Treatment Of Mycobacterium Tuberculosis Infection Research Articles

Isoniazid therapy for Mycobacterium tuberculosis infection in HIV clinics, Los Angeles, California.

Publicly funded human immunodeficiency virus (HIV) clinics in Los Angeles County, California, USA. HIV-infected persons are a high priority group for targeted testing and treatment for Mycobacterium tuberculosis infection in the United States. To describe rates of isoniazid (INH) initiation and completion among HIV-1 and M. tuberculosis co-infected persons in Los Angeles County. We conducted a cross-sectional study using routinely collected surveillance data from publicly funded HIV clinics. We examined differences in INH treatment initiation and completion between four clinic categories: the three largest clinics (Clinics A, B, and C) and 'Other' clinics (pooled data for the remaining 10 clinics). During 2010-2013, 802 (5.3%) of 15 029 HIV-1-infected persons tested positive for M. tuberculosis infection. INH was initiated in 581 (72.4%) persons, of whom 457 (78.7%) completed treatment. We found significant differences between clinics in terms of treatment initiation (range 59.1-93.4%) and completion (range 58.8-82.3%). Overall, 57% (457/802) of HIV and M. tuberculosis co-infected persons completed the recommended treatment (range across clinics 34.8-76.3%). We identified significant gaps in the treatment for M. tuberculosis infection among HIV-infected persons in Los Angeles County. Interventions are needed to improve initiation and completion of treatment for M. tuberculosis infection in this population.

Baeyer-Villiger Monooxygenases EthA and MymA Are Required for Activation of Replicating and Non-replicating Mycobacterium tuberculosis Inhibitors

Successful treatment of Mycobacterium tuberculosis infection typically requires a complex regimen administered over at least 6months. Interestingly, many of the antibiotics used to treat M.tuberculosis are prodrugs that require intracellular activation. Here, we describe three small molecules, active against both replicating and non-replicating M.tuberculosis, that require activation by Baeyer-Villiger monooxygenases (BVMOs). Two molecules require BVMO EthA (Rv3854c) for activation and the third molecule requires the BVMO MymA (Rv3083). While EthA is known to activate the antitubercular drug ethionamide, this is the first description of MymA as an activating enzyme of a prodrug. Furthermore, we found that MymA also plays a role in activating ethionamide, with loss of MymA function resulting in ethionamide-resistant M.tuberculosis. These findings suggest overlap in function and specificity of the BVMOs in M.tuberculosis.

Development of dual inhibitors targeting DprE1 and AHAS for treatment of Mycobacterium tuberculosis infection

Background The emerging multidrug resistant (MDR) and extensively drug resistant (XDR) Mycobacterium tuberculosis (MTB) infection is increasing with greater complexity, estimated 220000-400000 tuberculosis cases emerged in 2011 globally. A number of lead compounds have been developed for treatment of MDR and XDR TB, but no new chemical entity has emerged for clinical use. Recently DprE1 and AHAS have been identified as promising drug targets.

Characterization and classification of Bo4 as a cluster G mycobacteriophage that can infect and lyse M. tuberculosis

Mycobacteriophage therapy is a potential alternative treatment for Mycobacterium tuberculosis infection. Here, we further characterized a mycobacteriophage, Bo4, and evaluated its ability to infect and kill M. tuberculosis. We first found that Bo4 can infect M. tuberculosis and Mycobacterium smegmatis. The observed clear plaques created by Bo4 infection indicated that Bo4 might be a lytic phage able to lyse mycobacterial strains, which was confirmed by phage antimicrobial activity. Bo4 formed clear zones in a medium with pH values of 7.4 or 5.0, suggesting the possibility that Bo4 could lyse mycobacteria, such as M. tuberculosis, in blood as well as in lysosomal macrophages. Further investigation into the Bo4 genome revealed that Bo4 had a dsDNA genome. Moreover, Bo4 contained ~39,318bp comprised of 66.76% G+C content. Complete genome sequencing showed high nucleotide identity with cluster G mycobacteriophages, thus classifying Bo4 as a member of the cluster G family. Additionally, annotation of the Bo4 genome indicated that it was a lytic bacteriophage and did not contain any harmful genes that increased mycobacterial virulence or decreased human immunity. Overall, the results of investigation indicate that the Bo4 possesses the potential to destroy M. tuberculosis, making it a potentially useful tool for diagnosing and treating tuberculosis.

