39.9%) and patients native of Eastern Europe (40.7%). Genotyping detected 113 clusters involving 321 patients, giving a recent transmission index of 22.9%. Molecular-guided epidemiological investigations and routine surveillance activities revealed family transmission or social contact for patients distributed over 34 clusters. Most of the patients were foreign-born (75.7%). However, cluster analysis revealed only limited trans-national transmission. Comparison with the previous study shows a stable epidemiological situation except for the mean age difference between Belgian-born and foreign-born patients which has disappeared. This study confirms that molecular epidemiology has become an important determinant for TB control programs. However, sufficient financial means need to be available to perform all required epidemiological investigations.

VL - 12 CP - 2 U1 - https://www.ncbi.nlm.nih.gov/pubmed/28222189?dopt=Abstract M3 - 10.1371/journal.pone.0172554 ER - TY - JOUR T1 - Nontuberculous mycobacteria among pulmonary tuberculosis patients: a retrospective Belgian multicenter study. JF - Acta Clin Belg Y1 - 2017 A1 - De Keukeleire, Steven A1 - Vanessa Mathys A1 - Van den Wijngaert, Sigi A1 - Van De Vyvere, Martine A1 - Jonckheere, Stijn A1 - De Beenhouwer, Hans A1 - de Bel, Annelies A1 - Arrazola de Oñate, Wouter A1 - Wanlin, Maryse A1 - Piérard, Denis A1 - Nulens, Eric A1 - Saegeman, Veroniek KW - Adult KW - Belgium KW - Coinfection KW - Female KW - Humans KW - Male KW - middle aged KW - Mycobacterium Infections, Nontuberculous KW - Nontuberculous Mycobacteria KW - Retrospective Studies KW - Tuberculosis, Pulmonary AB -

OBJECTIVES: Currently, there are no European data about the frequency and clinical significance of nontuberculous mycobacteria (NTM) grown from respiratory samples during the treatment of tuberculosis (TB). We determined the frequency and clinical significance of NTM isolated before or during pulmonary tuberculosis treatment in Belgian laboratories.

METHODS: We conducted a nationwide retrospective multicenter cohort study on the co-isolation of TB and NTM in Belgium. Starting from laboratory data between 2006 and 2013, possible TB-NTM co-isolations were searched for.

RESULTS: A total of 2569 unique culture-positive pulmonary tuberculosis cases were included in the study. Only 35 (1.4%) of these TB cases had an NTM co-isolated, and two of these 35 fulfilled the ATS criteria for NTM lung disease.

CONCLUSION: A very low prevalence of 1.4% NTM co-isolations was found in Belgian patients with culture-proven pulmonary TB.

VL - 72 CP - 1 U1 - http://www.ncbi.nlm.nih.gov/pubmed/27345031?dopt=Abstract M3 - 10.1080/17843286.2016.1200823 ER - TY - JOUR T1 - Reversion of antibiotic resistance in Mycobacterium tuberculosis by spiroisoxazoline SMARt-420. JF - Science Y1 - 2017 A1 - Blondiaux, Nicolas A1 - Moune, Martin A1 - Desroses, Matthieu A1 - Frita, Rosangela A1 - Flipo, Marion A1 - Vanessa Mathys A1 - Karine Soetaert A1 - Kiass, Mehdi A1 - Delorme, Vincent A1 - Djaout, Kamel A1 - Trebosc, Vincent A1 - Kemmer, Christian A1 - Wintjens, René A1 - Wohlkönig, Alexandre A1 - Antoine, Rudy A1 - Huot, Ludovic A1 - Hot, David A1 - Coscolla, Mireia A1 - Feldmann, Julia A1 - Gagneux, Sebastien A1 - Locht, Camille A1 - Brodin, Priscille A1 - Gitzinger, Marc A1 - Déprez, Benoit A1 - Willand, Nicolas A1 - Baulard, Alain R AB -

Antibiotic resistance is one of the biggest threats to human health globally. Alarmingly, multidrug-resistant and extensively drug-resistant Mycobacterium tuberculosis have now spread worldwide. Some key antituberculosis antibiotics are prodrugs, for which resistance mechanisms are mainly driven by mutations in the bacterial enzymatic pathway required for their bioactivation. We have developed drug-like molecules that activate a cryptic alternative bioactivation pathway of ethionamide in M. tuberculosis, circumventing the classic activation pathway in which resistance mutations have now been observed. The first-of-its-kind molecule, named SMARt-420 (Small Molecule Aborting Resistance), not only fully reverses ethionamide-acquired resistance and clears ethionamide-resistant infection in mice, it also increases the basal sensitivity of bacteria to ethionamide.

VL - 355 CP - 6330 U1 - http://www.ncbi.nlm.nih.gov/pubmed/28302858?dopt=Abstract M3 - 10.1126/science.aag1006 ER - TY - JOUR T1 - Trend analysis suggested a change in subspecies among Mycobacterium avium isolated from pigs in Belgium, 1967–2013 JF - Veterinary Record Y1 - 2017 A1 - Karine Soetaert A1 - C. Vluggen A1 - L Duytschaever A1 - J. Denoël A1 - Virginie Roupie A1 - F. Smeets A1 - Nicolas Bruffaerts A1 - Huygen, K. A1 - David Fretin A1 - M. Diels A1 - L. Rigouts A1 - C. Saegerman A1 - Vanessa Mathys VL - 180 CP - 18 M3 - 10.1136/vr.103951 ER - TY - JOUR T1 - Trend analysis suggested a change in subspecies among isolated from pigs in Belgium, 1967-2013. JF - Vet Rec Y1 - 2017 A1 - Karine Soetaert A1 - Duytschaever, L A1 - Denoël, J A1 - Virginie Roupie A1 - Smeets, F A1 - Nicolas Bruffaerts A1 - Huygen, K A1 - David Fretin A1 - Diels, M A1 - Rigouts, L A1 - Saegerman, C A1 - Vanessa Mathys KW - Animals KW - Belgium KW - Genotyping Techniques KW - Mycobacterium avium KW - Swine KW - Swine Diseases KW - Tuberculosis VL - 180 CP - 18 U1 - https://www.ncbi.nlm.nih.gov/pubmed/28283669?dopt=Abstract M3 - 10.1136/vr.103951 ER - TY - Generic T1 - Tuberculose en antibioticaresistentie: het WIV draagt bij tot de ontwikkeling van een nieuw prototype van een geneesmiddel T2 - WIV-ISP website Y1 - 2017 A1 - Vanessa Mathys A1 - Karine Soetaert KW - antibioresistentie KW - geneesmiddelen KW - tuberculose JF - WIV-ISP website PB - WIV-ISP CY - Brussels, Belgium UR - https://www.wiv-isp.be/nl/pershoek/tuberculose-en-antibioticaresistentie-het-wiv-draagt-bij-tot-de-ontwikkeling-van-een-nieuw ER - TY - Generic T1 - Tuberculose et antibiorésistance : l’ISP contribue au développement d’un nouveau prototype de médicament T2 - WIV-ISP website Y1 - 2017 A1 - Vanessa Mathys A1 - Karine Soetaert KW - antibiorésistance KW - médicament KW - tuberculose JF - WIV-ISP website PB - WIV-ISP CY - Brussels, Belgium UR - https://www.wiv-isp.be/fr/coin-presse/tuberculose-et-antibioresistance-lisp-contribue-au-developpement-dun-nouveau-prototype ER - TY - JOUR T1 - Virulence and immunogenicity of genetically defined human and porcine isolates of M. avium subsp. hominissuis in an experimental mouse infection. JF - PLoS One Y1 - 2017 A1 - Bruffaerts, Nicolas A1 - Vluggen, Christelle A1 - Roupie, Virginie A1 - Duytschaever, Lucille A1 - Van den Poel, Christophe A1 - Denoël, Joseph A1 - Wattiez, Ruddy A1 - Letesson, Jean-Jacques A1 - Fretin, David A1 - Rigouts, Leen A1 - Chapeira, Ophélie A1 - Vanessa Mathys A1 - Saegerman, Claude A1 - Huygen, Kris KW - Adult KW - Animals KW - Child, Preschool KW - Female KW - Genome, Bacterial KW - Genotype KW - Humans KW - Interferon-gamma KW - Interleukins KW - Liver KW - Lung KW - Male KW - mice KW - Mice, Inbred BALB C KW - Mycobacterium avium KW - Spleen KW - Swine KW - Tuberculosis KW - virulence AB -

Mycobacterium avium subsp. hominissuis (Mah) represents a health concern for humans and to a lesser extent for pigs, but its zoonotic potential remains elusive. Using multispacer sequence typing (MST) we previously identified 49 different genotypes of Mah among Belgian clinical and porcine isolates, with 5 MSTs shared by both hosts. Using experimental intranasal infection of BALB/c mice, we compared the virulence and immunogenicity of porcine and clinical human isolates with shared genotype or with a genotype only found in humans or pigs. Bacterial replication was monitored for 20 weeks in lungs, spleen and liver and mycobacteria specific spleen cell IFN-γ, IL-10 and IL-17 production as well as serum antibody responses were analyzed. Isolates varied in virulence, with human and porcine isolates sharing MST22 genotype showing a thousand fold higher bacterial replication in lungs and more dissemination to spleen and liver than the human and porcine MST91 isolates. Virulent MST22 type was also associated with progressive suppression of IFN-γ and IL-17 responses, and increased IL-10 production. Whole genome sequencing of the two virulent isolates with MST22 genotype and two avirulent isolates of genotype MST91 and comparison with two well-studied M. avium subsp. hominissuis reference strains i.e. Mah 104 and Mah TH135, identified in the two MST22 isolates nine specific virulence factors of the mammalian cell entry family, that were identical with Mah 104 strain. Despite the obvious limitations of the mouse model, a striking link of virulence and identity at the genome level of porcine and human isolates with the same multisequence type, for which no correlation of place of residence (humans) or farm of origin (pigs) was observed, seems to point to the existence in the environment of certain genotypes of Mah which may be more infectious both for humans and pigs than other genotypes.

VL - 12 CP - 2 U1 - https://www.ncbi.nlm.nih.gov/pubmed/28182785?dopt=Abstract M3 - 10.1371/journal.pone.0171895 ER - TY - JOUR T1 - Virulence and immunogenicity of genetically defined human and porcine isolates of M. avium subsp. hominissuis in an experimental mouse infection JF - PLOS ONE Y1 - 2017 A1 - Nicolas Bruffaerts A1 - Vluggen, Christelle A1 - Virginie Roupie A1 - Duytschaever, Lucille A1 - Christophe Van den Poel A1 - Joseph Noël A1 - Wattiez, Ruddy A1 - Letesson, Jean-Jacques A1 - David Fretin A1 - Rigouts, Leen A1 - Elie Chapeira A1 - Vanessa Mathys A1 - Saegerman, Claude A1 - Huygen, Kris ED - Cloeckaert, Axel KW - Paratuberculosis AB -

Mycobacterium avium subsp. hominissuis (Mah) represents a health concern for humans and to a lesser extent for pigs, but its zoonotic potential remains elusive. Using multispacer sequence typing (MST) we previously identified 49 different genotypes of Mah among Belgian clinical and porcine isolates, with 5 MSTs shared by both hosts. Using experimental intranasal infection of BALB/c mice, we compared the virulence and immunogenicity of porcine and clinical human isolates with shared genotype or with a genotype only found in humans or pigs. Bacterial replication was monitored for 20 weeks in lungs, spleen and liver and mycobacteria specific spleen cell IFN-γ, IL-10 and IL-17 production as well as serum antibody responses were analyzed. Isolates varied in virulence, with human and porcine isolates sharing MST22 genotype showing a thousand fold higher bacterial replication in lungs and more dissemination to spleen and liver than the human and porcine MST91 isolates. Virulent MST22 type was also associated with progressive suppression of IFN-γ and IL-17 responses, and increased IL-10 production. Whole genome sequencing of the two virulent isolates with MST22 genotype and two avirulent isolates of genotype MST91 and comparison with two well-studied M. avium subsp. hominissuis reference strains i.e. Mah 104 and Mah TH135, identified in the two MST22 isolates nine specific virulence factors of the mammalian cell entry family, that were identical with Mah 104 strain. Despite the obvious limitations of the mouse model, a striking link of virulence and identity at the genome level of porcine and human isolates with the same multisequence type, for which no correlation of place of residence (humans) or farm of origin (pigs) was observed, seems to point to the existence in the environment of certain genotypes of Mah which may be more infectious both for humans and pigs than other genotypes.

