High-throughput screening to discover inhibitors of the CarD·RNA polymerase protein–protein interaction in Mycobacterium tuberculosis

AbstractMultidrug-resistant Mycobacterium tuberculosis (MDR-TB) accounts for 3.7% of new cases of TB annually worldwide and is a major threat to global public health. Due to the prevalence of the MDR-TB and extensively drug resistant tuberculosis (XDR-TB) cases, there is an urgent need for new drugs with novel mechanisms of action. CarD, a global transcription regulator in MTB, binds RNAP and activates transcription by stabilizing the transcription initiation open-promoter complex (RPo). CarD is required for MTB viability and it has highly conserved homologues in many eubacteria. A fluorescence polarization (FP) assay which monitors the association of MTB RNAP, native rRNA promoter DNA and CarD has been developed. Overall, our objective is to identify and characterize small molecule inhibitors which block the CarD/RNAP interaction and to understand the mechanisms by which CarD interacts with the molecules. We expect that the development of a new and improved anti-TB compound with a novel mechanism of action will relieve the burden of resistance. This CarD FP assay is amenable to HTS and is an enabling tool for future novel therapeutic discovery.

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Multidisciplinary advisory teams to manage multidrug-resistant tuberculosis: the example of the French Consilium

The International Journal of Tuberculosis and Lung Disease ◽

10.5588/ijtld.18.0779 ◽

2019 ◽

Vol 23 (10) ◽

pp. 1050-1054

Author(s):

L. Guglielmetti ◽

J. Jaffré ◽

C. Bernard ◽

F. Brossier ◽

N. El Helali ◽

...

Keyword(s):

Multidrug Resistant ◽

New Drugs ◽

World Health ◽

Advisory Committees ◽

Treatment Regimens ◽

Tb Treatment ◽

Resistant Tuberculosis ◽

Multidrug Resistant Tuberculosis ◽

Mdr Tb ◽

Health Organization

SETTING: The World Health Organization (WHO) recommends that multidrug-resistant tuberculosis (MDR-TB) treatment should be managed in collaboration with multidisciplinary advisory committees (consilia). A formal national Consilium has been established in France since 2005 to provide a centralised advisory service for clinicians managing MDR-TB and extensively drug-resistant (XDR-TB) cases.OBJECTIVE: Review the activity of the French TB Consilium since its establishment.DESIGN: Retrospective description and analysis of the activity of the French TB Consilium.RESULTS: Between 2005 and 2016, 786 TB cases or contacts of TB cases were presented at the French TB Consilium, including respectively 42% and 79% of all the MDR-TB and XDR-TB cases notified in France during this period. Treatment regimens including bedaquiline and/or delamanid were recommended for 42% of the cases presented at the French TB Consilium since 2009. Patients were more likely to be presented at the French TB Consilium if they were born in the WHO Europe Region, had XDR-TB, were diagnosed in the Paris region, or had resistance to additional drugs than those defining XDR-TB.CONCLUSION: The French TB Consilium helped supervise appropriate management of MDR/XDR-TB cases and facilitated implementation of new drugs for MDR/XDR-TB treatment.

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Molecular Characterization of Multidrug-Resistant Mycobacterium tuberculosis Isolated in Nepal

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.06418-11 ◽

2012 ◽

Vol 56 (6) ◽

pp. 2831-2836 ◽

Cited By ~ 38

Author(s):

Ajay Poudel ◽

Chie Nakajima ◽

Yukari Fukushima ◽

Haruka Suzuki ◽

Basu Dev Pandey ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Promoter Region ◽