Novel Tuberculostatic Agents Suitable for Treatment of Mycobacterium tuberculosis Infections of the Central Nervous System

Aims: To demonstrate the efficacy of five small molecule compounds for inhibiting the growth of Mycobacterium tuberculosis. To present evidence that these compounds will penetrate into the central nervous system.Study Design: Five small molecule compounds bearing a hydrazide group were synthesized utilizing microwave excitation. These compounds were then placed into tissue culture with Mycobacterium tuberculosis at various concentrations for evaluation of bacterial growth inhibition.Place and Duration of Study: The compounds to be tested were prepared at the University of Nebraska Chemistry Department August 2013. The evaluation of antibacterial activity was determined at the Texas A&M Health Science Center during October to December of 2013.Methodology: Applying microwave excitation for generation of hydrazide groups within the structure of small molecule carboxylic acids, five agents were prepared for evaluation of bacterial growth inhibition. These agents were dissolved into tissue culture media at various concentrations. Having various levels of tuberculostatic agents, then tuberculosis bacteria were added to determine level of growth inhibition. Growth inhibition of the bacteria was achieved and measured by compound concentration for comparison and evaluation.Results: Five compounds having a hydrazide functional group greatly inhibited the growth of Mycobacterium tuberculosis. All five agents had molecular weight less than 215 grams/mole and polar surface area of less than 70 Angstroms2. Values of Log P ranged from -0.226 to 0.998. Values of Log BB (Log [Cbrain/Cblood]) ranged from -0.711 to - 0.525, with a range in central nervous system penetration Cbrain/Cblood of 0.195 to 0.299. All compounds showed zero violations of the Rule of 5. Substantial inhibition of bacterial growth was observed at concentrations as low as 30 micrograms/mL, as measured by optical density and colony forming units.Conclusion: These five hydrazide compounds substantially decreased the proliferation of tuberculosis bacteria at concentrations as low as 30 micrograms/mL. In addition, their physicochemical properties are shown to allow high levels of penetration into the central nervous system.

Resistome analysis of Mycobacterium tuberculosis: Identification of aminoglycoside 2'-Nacetyltransferase (AAC) as co-target for drug desigining.

The emergence of multidrug resistant tuberculosis (MDRTB) highlights the urgent need to understand the mechanisms of resistance to the drugs and to develop a new arena of therapeutics to treat the disease. Ethambutol, isonazid, pyrazinamide, rifampicin are first line of drugs against TB, whereas aminoglycoside, polypeptides, fluoroquinolone, ethionamide are important second line of bactericidal drugs used to treat MDRTB, and resistance to one or both of these drugs are defining characteristic of extensively drug resistant TB. We retrieved 1,221 resistant genes from Antibiotic Resistance Gene Database (ARDB), which are responsible for resistance against first and second line antibiotics used in treatment of Mycobacterium tuberculosis infection. From network analysis of these resistance genes, 53 genes were found to be common. Phylogenetic analysis shows that more than 60% of these genes code for acetyltransferase. Acetyltransferases detoxify antibiotics by acetylation, this mechanism plays central role in antibiotic resistance. Seven acetyltransferase (AT-1 to AT-7) were selected from phylogenetic analysis. Structural alignment shows that these acetyltransferases share common ancestral core, which can be used as a template for structure based drug designing. From STRING analysis it is found that acetyltransferase interact with 10 different proteins and it shows that, all these interaction were specific to M. tuberculosis. These results have important implications in designing new therapeutic strategies with acetyltransferase as lead co-target to combat against MDR as well as Extreme drug resistant (XDR) tuberculosis.AbbreviationsAA - amino acid, AT - Acetyltransferase, AAC - Aminoglycoside 2'-N-acetyltransferase, XDR - Extreme drug-resistant, MDR - Multidrug-resistant, Mtb - Mycobacterium tuberculosis, TB - Tuberculosis.

