Maintenance of cell wall remodeling and vesicle production are connected in .

Pathogenic and nonpathogenic mycobacteria secrete extracellular vesicles (EVs) under various conditions. EVs produced by ( ) have raised significant interest for their potential in cell communication, nutrient acquisition, and immune evasion. However, the relevance of vesicle secretion during tuberculosis infection remains unknown due to the limited understanding of mycobacterial vesicle biogenesis.

A novel chemogenomic discovery platform identifies bioactive hits with rapid bactericidal activity against Mycobacteroides Abscessus.

Mycobacteroides abscessus (M. ab) infections are innately resistant to most currently available antibiotics and present a growing, poorly addressed medical need. The existing treatment regimens are lengthy and produce inadequate outcomes for many patients.

Feasibility of novel approaches to detect viable within the spectrum of the tuberculosis disease.

Tuberculosis (TB) is a global disease caused by () and is manifested as a continuum spectrum of infectious states. Both, the most common and clinically asymptomatic latent tuberculosis infection (LTBI), and the symptomatic disease, active tuberculosis (TB), are at opposite ends of the spectrum. Such binary classification is insufficient to describe the existing clinical heterogeneity, which includes incipient and subclinical TB.

Multiple genetic paths including massive gene amplification allow to overcome loss of ESX-3 secretion system substrates.

() possesses five type VII secretion systems (T7SS), virulence determinants that include the secretion apparatus and associated secretion substrates. strains deleted for the genes encoding substrates of the ESX-3 T7SS, or , require iron supplementation for in vitro growth and are highly attenuated in vivo. In a subset of infected mice, suppressor mutants of or deletions were isolated, which enabled growth to high titers or restored virulence.

Drivers and sites of diversity in the DNA adenine methylomes of 93 complex clinical isolates.

This study assembles DNA adenine methylomes for 93 complex (MTBC) isolates from seven lineages paired with fully-annotated, finished, de novo assembled genomes. Integrative analysis yielded four key results. First, methyltransferase allele-methylome mapping corrected methyltransferase variant effects previously obscured by reference-based variant calling.

Characterization of Large Deletion Mutants of Mycobacterium tuberculosis Selected for Isoniazid Resistance.

Large genomic deletions (LGDs) (6 to 63 kbp) were observed in isoniazid-resistant mutants derived from four strains. These LGDs had no growth defect but could be defective in intracellular growth and showed various sensitivities toward oxidative stress despite lacking The LGD regions comprise 74 genes, mostly of unknown function, that may be important for intracellular growth and protection against oxidative stress.

Genotyping of Mycobacterium tuberculosis Rifampin Resistance-Associated Mutations by Use of Data from Xpert MTB/RIF Ultra Enables Large-Scale Tuberculosis Molecular Epidemiology Studies.

Molecular epidemiology studies of tuberculosis have been empowered in recent years by the availability of whole-genome sequencing, which has allowed a new focus on the adaptive significance of drug resistance mutations. Genome sequencing technology remains expensive, however, limiting the potential for larger studies. Conversely, during this same time the GeneXpert molecular diagnostic method has been deployed globally and now serves as a cornerstone of tuberculosis diagnosis and drug sensitivity testing.

Arginine-deprivation-induced oxidative damage sterilizes .

Reactive oxygen species (ROS)-mediated oxidative stress and DNA damage have recently been recognized as contributing to the efficacy of most bactericidal antibiotics, irrespective of their primary macromolecular targets. Inhibitors of targets involved in both combating oxidative stress as well as being required for in vivo survival may exhibit powerful synergistic action. This study demonstrates that the de novo arginine biosynthetic pathway in () is up-regulated in the early response to the oxidative stress-elevating agent isoniazid or vitamin C.

Vaginal microbiome modulates topical antiretroviral drug pharmacokinetics.

Tenofovir gel and dapivirine ring provided variable HIV protection in clinical trials, reflecting poor adherence and possibly biological factors. We hypothesized that vaginal microbiota modulates pharmacokinetics and tested the effects of pH, individual bacteria, and vaginal swabs from women on pharmacokinetics and antiviral activity. Tenofovir, but not dapivirine, uptake by human cells was reduced as pH increased.

Rational Design of Biosafety Level 2-Approved, Multidrug-Resistant Strains of Mycobacterium tuberculosis through Nutrient Auxotrophy.