VL - 12 CP - 2 M3 - 10.1371/journal.pone.017189510.1371/ ER - TY - Generic T1 - Détermination des sous-espèces et génotypage des souches humaines et porcines de Mycobacterium avium isolées en Belgique Y1 - 2016 A1 - Christelle Vluggen A1 - Karine Soetaert A1 - Dutschaever,L. A1 - J. Denoel A1 - Smeets,F. A1 - Nicolas Bruffaerts A1 - K Huygen A1 - David Fretin A1 - Saegerman,C. A1 - Vanessa Mathys KW - Belgique KW - de KW - EN KW - ET KW - Mycobacterium JF - Belspo RT-project PB - NA CY - NA CP - Belspo U1 - 37202 U2 - 16/03/2016 ER - TY - Generic T1 - Evolution des espèces mycobactériennes identifiées au CNR au cours du temps. Y1 - 2016 A1 - Vanessa Mathys ED - FARES-VRGT KW - de KW - espèces KW - Evolution KW - Surveillance JF - Réunion des laboratoires du réseau de surveillance de la (multi)résistance. CP - FARES U1 - 37196 U2 - 18/02/2016 ER - TY - JOUR T1 - Frequency of Mycobacterium chimaera among Belgian patients, 2015. JF - J Med Microbiol Y1 - 2016 A1 - Karine Soetaert A1 - Vluggen, Christelle A1 - Emmanuel André A1 - Vanhoof, Raymond A1 - Vanfleteren, Brigitte A1 - Vanessa Mathys KW - Adult KW - Aged KW - Aged, 80 and over KW - Belgium KW - DNA, Bacterial KW - Female KW - Humans KW - Male KW - middle aged KW - Mycobacterium KW - Mycobacterium Infections KW - Phylogeny KW - RNA, Ribosomal, 16S KW - Young adult AB -

Mycobacterium chimaera arouses an increasing public health concern, as this non-tuberculous mycobacterium (NTM) has recently been associated with life-threatening cardiac infections. M. chimaera and Mycobacteriumintracellulare are genetically very close but recently appeared to present different epidemiological and clinical significance. Therefore, it has become important for laboratories to use adequate techniques allowing precise species identification. To date, most commercially available laboratory assays cannot distinguish them and erroneously identify M. chimaera as M. intracellulare. We performed a re-analysis of the 149 M. intracellulare strains received by the Belgian National Reference Laboratory using 16S rRNA gene sequencing, representing 25 % of all NTM collected in 2015. We found that M. chimaera represents the majority (n=94, 63 %) of the previous M. intracellulare. This study reports the large presence of M. intracellulare/chimaera among Belgian patients infected by an NTM and the predominance of the species M. chimaera among this group. This study also stresses the public health importance of M. chimaera and demonstrates the inability of commonly used laboratory techniques to correctly diagnose these infections.

VL - 65 CP - 11 U1 - http://www.ncbi.nlm.nih.gov/pubmed/27902393?dopt=Abstract M3 - 10.1099/jmm.0.000359 ER - TY - JOUR T1 - Genome Sequences of Four Strains of Mycobacterium avium subsp. hominissuis, Isolated from Swine and Humans, Differing in Virulence in a Murine Intranasal Infection Model. JF - Genome Announc Y1 - 2016 A1 - Nicolas Bruffaerts A1 - Duytschaever, L A1 - Vanessa Mathys A1 - Saegerman, C A1 - Chapeira, O A1 - Huygen, K KW - genome sequence KW - hominissuis KW - Humans KW - intranasal infection model KW - murine KW - Mycobacterium avium KW - Swine KW - virulence AB -

This paper announces the genome sequences of four strains of Mycobacterium avium subsp. hominissuis, isolated from cases of lymphadenopathy in swine and humans, differing in virulence in a murine intranasal infection model.

VL - 4 CP - 3 U1 - http://www.ncbi.nlm.nih.gov/pubmed/27313293?dopt=Abstract M3 - 10.1128/genomeA.00533-16 ER - TY - JOUR T1 - Genotyping and strain distribution of Mycobacterium avium subspecies hominissuis isolated from humans and pigs in Belgium, 2011-2013. JF - Euro Surveill Y1 - 2016 A1 - Vluggen, Christelle A1 - Karine Soetaert A1 - Duytschaever, Lucille A1 - Denoël, Joseph A1 - Fauville-Dufaux, Maryse A1 - Smeets, François A1 - Nicolas Bruffaerts A1 - Huygen, Kris A1 - David Fretin A1 - Rigouts, Leen A1 - Saegerman, Claude A1 - Vanessa Mathys KW - Animals KW - Belgium KW - Genetic Variation KW - Genotype KW - Humans KW - Minisatellite Repeats KW - Mycobacterium avium KW - Phylogeny KW - polymerase chain reaction KW - Polymorphism, Restriction Fragment Length KW - Retrospective Studies KW - Sequence Analysis, DNA KW - Swine KW - Swine Diseases KW - Tuberculosis AB -

Mycobacterium avium represents a health concern for both humans and pigs. The characterisation of its subspecies is an important step improving the understanding of the epidemiology and the control of this pathogen. Ninety-two human M. avium strains were selected for a retrospective study. Subspecies determination by rpoB sequencing and IS1245/IS901 analysis showed that 98.9% of Belgian human M. avium strains belong to the subspecies hominissuis (MAH). Some of these MAH strains present particular IS1245/IS901 profiles (absence of IS1245 and false IS901 detection provoked by the presence of ISMav6). In addition, 54 MAH strains isolated from submandibular lymph nodes of Belgian pigs with lymphadenitis were included in this study. Genotyping of human and porcine isolates was performed using multispacer sequence typing (MST). In total, 49 different MST types were identified among pig (n = 11) and human (n = 43) MA isolates, with only five shared by both hosts. Among these MST types, 34 were newly identified. Our findings demonstrate the extensive genetic diversity among MAH isolates. Some genotypes were more prevalent in human or pigs but no correlation was observed between MST type and place of residence or the farm of origin for human and porcine isolates respectively, suggesting an environmental source of infection.

VL - 21 CP - 3 U1 - http://www.ncbi.nlm.nih.gov/pubmed/26835872?dopt=Abstract M3 - 10.2807/1560-7917.ES.2016.21.3.30111 ER - TY - Generic T1 - Infections mycobactériennes chez l'homme et l'animal Y1 - 2016 A1 - Vanessa Mathys A1 - David Fretin KW - ET KW - health KW - INFECTION KW - infections JF - One Health Seminar PB - NA CY - NA CP - WIV-ISP U1 - 37199 U2 - 25/05/2016 ER - TY - Generic T1 - MALDI-TOF for drug resistance testing in mycobacterium tuberculosis Y1 - 2016 A1 - Pieter-Jan Ceyssens A1 - Karine Soetaert A1 - Koen De Cremer A1 - Sophie Bertrand A1 - Marijke Hendrickx A1 - Vanessa Mathys KW - DRUG KW - Drug Resistance KW - MALDI-TOF KW - Mycobacterium KW - resistance KW - TESTING JF - ECCMID 2016 PB - NA CY - NA CP - ECCMID U1 - 39257 U2 - 09/04/2016 ER - TY - Generic T1 - MALDI-TOF for the identification of Mycobacteria and their drug resistance profiles Y1 - 2016 A1 - An Van den Bossche A1 - Pieter-Jan Ceyssens A1 - Marijke Hendrickx A1 - Vanessa Mathys KW - drug susceptibility testing KW - identification KW - MALDI-TOF KW - Mycobacterium tuberculosis AB -

In the last years, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has been increasingly introduced as a valuable method for the identification of bacteria and yeast1. This technique, which generates mass spectra that are compared to reference spectra in databases, is accurate, fast and cost-effective compared to traditional biochemical and molecular techniques.

For the identification of Mycobacteria, this method is more challenging due to the special need for inactivation and protein extraction. Although several studies have been published, there is no clear consensus on the processing of Mycobacteria for MALDI-TOF MS and outcomes vary from 55% correct identifications to 97%2-4. In this study, we optimized an extraction protocol and evaluated its efficiency using the Brüker Biotyper system v3.0 for 194 cultures. In total 88.6% (172/194) of the samples were correctly identified on the species-level using a cut-off score of 1.8 and 91.75% (178/194) using a cut-off of 1.65. All samples of the Mycobacterium tuberculosis complex were correctly classified (52/52), while 126 of the 142 Non-tuberculosis Mycobacteria were accurately identified at a 1.65 cut-off.

Recently, it was described that MALDI-TOF MS can also be used for drug-susceptibility testing (DST) in bacteria. The MS-ASTRA technology is based on the parallel incubation of a culture in presence or absence of an antibiotic. By adding an internal standard during the protein extraction, semi-quantitative measurements of the bacterial biomass can be derived from the MALDI-TOF spectra7.

In this study, we show that this technology can be applied on Mycobacteria. 34 clinical M. tuberculosis isolates were investigated for their resistance profile to four different drugs (rifampicin, isoniazid, ethambutol and linezolid), yielding a 100% concordance to the BACTEC MGIT results. Moreover, incubation with serial dilutions allowed a correct determination of the Minimal Inhibitory Concentrations of isoniazid and rifampicin. Therefore, this method has the potential to provide a cost-effective and fast alternative for classical phenotypic DST, independent of the mechanism of drug resistance. However, a current lack of automation hinders implementation in diagnostic laboratories.

JF - 37th Annual Congress of the European Society of Mycobacteriology PB - ESM CY - Catania, Sicily, Italy ER - TY - Generic T1 - Mycobacteria: interactions between human, animals and environment Y1 - 2016 A1 - Vanessa Mathys KW - Animal KW - Animals KW - environment KW - Human KW - interaction KW - interactions KW - Mycobacterium KW - study JF - FSVEE study day PB - NA CY - NA CP - KULeuven U1 - 37197 U2 - 28/10/2016 ER - TY - JOUR T1 - Nontuberculous mycobacteria among pulmonary tuberculosis patients: a retrospective Belgian multicentre study JF - Acta Clinica Belgica Y1 - 2016 A1 - De Keukeleire,S. A1 - Vanessa Mathys A1 - Van Den Wijngaert,S. A1 - Van de Vyvere,M. A1 - S. Jonckheere A1 - De Beenhouwer,H. A1 - A. De Bel A1 - W. Arrazola de Onate A1 - Wanlin,M. A1 - Denis Piérard A1 - Nulens,E. A1 - Saegeman,V. KW - a KW - Belgian KW - Mycobacterium KW - Nontuberculous Mycobacteria KW - Patient KW - patients KW - Pulmonary KW - Tuberculosis AB - . VL - . U1 - 37111 M3 - http://dx.doi.org/10.1080/17843286.2016.1200823 ER - TY - JOUR T1 - Synthesis and anti-tubercular activity of N(2)-arylbenzo[g]isoquinoline-5,10-dione-3-iminium bromides. JF - Org Biomol Chem Y1 - 2016 A1 - Rotthier, G A1 - Davie Cappoen A1 - Nguyen, Quang Trung A1 - Dang Thi, Tuyet Anh A1 - Vanessa Mathys A1 - Nguyen, Van Tuyen A1 - Huygen, K A1 - Maes, L A1 - Cos, P A1 - Abbaspour Tehrani, K KW - Antitubercular Agents KW - Dose-Response Relationship, Drug KW - Hydrocarbons, Brominated KW - Isoquinolines KW - Macrophages KW - Microbial Sensitivity Tests KW - Molecular Conformation KW - Mycobacterium tuberculosis KW - Structure-Activity Relationship KW - Tuberculosis, Multidrug-Resistant AB -

Tuberculosis has remained a challenge for medicinal chemists worldwide. In the framework of a collaborative program to identify and evaluate novel antitubercular candidate compounds, the biological properties of benzo[g]isoquinoline-5,10-diones have been found to be very promising. In this paper we have further expanded the library by incorporation of an amidinium moiety into the benzo[g]isoquinoline-5,10-dione scaffold. The presence of this functional group also increased the solubility of the quinones in polar solvents. To this purpose N(2)-arylbenzo[g]isoquinoline-5,10-dione-3-iminium bromides were synthesized in a straightforward way by means of a reaction of anilines with 2-(bromomethyl)-3-(cyanomethyl)-1,4-dimethoxynaphthalene. Following the biological evaluation, N(2)-(4-chlorophenyl)-5,10-dioxobenzo[g]isoquinoline-3(2H)-iminium bromide (MIC = 1.16 μM, CC50 = 28.51 μM, SI = 24.58) was selected as the most promising representative. Apart from the nano-molar anti-mycobacterial activity, the compound was able to target intracellular residing Mycobacterium tuberculosis and the susceptibility of a multi-drug-resistant strain towards the compound was confirmed.