Hair-analysis for acetyl-isoniazid/isoniazid ratio and N-acetyl-transferase-2-genotype in patients on treatment for Mycobacterium tuberculosis infection

Hair-analysis for acetyl-isoniazid/isoniazid ratio and N-acetyl-transferase-2-genotype in patients on treatment for Mycobacterium tuberculosis infection

Fighting Infection with Liposomes

Asymmetric engineered liposomes are antimicrobial and anti-inflammatory.

Renewed focus on tuberculosis holds promise for new treatments

FDA approved rifampin for the treatment of tuberculosis (TB) in 1971, the same year Intel Corporation invented the first microprocessor. Since then, advances in computer technology have revolutionized the way people live and work. But no additional medications have been approved by FDA for the treatment of Mycobacterium tuberculosis infection—a problem that federal officials and others want to fix. “It can’t take 40 years for us to get our next drug candidates for tuberculosis developed,” said Janet Woodcock, chief of FDA’s Center for Drug Evaluation and Research, during a March 19 briefing in Washington, D.C., sponsored by the Critical Path to TB Drug Regimens partnership. Launched in 2010, the partnership was founded by the Bill and Melinda Gates Foundation, the Critical Path Institute, and the Global Alliance for TB Drug Development, also known as the TB Alliance. The partnership promotes innovative ways to speed the development of new TB drugs.

Identification of putative biomarkers for the serodiagnosis of drug-resistant Mycobacterium tuberculosis

BackgroundEarly diagnosis and treatment of Mycobacterium tuberculosis infection can prevent most deaths resulting from this pathogen; however, multidrug-resistant strains present serious threats to global tuberculosis control and prevention efforts. In this study, we identified antigens that could be used for the serodiagnosis of drug-resistant M. tuberculosis strains, using a proteomics-based analysis.ResultsSerum from patients infected with drug-resistant or drug-susceptible M. tuberculosis strains and healthy controls was subjected to two-dimensional gel electrophoresis using a western blot approach. This procedure identified nine immunoreactive proteins, which were subjected to MALDI-TOF-MS analysis. Six recombinant proteins, namely rRv2031c, rRv0444c, rRv2145c, rRv3692, rRv0859c, and rRv3040, were expressed and used to determine the immuno-reactivity of 100 serum samples. Antibody reactivity against rRv2031c, rRv3692, and rRv0444c was consistently observed. Among them, the best sensitivity and specificity of rRv3692 were 37% and 95% respectively. Furthermore, when rRv2031c and rRv3692 or rRv2031c, rRv3692, and rRv0444c were combined in 2:1 or equal amounts, the assay sensitivity and specificity were improved to 56.7% and 100% respectively.ConclusionsThese results suggest that Rv2031c, Rv3692, and Rv0444c are possible candidate biomarkers for effective use in the serodiagnosis of drug-resistant tuberculosis infections, and a combined formula of these antigens should be considered when designing a subunit assay kit.

Molecular Identification of <em>Mycobacterium tuberculosis</em> and Analysis of Its Resistance to Rifampin in Sputa From Tuberculosis Suspected Patients

