In VitroandIn VivoEfficacy of β-Lactams against Replicating and Slowly Growing/Nonreplicating Mycobacterium tuberculosis

ABSTRACTBeta-lactams, in combination with beta-lactamase inhibitors, are reported to have activity againstMycobacterium tuberculosisbacteria growing in broth, as well as inside the human macrophage. We tested representative beta-lactams belonging to 3 different classes for activity against replicatingM. tuberculosisin broth and nonreplicatingM. tuberculosisunder hypoxia, as well as against streptomycin-starvedM. tuberculosisstrain 18b (ss18b) in the presence or absence of clavulanate. Most of the combinations showed bactericidal activity against replicatingM. tuberculosis, with up to 200-fold improvement in potency in the presence of clavulanate. None of the combinations, including those containing meropenem, imipenem, and faropenem, killedM. tuberculosisunder hypoxia. However, faropenem- and meropenem-containing combinations killed strain ss18b moderately. We tested the bactericidal activities of meropenem-clavulanate and amoxicillin-clavulanate combinations in the acute and chronic aerosol infection models of tuberculosis in BALB/c mice. Based on pharmacokinetic/pharmacodynamic indexes reported for beta-lactams against other bacterial pathogens, a cumulative percentage of a 24-h period that the drug concentration exceeds the MIC under steady-state pharmacokinetic conditions (%TMIC) of 20 to 40% was achieved in mice using a suitable dosing regimen. Both combinations showed marginal reduction in lung CFU compared to the late controls in the acute model, whereas both were inactive in the chronic model.

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In VivoEfficacy of Apramycin in Murine Infection Models

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.03239-14 ◽

2014 ◽

Vol 58 (11) ◽

pp. 6938-6941 ◽

Cited By ~ 11

Author(s):

Martin Meyer ◽

Pietro Freihofer ◽

Michael Scherman ◽

Joanne Teague ◽

Anne Lenaerts ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Antibacterial Activity ◽

Mycobacterium Tuberculosis ◽

Antibacterial Efficacy ◽

Drug Resistant ◽

Murine Models ◽

Content Type ◽

Infection Models ◽

Aerosol Infection ◽

Further Development

ABSTRACTApramycin is a unique aminoglycoside with a dissociation of antibacterial activity and ototoxicity. We assessed the antibacterial efficacy of apramycin in two murine models of infection,Mycobacterium tuberculosisaerosol infection andStaphylococcus aureussepticemia. In both infection models, the efficacy of apramycin was comparable to that of amikacin. These results suggest that apramycin has the potential to become a clinically useful agent against drug-resistant pathogens and support further development of this promising unique aminoglycoside.

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Transcriptional Profiling of Mycobacterium tuberculosis Exposed toIn VitroLysosomal Stress

Infection and Immunity ◽

10.1128/iai.00072-16 ◽

2016 ◽

Vol 84 (9) ◽

pp. 2505-2523 ◽

Cited By ~ 22

Author(s):

Wenwei Lin ◽

Paola Florez de Sessions ◽

Garrett Hor Keong Teoh ◽

Ahmad Naim Nazri Mohamed ◽

Yuan O. Zhu ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Efflux Pump ◽

Metabolic Reprogramming ◽

Human Macrophage ◽

Host Immune System ◽

Activated Macrophages ◽

Intrinsic Resistance ◽

Macrophage Infection ◽

Content Type

Increasing experimental evidence supports the idea thatMycobacterium tuberculosishas evolved strategies to survive within lysosomes of activated macrophages. To further our knowledge ofM. tuberculosisresponse to the hostile lysosomal environment, we profiled the global transcriptional activity ofM. tuberculosiswhen exposed to the lysosomal soluble fraction (SF) prepared from activated macrophages. Transcriptome sequencing (RNA-seq) analysis was performed using various incubation conditions, ranging from noninhibitory to cidal based on the mycobacterial replication or killing profile. Under inhibitory conditions that led to the absence of apparent mycobacterial replication,M. tuberculosisexpressed a unique transcriptome with modulation of genes involved in general stress response, metabolic reprogramming, respiration, oxidative stress, dormancy response, and virulence. The transcription pattern also indicates characteristic cell wall remodeling with the possible outcomes of increased infectivity, intrinsic resistance to antibiotics, and subversion of the host immune system. Among the lysosome-specific responses, we identified theglgE-mediated 1,4 α-glucan synthesis pathway and a defined group of VapBC toxin/anti-toxin systems, both of which represent toxicity mechanisms that potentially can be exploited for killing intracellular mycobacteria. A meta-analysis including previously reported transcriptomic studies in macrophage infection andin vitrostress models was conducted to identify overlapping and nonoverlapping pathways. Finally, the Tap efflux pump-encoding geneRv1258cwas selected for validation. AnM. tuberculosis ΔRv1258cmutant was constructed and displayed increased susceptibility to killing by lysosomal SF and the antimicrobial peptide LL-37, as well as attenuated survival in primary murine macrophages and human macrophage cell line THP-1.