Multidrug Resistant ◽

Content Type ◽

Molecular Tests ◽

Resistant Tuberculosis ◽

Multidrug Resistant Tuberculosis ◽

Mdr Tb ◽

Substitution Mutations ◽

Drug Resistant Strains

ABSTRACTDespite the fact that Nepal is one of the first countries globally to introduce multidrug-resistant tuberculosis (MDR-TB) case management, the number of MDR-TB cases is continuing to rise in Nepal. Rapid molecular tests applicable in this setting to identify resistant organisms would be an effective tool in reversing this trend. To develop such tools, information about the frequency and distribution of mutations that are associated with phenotypic drug resistance inMycobacterium tuberculosisis required. In the present study, we investigated the prevalence of mutations inrpoBandkatGgenes and theinhApromoter region in 158M. tuberculosisisolates (109 phenotypically MDR and 49 non-MDR isolates collected in Nepal) by DNA sequencing. Mutations affecting the 81-bp rifampin (RIF) resistance-determining region (RRDR) ofrpoBwere identified in 106 of 109 (97.3%) RIF-resistant isolates. Codons 531, 526, and 516 were the most commonly affected, at percentages of 58.7, 15.6, and 15.6%, respectively. Of 113 isoniazid (INH)-resistant isolates, 99 (87.6%) had mutations in thekatGgene, with Ser315Thr being the most prevalent (81.4%) substitution. Mutations in theinhApromoter region were detected in 14 (12.4%) INH-resistant isolates. The results from this study provide an overview of the current situation of RIF and INH resistance inM. tuberculosisin Nepal and can serve as a basis for developing or improving rapid molecular tests to monitor drug-resistant strains in this country.

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Genotypic diversity of multi- and pre-extremely drug-resistant Mycobacterium tuberculosis isolates from Morocco

PLoS ONE ◽

10.1371/journal.pone.0253826 ◽

2021 ◽

Vol 16 (7) ◽

pp. e0253826

Author(s):

Amal Oudghiri ◽

Ghizlane Momen ◽

Achraf Aainouss ◽

Amin Laglaoui ◽

My Driss El Messaoudi ◽

...

Keyword(s):

Genetic Diversity ◽

Mycobacterium Tuberculosis ◽

Genotypic Diversity ◽

Multidrug Resistant ◽

Drug Resistant ◽

Control Programs ◽

Resistant Tuberculosis ◽

Mdr Tb ◽

Database Clustering ◽

High Level

In Morocco, the prevalence of multidrug resistant tuberculosis (MDR-TB) continues to increase especially within previously treated cases; these MDR cases may evolve to extensively drug resistant tuberculosis (XDR-TB) raising major concern to TB control programs. From an epidemiological window, scarce informations are available about the genetic diversity of Mycobacterium tuberculosis (MTB) strains fueling these forms of resistance. The aim of this study was to assess to genetic diversity of MDR-MTB strains. Hence, this prospective study was conducted on patients diagnosed with MDR-TB at Pasteur Institute of Casablanca from 2010 to 2013. A total of 70 MDR-MTB isolates were genotyped by spoligotyping and 15-loci MIRU-VNTR methods. Spoligotyping generated four orphan patterns, five unique profiles whereas 61 strains were grouped in nine clusters (2 to 25 strains per cluster), the clustering rates being 87.1%. Subtyping by 15 loci MIRU-VNTR splitted all clusters already established by spoligotyping and generated 70 unique profiles not recognized in SITVIT2 database; clustering rate was equal to zero. HGDI analysis of 15 loci MIRU demonstrated that eight out of 15 loci were highly discriminant. Of note, all pre-XDR strains belongs to many clades, meaning that there no association between gyrA mutants and particular clade. Overall, the data generated by this study (i) describe the population structure of MDR MTBC in Morocco which is highly homogenous, (ii) confirm that TB in Morocco is almost exclusively transmitted by modern and evolutionary lineages with high level of biodiversity seen by MIRU, and (iii) validate the use of optimized 15-loci MIRU-VNTR format for future investigations in Morocco.

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Epidemiology of Rifampicin Resistant Tuberculosis and Common Mutations in rpoB Gene of Mycobacterium tuberculosis: A Retrospective Study from Six Districts of Punjab (India) Using Xpert MTB/RIF Assay

Journal of Laboratory Physicians ◽

10.4103/0974-2727.180789 ◽

2016 ◽

Vol 8 (02) ◽

pp. 096-100 ◽

Cited By ~ 4

Author(s):

Ramandeep Kaur ◽

Neerja Jindal ◽

Shilpa Arora ◽

Shajla Kataria

Keyword(s):