An accurate identification of different species of Mycobacterium provides to allow appropriate treatment for Mycobacterium tuberculosis infection. Beside that, drug resistance of M. tuberculosis strains to rifampin is not clearly understood in contributing to the spread of tuberculosis in Indonesia. To assess the molecular mechanism of rifampin resistance, a number of clinical specimens of M. tuberculosis were analyzed their molecular nature of a part of the rpoB gene using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) methods. DNA’s extracted from sputum samples were amplified and 32 P-labeled by PCR with the specific primers and the product was analyzed their mutation conferring resistance by MDE gel electrophoresis. Of the 70 specimens tested, 57 specimens were positive for M. tuberculos is organism only, three specimens contained a mixture of M. tuberculosis and non tuberculosis mycobacteria (NTM), and 10 specimens were negative approved by Duplex PCR. Of these sixty DNA positive samples (thus the sensitivity of PCR was 85.71%), 5 (8.3%) of them suspected to contain mutations in rpoB which were associated with rifampin resistance. Even though the frequency of mutation was low, the results from our study clearly indicate that the molecular mechanism of rifampin resistance in M. tuberculosis isolates from Indonesia involves alterations in the rpoB gene. Molecular diagnosis by PCR which is fast and easy to perform is useful for early and rapid detection of TB in sputum specimen. Received: 27 February 2009; Revised: 28 August 2010; Accepted: 30 August 2010

Interleukin 24 as a novel potential cytokine immunotherapy for the treatment of Mycobacterium tuberculosis infection

Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) remains a major global health problem. Interleukin 24 (IL-24) is a novel tumor suppressor and a unique member of the IL-10 family of cytokines. However, the in vivo immunological consequences of this cytokine's activity during Mtb infection are still unknown. We found that IL-24 concentration was significantly decreased in the sera of TB patients, and Mtb infection suppressed IL-24 expression of human peripheral blood mononuclear cells (PBMCs) in vitro. Furthermore, we used a mouse infection model utilizing the virulent Mtb H37Rv strain to demonstrate that the administration of exogenous IL-24 had a protective effect against the bacteria. We found that IL-24 could activate human CD8 + T cells, driving CD8 + T cells to produce interferon-γ (IFN-γ) and counteract TB. This activity was found to be dependent on early involvement of neutrophils in the mouse model. IL-24 strongly stimulated IFN-γ production mainly by signaling through the IL-24 receptors of human CD8 + T cells. IL-12 secretion from neutrophils in response to IL-24 treatment might be a minor factor in activating human CD8 + T cells to secrete IFN-γ. Suppression of IL-24 expression by Mtb infection might enhance susceptibility to infection and promote the development of chronic TB. This new information could potentially stimulate the development of a new cytokine-based immunotherapeutic approach using IL-24 to stimulate immunity against TB.

Properties and Potency of Small Molecule Agents for Treatment of Mycobacterium Tuberculosis Infections of the Central Nervous System

Tuberculous meningitis is caused by Mycobacterium tuberculosis, which is the bacteria that causes tuberculosis. This bacteria spreads to the brain from another site within the body. If untreated, tuberculous meningitis can be life-threatening. Substantial brain damage can result from infection which results in mental impairment, motor paralysis, seizures, and abnormal behavior. This study presents the analysis of the effectiveness of 13 novel tuberculostatic agents, along with isoniazid (a first-line drug utilized for treatment of Mycobacterium tuberculosis). All 13 agents retain a diverse character of molecular structure effectuating a range of molecular properties such as in polar surface area, Log P, and formula weight. Properties such as polar surface area, formula weight, and Log P were determined for these agents as well as values of BB (Cbrain/Cblood), and Log BB. Values of BB varied from 0.0681 to 1.16. Only three of the novel drug structures showed one violation of the Rule of 5, while the remaining showed zero violations (an outcome favoring bioavailability). Values of polar surface area ranged from 55.121 Angstroms(2) to 110.24 Angstroms(2), and Log P from -3.52 to 4.965. This group of novel tuberculostatic agents reveals examples of three homologous series of tuberculostatic drugs which purposes the likelihood of as many as 20 effective tuberculostatic drugs. Determination of MIC60 disclosed eight novel structures matching the effectiveness of isoniazid, while two showed superior ability to deter Mycobacterium tuberculosis proliferation. These novel drug designs are shown to have substantial potential for the beneficial treatment of patients having infections of Mycobacterium tuberculosis within the central nervous system.

