scholarly journals Rapid, Semiquantitative Assay To Discriminate among Compounds with Activity against Replicating or Nonreplicating Mycobacterium tuberculosis

2015 ◽  
Vol 59 (10) ◽  
pp. 6521-6538 ◽  
Author(s):  
Ben Gold ◽  
Julia Roberts ◽  
Yan Ling ◽  
Landys Lopez Quezada ◽  
Jou Glasheen ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Bactericidal Activity ◽  
Activated Charcoal ◽  
False Positive ◽  
Second Phase ◽  
Bacteriostatic Activity ◽  
Content Type ◽  
Resazurin Assay ◽  
Resazurin Dye

ABSTRACTThe search for drugs that can kill replicating and nonreplicatingMycobacterium tuberculosisfaces practical bottlenecks. Measurement of CFU and discrimination of bacteriostatic from bactericidal activity are costly in compounds, supplies, labor, and time. Testing compounds againstM. tuberculosisunder conditions that prevent the replication ofM. tuberculosisoften involves a second phase of the test in which conditions are altered to permit the replication of bacteria that survived the first phase. False-positive determinations of activity against nonreplicatingM. tuberculosismay arise from carryover of compounds from the nonreplicating stage of the assay that act in the replicating stage. We mitigate these problems by carrying out a 96-well microplate liquid MIC assay and then transferring an aliquot of each well to a second set of plates in which each well contains agar supplemented with activated charcoal. After 7 to 10 days—about 2 weeks sooner than required to count CFU—fluorometry reveals whetherM. tuberculosisbacilli in each well have replicated extensively enough to reduce a resazurin dye added for the final hour. This charcoal agar resazurin assay (CARA) distinguishes between bacterial biomasses in any two wells that differ by 2 to 3 log10CFU. The CARA thus serves as a pretest and semiquantitative surrogate for longer, more laborious, and expensive CFU-based assays, helps distinguish bactericidal from bacteriostatic activity, and identifies compounds that are active under replicating conditions, nonreplicating conditions, or both. Results for 14 antimycobacterial compounds, including tuberculosis (TB) drugs, revealed that PA-824 (pretomanid) and TMC207 (bedaquiline) are largely bacteriostatic.

scholarly journals Transcriptional Inhibition of the F1F0-Type ATP Synthase Has Bactericidal Consequences on the Viability of Mycobacteria

2020 ◽  
Vol 64 (8) ◽  
Author(s):  
Matthew B. McNeil ◽  
Heath W. K. Ryburn ◽  
Liam K. Harold ◽  
Justin F. Tirados ◽  
Gregory M. Cook
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Atp Synthase ◽  
Bactericidal Activity ◽  
Mycobacterium Smegmatis ◽  
Drug Target ◽  
Potent Inhibitor ◽  
Time Dependent ◽  
Content Type ◽  
Transcriptional Inhibition ◽  
Delayed Time

ABSTRACT Bedaquiline, an inhibitor of the mycobacterial ATP synthase, has revolutionized the treatment of Mycobacterium tuberculosis infection. Although a potent inhibitor, it is characterized by poorly understood delayed time-dependent bactericidal activity. Here, we demonstrate that in contrast to bedaquiline, the transcriptional inhibition of the ATP synthase in M. tuberculosis and Mycobacterium smegmatis has rapid bactericidal activity. These results validate the mycobacterial ATP synthase as a drug target with the potential for rapid bactericidal activity.

scholarly journals Preclinical Development of Inhalable D-Cycloserine and Ethionamide To Overcome Pharmacokinetic Interaction and Enhance Efficacy against Mycobacterium tuberculosis

2019 ◽  
Vol 63 (6) ◽  
Author(s):  
Rajeev Ranjan ◽  
Ashish Srivastava ◽  
Reena Bharti ◽  
Trisha Roy ◽  
Sonia Verma ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Bactericidal Activity ◽  
Half Life ◽  
Pharmacokinetic Interaction ◽  
Preclinical Development ◽  
Time Curve ◽  
Content Type ◽  
Concentration Time ◽  
Oral Doses ◽  
Plasma Half Life

ABSTRACT We compared the pharmacokinetics and efficacy of a combination of d-cycloserine (DCS) and ethionamide (ETO) via oral and inhalation routes in mice. The plasma half-life (t1/2) of oral ETO at a human-equivalent dose decreased from 4.63 ± 0.61 h to 1.64 ± 0.40 h when DCS was coadministered. The area under the concentration-time curve from 0 h to time t (AUC0–t) was reduced to one-third. Inhalation overcame the interaction. Inhalation, but not oral doses, reduced the lung CFU/g of Mycobacterium tuberculosis H37Rv from 6 to 3 log10 in 4 weeks, indicating bactericidal activity.