Mycobacterium Tuberculosis ◽

Surrogate Marker ◽

Rpob Gene ◽

Multidrug Resistant ◽

Resistance Pattern ◽

Resistance Mutations ◽

Resistant Tuberculosis ◽

Mdr Tb ◽

Previously Treated Patients ◽

User Friendly

ABSTRACT Background: Xpert MTB/RIF assay has revolutionized the diagnosis of tuberculosis (TB) by simultaneously detecting the bacteria and resistance to rifampicin (RIF), a surrogate marker for multidrug-resistant TB (MDR-TB) in <2 h. The RIF resistance pattern in Malwa region of Punjab, India, is not documented. Here, we report the epidemiology of RIF-resistant TB and mutations in rpoB gene of Mycobacterium tuberculosis (MTB). Materials and Methods: A total of 1612 specimens received between October 2013 and February 2015 were tested by Xpert MTB/RIF assay following manufacturer’s instructions. The results thus obtained were analyzed using SPSS version 20.0.0 (SPSS Inc., Chicago, IL, USA) statistical software. Result: RIF resistance was statistically higher in previously treated patients in comparison to the new patients (P = 0.006) and in patients with acid fast-Bacilli (AFB) positive smears to AFB-negative smears (P = 0.048). RIF resistance mutations in 130 specimens revealed frequency of E 73/130 (56%), B 28/130 (21.5%), D 18/130 (13.8%), A 11/130 (8.4%), and C 1/130 (0.7%) while in one specimen, mutation combination, i.e., mutations associated with more than one probe (A and B both) was present. Conclusion: Xpert MTB/RIF assay is a user-friendly screening tool for detection of MTB and RIF resistance from suspected TB/MDR cases in a shorter period of time. It could also serve as a useful technique to have simultaneous preliminary information regarding the mutation pattern of RIF resistance in MTB isolates.

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Draft Genome Sequences of Two Drug-Resistant Mycobacterium tuberculosis Isolates from Myanmar

Genome Announcements ◽

10.1128/genomea.00850-16 ◽

2016 ◽

Vol 4 (5) ◽

Cited By ~ 1

Author(s):

Htin Lin Aung ◽

Thanda Tun ◽

Elizabeth Permina ◽

Wint Wint Nyunt ◽

Si Thu Aung ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Draft Genome ◽

Multidrug Resistant ◽

Drug Resistant ◽

Genome Sequences ◽

Health Concerns ◽

Resistant Tuberculosis ◽

Multidrug Resistant Tuberculosis ◽

Extensively Drug Resistant ◽

Mdr Tb

Multidrug-resistant tuberculosis (MDR-TB) and lately, extensively drug-resistant TB (XDR-TB) are increasing global health concerns. Here, we present the genome sequences of two MDR-TB isolates from Myanmar, one of 27 countries with a high MDR-TB burden, and describe a number of mutations consistent with these being XDR-TB isolates.

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Regimens to treat multidrug-resistant tuberculosis: past, present and future perspectives

European Respiratory Review ◽

10.1183/16000617.0035-2019 ◽

2019 ◽

Vol 28 (152) ◽

pp. 190035 ◽

Cited By ~ 23

Author(s):

Emanuele Pontali ◽

Mario C. Raviglione ◽

Giovanni Battista Migliori

Keyword(s):

Multidrug Resistant ◽

New Drugs ◽

World Health ◽

Drug Resistant ◽

Optimal Outcomes ◽

Resistant Tuberculosis ◽

Prolonged Duration ◽

Mdr Tb ◽

Revised Definitions ◽

Health Organization

Over the past few decades, treatment of multidrug-resistant (MDR)/extensively drug-resistant (XDR) tuberculosis (TB) has been challenging because of its prolonged duration (up to 20–24 months), toxicity, costs and sub-optimal outcomes.After over 40 years of neglect, two new drugs (bedaquiline and delamanid) have been made available to manage difficult-to-treat MDR-/XDR-TB cases. World Health Organization (WHO) guidelines published in March 2019 endorsed the possibility of treating MDR-TB patients with a full oral regimen, following previous guidelines published in 2016 which launched a shorter regimen lasting 9–10 months.The objectives of this article are to review the main achievements in MDR-TB treatment through the description of the existing WHO strategies, to discuss the main ongoing trials and to shed light on potential future scenarios and revised definitions necessary to manage drug-resistant TB.