VL - 14 CP - 6 U1 - http://www.ncbi.nlm.nih.gov/pubmed/26763748?dopt=Abstract M3 - 10.1039/c5ob02138c ER - TY - Generic T1 - TB-BRU-NET 2010-2013 Y1 - 2016 A1 - Vanessa Mathys A1 - Christelle Vluggen A1 - Karine Soetaert ED - FARES-VRGT KW - meeting KW - TB-BRU-NET JF - MDR meeting CP - FARES U1 - 37200 U2 - 9/06/2016 ER - TY - Generic T1 - Time and motion exercise Y1 - 2016 A1 - Vanessa Mathys ED - ERL-TB KW - Exercise KW - meeting KW - time JF - ERL-TB annual meeting CP - ERL-TB U1 - 37198 U2 - 27/04/2016 ER - TY - Generic T1 - Amélioration de l'activité thérapeutique de l'ethionamide. Y1 - 2015 A1 - Vanessa Mathys ED - FARES-VRGT KW - de JF - FARES annual meeting CP - FARES U1 - 39186 U2 - 26/02/2015 ER - TY - Generic T1 - Analysis of the virulence and immunogenicity of porcine and human M. avium ssp. hominissuis isolates in an experimental mouse model. Y1 - 2015 A1 - Nicolas Bruffaerts A1 - Christelle Vluggen A1 - L Duytschaever A1 - J. Denoel A1 - Wattiez,R. A1 - J.J. Letesson A1 - Vanessa Mathys A1 - Virginie Roupie A1 - David Fretin A1 - Saegerman,C. A1 - K Huygen ED - Research Center Borstel KW - an KW - analysi KW - analysis KW - European KW - Human KW - M KW - MODEL KW - ON KW - symposium KW - virulence JF - European Symposium on Non-tuberculous Mycobacteria CP - Research Center Borstel U1 - 37032 U2 - 24/06/2015;27/06/2015 ER - TY - Generic T1 - Antimicrobial resistance testing in 2020: Will we still be looking at the phenotype? Y1 - 2015 A1 - Pieter-Jan Ceyssens A1 - Wesley Mattheus A1 - Vanessa Mathys A1 - R. Vanhoof A1 - Michael Kalai A1 - Sophie Bertrand ED - Institut Pasteur KW - Antimicrobial KW - Antimicrobial resistance KW - at KW - Phenotype KW - resistance KW - Still KW - TESTING JF - RIIP Meeting CP - Institut Pasteur U1 - 37023 U2 - 04/2015 ER - TY - JOUR T1 - Case report of a false positive result of the Xpert MTB/RIF assay for rifampicin resistance in mycobacterium tuberculosis complex JF - Acta Clin.Belg. Y1 - 2015 A1 - Claessens,J. A1 - Vanessa Mathys A1 - Derdelinckx,I. A1 - Saegeman,V. KW - a KW - Abdomen KW - article KW - AS KW - Case KW - conventional KW - culture KW - detection KW - Diagnosis KW - DRUG KW - health KW - INFECTION KW - IS KW - IT KW - journal KW - Microscopy KW - Molecular KW - Mycobacterium KW - Mycobacterium tuberculosis KW - old KW - ON KW - organisation KW - Patient KW - present KW - Print KW - report KW - resistance KW - result KW - results KW - specificity KW - Test KW - TESTING KW - tests KW - Tuberculosis KW - use KW - Version KW - WHO KW - world KW - World Health AB - In the present case, we report a false positive result for the detection of rifampicin (RIF) resistance by the Xpert(R) MTB/RIF assay, version G4.Miliary Mycobacterium tuberculosis infection (miliary TB) was suspected in a 50-year old Angolan woman. Imaging of the thorax and abdomen displayed diffuse lesions. The Xpert(R) MTB/RIF assay conducted on the broncho-alveolar lavage (BAL) fluid was positive for TB and positive for RIF resistance. Confirmatory molecular tests and the phenotypic drug susceptibility determination supported the diagnosis of TB but not RIF resistance. The patient was treated successfully with a conventional therapeutic scheme. Because, the Xpert(R) MTB/RIF assay allows the simultaneous detection of TB and RIF resistance, the World Health Organisation (WHO) recommends its use as initial diagnostic test, over microscopy, culture and phenotypic drug susceptibility testing. Even though specificity of the Xpert(R) MTB/RIF assay version G4 is high, false positive test results remain possible and have to be considered for the interpretation of the RIF resistance detection by Xpert(R) MTB/RIF assay VL - . U1 - 37103 M3 - http://dx.doi.org/10.1179/2295333715Y.0000000072 ER - TY - JOUR T1 - Evaluation of the Speed-Oligo Mycobacteria assay for the identification of nontuberculous mycobacteria. JF - J Med Microbiol Y1 - 2015 A1 - Ivy Bastos Ramis A1 - Cnockaert, Margo A1 - Von Groll, Andrea A1 - Vanessa Mathys A1 - Simon, Anne A1 - Tortoli, Enrico A1 - Palomino, Juan Carlos A1 - Almeida da Silva, Pedro Eduardo A1 - Vandamme, Peter A1 - Emmanuel André A1 - Martin, Anandi KW - DNA Probes KW - DNA, Ribosomal KW - DNA, Ribosomal Spacer KW - Humans KW - Mycobacterium Infections, Nontuberculous KW - Nontuberculous Mycobacteria KW - Nucleic Acid Hybridization KW - Oligonucleotides KW - polymerase chain reaction KW - Reagent Kits, Diagnostic KW - RNA, Ribosomal, 16S KW - Sensitivity and Specificity KW - Sequence Analysis, DNA KW - Species Specificity KW - Time Factors AB -

Nontuberculous mycobacteria (NTM) causing human infectious disease have become increasingly common. Rapid and accurate identification to the species level is, therefore, critical. The Speed-Oligo Mycobacteria assay is an oligochromatographic method that was made available recently for the identification and differentiation of mycobacteria. The present study aimed to evaluate the performance of the Speed-Oligo Mycobacteria assay for the identification of NTM. We examined a total of 62 strains (9 type strains, 19 reference strains and 34 clinical isolates) belonging to 13 different species (Mycobacterium intracellulare, M. fortuitum, M. gordonae, M. kansasii, M. marinum, M. peregrinum, M. scrofulaceum, M. abscessus, M. bovis BCG, M. chelonae, M. avium, M. malmoense and M. xenopi). The Speed-Oligo Mycobacteria assay was performed according to the manufacturer's instructions. Discrepant results between Speed-Oligo Mycobacteria and the original identification were reassessed by the Speed-Oligo Mycobacteria assay and resolved by the GenoType Mycobacterium CM assay and by sequencing of 16S rRNA and protein-encoding genes. We found 93.5 % (58/62) concordance for the identification of NTM as compared with the original identification. Three strains were erroneously identified by Speed-Oligo Mycobacteria: one M. kansasii strain was identified as Mycobacterium tuberculosis complex, and one M. chelonae strain and one M. peregrinum strain were both identified as Mycobacterium abscessus. Moreover, one M. chelonae strain was not identified by Speed-Oligo Mycobacteria since it did not react with any species-specific probe. For these strains, sequencing of the genes hsp65, 16S rRNA and rpoB and the GenoType Mycobacterium CM assay were performed. The Speed-Oligo Mycobacteria assay can be a useful tool for the rapid and easy identification of the most common NTM. If applied in clinical practice it could reduce diagnostic delays and contribute to correct clinical and better management of infections caused by NTM.