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Susceptibility Testing of Extensively Drug-Resistant and Pre-Extensively Drug-Resistant Mycobacterium tuberculosis against Levofloxacin, Linezolid, and Amoxicillin-Clavulanate

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02020-12 ◽

2013 ◽

Vol 57 (6) ◽

pp. 2522-2525 ◽

Cited By ~ 20

Author(s):

Imran Ahmed ◽

Kauser Jabeen ◽

Raunaq Inayat ◽

Rumina Hasan

Keyword(s):

Mycobacterium Tuberculosis ◽

Susceptibility Testing ◽

Critical Concentration ◽

Drug Susceptibility ◽

Drug Susceptibility Testing ◽

Fluoroquinolone Resistance ◽

Drug Resistant ◽

Content Type ◽

Extensively Drug Resistant ◽

Amoxicillin Clavulanate

ABSTRACTPakistan is a high-burden country for tuberculosis (TB). The emergence and increasing incidence of extensively drug-resistant (XDR) TB has been reported in Pakistan. Similarly, the prevalence of multidrug-resistant TB infections with fluoroquinolone resistance (pre-XDR) is also increasing. To treat these infections, local drug susceptibility patterns of alternate antituberculosis agents, including levofloxacin (LVX), linezolid (LZD), and amoxicillin-clavulanate (AMC), is urgently needed. The aim of this study was to determine the susceptibility frequencies of drug-resistant (DR)Mycobacterium tuberculosisagainst LVX, LZD, and AMC. All susceptibilities were determined on Middlebrook 7H10 agar. A critical concentration was used for LVX (1 μg/ml), whereas MICs were determined for LZD and AMC.M. tuberculosisH37Rv was used as a control strain. A total of 102M. tuberculosisisolates (XDR,n= 59; pre-XDR,n= 43) were tested. Resistance to LVX was observed in 91.2% (93/102). Using an MIC value of 0.5 μg/ml as a cutoff, resistance to LZD (MIC ≥ 1 μg/ml) was noted in 5.9% (6/102). Although the sensitivity breakpoints are not established for AMC, the MIC values were high (>16 μg/ml) in 97.1% (99/102). Our results demonstrate that LZD may be effective for the treatment of XDR and pre-XDR cases from Pakistan. High resistance rates against LVX in our study suggest the use of this drug with caution for DR-TB cases from this area. Drug susceptibility testing against LVX and AMC may be helpful in complicated and difficult-to-manage cases.

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Effect of Coadministration of Moxifloxacin and Rifampin on Mycobacterium tuberculosis in a Murine Aerosol Infection Model

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.06383-11 ◽

2012 ◽

Vol 56 (6) ◽

pp. 3054-3057 ◽

Cited By ~ 18

Author(s):

V. Balasubramanian ◽

S. Solapure ◽

S. Gaonkar ◽

K. N. Mahesh Kumar ◽

R. K. Shandil ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Hollow Fiber ◽

Lung Model ◽

Control Group ◽

Infection Model ◽

Content Type ◽

Days Of Therapy ◽

Cell Kill ◽

Aerosol Infection

ABSTRACTCoadministration of moxifloxacin and rifampin was evaluated in a murine model ofMycobacterium tuberculosispulmonary infection to determine whether the finding of antagonism documented in a hollow-fiber infection model could be recapitulatedin vivo. Colony counts were followed in a no-treatment control group, groups administered moxifloxacin or rifampin monotherapy, and a group administered a combination of the two agents. Following 18 days of once-daily oral administration to mice infected withM. tuberculosis, there was a reduction in the plasma exposure to rifampin that decreased further when rifampin was coadministered with moxifloxacin. Pharmacodynamic analysis demonstrated a mild antagonistic interaction between moxifloxacin and rifampin with respect to cell kill in the mouse model for tuberculosis (TB). No emergence of resistance was noted over 28 days of therapy, even with monotherapy. This was true even though one of the agents in the combination (moxifloxacin) induces error-prone replication. The previously noted antagonism with respect to cell kill shown in the hollow-fiber infection model was recapitulated in the murine TB lung model, although to a lesser extent.