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Treatment of Multidrug-Resistant and Extensively Drug-Resistant Tuberculosis in Children: The Role of Bedaquiline and Delamanid

Microorganisms ◽

10.3390/microorganisms9051074 ◽

2021 ◽

Vol 9 (5) ◽

pp. 1074

Author(s):

Francesco Pecora ◽

Giulia Dal Canto ◽

Piero Veronese ◽

Susanna Esposito

Keyword(s):

Adverse Effects ◽

Pediatric Population ◽

Multidrug Resistant ◽

New Drugs ◽

Drug Resistant ◽

Standard Regimen ◽

Drug Resistant Tuberculosis ◽

Resistant Tuberculosis ◽

Extensively Drug Resistant ◽

Mdr Tb

Multidrug-resistant (MDR) tuberculosis (TB) has been emerging at an alarming rate over the last few years. It has been estimated that about 3% of all pediatric TB is MDR, meaning about 30,000 cases each year. Although most children with MDR-TB can be successfully treated, up to five years ago effective treatment was associated with a high incidence of severe adverse effects and patients with extensively drug-resistant (XDR) TB had limited treatment options and no standard regimen. The main objective of this manuscript is to discuss our present knowledge of the management of MDR- and XDR-TB in children, focusing on the characteristics and available evidence on the use of two promising new drugs: bedaquiline and delamanid. PubMed was used to search for all of the studies published up to November 2020 using key words such as “bedaquiline” and “delamanid” and “children” and “multidrug-resistant tuberculosis” and “extensively drug-resistant tuberculosis”. The search was limited to articles published in English and providing evidence-based data. Although data on pediatric population are limited and more studies are needed to confirm the efficacy and safety of bedaquiline and delamanid, their use in children with MDR-TB/XDR-TB appears to have good tolerability and efficacy. However, more evidence on these new anti-TB drugs is needed to better guide their use in children in order to design effective shorter regimens and reduce adverse effects, drug interactions, and therapeutic failure.

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Effectiveness and safety of delamanid- or bedaquiline-containing regimens among children and adolescents with multidrug resistant or extensively drug resistant tuberculosis: A nationwide study from Belarus, 2015-19

Monaldi Archives for Chest Disease ◽

10.4081/monaldi.2021.1646 ◽

2021 ◽

Vol 91 (1) ◽

Author(s):

Varvara Solodovnikova ◽

Ajay M.V. Kumar ◽

Hennadz Hurevich ◽

Yuliia Sereda ◽

Vera Auchynka ◽

...

Keyword(s):

Children And Adolescents ◽

Multidrug Resistant ◽

Post Treatment ◽

New Drugs ◽

Drug Resistant ◽

Drug Resistant Tuberculosis ◽

Resistant Tuberculosis ◽

Extensively Drug Resistant ◽

Mdr Tb ◽

Culture Conversion

There is limited evidence describing the safety and effectiveness of bedaquiline and delamanid containing regimens in children and adolescents with Multidrug-Resistant Tuberculosis (MDR-TB) and Extensively Drug-Resistant Tuberculosis (XDR-TB) globally. In this nationwide descriptive cohort study from Belarus, we examined adverse drug events, time to culture conversion, treatment outcomes including post-treatment recurrence among children and adolescents (<18 years of age) treated with bedaquiline and/or delamanid containing regimens from 2015 to 2019. Of the 40 participants included (55% females; age range 10-17 years), 20 (50%) had XDR-TB and 15 (38%) had resistance to either fluoroquinolone or second-line injectable. Half of the patients received delamanid and another half received bedaquiline with one patient receiving both drugs. AEs were reported in all the patients. A total of 224 AEs were reported, most of which (76%) were mild in nature. Only 10 (5%) AEs were graded severe and one AE was graded life-threatening. A total of 7 AEs (3%) were classified as ‘serious’ and only one patient required permanent discontinuation of the suspected drug (linezolid). Most of the AEs (94%) were resolved before the end of treatment. All patients culture-positive at baseline (n=34) became culture-negative within three months of treatment. Median time to culture conversion was 1.1 months (interquartile range: 0.9-1.6). Two patients were still receiving treatment at the time of analysis. The remaining 38 patients successfully completed treatment. Among those eligible and assessed at 6 (n=32) and 12 months (n=27) post-treatment, no recurrences were detected. In conclusion, treatment of children and adolescents with MDR-TB and XDR-TB using bedaquiline and/or delamanid containing regimens was effective and had favourable safety profile. Achieving such excellent outcomes under programmatic settings is encouraging for other national tuberculosis programmes, which are in the process of introducing or scaling-up the use of these new drugs in their countries.