VL - 64 CP - Pt 3 U1 - http://www.ncbi.nlm.nih.gov/pubmed/25596120?dopt=Abstract M3 - 10.1099/jmm.0.000025 ER - TY - Generic T1 - Improvement of the ETH therapeutic activity. Y1 - 2015 A1 - Vanessa Mathys KW - Activity KW - Improvement JF - MDR meeting PB - NA CY - NA CP - MDR-TB comity U1 - 39185 U2 - 11/06/2015 ER - TY - Generic T1 - Mycobacteria : current molecular laboratory techniques. Y1 - 2015 A1 - Vanessa Mathys KW - Diagnosis KW - future KW - Laboratories KW - Molecular KW - Molecular diagnosis KW - Mycobacterium KW - Technique JF - The future of the molecular diagnosis. PB - NA CY - NA CP - VAKB UZ Leuven U1 - 39187 U2 - 05/02/2015 ER - TY - Generic T1 - Mycobacterium tuberculosis & les mycobactéries atypiques: techniques de laboratoire. Y1 - 2015 A1 - Vanessa Mathys ED - FARES-VRGT KW - de KW - LE KW - Mycobactéries KW - Mycobacterium KW - Mycobacterium tuberculosis KW - Technique KW - Tuberculosis JF - Formation infirmière CP - FARES U1 - 39183 U2 - 25/09/2015 ER - TY - JOUR T1 - Revisiting susceptibility testing in MDR-TB by a standardized quantitative phenotypic assessment in a European multicentre study9 JF - J.Antimicrob.Chemother. Y1 - 2015 A1 - Cambau,E. A1 - Viveiros,M. A1 - Machado,D. A1 - Raskine,L. A1 - Ritter,C. A1 - Tortoli,E. A1 - Vanessa Mathys A1 - S Hoffner A1 - Richter,E. A1 - Perez Del Molino,M.L. A1 - D M. Cirillo A1 - D. van Soolingen A1 - Bottger,E.C. KW - 0 KW - a KW - acid KW - Agent KW - Agents KW - Amikacin KW - an KW - Antitubercular Agents KW - article KW - AS KW - assessment KW - at KW - Belgium KW - Biology KW - Brussels KW - care KW - Cell KW - Clinical KW - Concordance KW - de KW - detection KW - disease KW - Diseases KW - DRUG KW - drug effects KW - Drug Resistance KW - drugs KW - e KW - electronic KW - environment KW - Establish KW - estimation KW - ET KW - Ethionamide KW - Europe KW - European KW - EVALUATION KW - France KW - gene KW - genetics KW - Genotype KW - Genotyping Techniques KW - Germany KW - growth & development KW - health KW - Help KW - Humans KW - im KW - Impact KW - Infectious KW - Infectious diseases KW - Institute KW - IS KW - isolation & purification KW - ITALY KW - journal KW - Laboratories KW - LEVEL KW - levels KW - medical KW - method KW - methods KW - Microbial Sensitivity Tests KW - microbiology KW - Mutation KW - Mycobacterium KW - Mycobacterium tuberculosis KW - national KW - Netherlands KW - objectives KW - observed KW - ON KW - outcome KW - paris KW - pathogen KW - Patient KW - Patient Care KW - patients KW - pharmacology KW - portugal KW - proportion KW - protocol KW - public KW - public health KW - Public-health KW - Pulmonary KW - Pyrazinamide KW - region KW - resistance KW - result KW - results KW - SB - IM KW - SCREENING KW - Service KW - Short KW - SOFTWARE KW - Spain KW - standards KW - Streptomycin KW - study KW - Sweden KW - Switzerland KW - Technique KW - TESTING KW - The Netherlands KW - therapeutic use KW - time KW - treatment KW - Treatment Outcome KW - Tuberculosis KW - Tuberculosis,Multidrug-Resistant KW - tumor KW - Universities KW - university KW - use KW - VALIDATION KW - WIV-ISP AB - OBJECTIVES: Treatment outcome of MDR-TB is critically dependent on the proper use of second-line drugs as per the result of in vitro drug susceptibility testing (DST). We aimed to establish a standardized DST procedure based on quantitative determination of drug resistance and compared the results with those of genotypes associated with drug resistance. METHODS: The protocol, based on MGIT 960 and the TB eXiST software, was evaluated in nine European reference laboratories. Resistance detection at a screening drug concentration was followed by determination of resistance levels and estimation of the resistance proportion. Mutations in 14 gene regions were investigated using established techniques. RESULTS: A total of 139 Mycobacterium tuberculosis isolates from patients with MDR-TB and resistance beyond MDR-TB were tested for 13 antituberculous drugs: isoniazid, rifampicin, rifabutin, ethambutol, pyrazinamide, streptomycin, para-aminosalicylic acid, ethionamide, amikacin, capreomycin, ofloxacin, moxifloxacin and linezolid. Concordance between phenotypic and genotypic resistance was >80%, except for ethambutol. Time to results was short (median 10 days). High-level resistance, which precludes the therapeutic use of an antituberculous drug, was observed in 49% of the isolates. The finding of a low or intermediate resistance level in 16% and 35% of the isolates, respectively, may help in designing an efficient personalized regimen for the treatment of MDR-TB patients. CONCLUSIONS: The automated DST procedure permits accurate and rapid quantitative resistance profiling of first- and second-line antituberculous drugs. Prospective validation is warranted to determine the impact on patient care VL - 70 CP - 3 U1 - 9 M3 - dku438 [pii];10.1093/jac/dku438 [doi] ER - TY - Generic T1 - Time and motion exercise: MIRU-VNTR typing. Y1 - 2015 A1 - Vanessa Mathys ED - ERL-TB KW - Exercise KW - time JF - ERL-TB net annual meeting CP - ERL-TB U1 - 38293 U2 - 12/05/2015 ER - TY - Generic T1 - Tuberculose humaine Y1 - 2015 A1 - Vanessa Mathys ED - Scicom ED - FAVV-AFSCA KW - tuberculose JF - Scicom CP - Scicom, AFSCA U1 - 37015 U2 - 26/08/2015 ER - TY - JOUR T1 - 1,2,3,4,8,9,10,11-Octahydrobenzo[j]phenanthridine-7,12-diones as New Leads against Mycobacterium tuberculosis JF - J.Med.Chem. Y1 - 2014 A1 - Davie Cappoen A1 - Claes,P. A1 - J. Jacobs A1 - R. Anthonissen A1 - Vanessa Mathys A1 - Luc Verschaeve A1 - K Huygen A1 - Kimpe,N.D. KW - Activity KW - acute KW - Agent KW - article KW - AS KW - at KW - Belgium KW - continue KW - Death KW - Design KW - Diagnostics KW - disease KW - Diseases KW - DRUG KW - Drug Resistance KW - drugs KW - effective KW - electronic KW - Exploration KW - genotoxicity KW - global KW - health KW - HIV KW - immunology KW - Infectious KW - Infectious diseases KW - Institute KW - IS KW - journal KW - Lead KW - Mycobacterium KW - Mycobacterium tuberculosis KW - pandemic KW - PROGRAM KW - public KW - public health KW - Public-health KW - Research KW - resistance KW - Service KW - strain KW - study KW - Target KW - toxicity KW - Tuberculosis KW - vaccine KW - vaccines AB - Tuberculosis (TB) continues to be a worldwide health problem with over 1.4 million deaths each year. Despite efforts to develop more effective vaccines, more reliable diagnostics, and chemotherapeutics, tuberculosis remains a threat to global health, fueled by the HIV pandemic and the rapid generation of drug resistance. The exploration of novel drugs to serve as a companion drug for existing drugs is of paramount importance. As part of our program to design new 2-aza-anthraquinones with antimycobacterial activity, various tetrahydro- and octahydrobenzo[j]phenanthridinediones were synthesized. These compounds showed high in vitro potency against Mycobacterium tuberculosis, the etiological agent of TB and against other clinically relevant mycobacterial species at submicromolar concentrations. The susceptibility of a multidrug resistant strain toward these compounds and their ability to target intracellular replicating Mycobacterium tuberculosis was demonstrated. Next to the acute toxicity, the genotoxicity of these compounds was investigated. Often overlooked in studies, genotoxicity could be dismissed for the investigated compounds, making them a promising scaffold in TB drug research VL - 57 CP - 7 U1 - 37497 M3 - http://dx.doi.org/10.1021/jm401735w ER - TY - JOUR T1 - 2,4-Dialkyl-8,9,10,11-tetrahydrobenzo[g]pyrimido[4,5-c]isoquinoline-1,3,7,12(2H,4H)-tetraones as new leads against Mycobacterium tuberculosis. JF - Eur J Med Chem Y1 - 2014 A1 - Claes, Pieter A1 - Davie Cappoen A1 - Uythethofken, Cynthia A1 - Jacobs, Jan A1 - Birgit Mertens A1 - Vanessa Mathys A1 - Luc Verschaeve A1 - Huygen, Kris A1 - De Kimpe, Norbert KW - Animals KW - Antitubercular Agents KW - Cell Line KW - Dose-Response Relationship, Drug KW - Isoquinolines KW - Macrophages KW - mice KW - Microbial Sensitivity Tests KW - Molecular Structure KW - Mycobacterium tuberculosis KW - Pyrimidinones KW - Quinones KW - Structure-Activity Relationship KW - Tuberculosis, Multidrug-Resistant AB -

Given the re-emergence of tuberculosis in Europe and beyond, the search for novel bio-active compound classes against this disease is of utmost importance. As a result of a high intrinsic tolerance of the etiological agent, Mycobacterium tuberculosis, towards most antibiotics and xenobiotics, the search for such new compounds is far from trivial. Further exacerbated by the rapid generation and spread of drug resistant M. tuberculosis and fuelled by the HIV/AIDS pandemic, halting the tuberculosis epidemic is of paramount importance. As part of our program to design new 2-aza-anthraquinones with anti-mycobacterial activity, various dialkyltetrahydrobenzo[g]pyrimido[4,5-c]isoquinolinetetraones were designed and synthesised. The compounds were submitted to a biological evaluation in which the activity against M.tb H37Rv(lux) was observed, as well as the acute toxicity towards J774 A.1 macrophages. From these results, the selectivity index was calculated. Furthermore, the activity of the most promising compounds was further studied against a multi-drug resistant LAM-1 strain and against intracellular replicating M.tb. The study was further extended with a comet assay and a VITOTOX™ assay to investigate the possibility of observable genotoxic effects caused by these compounds.

VL - 77 U1 - http://www.ncbi.nlm.nih.gov/pubmed/24681334?dopt=Abstract M3 - 10.1016/j.ejmech.2014.03.024 ER - TY - JOUR T1 - Anti-mycobacterial activity of 1,3-diaryltriazenes. JF - Eur J Med Chem Y1 - 2014 A1 - Davie Cappoen A1 - Vajs, Jure A1 - Uythethofken, Cynthia A1 - Virag, Andrej A1 - Vanessa Mathys A1 - Kočevar, Marijan A1 - Luc Verschaeve A1 - Gazvoda, Martin A1 - Polanc, Slovenko A1 - Huygen, Kris A1 - Košmrlj, Janez KW - Animals KW - Antitubercular Agents KW - Cell Line KW - Cell Survival KW - Dose-Response Relationship, Drug KW - Macrophages KW - mice KW - Microbial Sensitivity Tests KW - Molecular Structure KW - Mycobacterium avium KW - Mycobacterium bovis KW - Mycobacterium ulcerans KW - Structure-Activity Relationship KW - Triazenes AB -

The rapid generation and spread of the drug resistant tuberculosis has led to an ongoing demand for novel compounds for therapeutic use. Identification and study of compounds with the ability to inhibit Mycobacterium tuberculosis is of paramount importance. For this reason, a library of substituted 1,3-diaryltriazenes based on the acting component of the anti-trypanosomal drug, diminazene aceturate was created and evaluated for its potential as anti-tubercular agent. Several compounds were identified with sub-micro molar inhibitory concentrations against M. tuberculosis and other clinically relevant mycobacterial species such as Mycobacterium bovis, Mycobacterium avium and Mycobacterium ulcerans. Although the library of the compounds showed a considerable acute cytotoxicity, a genotoxicity could not be observed. Finally, the triazene 14 was selected with the best biological properties (IC50 = 3.26 μM, NI50 = 24.22 μM, SI = 7.44). The compound 14 showed the ability to inhibit the growth of intracellular replicating and multi-drug resistant M. tuberculosis. The results suggest the molecule to be an interesting scaffold for further study and optimization.

VL - 77 U1 - http://www.ncbi.nlm.nih.gov/pubmed/24631899?dopt=Abstract M3 - 10.1016/j.ejmech.2014.02.065 ER - TY - JOUR T1 - Biological evaluation of diazene derivatives as anti-tubercular compounds. JF - Eur J Med Chem Y1 - 2014 A1 - Davie Cappoen A1 - Majce, Vita A1 - Uythethofken, Cynthia A1 - Urankar, Damijana A1 - Vanessa Mathys A1 - Kočevar, Marijan A1 - Luc Verschaeve A1 - Polanc, Slovenko A1 - Huygen, Kris A1 - Košmrlj, Janez KW - Antitubercular Agents KW - Drug Screening Assays, Antitumor KW - Imides AB -

Despite efforts made in chemotherapeutic research in the past and present, Mycobacterium tuberculosis (M.tb), the etiological agent of tuberculosis, still causes more than a million deadly casualties each year, second only to HIV. The rapid generation and spread of drug resistant strains, a problem exacerbated by co-infection with HIV demands further efforts in the investigation of novel classes of anti-tubercular compounds. A library of eight substituted diazenecarboxamides, three carbamoyldiazenecarboxylates and four diazene-1,2-dicarboxamides was synthesized in a straightforward manner followed by a biological evaluation of the compounds. We observed minimal inhibitory concentrations below 10 μg/mL against the H37Rv lab strain of M.tb. Three compounds that showed a potency of 90% growth inhibition of M.tb at a concentration lower than 10 μg/mL were further evaluated and showed potency against other clinically relevant mycobacterial species such as Mycobacterium bovis, Mycobacterium avium and Mycobacterium ulcerans. The selected compounds were examined for acute cell toxicity on a murine macrophage like monocyte cell line J774 A.1 in which the cell viability was reduced by 50% at concentrations ranging from 7.4 μg/mL to 20.7 μg/mL. Neither of the three compounds showed signs of genotoxicity by VITOTOX or by Comet assay. The study was complemented by demonstration of the inhibition of intracellular replication of M.tb H37Rv inside J774 A.1 cells at 2 μg/mL concentration and the susceptibility of a MDR LAM-1 strain at concentrations between 5 and 1 μg/mL of the most active compound.

VL - 74 U1 - http://www.ncbi.nlm.nih.gov/pubmed/24448419?dopt=Abstract M3 - 10.1016/j.ejmech.2013.12.057 ER - TY - JOUR T1 - False-positive rifampicin resistance on Xpert® MTB/RIF caused by a silent mutation in the rpoB gene. JF - Int J Tuberc Lung Dis Y1 - 2014 A1 - Vanessa Mathys A1 - van de Vyvere, M A1 - de Droogh, E A1 - Groenen, G KW - Aged KW - Amikacin KW - Amino Acid Sequence KW - Antibiotics, Antitubercular KW - Bacterial Proteins KW - DNA-Directed RNA Polymerases KW - Drug Resistance, Multiple, Bacterial KW - Ethambutol KW - Ethionamide KW - Fluoroquinolones KW - Genotype KW - Humans KW - Isoniazid KW - Male KW - Molecular Sequence Data KW - Mutation KW - Mycobacterium tuberculosis KW - Phenotype KW - Pyrazinamide KW - Rifampin KW - Sensitivity and Specificity KW - sputum KW - Tuberculosis, Pulmonary AB -

The Xpert® MTB/RIF assay detects the presence of Mycobacterium tuberculosis and its resistance to rifampicin (RMP) directly in sputum samples. Discrepant results were observed in a case of smear-positive pulmonary tuberculosis that was Xpert-resistant but phenotypically susceptible to RMP. Complementary investigations (repeat Xpert, Genotype®MTBDRplus assay and sequencing of the rpoB gene) revealed the presence of a silent mutation in the rpoB gene, leading to the conclusion of a false-positive Xpert result. As misinterpretation of Xpert results may lead to inappropriate treatment, the presence of rpoB mutations should be confirmed by sequencing the rpoB gene.

