Edwards R.L.,University of Michigan |
Edwards R.L.,Tuberculosis Research Section |
Jules M.,Institute Pasteur Paris |
Jules M.,French National Center for Scientific Research |
And 6 more authors.
Infection and Immunity | Year: 2010
When confronted with metabolic stress, replicative Legionella pneumophila bacteria convert to resilient, infectious cells equipped for transmission. Differentiation is promoted by the LetA/LetS two-component system, which belongs to a family of signal-transducing proteins that employ a four-step phosphorelay to regulate gene expression. Histidine 307 of LetS was essential to switch on the transmission profile, but a threonine substitution at position 311 (T311M) suggested a rheostat-like function. The letS(T311M) bacteria resembled the wild type (WT) for some traits and letS null mutants for others, whereas they displayed intermediate levels of infectivity, cytotoxicity, and lysosome evasion. Although only 30 to 50% of letS(T311M) mutants became motile, flow cytometry determined that every cell eventually activated the flagellin promoter to WT levels, but expression was delayed. Likewise, letS(T311M) mutants exhibited delayed induction of RsmY and RsmZ, regulatory RNAs that relieve CsrA repression of transmission traits. Transcriptional profile analysis revealed that letS(T311M) mutants expressed the flagellar regulon and multiple other transmissive-phase loci at a higher cell density than the WT. Accordingly, we postulate that the letS(T311M) mutant may relay phosphate less efficiently than the WT LetS sensor protein, leading to sluggish gene expression and a variety of phenotypic profiles. Thus, as first described for BvgA/BvgS, rather than acting as on/off switches, this family of two-component systems exhibit rheostat activity that likely confers versatility as microbes adapt to fluctuating environments. Copyright © 2010, American Society for Microbiology. All Rights Reserved.
Arnold M.F.F.,University of Aberdeen |
Haag A.F.,University of Aberdeen |
Capewell S.,University of Edinburgh |
Boshoff H.I.,Tuberculosis Research Section |
And 13 more authors.
Journal of Bacteriology | Year: 2013
The Sinorhizobium meliloti BacA ABC transporter protein plays an important role in its nodulating symbiosis with the legume alfalfa (Medicago sativa). The Mycobacterium tuberculosis BacA homolog was found to be important for the maintenance of chronic murine infections, yet its in vivo function is unknown. In the legume plant as well as in the mammalianhost, bacteria encounter host antimicrobial peptides (AMPs). We found that the M. tuberculosis BacA protein was able to partially complement the symbiotic defect of an S. meliloti BacA-deficient mutant on alfalfa plants and to protect this mutant in vitro from the antimicrobial activity of a synthetic legume peptide, NCR247, and a recombinant human β-defensin 2 (HBD2). This finding was also confirmed using an M. tuberculosis insertion mutant. Furthermore, M. tuberculosis BacA-mediated protection of the legume symbiont S. meliloti against legume defensins as well as HBD2 is dependent on its attached ATPase domain. In addition, we show that M. tuberculosis BacA mediates peptide uptake of the truncated bovine AMP, Bac71-16. This process required a functional ATPase domain. We therefore suggest that M. tuberculosis BacA is important for the transport of peptides across the cytoplasmic membrane and is part of a complete ABC transporter. Hence, BacA-mediated protection against host AMPs might be important for the maintenance of latent infections. © 2013, American Society for Microbiology.
Comparative evaluation of Sloppy Molecular Beacon and Dual-labeled probe melting temperature assays to identify mutations in Mycobacterium tuberculosis resulting in Rifampin, Fluoroquinolone and Aminoglycoside resistance
Roh S.S.,Rutgers University |
Smith L.E.,Rutgers University |
Lee J.S.,International Tuberculosis Research Center |
Via L.E.,Tuberculosis Research Section |
And 3 more authors.