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Drug susceptibility patterns of Mycobacterium tuberculosis from adults with multidrug-resistant tuberculosis and implications for a household contact preventive therapy trial

BMC Infectious Diseases ◽

10.1186/s12879-021-05884-4 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Anne-Marie Demers ◽

◽

Soyeon Kim ◽

Sara McCallum ◽

Kathleen Eisenach ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Sputum Sample ◽

Drug Susceptibility ◽

Multidrug Resistant ◽

Preventive Therapy ◽

Drug Susceptibility Testing ◽

Study Entry ◽

Resistant Tuberculosis ◽

Mdr Tb ◽

Therapy Trial

Abstract Background Drug susceptibility testing (DST) patterns of Mycobacterium tuberculosis (MTB) from patients with rifampicin-resistant tuberculosis (RR-TB) or multidrug-resistant TB (MDR-TB; or resistant to rifampicin and isoniazid (INH)), are important to guide preventive therapy for their household contacts (HHCs). Methods As part of a feasibility study done in preparation for an MDR-TB preventive therapy trial in HHCs, smear, Xpert MTB/RIF, Hain MTBDRplus, culture and DST results of index MDR-TB patients were obtained from routine TB programs. A sputum sample was collected at study entry and evaluated by the same tests. Not all tests were performed on all specimens due to variations in test availability. Results Three hundred eight adults with reported RR/MDR-TB were enrolled from 16 participating sites in 8 countries. Their median age was 36 years, and 36% were HIV-infected. Routine testing on all 308 were confirmed as having RR-TB, but only 75% were documented as having MDR-TB. The majority of those not classified as having MDR-TB were because only rifampicin resistance was tested. At study entry (median 59 days after MDR-TB treatment initiation), 280 participants (91%) were able to produce sputum for the study, of whom 147 (53%) still had detectable MTB. All but 2 of these 147 had rifampicin DST done, with resistance detected in 89%. Almost half (47%) of the 147 specimens had INH DST done, with 83% resistance. Therefore, 20% of the 280 study specimens had MDR-TB confirmed. Overall, DST for second-line drugs were available in only 35% of the 308 routine specimens and 15% of 280 study specimens. Conclusions RR-TB was detected in all routine specimens but only 75% had documented MDR-TB, illustrating the need for expanded DST beyond Xpert MTB/RIF to target preventive therapy for HHC.

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In Vitro Activity of Tetracycline Analogs against Multidrug-resistant and Extensive drug resistance Clinical Isolates of Mycobacterium tuberculosis

10.21203/rs.2.17754/v1 ◽

2019 ◽

Author(s):

Qi Ouyang ◽

Dachuan Lin ◽

Guofang Deng ◽

Zhihua Wen ◽

Houming Liu ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Peak Plasma ◽

Peak Plasma Concentration ◽

Underlying Mechanism ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Ammonium Bromide ◽

Resistant Tuberculosis ◽

Mdr Tb

Abstract Background Multidrug-resistant tuberculosis (MDR-TB) has become a big threaten to global health . The current strategy for treatment of MDR-TB and extensive drug resistant tuberculosis (XDR-TB) is with low efficacy and high side effect. While new drug is fundamental for cure MDR-TB, repurposing the Food and Drug Administration (FDA)-approved drugs represents an alternative soluation with less cost.Methods The activity of 8 tetracycline-class antibiotics against mycobacterium tuberculosis ( M.tb ) were determined by Minimum Inhibitory Concentration (MIC) in vitro. A transposon M.smeg libraries was generated by using the Harm phage and then used to isolate the conditional growth mutants in doxycycline containing plate. 11 mutants were isolated and genomic DNAs were extracted using the cetyltrimethyl ammonium bromide (CTAB) method and analyzed by whole genome sequencing.Results We found that three of eight drugs efficiently inhibited mycobacteria growth under the peak plasma concentration in the human body. Further tests showed these three tetracycline analogs (demeclocycline, doxycycline and methacycline) had antimicrobial activity against seven clinical isolates, including MDR and XDR strains. Among them, Doxycycline had the lowest MICs in all mycobacteria strains tested in this study. By using a transposon library, we identify the insertion of transposon in two genes, porin and MshA, associate with the resistant to doxycycline.Conclusions Our findings show that tetracycline analogs such as doxycycline, has bactericidal activity against not only drug sensitive M.tb , but also clinical MDR and XDR strains, provided proof of concept to repurpose doxycycline to fight MDR-TB and XDR-TB. Further investigations are warranted to clarify the underlying mechanism and optimize the strategy in combination with other anti-TB drugs.

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