VL - 18 CP - 10 U1 - http://www.ncbi.nlm.nih.gov/pubmed/25216843?dopt=Abstract M3 - 10.5588/ijtld.14.0297 ER - TY - Generic T1 - Simultaneous determination of anti-tuberculosis drug levels in human plasma by ultra-performance liquid chromatography-tandem mass spectrometry for subsequent Therapeutic Drug Monitoring Y1 - 2014 A1 - Tim Reyns A1 - Vanessa Mathys A1 - Groenen,G. A1 - Mirjana Andjelkovic A1 - Joris Van Loco KW - alternative KW - Anti tubercolosis drugs KW - DRUG KW - Drug Monitoring KW - Human KW - LC-MS/MS KW - LEVEL KW - levels KW - Liquid chromatography-tandem mass spectrometry KW - Mass KW - Mass Spectrometry KW - meeting KW - Monitoring KW - plasma KW - Strategies KW - Strategy KW - TDM KW - Therapeutic drug monitoring KW - Toxicology KW - ultra-performance liquid chromatography-tandem mass spectrometry JF - Alternative Sampliong Strategies in Toxicology and Therapeutic Drug Monitoring - IATDMCT Satellite Meeting 2014 PB - NA CY - NA CP - IATDMCT U1 - 2270 U2 - 18/09/2014 - 19/09/2014 ER - TY - JOUR T1 - Biological evaluation of bisbenzaldehydes against four Mycobacterium species. JF - Eur J Med Chem Y1 - 2013 A1 - Davie Cappoen A1 - Forge, Delphine A1 - Vercammen, Frank A1 - Vanessa Mathys A1 - Kiass, Mehdi A1 - Virginie Roupie A1 - Anthonissen, Roel A1 - Luc Verschaeve A1 - Vanden Eynde, Jean Jacques A1 - Huygen, Kris KW - Animals KW - Anti-Bacterial Agents KW - Antitubercular Agents KW - Benzaldehydes KW - Cell Line KW - Cell Survival KW - Cells, Cultured KW - Dose-Response Relationship, Drug KW - Drug Resistance, Bacterial KW - Hepatocytes KW - Host-Pathogen Interactions KW - Humans KW - Macrophages KW - mice KW - Microbial Sensitivity Tests KW - Molecular Structure KW - Mycobacterium KW - Mycobacterium avium KW - Mycobacterium bovis KW - Mycobacterium tuberculosis KW - Mycobacterium ulcerans KW - Species Specificity AB -

A series of bisbenzaldehydes and structurally related analogs, conveniently synthesized via microwave-assisted reactions, were evaluated in vitro against drug susceptible and multi-drug resistant Mycobacterium tuberculosis, against virulent Mycobacterium bovis, against Mycobacterium ulcerans and against two Mycobacterium avium subspecies. Among the 33 substances that were tested, compound 12, i.e. 4,4'-[1,12-dodecanediyl(oxy)]bisbenzaldehyde, emerged as the most promising hit. Its activity was further confirmed in an intracellular growth inhibition assay of M. tb in murine J774 A.1 macrophages. None of the compounds showed significant cytotoxicity on human C3A hepatocytes in a neutral red dye uptake assay and no genotoxicity or mutagenicity was observed as demonstrated by a VITOTOX™ test and confirmed with a comet assay.

VL - 63 U1 - http://www.ncbi.nlm.nih.gov/pubmed/23567963?dopt=Abstract M3 - 10.1016/j.ejmech.2013.03.023 ER - TY - RPRT T1 - Evaluation externe de qualité 2013 en microbiologie. Identification de Mycobactéries et test de sensibilité de Mycobacterium tuberculosis à l'isoniazide (INH), la rifampicine (RMP), l'éthambutol (EMB) et facultatif la pyrazinamide (PZA). Y1 - 2013 A1 - Vanessa Mathys A1 - Bernard China A1 - Kris Vernelen KW - antibiotics KW - bactériologie KW - biologie clinique KW - de KW - EEQ KW - EVALUATION KW - identification KW - microbiologie KW - Mycobactéries KW - Mycobacterium KW - Pyrazinamide KW - Test KW - Tuberculosis PB - WIV-ISP CY - Bruxelles U1 - 567 ER - TY - JOUR T1 - From multidrug- to extensively drug-resistant tuberculosis: upward trends as seen from a 15-year nationwide study. JF - PLoS One Y1 - 2013 A1 - Stoffels, Karolien A1 - Allix-Béguec, Caroline A1 - Groenen, Guido A1 - Wanlin, Maryse A1 - Berkvens, Dirk A1 - Vanessa Mathys A1 - Supply, Philip A1 - Fauville-Dufaux, Maryse KW - ADOLESCENT KW - Adult KW - Antitubercular Agents KW - Belgium KW - Extensively Drug-Resistant Tuberculosis KW - Female KW - health surveys KW - Humans KW - incidence KW - Male KW - Mutation KW - Mycobacterium tuberculosis KW - REGISTRIES KW - Treatment Outcome KW - Tuberculosis, Multidrug-Resistant KW - Young adult AB -

BACKGROUND: Emergence of extensively drug-resistant tuberculosis (XDR-TB) represents an enormous challenge to Public Health globally.

METHODS: Progression towards XDR-TB was investigated in Belgium, a country with a typically low TB incidence, by analyzing the magnitude, characteristics, and treatment success of multidrug-resistant tuberculosis (MDR-TB) through a population-based study from 1994 to 2008.

RESULTS: Among the 174 MDR-TB patients, 81% were foreign-born, 48% of these being asylum seekers. Although the number of MDR-TB patients remained stable through the study period at around 15 new cases annually, frequencies of resistance of the patients' first MDR-TB isolate to second-line drugs increased, as well as the total number of antibiotics it was resistant to (p<0.001). XDR-TB cases were detected from 2002 onwards. For 24 patients, additional resistance to several second-line drugs was acquired during treatment. Molecular-guided investigations indicated little to no contribution of in-country clonal spread or exogenous re-infection. The increase of pre-XDR and XDR cases could be attributed to rising proportions of patients from Asia and Central and Eastern Europe (p<0.001) and an increase in the isolation of Beijing strains in these groups (p<0.001). Despite augmented resistance, the treatment success rate improved from 63.0% to 75.8% (p = 0.080) after implementation in 2005 of improved surveillance measures and therapeutic access.

CONCLUSIONS: Increasing severity in drug resistance patterns leading to more XDR- and "panresistant" TB cases in a country with a low TB incidence like Belgium represents a strong alert on worsening situations in other world regions and requires intense public health measures.

VL - 8 CP - 5 U1 - http://www.ncbi.nlm.nih.gov/pubmed/23671662?dopt=Abstract M3 - 10.1371/journal.pone.0063128 ER - TY - JOUR T1 - Para-Aminosalicylic acid is a prodrug targeting dihydrofolate reductase in Mycobacterium tuberculosis33991 JF - J.Biol.Chem. Y1 - 2013 A1 - Zheng,J. A1 - Rubin,E.J. A1 - Bifani,P. A1 - Vanessa Mathys A1 - Lim,V. A1 - Au,M. A1 - Jang,J. A1 - Nam,J. A1 - Dick,T. A1 - Walker,J.R. A1 - Pethe,K. A1 - L. Camacho KW - 0 KW - a KW - acid KW - Activity KW - Agent KW - Agents KW - Aminosalicylic Acid KW - an KW - antagonists & inhibitors KW - Antitubercular Agents KW - article KW - AS KW - Bacterial Proteins KW - Benzoic Acid KW - biosynthesis KW - Clinical KW - Dihydropteroate Synthase KW - disease KW - Diseases KW - DRUG KW - drugs KW - electronic KW - evidence KW - Folate KW - Folic Acid KW - Folic Acid Antagonists KW - Folic-acid KW - Functional KW - Gene Knockdown Techniques KW - genetics KW - identification KW - im KW - Institute KW - IS KW - IT KW - journal KW - Laboratories KW - mechanism KW - metabolism KW - Mutation KW - Mycobacterium KW - Mycobacterium tuberculosis KW - Pa KW - PAS KW - pharmacokinetics KW - pharmacology KW - prevent KW - Prodrugs KW - protein KW - Proteins KW - report KW - Research KW - Research Support KW - resistance KW - SB - IM KW - strain KW - Tetrahydrofolate Dehydrogenase KW - Tuberculosis KW - use AB - para-Aminosalicylic acid (PAS) is one of the antimycobacterial drugs currently used for multidrug-resistant tuberculosis. Although it has been in clinical use for over 60 years, its mechanism(s) of action remains elusive. Here we report that PAS is a prodrug targeting dihydrofolate reductase (DHFR) through an unusual and novel mechanism of action. We provide evidences that PAS is incorporated into the folate pathway by dihydropteroate synthase (DHPS) and dihydrofolate synthase (DHFS) to generate a hydroxyl dihydrofolate antimetabolite, which in turn inhibits DHFR enzymatic activity. Interestingly, PAS is recognized by DHPS as efficiently as its natural substrate para-amino benzoic acid. Chemical inhibition of DHPS or mutation in DHFS prevents the formation of the antimetabolite, thereby conferring resistance to PAS. In addition, we identified a bifunctional enzyme (riboflavin biosynthesis protein (RibD)), a putative functional analog of DHFR in a knock-out strain. This finding is further supported by the identification of PAS-resistant clinical isolates encoding a RibD overexpression mutation displaying cross-resistance to genuine DHFR inhibitors. Our findings reveal that a metabolite of PAS inhibits DHFR in the folate pathway. RibD was shown to act as a functional analog of DHFR, and as for DHFS, both were shown to be associated in PAS resistance in laboratory strains and clinical isolates VL - 288 CP - 32 U1 - 38480 M3 - http://dx.doi.org/10.1074/jbc.M113.475798 ER - TY - JOUR T1 - Synthesis and antimycobacterial activity of analogues of the bioactive natural products sampangine and cleistopholine36497 JF - Eur.J.Med.Chem. Y1 - 2013 A1 - Claes,P. A1 - Davie Cappoen A1 - Mbala,B.M. A1 - J. Jacobs A1 - Birgit Mertens A1 - Vanessa Mathys A1 - Luc Verschaeve A1 - K Huygen A1 - N. De Kimpe KW - acute toxicity KW - antibiotics KW - Cleistopholine KW - Pyridinium ylids KW - Sampangine KW - Tuberculosis AB -

Identification and investigation of novel classes and compounds for the treatment of tuberculosis remains of utmost importance in the fight against the disease. Despite many efforts, the weakly gram positive Mycobacterium tuberculosis keeps demanding its toll in human lives. For this reason a small library of substituted and unsubstituted aza analogues of cleistopholine and sampangine were synthesized in a short and straightforward manner and tested in vitro against M.tb. The compounds showed promising activity against the M.tb H37Rv strain and Minimal Inhibitory Concentrations (MIC) could be observed as low as 0.88 muM. Accompanied by moderate acute toxicity against C3A hepatocytes, the therapeutic index showed an acceptable range. Further tests confirmed the inhibition by up to 74% of intracellular growth of M.tb inside macrophages conferred by 1-hydroxybenzo[g]isoquinoline-5,10-diones. Activity of the library against other clinically relevant mycobacterial species such as Mycobacterium bovis, Mycobacterium avium and Mycobacterium ulcerans was confirmed. Furthermore the activity against a multi-drug-resistant MDR LAM-1 M.tb strain was tested and the MIC value situated around 1 muM. The lacking genotoxicity of a group of enamine substituted cleistopholine analogues indicates this group as a hit and encourages their use as a scaffold for further studies