Anti money laundering using a two-phase system

2015 ◽  
Vol 18 (3) ◽  
pp. 304-329 ◽  
Author(s):  
Tamer Hossam Moustafa ◽  
Mohamed Zaki Abd El-Megied ◽  
Tarek Salah Sobh ◽  
Khaled Mohamed Shafea
Keyword(s):  
Money Laundering ◽  
False Positive ◽  
Phase System ◽  
Second Phase ◽  
Two Phase ◽  
Content Type ◽  
Risk Scoring ◽  
Two Phases ◽  
Clear Shift ◽  
Monitoring Phase

Purpose – This paper aims to compete and detect suspicious transactions that can lead to detecting money laundering cases. Design/methodology/approach – This paper presents a plan-based framework for anti-money laundering systems (PBAMLS). Such a framework is novel and consists of two phases, in addition to several supporting modules. The first phase, the monitoring phase, utilizes an automata approach as a formalism to detect probable money laundering. The detection process is based on a money laundering deterministic finite automaton that has been obtained from the corresponding regular expressions which specify different money laundering processes. The second phase is STRIPS-based planning phase that aims at strengthening the belief in the probable problems discovered in the first (monitoring) phase. In addition, PBAMLS contains several supporting modules for data collection and mediation, link analysis and risk scoring. To assess the applicability of PBAMLS, it has been tested using different cases studies. Findings – This framework provides a clear shift of anti-money laundering systems (AML) from depending heuristic and human expertise to making use of a rigorous formalism to accomplish concrete decisions. It minimizes the possibilities of false positive alarms and increases the certainty in decision-making. Practical implications – This framework enhances the detection of money laundering cases. It also minimizes the number of false-positive alarms that waste the investigators’ efforts and time; it decreases the efforts presented by the investigators. Originality/value – This work proposes PBAMLS as a novel plan-based framework for AML systems.

scholarly journals Identification of Mycobacterial Genes Involved in Antibiotic Sensitivity: Implications for the Treatment of Tuberculosis with β-Lactam-Containing Regimens

2017 ◽  
Vol 61 (7) ◽  
Author(s):  
Gopinath Viswanathan ◽  
Sangya Yadav ◽  
Tirumalai R. Raghunand
Keyword(s):  
Antibiotic Resistance ◽  
Mycobacterium Tuberculosis ◽  
Bactericidal Activity ◽  
Mycobacterium Smegmatis ◽  
Antibiotic Sensitivity ◽  
Mutant Library ◽  
Content Type ◽  
Factors Associated ◽  
Library Screen ◽  
Transposon Mutants

ABSTRACT In a Mycobacterium smegmatis mutant library screen, transposon mutants with insertions in fhaA, dprE2, rpsT, and parA displayed hypersusceptibility to antibiotics, including the β-lactams meropenem, ampicillin, amoxicillin, and cefotaxime. Sub-MIC levels of octoclothepin, a psychotic drug inhibiting ParA, phenocopied the parA insertion and enhanced the bactericidal activity of meropenem against Mycobacterium tuberculosis in combination with clavulanate. Our study identifies novel factors associated with antibiotic resistance, with implications in repurposing β-lactams for tuberculosis treatment.

scholarly journals Efficacy of Nitazoxanide against Clinical Isolates of Mycobacterium tuberculosis

2013 ◽  
Vol 57 (6) ◽  
pp. 2834-2837 ◽  
Author(s):  
Kristina Shigyo ◽  
Oksana Ocheretina ◽  
Yves Mary Merveille ◽  
Warren D. Johnson ◽  
Jean William Pape ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Bactericidal Activity ◽  
Laboratory Strain ◽  
Clinical Isolates ◽  
Second Line ◽  
Antituberculosis Drugs ◽  
Content Type

ABSTRACTNitazoxanide (NTZ) has bactericidal activity against theH37Rvlaboratory strain ofMycobacterium tuberculosiswith a MIC of 16 μg/ml. However, its efficacy against clinical isolates ofM. tuberculosishas not been determined. We found that NTZ's MIC against 50 clinical isolates ranged from 12 to 28 μg/ml with a median of 16 μg/ml and was unaffected by resistance to first- or second-line antituberculosis drugs or a diversity of spoligotypes.

scholarly journals Mutations in fbiD (Rv2983) as a Novel Determinant of Resistance to Pretomanid and Delamanid in Mycobacterium tuberculosis

2020 ◽  
Vol 65 (1) ◽  
pp. e01948-20
Author(s):  
Dalin Rifat ◽  
Si-Yang Li ◽  
Thomas Ioerger ◽  
Keshav Shah ◽  
Jean-Philippe Lanoix ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Clinical Evidence ◽  
Clinical Samples ◽  
Loss Of Function ◽  
Wild Type ◽  
Content Type ◽  
Solid Media ◽  
High Level ◽  
Level Resistance