PLoS ONE | Year: 2015
Several molecular assays to detect resistance to Rifampin, the Fluoroquinolones, and Aminoglycosides in Mycobacterium tuberculosis (M. tuberculosis) have been recently described. A systematic approach for comparing these assays in the laboratory is needed in order to determine the relative advantage of each assay and to decide which ones should be advanced to evaluation. We performed an analytic comparison of a Sloppy Molecular Beacon (SMB) melting temperature (Tm) assay and a Dual labeled probe (DLP) Tm assay. Both assays targeted the M. tuberculosis rpoB, gyrA, rrs genes and the eis promoter region. The sensitivity and specificity to detect mutations, analytic limit of detection (LOD) and the detection of heteroresistance were tested using a panel of 56 clinical DNA samples from drug resistant M. tuberculosis strains. Both SMB and DLP assays detected 29/29 (100%) samples with rpoB RRDR mutations and 3/3 (100%) samples with eis promoter mutations correctly. The SMB assay detected all 17/17 gyrA mutants and 22/22 rrs mutants, while the DLP assay detected 16/17 (94%) gyrA mutants and 12/22 (55%) rrs mutants. Both assays showed comparable LODs for detecting rpoB and eis mutations; however, the SMB assay LODs were at least two logs better for detecting wild type and mutants in gyrA and rrs targets. The SMB assay was also moderately better at detecting heteroresistance. In summary, both assays appeared to be promising methods to detect drug resistance associated mutations in M. tuberculosis; however, the relative advantage of each assay varied under each test condition. © 2015, Public Library of Science. All rights reserved.
Yang X.,State University of New York at Stony Brook |
Yang X.,Tuberculosis Research Section |
Gao J.,State University of New York at Stony Brook |
Gao J.,Boston University |
And 3 more authors.
Journal of Bacteriology | Year: 2011
Rv1106c (hsd; 3β-hydroxysteroid dehydrogenase) is required by Mycobacterium tuberculosis for growth on cholesterol as a sole carbon source, whereas Rv3409c is not. Mutation of Rv1106c does not reduce Mycobacterium tuberculosis growth in infected macrophages or guinea pigs. We conclude that cholesterol is not required as a nutritional source during infection. Copyright © 2011, American Society for Microbiology. All Rights Reserved.
Boshoff H.I.M.,Tuberculosis Research Section |
Lun D.S.,University of South Australia
Drug Discovery Today: Disease Mechanisms | Year: 2010
The advent of high-throughput platforms for the interrogation of biological systems at the cellular and molecular levels has allowed living cells to be observed and understood at a hitherto unprecedented level of detail and has enabled the construction of comprehensive, predictive in silico models. Here, we review the application of such high-throughput, systems-biological techniques to mycobacteria - specifically to the pernicious human pathogen Mycobacterium tuberculosis (MTb) and its ability to survive in human hosts. We discuss the development and application of transcriptomic, proteomic, regulomic, and metabolomic techniques for MTb as well as the development and application of genome-scale in silico models. Thus far, systems-biological approaches have largely focused on in vitro models of MTb growth; reliably extending these approaches to in vivo conditions relevant to infection is a significant challenge for the future that holds the ultimate promise of novel chemotherapeutic interventions. © 2010 Elsevier Ltd. All rights reserved.
Kostova M.B.,American University of Washington |
Myers C.J.,American University of Washington |
Beck T.N.,American University of Washington |
Plotkin B.J.,Midwestern University |
And 5 more authors.
Bioorganic and Medicinal Chemistry | Year: 2011
Antimicrobial resistance represents a global threat to healthcare. The ability to adequately treat infectious diseases is increasingly under siege due to the emergence of drug-resistant microorganisms. New approaches to drug development are especially needed to target organisms that exhibit broad antibiotic resistance due to expression of β-lactamases which is the most common mechanism by which bacteria become resistant to β-lactam antibiotics. We designed and synthesized 20 novel monocyclic β-lactams with alkyl- and aryl-thio moieties at C4, and subsequently tested these for antibacterial activity. These compounds demonstrated intrinsic activity against serine β-lactamase producing Mycobacterium tuberculosis wild type strain (Mtb) and multiple (n = 6) β-lactamase producing Moraxella catarrhalis clinical isolates. © 2011 Elsevier Ltd. All rights reserved.
Lee I.-Y.,Korea Research Institute of Chemical Technology |
Gruber T.D.,Tuberculosis Research Section |
Samuels A.,Tuberculosis Research Section |
Yun M.,Korea Research Institute of Chemical Technology |
And 7 more authors.