VL - 67 U1 -

36497

M3 - http://dx.doi.org/10.1016/j.ejmech.2013.06.010 ER - TY - JOUR T1 - Ethionamide boosters. 2. Combining bioisosteric replacement and structure-based drug design to solve pharmacokinetic issues in a series of potent 1,2,4-oxadiazole EthR inhibitors31058 JF - J.Med.Chem. Y1 - 2012 A1 - Flipo,M. A1 - Desroses,M. A1 - Lecat-Guillet,N. A1 - Villemagne,B. A1 - Blondiaux,N. A1 - Leroux,F. A1 - Piveteau,C. A1 - Vanessa Mathys A1 - Flament,M.P. A1 - Siepmann,J. A1 - Villeret,V. A1 - Wohlkonig,A. A1 - Wintjens,R. A1 - Soror,S.H. A1 - Christophe,T. A1 - Jeon,H.K. A1 - Locht,C. A1 - Brodin,P. A1 - Deprez,B. A1 - Baulard,A.R. A1 - Willand,N. KW - 0 KW - a KW - Administration,Oral KW - Agent KW - Agents KW - Animals KW - antagonists & inhibitors KW - Antibiotic KW - Antitubercular Agents KW - article KW - Cell Line KW - chemical synthesis KW - chemistry KW - Combination KW - Control KW - Crystallography,X-Ray KW - de KW - Design KW - DRUG KW - Drug Design KW - drug effects KW - Drug Synergism KW - electronic KW - Ethionamide KW - EVALUATION KW - exposure KW - expression KW - France KW - Human KW - im KW - IS KW - journal KW - Macrophages KW - metabolism KW - mice KW - microbiology KW - Microsomes,Liver KW - Models,Molecular KW - Mycobacterium KW - Mycobacterium tuberculosis KW - ON KW - Oxadiazoles KW - pathogen KW - Pharmacokinetic KW - pharmacokinetics KW - Piperidines KW - PROCESSES KW - Prodrugs KW - protein KW - Proteins KW - Repressor Proteins KW - Research KW - Research Support KW - SB - IM KW - SENSITIVITY KW - series KW - stability KW - Stereoisomerism KW - Structure-Activity Relationship KW - study KW - Tuberculosis KW - work AB - Mycobacterial transcriptional repressor EthR controls the expression of EthA, the bacterial monooxygenase activating ethionamide, and is thus largely responsible for the low sensitivity of the human pathogen Mycobacterium tuberculosis to this antibiotic. We recently reported structure-activity relationships of a series of 1,2,4-oxadiazole EthR inhibitors leading to the discovery of potent ethionamide boosters. Despite high metabolic stability, pharmacokinetic evaluation revealed poor mice exposure; therefore, a second phase of optimization was required. Herein a structure-property relationship study is reported according to the replacement of the two aromatic heterocycles: 2-thienyl and 1,2,4-oxadiazolyl moieties. This work was done using a combination of structure-based drug design and in vitro/ex vivo evaluations of ethionamide boosters on the targeted protein EthR and on the human pathogen Mycobacterium tuberculosis. Thanks to this process, we identified compound 42 (BDM41906), which displays improved efficacy in addition to high exposure to mice after oral administration VL - 55 CP - 1 U1 - 38193 M3 - http://dx.doi.org/10.1021/jm200825u ER - TY - JOUR T1 - Evolutionary changes in antimicrobial resistance of invasive Neisseria meningitidis isolates in Belgium from 2000 to 2010: increasing prevalence of penicillin nonsusceptibility. JF - Antimicrob Agents Chemother Y1 - 2012 A1 - Sophie Bertrand A1 - Carion, Françoise A1 - Wintjens, René A1 - Vanessa Mathys A1 - Vanhoof, Raymond KW - 3' Flanking Region KW - alleles KW - Anti-Bacterial Agents KW - Bacterial Typing Techniques KW - Belgium KW - Biological Evolution KW - Ceftriaxone KW - Drug Resistance, Bacterial KW - Gene Frequency KW - Genes, Bacterial KW - Humans KW - Longitudinal Studies KW - Meningococcal Infections KW - Microbial Sensitivity Tests KW - Neisseria meningitidis KW - Penicillin G KW - Penicillin-Binding Proteins KW - Sequence Analysis, DNA AB -

This study was conducted to evaluate the evolution of the antimicrobial susceptibility of Neisseria meningitidis causing invasive diseases in Belgium in the period of January 2000 to December 2010. A total of 1,933 cases of N. meningitidis from invasive infections were analyzed by antimicrobial susceptibility testing at the Belgian Meningococcal Reference Centre. The majority of strains were susceptible to antibiotics that are currently used for the treatment and prophylaxis of meningococcal disease, but the prevalence of clinical isolates with reduced susceptibility to penicillin increased over the years. The phenotyping, genotyping, and determination of MICs of penicillin G were performed. The systematic shift of the curves toward higher penicillin MICs in the susceptible population indicated that this population became less sensitive to penicillin in this period. A 402-bp DNA fragment in the 3' end of penA was sequenced for the 296 nonsusceptible meningococcal strains isolated between 2000 and 2010 to examine the genetic diversity and evolution of their penA gene. In conclusion, the data obtained in our study support the statement that the position of penicillin G as a first choice in the treatment of invasive meningococcal diseases in Belgium should be reexamined. Despite an important number of isolates displaying a reduced susceptibility to penicillin, at present the expanded-spectrum cephalosporins, such as ceftriaxone, are not affected. The follow-up of the evolutionary changes in antimicrobial resistance has also proved to be essential for the recommendation of an appropriate antimicrobial treatment for invasive meningococcal diseases.

VL - 56 CP - 5 U1 - http://www.ncbi.nlm.nih.gov/pubmed/22290951?dopt=Abstract M3 - 10.1128/AAC.06310-11 ER - TY - Generic T1 - The fluoropyrimidine chemotherapeutic agent 5-fluorouracil displays strong anti-tubercular activity in vitro Y1 - 2012 A1 - Vanessa Mathys A1 - Singhal,A. A1 - P. Lefevre A1 - Kiass,M. A1 - Wang,X.M. A1 - Mathema,B. A1 - Kurepina,N. A1 - B.N. Kreiswirth A1 - Bifani,P. KW - Activity KW - Agent KW - general KW - meeting JF - 107th General Meeting PB - NA CY - NA CP - American Society for Microbiology (ASM) U1 - 38291 U2 - 05/06/2008 ER - TY - Generic T1 - Le rat Wistar : modèle pour la tuberculose latente? Y1 - 2012 A1 - Vanessa Mathys ED - WIV-ISP KW - Antibiotic KW - antibiotics KW - journal KW - latence KW - LE KW - tuberculose KW - Wistar rat JF - Journal Club Antibiotics (JCA) PB - NA CY - NA CP - Scientific Institute of Public Health, Service Maladies Bactériennes U1 - 38292 U2 - 21/09/2011 ER - TY - JOUR T1 - Systematic analysis of pyrazinamide-resistant spontaneous mutants and clinical isolates of Mycobacterium tuberculosis. JF - Antimicrob Agents Chemother Y1 - 2012 A1 - Stoffels, Karolien A1 - Vanessa Mathys A1 - Fauville-Dufaux, Maryse A1 - Wintjens, René A1 - Bifani, Pablo KW - Amidohydrolases KW - Antitubercular Agents KW - Crystallography, X-Ray KW - Microbial Sensitivity Tests KW - Mutation KW - Mycobacterium tuberculosis KW - Pyrazinamide AB -

Pyrazinamide (PZA) is a first-line antitubercular drug known for its activity against persistent Mycobacterium tuberculosis bacilli. We set out to systematically determine the PZA susceptibility profiles and mutations in the pyrazinamidase (pncA) gene of a collection of multidrug-resistant tuberculosis (MDR-TB) clinical isolates and PZA-resistant (PZA(r)) spontaneous mutants. The frequency of acquired resistance to PZA was determined to be 10(-5) bacilli in vitro. Selection at a lower concentration of PZA yielded a significantly larger number of spontaneous mutants. The methodical approach employed allowed for determination of the frequency of the PZA(r) phenotype correlated with mutations in the pncA gene, which was 87.5% for the laboratory-selected spontaneous mutants examined in this study. As elucidated by structural analysis, most of the identified mutations were foreseen to affect protein activity through either alteration of an active site residue or destabilization of protein structure, indicating some preferential mutation site rather than random scattering. Twelve percent of the PZA(r) mutants did not have a pncA mutation, strongly indicating the presence of at least one other mechanism(s) of PZA(r).