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The Mycobacterium tuberculosis sRNA F6 Modifies Expression of Essential Chaperonins, GroEL2 and GroES

Microbiology Spectrum ◽

10.1128/spectrum.01095-21 ◽

2021 ◽

Author(s):

Joanna Houghton ◽

Angela Rodgers ◽

Graham Rose ◽

Alexandre D’Halluin ◽

Terry Kipkorir ◽

...

Keyword(s):

Gene Expression ◽

Mycobacterium Tuberculosis ◽

Differential Gene Expression ◽

Control Of Gene Expression ◽

Content Type ◽

Infection Models ◽

Small Regulatory Rnas ◽

Differential Gene ◽

Regulatory Rnas

Control of gene expression via small regulatory RNAs (sRNAs) is poorly understood in one of the most successful pathogens, Mycobacterium tuberculosis . Here, we present an in-depth characterization of the sRNA F6, including its expression in different infection models and the differential gene expression observed upon deletion of the sRNA.

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In Vitro and In Vivo Activities of the Riminophenazine TBI-166 against Mycobacterium tuberculosis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02155-18 ◽

2019 ◽

Vol 63 (5) ◽

Cited By ~ 9

Author(s):

Jian Xu ◽

Bin Wang ◽

Lei Fu ◽

Hui Zhu ◽

Shaochen Guo ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Skin Pigmentation ◽

Multidrug Resistant ◽

Content Type ◽

Resistant Tuberculosis ◽

Wild Strains ◽

Skin Discoloration ◽

Aerosol Infection

ABSTRACT The riminophenazine agent clofazimine (CFZ) is repurposed as an important component of the new short-course multidrug-resistant tuberculosis regimen and significantly shortens first-line regimen for drug-susceptible tuberculosis in mice. However, CFZ use is hampered by its unwelcome skin discoloration in patients. A new riminophenazine analog, TBI-166, was selected as a potential next-generation antituberculosis riminophenazine following an extensive medicinal chemistry effort. Here, we evaluated the activity of TBI-166 against Mycobacterium tuberculosis and its potential to accumulate and discolor skin. The in vitro activity of TBI-166 against both drug-sensitive and drug-resistant M. tuberculosis is more potent than that of CFZ. Spontaneous mutants resistant to TBI-166 were found at a frequency of 2.3 × 10−7 in wild strains of M. tuberculosis. TBI-166 demonstrates activity at least equivalent to that of CFZ against intracellular M. tuberculosis and in low-dose aerosol infection models of acute and chronic murine tuberculosis. Most importantly, TBI-166 causes less skin discoloration than does CFZ despite its higher tissue accumulation. The efficacy of TBI-166, along with its decreased skin pigmentation, warrants further study and potential clinical use.

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Simple and Rapid Method To Determine Antimycobacterial Potency of Compounds by Using Autoluminescent Mycobacterium tuberculosis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.03205-14 ◽

2014 ◽

Vol 58 (10) ◽

pp. 5801-5808 ◽

Cited By ~ 16

Author(s):

Sreevalli Sharma ◽

Ekaterina Gelman ◽

Chandan Narayan ◽

Deepa Bhattacharjee ◽

Vijayashree Achar ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Slow Growth ◽

Gc Content ◽

Content Type ◽

Slow Growth Rate ◽

Drug Candidates ◽

Luminescence Assay ◽

Bactericidal Activities ◽

Time Required

ABSTRACTA major obstacle in the process of discovery of drugs againstMycobacterium tuberculosisis its extremely slow growth rate and long generation time (∼20 to 24 h). Consequently, determination of MICs and minimum bactericidal concentrations (MBCs) of potential drug candidates using current methods requires 7 days (resazurin-based MIC assay [REMA]) and 1 month (CFU enumeration), respectively. We employed a synthetic luciferase operon optimized for expression in high-GC-content bacteria and adapted it for use in mycobacteria. Using luminescence-based readouts, we were able to determine the MICs and bactericidal activities of approved tuberculosis (TB) drugs, which correlated well with currently used methods. Although luminescence-based readouts have been used previously to determine the MICs and bactericidal activities of approved TB drugs, in this study we adapted this assay to carry out a pilot screen using a library of 1,114 compounds belonging to diverse chemical scaffolds. We found that MICs derived from a 3-day luminescence assay matched well with REMA-based MIC values. To determine the bactericidal potencies of compounds, a 1:10 dilution of the cultures from the MIC plate was carried out on day 7, and the bactericidal concentrations determined based on time to positivity in 2 weeks were found to be comparable with MBC values determined by the conventional CFU approach. Thus, the luminescent mycobacterium-based approach not only is very simple and inexpensive but also allowed us to generate the information in half the time required by conventional methods.