Bioorganic and Medicinal Chemistry | Year: 2013
A series of salicylanilides was synthesized based on a high-throughput screening hit against Mycobacterium tuberculosis. A free phenolic hydroxyl on the salicylic acid moeity is required for activity, and the structure-activity relationship of the aniline ring is largely driven by the presence of electron withdrawing groups. We synthesized 94 analogs exploring substitutions of both rings and the linker region in this series and we have identified multiple compounds with low micromolar potency. Unfortunately, cytotoxicity in a murine macrophage cell line trends with antimicrobial activity, suggesting a similar mechanism of action. We propose that salicylanilides function as proton shuttles that kill cells by destroying the cellular proton gradient, limiting their utility as potential therapeutics. © 2012 Elsevier Ltd. All rights reserved.
Varshney S.K.,Tuberculosis Research Section |
Shukla I.,Tuberculosis Research Section |
Raza A.,Tuberculosis Research Section |
Ahmad Z.,Jn Medical College
Asian Pacific Journal of Tropical Disease | Year: 2014
Objective: To evaluate in vitro resistance pattern of the first line anti-tubercular drugs in new and previously treated cases of pulmonary tuberculosis patients in Aligarh region. Methods: This study was carried out involving 975 suspected tuberculosis patients. All the specimens of patients were subjected to Ziehl-Neelsen staining, cultured on Lowenstein-Jensen medium and resistance pattern was evaluated by standard proportion method. All patients diagnosed with pulmonary tuberculosis were placed in CAT I and II under Revised National Tuberculosis Control Programme guidelines. Result: Out of 220 patients, 129 (58.7%) were from CAT I and 91 (41.3%) were from CAT II. Totally 44.5% were resistant to one or more than two drugs and 18.6% patients showed resistance to both isoniazid and rifampicin. The individual resistance pattern of these first line drugs were as follows: 37.7% patients were resistant to isoniazid, 22.2% to rifampicin, 8.6% to streptomycin and 10% were resistant to ethambutol. Conclusions: Our findings concluded a high prevalence of in vitro drug resistance of Mycobacterium tuberculosis isolates, especially multidrug resistant tuberculosis, in both the categories. So there is an urgent need to further study the risk factors for transmission and multidrug resistant tuberculosis in these settings. © 2014 by the Asian Pacific Journal of Tropical Disease.
Mukherjee T.,Tuberculosis Research Section |
Boshoff H.,Tuberculosis Research Section
Future Medicinal Chemistry | Year: 2011
Tuberculosis remains a leading cause of death resulting from an infectious agent, and the spread of multi- and extensively drug-resistant strains of Mycobacterium tuberculosis poses a threat to management of global health. New drugs that effectively shorten the duration of treatment and are active against drug-resistant strains of this pathogen are urgently required to develop effective chemotherapies to combat this disease. Two nitroimidazoles, PA-824 and OPC-67683, are currently in Phase II clinical trials for the treatment of TB and the outcome of these may determine the future directions of drug development for anti-tubercular nitroimidazoles. In this review we summarize the development of these nitroimidazoles and alternative analogs in these series that may offer attractive alternatives to PA-824 and OPC-67683 for further development in the drug-discovery pipeline. Lastly, the potential pitfalls in the development of nitroimidazoles as drugs for TB are discussed. © 2011 Future Science Ltd.
PubMed | Tuberculosis Research Section
Type: Journal Article | Journal: Drug discovery today. Disease mechanisms | Year: 2010
The advent of high-throughput platforms for the interrogation of biological systems at the cellular and molecular level have allowed living cells to be observed and understood at a hitherto unprecedented level of detail and have enabled the construction of comprehensive, predictive in silico models. Here, we review the application of such high-throughput, systems-biological techniques to mycobacteria-specifically to the pernicious human pathogen Mycobacterium tuberculosis (MTb) and its ability to survive in human hosts. We discuss the development and application of transcriptomic, proteomic, regulomic, and metabolomic techniques for MTb as well as the development and application of genome-scale in silico models. Thus far, systems-biological approaches have largely focused on in vitro models of MTb growth; reliably extending these approaches to in vivo conditions relevant to infection is a significant challenge for the future that holds the ultimate promise of novel chemotherapeutic interventions.