VL - 56 CP - 10 U1 - http://www.ncbi.nlm.nih.gov/pubmed/22825123?dopt=Abstract M3 - 10.1128/AAC.05385-11 ER - TY - JOUR T1 - Urease activity represents an alternative pathway for Mycobacterium tuberculosis nitrogen metabolism31064 JF - Infect.Immun. Y1 - 2012 A1 - Lin,W. A1 - Vanessa Mathys A1 - Ang,E.L. A1 - Koh,V.H. A1 - J.M. Martinez Gomez A1 - Ang,M.L. A1 - S.Z. Zainul Rahim A1 - Tan,M.P. A1 - Pethe,K. A1 - Alonso,S. KW - a KW - Activity KW - alternative KW - an KW - article KW - AS KW - bacteria KW - Carbon KW - data KW - effect KW - electronic KW - environment KW - Functional KW - growth KW - Help KW - immunology KW - INFECTION KW - IS KW - IT KW - journal KW - Liver KW - Lung KW - M KW - Macrophage KW - Macrophages KW - Medicine KW - metabolism KW - mice KW - microbiology KW - Mycobacterium KW - Mycobacterium tuberculosis KW - national KW - Nitrogen KW - pathogen KW - pathogenic KW - profile KW - Profiles KW - programme KW - Role KW - School KW - Spleen KW - strain KW - survival KW - Tuberculosis KW - Type KW - Universities KW - university KW - Urea KW - urease KW - virulence KW - work AB - Urease represents a critical virulence factor for some bacteria species through its alkalizing effect that helps neutralize the acidic microenvironment of the pathogen. In addition, urease serves as a nitrogen source provider for bacterial growth. Pathogenic mycobacteria express a functional urease but its role during infection has yet to be characterized. Here, we constructed a urease-deficient M. tuberculosis (Mtb) strain and confirmed the alkalizing effect of the urease activity within the mycobacteria-containing vacuole in resting macrophages but not in the more acidic phagolysosomal compartment of activated macrophages. However, the urease-mediated alkalizing effect did not confer any growth advantage to Mtb in macrophage as evidenced by comparable growth profiles for the mutant, wild type (WT) and complemented strains. In contrast, the urease-deficient mutant exhibited impaired in vitro growth compared to the WT and complemented strains when urea is the sole source of nitrogen. Substantial amounts of ammonia were produced by the WT and complemented strains, but not with the urease-deficient mutant, which represents the actual nitrogen source for mycobacterial growth. However, the urease-deficient mutant displayed parental colonization profiles in the lungs, spleen and liver in mice. Together, our data demonstrate a role of the urease activity in Mtb nitrogen metabolism that could be crucial for the pathogen's survival in nutrient-limited microenvironments where urea is the sole nitrogen source. Our work supports the notion that Mtb virulence correlates with its unique metabolic versatility and ability to utilize virtually any carbon and nitrogen sources available in its environment VL - 80 CP - 8 U1 - 31064 M3 - http://dx.doi.org/10.1128/IAI.06195-11 ER - TY - JOUR T1 - BCG induces protection against Mycobacterium tuberculosis infection in the Wistar rat model36573 JF - PLoS.One. Y1 - 2011 A1 - Singhal,A. A1 - Vanessa Mathys A1 - Kiass,M. A1 - Creusy,C. A1 - Delaire,B. A1 - M. El Aliouat A1 - Dartois,V. A1 - Kaplan,G. A1 - Bifani,P. KW - a KW - analysi KW - analysis KW - Animal KW - Animals KW - article KW - at KW - BALANCE KW - BCG KW - Biology KW - Cell KW - cells KW - challenge KW - Control KW - Correlation KW - data KW - disease KW - Diseases KW - electronic KW - EVALUATION KW - expression KW - gene KW - Genes KW - Granuloma KW - growth KW - i KW - im KW - immune response KW - immunohistochemistry KW - INFECTION KW - Inflammation KW - Institute KW - IS KW - journal KW - Lung KW - MODEL KW - Mycobacterium KW - Mycobacterium bovis KW - Mycobacterium tuberculosis KW - observed KW - PCR KW - present KW - protection KW - Pulmonary KW - rats KW - real time PCR KW - regulation KW - Research KW - Research Support KW - response KW - Responses KW - SB - IM KW - specific KW - Still KW - study KW - time KW - Tuberculosis KW - Type KW - Vaccination KW - Wistar rat AB - Our understanding of the correlation of Mycobacterium bovis Bacille Calmette-Guerin (BCG)-mediated immune responses and protection against Mycobacterium tuberculosis (Mtb) infection is still limited. We have recently characterized a Wistar rat model of experimental tuberculosis (TB). In the present study, we evaluated the efficacy of BCG vaccination in this model. Upon Mtb challenge, BCG vaccinated rats controlled growth of the bacilli earlier than unvaccinated rats. Histopathology analysis of infected lungs demonstrated a reduced number of granulomatous lesions and lower parenchymal inflammation in vaccinated animals. Vaccine-mediated protection correlated with the rapid accumulation of antigen specific CD4(+) and CD8(+) T cells in the infected lungs. Immunohistochemistry further revealed higher number of CD8(+) cells in the pulmonary granulomas of vaccinated animals. Evaluation of pulmonary immune responses in vaccinated and Mtb infected rats by real time PCR at day 15 post-challenge showed reduced expression of genes responsible for negative regulation of Th1 immune responses. Thus, early protection observed in BCG vaccinated rats correlated with a similarly timed shift of immunity towards the Th1 type response. Our data support the importance of (i) the Th1-Th2 balance in the control of mycobacterial infection and (ii) the value of the Wistar rats in understanding the biology of TB VL - 6 CP - 12 U1 - 38349 M3 - http://dx.doi.org/10.1371/journal.pone.0028082 ER - TY - JOUR T1 - Experimental tuberculosis in the Wistar rat: a model for protective immunity and control of infection36574 JF - PLoS.One. Y1 - 2011 A1 - Singhal,A. A1 - M. El Aliouat A1 - Herve,M. A1 - Vanessa Mathys A1 - Kiass,M. A1 - Creusy,C. A1 - Delaire,B. A1 - Tsenova,L. A1 - Fleurisse,L. A1 - Bertout,J. A1 - L. Camacho A1 - Foo,D. A1 - Tay,H.C. A1 - Siew,J.Y. A1 - Boukhouchi,W. A1 - Marta Romano A1 - Mathema,B. A1 - Dartois,V. A1 - Kaplan,G. A1 - Bifani,P. KW - a KW - Analyses KW - analysi KW - analysis KW - Animal KW - Animals KW - article KW - Cell KW - cells KW - Control KW - Cost KW - disease KW - Disease Models,Animal KW - Diseases KW - DRUG KW - electronic KW - Granuloma KW - growth KW - Host-Pathogen Interactions KW - Human KW - Humans KW - im KW - immunology KW - INFECTION KW - Institute KW - interaction KW - interactions KW - IS KW - journal KW - Kinetics KW - Lung KW - lymphocyte KW - Lymphocytes KW - Macrophage KW - Macrophages KW - MODEL KW - models KW - Mycobacterium KW - Mycobacterium tuberculosis KW - need KW - observed KW - ON KW - pathology KW - Pharmacokinetic KW - prevention & control KW - rats KW - Rats,Wistar KW - Research KW - Research Support KW - result KW - SB - IM KW - Size KW - Still KW - strain KW - study KW - Toxicology KW - Tuberculosis KW - Type KW - use KW - virulence KW - Wistar rat AB - BACKGROUND: Despite the availability of many animal models for tuberculosis (TB) research, there still exists a need for better understanding of the quiescent stage of disease observed in many humans. Here, we explored the use of the Wistar rat model for the study of protective immunity and control of Mycobacterium tuberculosis (Mtb) infection. METHODOLOGY/PRINCIPAL FINDINGS: The kinetics of bacillary growth, evaluated by the colony stimulating assay (CFU) and the extent of lung pathology in Mtb infected Wistar rats were dependent on the virulence of the strains and the size of the infecting inoculums. Bacillary growth control was associated with induction of T helper type 1 (Th1) activation, the magnitude of which was also Mtb strain and dose dependent. Histopathology analysis of the infected lungs demonstrated the formation of well organized granulomas comprising epithelioid cells, multinucleated giant cells and foamy macrophages surrounded by large numbers of lymphocytes. The late stage subclinical form of disease was reactivated by immunosuppression leading to increased lung CFU. CONCLUSION: The Wistar rat is a valuable model for better understanding host-pathogen interactions that result in control of Mtb infection and potentially establishment of latent TB. These properties together with the ease of manipulation, relatively low cost and well established use of rats in toxicology and pharmacokinetic analyses make the rat a good animal model for TB drug discovery VL - 6 CP - 4 U1 - 38350 M3 - http://dx.doi.org/10.1371/journal.pone.0018632 ER - TY - JOUR T1 - Extremely high prevalence of multidrug resistant tuberculosis in Murmansk, Russia: a population-based study36555 JF - Eur.J.Clin.Microbiol.Infect.Dis. Y1 - 2011 A1 - Makinen,J. A1 - Marjamaki,M. A1 - Haanpera-Heikkinen,M. A1 - Marttila,H. A1 - Endourova,L.B. A1 - Presnova,S.E. A1 - Vanessa Mathys A1 - Bifani,P. A1 - Ruohonen,R. A1 - Viljanen,M.K. A1 - Soini,H. KW - 0 KW - 2004 KW - a KW - Agent KW - Agents KW - ALL KW - Antimicrobial KW - Antimicrobial resistance KW - Antitubercular Agents KW - article KW - AS KW - at KW - Case KW - classification KW - DRUG KW - drug effects KW - Drug Resistance KW - electronic KW - epidemiology KW - Finland KW - gene KW - Genetic KW - Genotype KW - health KW - High prevalence KW - Humans KW - im KW - Institute KW - IS KW - isolation & purification KW - journal KW - Laboratories KW - Microbial Sensitivity Tests KW - Molecular KW - Molecular Epidemiology KW - Molecular Typing KW - Mutation KW - Mycobacterium tuberculosis KW - n KW - national KW - pharmacology KW - Polymorphism,Genetic KW - POPULATION KW - population based KW - population-based KW - Population-based studies KW - population-based study KW - prevalence KW - region KW - resistance KW - Rifampin KW - Russia KW - SB - IM KW - strain KW - study KW - Surveillance KW - Target KW - Targets KW - Tuberculosis KW - Tuberculosis,Multidrug-Resistant KW - welfare AB - Drug resistance and molecular epidemiology of tuberculosis (TB) in the Murmansk region was investigated in a 2-year, population-based surveillance of the civilian population. During 2003 and 2004, isolates from all culture-positive cases were collected (n = 1,226). Prevalence of multi-drug resistance (MDR) was extremely high, as 114 out of 439 new cases (26.0%), and 574 out of 787 previously treated cases (72.9%) were resistant to at least isoniazid (INH) and rifampin (RIF). Spoligotyping of the primary MDR-TB isolates revealed that most isolates grouped to the Beijing SIT1 genotype (n = 91, 79.8%). Isolates of this genotype were further analyzed by IS6110 RFLP. Sequencing of gene targets associated with INH and RIF resistance further showed that the MDR-TB strains are highly homogeneous as 78% of the MDR, SIT1 strains had the same resistance-conferring mutations. The genetic homogeneity of the MDR-TB strains indicates that they are actively transmitted in Murmansk VL - 30 CP - 9 U1 - 38273 M3 - http://dx.doi.org/10.1007/s10096-011-1200-7 ER - TY - Generic T1 - Le rat Wistar: un modèle pour la tuberculose latente? Y1 - 2011 A1 - Singhal,A. A1 - M. el Aliouat A1 - Herve,M. A1 - Vanessa Mathys A1 - Kiass,M. A1 - Creusy,C. A1 - Delaire,B. A1 - Tsenova,L. A1 - Fleurisse,L. A1 - Boukhouchi,W. A1 - Marta Romano A1 - Dartois,V. A1 - Kaplan,G. A1 - Bifani,P. ED - Mycoclub KW - latence KW - LE KW - tuberculose KW - Wistar rat JF - Mycoclub2 CP - Mycoclub U1 - 38348 U2 - 26/09/2011 ER - TY - Generic T1 - M. tuberculosis : traitement et résistance aux antibiotiques Y1 - 2011 A1 - Vanessa Mathys A1 - M. Fauville KW - Antibiotique KW - de KW - ET KW - M KW - resistance KW - Surveillance KW - traitement KW - Tuberculosis JF - Réunion des laboratoires du réseau de surveillance de la (multi)résistance PB - NA CY - NA CP - FARES-VRGT U1 - 38289 U2 - 15/02/2011 ER - TY - Generic T1 - Molecular epidemiology of tuberculosis in Brussels - 1 year results (2010) Y1 - 2011 A1 - Vanessa Mathys A1 - Vanfleteren,B. A1 - M. Fauville KW - 2010 KW - Brussels KW - epidemiology KW - meeting KW - Molecular KW - Molecular Epidemiology KW - result KW - results KW - Tuberculosis JF - TB-PANNET Mid-Term Meeting PB - NA CY - NA CP - TB-PANNET U1 - 38290 U2 - 30/05/2011 ER - TY - JOUR T1 - Mycobacterium tuberculosis infection induces hypoxic lung lesions in the rat36548 JF - Tuberculosis.(Edinb.) Y1 - 2011 A1 - Heng,Y. A1 - Seah,P.G. A1 - Siew,J.Y. A1 - Tay,H.C. A1 - Singhal,A. A1 - Vanessa Mathys A1 - Kiass,M. A1 - Bifani,P. A1 - Dartois,V. A1 - Herve,M. KW - 0 KW - Activity KW - Animals KW - Anoxia KW - article KW - cause KW - disease KW - Disease Models,Animal KW - Diseases KW - DRUG KW - Drug Resistance KW - electronic KW - Evaluating KW - Female KW - Granuloma KW - im KW - immunohistochemistry KW - in vivo KW - INFECTION KW - Institute KW - IS KW - journal KW - Lung KW - microbiology KW - MODEL KW - Mycobacterium KW - Mycobacterium tuberculosis KW - Nitroimidazoles KW - Oxygen KW - pathogenicity KW - pathology KW - pharmacology KW - rats KW - region KW - Research KW - Research Support KW - resistance KW - result KW - results KW - SB - IM KW - State KW - Tension KW - Tuberculosis KW - Tuberculosis,Pulmonary KW - use AB - Hypoxia is believed to influence the metabolic state of Mycobacterium tuberculosis and cause phenotypic drug resistance. Using pimonidazole adduct staining, we show that lung lesions of infected rats contain regions of low oxygen tension. Our results support the use of the rat model for evaluating anaerobic drug activity in vivo VL - 91 CP - 4 U1 - 38225 M3 - http://dx.doi.org/10.1016/j.tube.2011.05.003 ER - TY - Generic T1 - TB BRU-NET…une partie de TB-PAN-NET Y1 - 2011 A1 - Vanessa Mathys A1 - M. Fauville KW - de KW - Surveillance KW - TB-BRU-NET JF - Réunion des laboratoires du réseau de surveillance de la (multi)résistance PB - NA CY - NA CP - FARES-VRGT U1 - 38288 U2 - 15/02/2011 ER - TY - JOUR T1 - Effect of PstS sub-units or PknD deficiency on the survival of Mycobacterium tuberculosis. JF - Tuberculosis (Edinb) Y1 - 2010 A1 - Vanzembergh, Frederic A1 - Peirs, Priska A1 - Lefèvre, Philippe A1 - Celio, Nathalie A1 - Vanessa Mathys A1 - Content, Jean A1 - Kalai, Michael KW - Animals KW - ATP-Binding Cassette Transporters KW - Bacterial Proteins KW - Blotting, Western KW - Cell Proliferation KW - Gene Expression Regulation, Bacterial KW - mice KW - Molecular Sequence Data KW - Mycobacterium tuberculosis KW - Protein Kinases KW - Reverse Transcriptase Polymerase Chain Reaction KW - Tuberculosis AB -

The membrane-associated phosphate-specific transporter (Pst) complex is composed of four different proteins: PstS, PstC, PstA and PstB. The PstS component detects and binds Pi with high affinity; the PstA and PstC form transmembrane pores for Pi entry, while PstB provides energy through ATP hydrolysis. In the Mycobacterium tuberculosis genome, four different gene clusters encode three PstS, and two of each of the other sub-units. We used RT-PCR to show that these clusters represent at least three distinct operons. The pstS3-containing operon was the only one induced by lack of environmental Pi. To study the physiologic role of the different PstS sub-units and that of another potential Pi receptor, PknD, we constructed and complemented their knockout (KO) mutants. In Sauton medium, the PstS1-3 KO grew faster than the Wt or the PknD KO. Following 24 h of complete starvation, the PstS3 or PknD deficient strains died if exposed to Pi poor conditions while the PstS1 and PstS2 KO survived and still grew faster than the Wt strain. These results suggest that PstS1-3 may play a role in the regulation of M. tuberculosis growth or metabolism while PstS3 and PknD contribute to the survival of the bacteria in phosphate poor conditions.