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Mycobacterium tuberculosis Lacking All Mycolic Acid Cyclopropanation Is Viable but Highly Attenuated and Hyperinflammatory in Mice

Infection and Immunity ◽

10.1128/iai.00021-12 ◽

2012 ◽

Vol 80 (6) ◽

pp. 1958-1968 ◽

Cited By ~ 47

Author(s):

Daniel Barkan ◽

Dorsaf Hedhli ◽

Han-Guang Yan ◽

Kris Huygen ◽

Michael S. Glickman

Keyword(s):

Mycobacterium Tuberculosis ◽

Mycolic Acid ◽

Acid Modification ◽

Content Type ◽

Mycolic Acids ◽

Trehalose Dimycolate ◽

Cord Factor ◽

Aerosol Infection ◽

Enzyme Class

ABSTRACTMycolic acids, the major lipid of theMycobacterium tuberculosiscell wall, are modified by cyclopropane rings, methyl branches, and oxygenation through the action of eightS-adenosylmethionine (SAM)-dependent mycolic acid methyltransferases (MAMTs), encoded at four genetic loci. Mycolic acid modification has been shown to be important forM. tuberculosispathogenesis, in part through effects on the inflammatory activity of trehalose dimycolate (cord factor). Studies using the MAMT inhibitor dioctylamine have suggested that the MAMT enzyme class is essential forM. tuberculosisviability. However, it is unknown whether a cyclopropane-deficient strain ofM. tuberculosiswould be viable and what the effect of cyclopropane deficiency on virulence would be. We addressed these questions by creating and characterizingM. tuberculosisstrains lacking all functional MAMTs. Our results show thatM. tuberculosisis viable either without cyclopropanation or without cyclopropanation and any oxygenated mycolates. Characterization of these strains revealed that MAMTs are required for acid fastness and resistance to detergent stress. Complete lack of cyclopropanation confers severe attenuation during the first week after aerosol infection of the mouse, whereas complete loss of MAMTs confers attenuation in the second week of infection. Characterization of immune responses to the cyclopropane- and MAMT-deficient strains indicated that the net effect of mycolate cyclopropanation is to dampen host immunity. Taken together, our findings establish the immunomodulatory function of the mycolic acid modification pathway in pathogenesis and buttress this enzyme class as an attractive target for antimycobacterial drug development.

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Isoniazid Pharmacokinetics-Pharmacodynamics in an Aerosol Infection Model of Tuberculosis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.48.8.2951-2957.2004 ◽

2004 ◽

Vol 48 (8) ◽

pp. 2951-2957 ◽

Cited By ~ 92

Author(s):

Ramesh Jayaram ◽

Radha. K. Shandil ◽

Sheshagiri Gaonkar ◽

Parvinder Kaur ◽

B. L. Suresh ◽

...

Keyword(s):

Dose Fractionation ◽

Maximum Effect ◽

Infection Model ◽

Limited Data ◽

Time Curve ◽

Bactericidal Efficacy ◽

Infection Models ◽

Bactericidal Activities ◽

Aerosol Infection

ABSTRACT Limited data exist on the pharmacokinetic-pharmacodynamic (PK-PD) parameters of the bactericidal activities of the available antimycobacterial drugs. We report on the PK-PD relationships for isoniazid. Isoniazid exhibited concentration (C)-dependent killing of Mycobacterium tuberculosis H37Rv in vitro, with a maximum reduction of 4 log10 CFU/ml. In these studies, 50% of the maximum effect was achieved at a C/MIC ratio of 0.5, and the maximum effect did not increase with exposure times of up to 21 days. Conversely, isoniazid produced less than a 0.5-log10 CFU/ml reduction in two different intracellular infection models (J774A.1 murine macrophages and whole human blood). In a murine model of aerosol infection, isoniazid therapy for 6 days produced a reduction of 1.4 log10 CFU/lung. Dose fractionation studies demonstrated that the 24-h area under the concentration-time curve/MIC (r 2 = 0.83) correlated best with the bactericidal efficacy, followed by the maximum concentration of drug in serum/MIC (r 2 = 0.73).

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

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01948-20 ◽

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.

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