A rapid approach to lipid profiling of mycobacteria using 2D HSQC NMR maps

Mycobacteria, including Mycobacterium tuberculosis, are characterized by a unique cell wall rich in complex lipids, glycolipids, polyketides, and terpenoids. Many of these metabolites have been shown to play important roles in mycobacterial virulence and their inherent resistance to many antibiotics. Here, we report the development of a new simple method for global analysis of these metabolites using two-dimensional (1)H-(13)C heteronuclear single quantum coherence nuclear magnetic resonance. The major advantages of this method are as follows: the small amount of sample and the minimal sample manipulation required; a relatively short procedural time; and the ability to rapidly attain a qualitative and quantitative lipid profile of a mycobacterial sample in which the majority of the clinically relevant lipids can be observed simultaneously. The effectiveness of this method is demonstrated in four different areas of major concern to the mycobacterial research community: i) adaptive changes in cell wall lipids as a result of drug treatment; ii) analysis of gene function; iii) characterization of new mycobacterial species; and iv) analysis of the production of virulence factors in clinical isolates of M. tuberculosis. This method is complementary to mass spectrometry-based lipidomic technologies and provides an urgently needed tool to gain a better understanding of the role of lipids in mycobacteria pathogenesis.

The Mechanism of Isoniazid Killing: Clarity Through the Scope of Genetics

Isoniazid (INH) is one of the most efficient drugs for the treatment of Mycobacterium tuberculosis infections. Despite its rather simple chemical structure, the mechanism by which INH kills M. tuberculosis is complex. A full understanding of the mechanisms of action of INH required the development of genetic tools in M. tuberculosis. Herein, we discuss the different hypotheses that have been used to describe INH action against M. tuberculosis over the past 50 years in terms of the pregenetic and genetic era. We also review the different mechanisms of INH resistance and propose what we think is the means by which INH kills M. tuberculosis.

Irreversible Inhibition of the Mycobacterium tuberculosis β-Lactamase by Clavulanate

Members of the beta-lactam class of antibiotics, which inhibit the bacterial d,d-transpeptidases involved in cell wall biosynthesis, have never been used systematically in the treatment of Mycobacterium tuberculosis infections because of this organism's resistance to beta-lactams. The critical resistance factor is the constitutive production of a chromosomally encoded, Ambler class A beta-lactamase, BlaC in M. tuberculosis. We show that BlaC is an extended spectrum beta-lactamase (ESBL) with high levels of penicillinase and cephalosporinase activity as well as measurable activity with carbapenems, including imipenem and meropenem. We have characterized the enzyme's inhibition by three FDA-approved beta-lactamase inhibitors: sulbactam, tazobactam, and clavulanate. Sulbactam inhibits the enzyme competitively and reversibly with respect to nitrocefin. Tazobactam inhibits the enzyme in a time-dependent manner, but the activity of the enzyme reappears due to the slow hydrolysis of the covalently acylated enzyme. In contrast, clavulanate reacts with the enzyme quickly to form hydrolytically stable, inactive forms of the enzyme that have been characterized by mass spectrometry. Clavulanate has potential to be used in combination with approved beta-lactam antibiotics to treat multi-drug resistant (MDR) and extremely drug resistant (XDR) strains of M. tuberculosis.

Characterization of pncA mutations of pyrazinamide-resistant Mycobacterium tuberculosis in Korea.

Pyrazinamide (PZA) is one of the most important drugs for the treatment of Mycobacterium tuberculosis infection. However, the increasing frequency of PZA-resistant strains limits its effectiveness. In Korea, most PZA-resistant strains also exhibit both isoniazid and rifampin resistance making it essential to identify these resistant strains accurately and rapidly for effective treatment of mycobacterial infection. In this study, the characteristics and frequency of mutations of the pncA gene encoding pyrazinamidase were investigated in PZA-resistant clinical isolates from Korea. Automated DNA sequencing was used to evaluate the usefulness of DNA-based detection of PZA resistance. Among 95 PZA-resistant clinical isolates, 92 (97%) exhibited mutations potentially affecting either the production or the activity of the enzyme. Mutations were found throughout the pncA gene including the upstream region. Single nucleotide replacement appeared to be the major mutational event (69/92), although multiple substitutions as well as insertion and deletion of nucleotides were also identified. The high frequency of pncA mutations observed in this study supports the usefulness of DNA-based detection of PZA-resistant M. tuberculosis. Having verified the scattered and diverse mutational characteristics of the pncA gene, automated DNA sequencing seems to be the best strategy for rapid detection of PZA-resistant M. tuberculosis.