VL - 90 CP - 6 U1 - http://www.ncbi.nlm.nih.gov/pubmed/20933472?dopt=Abstract M3 - 10.1016/j.tube.2010.09.004 ER - TY - Generic T1 - Etude des mécanismes de résistance de M. tuberculosis aux agents antituberculeux Y1 - 2010 A1 - Vanessa Mathys KW - Agent KW - Agents KW - Antibiotic KW - antibiotics KW - de KW - journal KW - M KW - resistance KW - Tuberculosis JF - Journal Club Antibiotics PB - NA CY - NA CP - WIV-ISP U1 - 36880 U2 - 23/09/2012 ER - TY - Generic T1 - Identification d'espèce des mycobactéries non tuberculeuses: quel fragment génique séquencer et dans quelle(s) base(s) de séquences analyser le résultat? Y1 - 2010 A1 - Vanessa Mathys A1 - Vanfleteren,B. A1 - Stoffels,K. A1 - M. Fauville ED - Mycoclub KW - de KW - espèces KW - ET KW - gene KW - identification KW - LE KW - Mycobactéries KW - tuberculose JF - Mycoclub CP - Mycoclub U1 - 38286 U2 - 28/10/2010 ER - TY - Generic T1 - Identification of non-tuberculous mycobacteria by sequencing a fragment of the 16S rRNA gene : which internet database use to analyze the sequence? Y1 - 2010 A1 - Vanessa Mathys A1 - M. Fauville KW - 16S rRNA KW - a KW - Database KW - gene KW - identification KW - Internet KW - meeting KW - Mycobacterium KW - Nontuberculous Mycobacteria KW - use JF - ESM annual Meeting PB - NA CY - NA CP - European Society of Mycobacteriology (ESM) U1 - 36881 U2 - 30/06/2010 ER - TY - Generic T1 - Molecular Genetics of para -Aminosalicylic Acid Resistance in Clinical Isolates and Spontaneous Mutants of Mycobacterium tuberculosis Y1 - 2010 A1 - Vanessa Mathys KW - acid KW - Clinical KW - Genetic KW - genetics KW - meeting KW - Molecular KW - Mycobacterium KW - Mycobacterium tuberculosis KW - resistance KW - Tuberculosis JF - TB-PAN-NET 1st Anual Meeting PB - NA CY - NA CP - TB-PANNET U1 - 38285 U2 - 26/05/2010 ER - TY - Generic T1 - Mycobacterium tuberculosis : treatment and resistance to antibiotics Y1 - 2010 A1 - Vanessa Mathys KW - Antibiotic KW - antibiotics KW - Mycobacterium KW - Mycobacterium tuberculosis KW - resistance KW - treatment KW - Tuberculosis JF - Tuesday Seminar PB - NA CY - NA CP - WIV-ISP U1 - 38287 U2 - 19/10/2010 ER - TY - RPRT T1 - Contribution à la compréhension des mécanismes moléculaires de résistance de Mycobacterium tuberculosis aux agents anti-tuberculeux Y1 - 2009 A1 - Vanessa Mathys KW - Agent KW - Agent anti-tuberculeux KW - Agents KW - contribution KW - de KW - Mycobacterium KW - Mycobacterium tuberculosis KW - resistance KW - Tuberculosis PB - Univeristé Libre de Bruxelles (ULB) CY - Brussels, Belgium U1 - 38283 ER - TY - JOUR T1 - Molecular genetics of para-aminosalicylic acid resistance in clinical isolates and spontaneous mutants of Mycobacterium tuberculosis. JF - Antimicrob Agents Chemother Y1 - 2009 A1 - Vanessa Mathys A1 - Wintjens, René A1 - Lefèvre, Philippe A1 - Bertout, Julie A1 - Singhal, Amit A1 - Kiass, Mehdi A1 - Kurepina, Natalia A1 - Wang, Xiao-Ming A1 - Mathema, Barun A1 - Baulard, Alain A1 - Kreiswirth, Barry N A1 - Bifani, Pablo KW - Aminosalicylic Acid KW - Antitubercular Agents KW - Bacterial Proteins KW - Drug Resistance, Bacterial KW - Folic Acid KW - Humans KW - Microbial Sensitivity Tests KW - Mutation KW - Mycobacterium tuberculosis KW - Structure-Activity Relationship KW - Thymidylate Synthase KW - Thymine AB -

The emergence of Mycobacterium tuberculosis resistant to first-line antibiotics has renewed interest in second-line antitubercular agents. Here, we aimed to extend our understanding of the mechanisms underlying para-aminosalicylic acid (PAS) resistance by analysis of six genes of the folate metabolic pathway and biosynthesis of thymine nucleotides (thyA, dfrA, folC, folP1, folP2, and thyX) and three N-acetyltransferase genes [nhoA, aac(1), and aac(2)] among PAS-resistant clinical isolates and spontaneous mutants. Mutations in thyA were identified in only 37% of the clinical isolates and spontaneous mutants. Overall, 24 distinct mutations were identified in the thyA gene and 3 in the dfrA coding region. Based on structural bioinformatics techniques, the altered ThyA proteins were predicted to generate an unfolded or dysfunctional polypeptide. The MIC was determined by Bactec/Alert and dilution assay. Sixty-three percent of the PAS-resistant isolates had no mutations in the nine genes considered in this study, revealing that PAS resistance in M. tuberculosis involves mechanisms or targets other than those pertaining to the biosynthesis of thymine nucleotides. The alternative mechanism(s) or pathway(s) associated with PAS resistance appears to be PAS concentration dependent, in marked contrast to thyA-mutated PAS-resistant isolates.

VL - 53 CP - 5 U1 - http://www.ncbi.nlm.nih.gov/pubmed/19237648?dopt=Abstract M3 - 10.1128/AAC.01197-08 ER - TY - JOUR T1 - Synthetic EthR inhibitors boost antituberculous activity of ethionamide36849 JF - Nat.Med. Y1 - 2009 A1 - Willand,N. A1 - Dirie,B. A1 - Carette,X. A1 - Bifani,P. A1 - Singhal,A. A1 - Desroses,M. A1 - Leroux,F. A1 - Willery,E. A1 - Vanessa Mathys A1 - Deprez-Poulain,R. A1 - Delcroix,G. A1 - Frenois,F. A1 - Aumercier,M. A1 - Locht,C. A1 - Villeret,V. A1 - Deprez,B. A1 - Baulard,A.R. KW - 0 KW - a KW - Activity KW - Agent KW - Agents KW - Animals KW - antagonists & inhibitors KW - Antitubercular Agents KW - article KW - AS KW - Binding Sites KW - chemistry KW - concept KW - conventional KW - culture KW - de KW - DNA-Binding Proteins KW - DRUG KW - Drug Resistance KW - Drug Synergism KW - drug therapy KW - drugs KW - effect KW - effects KW - electronic KW - ET KW - Ethionamide KW - France KW - Hydrogen Bonding KW - identify KW - im KW - improve KW - index KW - interaction KW - IS KW - journal KW - Ligands KW - metabolism KW - mice KW - Models,Molecular KW - Mycobacterium KW - Mycobacterium tuberculosis KW - national KW - Oxadiazoles KW - production KW - protein KW - Protein Conformation KW - Proteins KW - Repressor Proteins KW - Research KW - Research Support KW - resistance KW - risk KW - SB - IM KW - Side effects KW - Side-effects KW - structure KW - Synthetic KW - therapeutic use KW - Therapy KW - Thiophenes KW - treatment KW - Tuberculosis KW - use AB - The side effects associated with tuberculosis therapy bring with them the risk of noncompliance and subsequent drug resistance. Increasing the therapeutic index of antituberculosis drugs should thus improve treatment effectiveness. Several antituberculosis compounds require in situ metabolic activation to become inhibitory. Various thiocarbamide-containing drugs, including ethionamide, are activated by the mycobacterial monooxygenase EthA, the production of which is controlled by the transcriptional repressor EthR. Here we identify drug-like inhibitors of EthR that boost the bioactivation of ethionamide. Compounds designed and screened for their capacity to inhibit EthR-DNA interaction were co-crystallized with EthR. We exploited the three-dimensional structures of the complexes for the synthesis of improved analogs that boosted the ethionamide potency in culture more than tenfold. In Mycobacterium tuberculosis-infected mice, one of these analogs, BDM31343, enabled a substantially reduced dose of ethionamide to lessen the mycobacterial load as efficiently as the conventional higher-dose treatment. This provides proof of concept that inhibiting EthR improves the therapeutic index of thiocarbamide derivatives, which should prompt reconsideration of their use as first-line drugs VL - 15 CP - 5 U1 - 38473 M3 - http://dx.doi.org/10.1038/nm.1950 ER - TY - Generic T1 - The anti-cancer drug 5-fluorouracil is highly active against Mycobacterium tuberculosis in vitro but not in vivo Y1 - 2008 A1 - Vanessa Mathys A1 - Singhal,A. A1 - Kiass,M. A1 - P. Lefevre A1 - Kurepina,N. A1 - Wang,X.M. A1 - Mathema,B. A1 - Baulard,A. A1 - Bifani,P. KW - Control KW - DRUG KW - in vivo KW - INFECTION KW - infections KW - IS KW - Mycobacterium KW - Mycobacterium tuberculosis KW - Tuberculosis JF - Pathogenesis and Control of Emerging Infections and Drug-Resistant Organisms PB - NA CY - NA CP - Keystone Symposia U1 - 38282 U2 - 24/10/2008 ER - TY - Generic T1 - Mutations associated with para-aminosalicylic acid (PAS) resistance in clinical isolates and spontaneous mutants of Mycobacterium tuberculosis Y1 - 2007 A1 - Vanessa Mathys A1 - Quatannens,J. A1 - Wintjens,R. A1 - Kurepina,N. A1 - Singhal,A. A1 - Kiass,M. A1 - Mathema,B. A1 - Baulard,A. A1 - B.N. Kreiswirth A1 - Bifani,P. KW - acid KW - Clinical KW - de KW - Mutation KW - Mycobacterium KW - Mycobacterium tuberculosis KW - Pa KW - PAS KW - resistance KW - Tuberculosis JF - 7ème journée des Doctorants PB - NA CY - NA CP - Université Libre de Bruxelles (ULB) U1 - 36787 U2 - 18/12/2007 ER - TY - Generic T1 - Molecular characterization of genes associated with ethionamide resistance in Mycobacterium tuberculosis clinical isolates and spontaneous mutants Y1 - 2006 A1 - Vanessa Mathys A1 - Baulard,A. A1 - Kurepina,N. A1 - M. Fauville A1 - B.N. Kreiswirth A1 - Bifani,P. KW - Clinical KW - Ethionamide KW - gene KW - general KW - Genes KW - meeting KW - Molecular KW - Mycobacterium KW - Mycobacterium tuberculosis KW - resistance KW - Tuberculosis JF - 106th General Meeting PB - NA CY - NA CP - American Society for Microbiology (ASM) U1 - 38281 U2 - 24/05/2006 ER - TY - Generic T1 - Mutations associated with PAS resistance in clinical isolates and spontaneous mutants of Mycobaterium tuberculosis Y1 - 2006 A1 - Vanessa Mathys A1 - Quatannens,J. A1 - Kurepina,N. A1 - Wintjens,R. A1 - Mathema,B. A1 - Baulard,A. A1 - B.N. Kreiswirth A1 - Bifani,P. ED - Belgian Society for Microbiology KW - Clinical KW - meeting KW - Mutation KW - Pa KW - PAS KW - resistance KW - Tuberculosis JF - BSM meeting CP - Belgian Society of Microbiology (BSM) U1 - 38280 U2 - 24/11/2006 ER - TY - Generic T1 - Rapport annuel - CNR Mycobacterium Y1 - 0 A1 - Vanessa Mathys ER -