ABSTRACTThe nitroimidazole prodrugs delamanid and pretomanid comprise one of only two new antimicrobial classes approved to treat tuberculosis (TB) in 50 years. Prior in vitro studies suggest a relatively low barrier to nitroimidazole resistance in Mycobacterium tuberculosis, but clinical evidence is limited to date. We selected pretomanid-resistant M. tuberculosis mutants in two mouse models of TB using a range of pretomanid doses. The frequency of spontaneous resistance was approximately 10−5 CFU. Whole-genome sequencing of 161 resistant isolates from 47 mice revealed 99 unique mutations, of which 91% occurred in 1 of 5 genes previously associated with nitroimidazole activation and resistance, namely, fbiC (56%), fbiA (15%), ddn (12%), fgd (4%), and fbiB (4%). Nearly all mutations were unique to a single mouse and not previously identified. The remaining 9% of resistant mutants harbored mutations in Rv2983 (fbiD), a gene not previously associated with nitroimidazole resistance but recently shown to be a guanylyltransferase necessary for cofactor F420 synthesis. Most mutants exhibited high-level resistance to pretomanid and delamanid, although Rv2983 and fbiB mutants exhibited high-level pretomanid resistance but relatively small changes in delamanid susceptibility. Complementing an Rv2983 mutant with wild-type Rv2983 restored susceptibility to pretomanid and delamanid. By quantifying intracellular F420 and its precursor Fo in overexpressing and loss-of-function mutants, we provide further evidence that Rv2983 is necessary for F420 biosynthesis. Finally, Rv2983 mutants and other F420H2-deficient mutants displayed hypersusceptibility to some antibiotics and to concentrations of malachite green found in solid media used to isolate and propagate mycobacteria from clinical samples.

scholarly journals Mutation ofRv2887, amarR-Like Gene, Confers Mycobacterium tuberculosis Resistance to an Imidazopyridine-Based Agent

2015 ◽  
Vol 59 (11) ◽  
pp. 6873-6881 ◽  
Author(s):  
Kathryn Winglee ◽  
Shichun Lun ◽  
Marco Pieroni ◽  
Alan Kozikowski ◽  
William Bishai
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Drug Targets ◽  
Efflux Pump ◽  
Transcriptional Regulator ◽  
Cross Resistance ◽  
Loss Of Function ◽  
Strong Activity ◽  
Content Type ◽  
Novel Drug

ABSTRACTDrug resistance is a major problem inMycobacterium tuberculosiscontrol, and it is critical to identify novel drug targets and new antimycobacterial compounds. We have previously identified an imidazo[1,2-a]pyridine-4-carbonitrile-based agent, MP-III-71, with strong activity againstM. tuberculosis. In this study, we evaluated mechanisms of resistance to MP-III-71. We derived three independentM. tuberculosismutants resistant to MP-III-71 and conducted whole-genome sequencing of these mutants. Loss-of-function mutations inRv2887were common to all three MP-III-71-resistant mutants, and we confirmed the role ofRv2887as a gene required for MP-III-71 susceptibility using complementation. The Rv2887 protein was previously unannotated, but domain and homology analyses suggested it to be a transcriptional regulator in the MarR (multiple antibiotic resistance repressor) family, a group of proteins first identified inEscherichia colito negatively regulate efflux pumps and other mechanisms of multidrug resistance. We found that two efflux pump inhibitors, verapamil and chlorpromazine, potentiate the action of MP-III-71 and that mutation ofRv2887abrogates their activity. We also used transcriptome sequencing (RNA-seq) to identify genes which are differentially expressed in the presence and absence of a functional Rv2887 protein. We found that genes involved in benzoquinone and menaquinone biosynthesis were repressed by functional Rv2887. Thus, inactivating mutations ofRv2887, encoding a putative MarR-like transcriptional regulator, confer resistance to MP-III-71, an effective antimycobacterial compound that shows no cross-resistance to existing antituberculosis drugs. The mechanism of resistance ofM. tuberculosisRv2887mutants may involve efflux pump upregulation and also drug methylation.

scholarly journals Evaluation of a New Immunochromatography Technology Test (LDBio Diagnostics) To Detect Toxoplasma IgG and IgM: Comparison with the Routine Architect Technique

2017 ◽  
Vol 55 (12) ◽  
pp. 3395-3404 ◽  
Author(s):  
Caroline Mahinc ◽  
Pierre Flori ◽  
Edouard Delaunay ◽  
Cécile Guillerme ◽  
Sana Charaoui ◽  
...  
Keyword(s):  
Western Blot ◽  
Sensitivity And Specificity ◽  
Blot Analysis ◽  
False Positive ◽  
Automated System ◽  
University Hospitals ◽  
Content Type ◽  
Automated Screening ◽  
Commercial Kits ◽  
Fully Automated System