Compensatory ahpC gene expression in isoniazid-resistant Mycobacterium tuberculosis.

Mutations that eliminate KatG catalase-peroxidase activity prevent activation of isoniazid and are a major mechanism of resistance to this principal drug for the treatment of Mycobacterium tuberculosis infections. However, the loss of KatG activity in clinical isolates seemed paradoxical because KatG is considered an important factor for the survival of the organism. Expression of either KatG or the recently identified alkyl hydroperoxidase AhpC was sufficient to protect bacilli against the toxic effects of organic peroxides. To survive during infection, isoniazid-resistant KatG mutants have apparently compensated for the loss of KatG catalase-peroxidase activity by a second mutation, resulting in hyperexpression of AhpC.

Identification, cloning, and expression of the Escherichia coli pyrazinamidase and nicotinamidase gene, pncA

Pyrazinamide (PZA) is one of the three most important drugs for treatment of Mycobacterium tuberculosis infections. The antibacterial activity of PZA requires a bacterial enzyme, pyrazinamidase (PZAase), which hydrolyzes PZA to form pyrazinoic acid and ammonia. Most PZA-resistant clinical M. tuberculosis isolates lack PZAase activity. With the goal of eventually identifying and characterizing the M.tuberculosis PZAase gene, we began with the more tractable organism, Escherichia coli, which also has PZAase activity. We screened a transposon-generated E. coli insertion mutant library, using a qualitative PZAase assay. Two PZAase-negative mutants out of 4,000 colonies screened were identified. In each mutant, the transposon interrupted the same 639-bp open reading frame (ORF), ORF1. The expression of ORF1 on a multicopy plasmid complemented a PZAase-negative mutant, leading to PZAase activity levels approximately 10-fold greater than those of the wild type. PZA has a structure similar to that of nicotinamide, a pyridine nucleotide cycle intermediate, so we tested our strains for nicotinamidase activity (EC 3.5.1.19) (genetic locus pncA). The construct with multiple plasmid copies of ORF1 had an approximately 10-fold increase in levels of nicotinamidase activity. This overexpressing strain could utilize nicotinamide as a sole nitrogen source, through wild-type E. coli cannot. We conclude that a single E. coli enzyme accounts for both PZAase and nicotinamidase activities and that ORF1 is the E.coli PZAase and nicotinamidase gene, pncA.

Tuberculosis in the AIDS era

A resurgence of tuberculosis has occurred in recent years in the United States and abroad. Deteriorating public health services, increasing numbers of immigrants from countries of endemicity, and coinfection with the human immunodeficiency virus (HIV) have contributed to the rise in the number of cases diagnosed in the United States. Outbreaks of resistant tuberculosis, which responds poorly to therapy, have occurred in hospitals and other settings, affecting patients and health care workers. This review covers the pathogenesis, epidemiology, clinical presentation, laboratory diagnosis, and treatment of Mycobacterium tuberculosis infection and disease. In addition, public health and hospital infection control strategies are detailed. Newer approaches to epidemiologic investigation, including use of restriction fragment length polymorphism analysis, are discussed. Detailed consideration of the interaction between HIV infection and tuberculosis is given. We also review the latest techniques in laboratory evaluation, including the radiometric culture system, DNA probes, and PCR. Current recommendations for therapy of tuberculosis, including multidrug-resistant tuberculosis, are given. Finally, the special problem of prophylaxis of persons exposed to multidrug-resistant tuberculosis is considered.