ABSTRACTA study comparing the ICT (immunochromatography technology)ToxoplasmaIgG and IgM rapid diagnostic test (LDBio Diagnostics, France) with a fully automated system, Architect, was performed on samples from university hospitals of Marseille and Saint-Etienne. A total of 767 prospective sera and 235 selected sera were collected. The panels were selected to test various IgG and IgM parameters. The reference technique,ToxoplasmaIgGII Western blot analysis (LDBio Diagnostics), was used to confirm the IgG results, and commercial kits Platelia Toxo IgM (Bio-Rad) and Toxo-ISAgA (bioMérieux) were used in Saint-Etienne and Marseille, respectively, as the IgM reference techniques. Sensitivity and specificity of the ICT and the Architect IgG assays were compared using a prospective panel. Sensitivity was 100% for the ICT test and 92.1% for Architect (cutoff at 1.6 IU/ml). The low-IgG-titer serum results confirmed that ICT sensitivity was superior to that of Architect. Specificity was 98.7% (ICT) and 99.8% (Architect IgG). The ICT test is also useful for detecting IgM without IgG and is both sensitive (100%) and specific (100%), as it can distinguish nonspecific IgM from specificToxoplasmaIgM. In comparison, IgM sensitivity and specificity on Architect are 96.1% and 99.6%, respectively (cutoff at 0.5 arbitrary units [AU]/ml). To conclude, this new test overcomes the limitations of automated screening techniques, which are not sensitive enough for IgG and lack specificity for IgM (rare IgM false-positive cases).

scholarly journals A Novel 6-Benzyl Ether Benzoxaborole Is Active against Mycobacterium tuberculosisIn Vitro

2017 ◽  
Vol 61 (9) ◽  
Author(s):  
Nipul Patel ◽  
Theresa O'Malley ◽  
Yong-Kang Zhang ◽  
Yi Xia ◽  
Bjorn Sunde ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Liquid Medium ◽  
Inhibitory Concentration ◽  
Solid Medium ◽  
Eukaryotic Cells ◽  
Intracellular Bacteria ◽  
Benzyl Ether ◽  
Content Type ◽  
Resistant Mutants

ABSTRACT We identified a novel 6-benzyl ether benzoxaborole with potent activity against Mycobacterium tuberculosis. The compound had an MIC of 2 μM in liquid medium. The compound was also able to prevent growth on solid medium at 0.8 μM and was active against intracellular bacteria (50% inhibitory concentration [IC50] = 3.6 μM) without cytotoxicity against eukaryotic cells (IC50 > 100 μM). We isolated resistant mutants (MIC ≥ 100 μM), which had mutations in Rv1683, Rv3068c, and Rv0047c.

scholarly journals Characterization of T Cells Specific for CFP-10 and ESAT-6 in Mycobacterium tuberculosis-Infected Mauritian Cynomolgus Macaques

2017 ◽  
Vol 85 (4) ◽  
Author(s):  
Amy Ellis ◽  
Alexis Balgeman ◽  
Mark Rodgers ◽  
Cassaundra Updike ◽  
Jaime Tomko ◽  
...  
Keyword(s):  
T Cells ◽  
Mycobacterium Tuberculosis ◽  
Mhc Class Ii ◽  
Nonhuman Primates ◽  
Content Type ◽  
Host Immune Responses ◽  
Cell Responses ◽  
Cynomolgus Macaques ◽  
Mhc Haplotype

ABSTRACT Nonhuman primates can be used to study host immune responses to Mycobacterium tuberculosis. Mauritian cynomolgus macaques (MCMs) are a unique group of animals that have limited major histocompatibility complex (MHC) genetic diversity, such that MHC-identical animals can be infected with M. tuberculosis. Two MCMs homozygous for the relatively common M1 MHC haplotype were bronchoscopically infected with 41 CFU of the M. tuberculosis Erdman strain. Four other MCMs, which had at least one copy of the M1 MHC haplotype, were infected with a lower dose of 3 CFU M. tuberculosis. All animals mounted similar T-cell responses to CFP-10 and ESAT-6. Two epitopes in CFP-10 were characterized, and the MHC class II alleles restricting them were determined. A third epitope in CFP-10 was identified but exhibited promiscuous restriction. The CFP-10 and ESAT-6 antigenic regions targeted by T cells in MCMs were comparable to those seen in cases of human M. tuberculosis infection. Our data lay the foundation for generating tetrameric molecules to study epitope-specific CD4 T cells in M. tuberculosis-infected MCMs, which may guide future testing of tuberculosis vaccines in nonhuman primates.

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