Multidrug-resistant (MDR) tuberculosis, defined as tuberculosis resistant to the two first-line drugs isoniazid and rifampin, poses a serious problem for global tuberculosis control strategies. Lack of a safe and convenient model organism hampers progress in combating the spread of MDR strains of We reasoned that auxotrophic MDR mutants of would provide a safe means for studying MDR without the need for a biosafety level 3 (BSL3) laboratory. Two different sets of triple auxotrophic mutants of were generated, which were auxotrophic for the nutrients leucine, pantothenate, and arginine or for leucine, pantothenate, and methionine.

Plasticity of NADH dehydrogenases and their role in virulence.

Worldwide control of the tuberculosis (TB) epidemic has not been achieved, and the latest statistics show that the TB problem might be more endemic than previously thought. Although drugs and a TB vaccine are available, TB eradication faces the challenges of increasing occurrences of multidrug-resistant and extensively drug-resistant () strains. To forestall this trend, the development of drugs targeting novel pathways is actively pursued.

Enhanced respiration prevents drug tolerance and drug resistance in .

Persistence, manifested as drug tolerance, represents a significant obstacle to global tuberculosis control. The bactericidal drugs isoniazid and rifampicin kill greater than 99% of exponentially growing () cells, but the remaining cells are persisters, cells with decreased metabolic rate, refractory to killing by these drugs, and able to generate drug-resistant mutants. We discovered that the combination of cysteine or other small thiols with either isoniazid or rifampicin prevents the formation of drug-tolerant and drug-resistant cells in cultures.

Enhanced control of Mycobacterium tuberculosis extrapulmonary dissemination in mice by an arabinomannan-protein conjugate vaccine.

Currently there are a dozen or so of new vaccine candidates in clinical trials for prevention of tuberculosis (TB) and each formulation attempts to elicit protection by enhancement of cell-mediated immunity (CMI). In contrast, most approved vaccines against other bacterial pathogens are believed to mediate protection by eliciting antibody responses. However, it has been difficult to apply this formula to TB because of the difficulty in reliably eliciting protective antibodies.

Dual-Reporter Mycobacteriophages (Φ2DRMs) Reveal Preexisting Mycobacterium tuberculosis Persistent Cells in Human Sputum.

Unlabelled: Persisters are the minor subpopulation of bacterial cells that lack alleles conferring resistance to a specific bactericidal antibiotic but can survive otherwise lethal concentrations of that antibiotic. In infections with Mycobacterium tuberculosis, such persisters underlie the need for long-term antibiotic therapy and contribute to treatment failure in tuberculosis cases. Here, we demonstrate the value of dual-reporter mycobacteriophages (ΦDRMs) for characterizing M.

HSV-2 ΔgD elicits FcγR-effector antibodies that protect against clinical isolates.

A single-cycle herpes simplex virus (HSV) deleted in glycoprotein D (ΔgD-2) elicited high titer HSV-specific antibodies (Abs) that (i) were rapidly transported into the vaginal mucosa; (ii) elicited antibody-dependent cell-mediated cytotoxicity but little neutralization; (iii) provided complete protection against lethal intravaginal challenge; and (iv) prevented establishment of latency in mice. However, clinical isolates may differ antigenically and impact vaccine efficacy. To determine the breadth and further define mechanisms of protection of this vaccine candidate, we tested ΔgD-2 against a panel of clinical isolates in a murine skin challenge model.

The Type of Growth Medium Affects the Presence of a Mycobacterial Capsule and Is Associated With Differences in Protective Efficacy of BCG Vaccination Against Mycobacterium tuberculosis.

Background: Bacillus Calmette-Guerin (BCG) vaccine is widely used for the prevention of tuberculosis, despite limited efficacy. Most immunological studies of BCG or Mycobacterium tuberculosis strains grow bacteria in the presence of detergent, which also strips the mycobacterial capsule. The impact of the capsule on vaccine efficacy has not been explored.

Structural characterization of muropeptides from Chlamydia trachomatis peptidoglycan by mass spectrometry resolves "chlamydial anomaly".

The "chlamydial anomaly," first coined by James Moulder, describes the inability of researchers to detect or purify peptidoglycan (PG) from pathogenic Chlamydiae despite genetic and biochemical evidence and antibiotic susceptibility data that suggest its existence. We recently detected PG in Chlamydia trachomatis by a new metabolic cell wall labeling method, however efforts to purify PG from pathogenic Chlamydiae have remained unsuccessful. Pathogenic chlamydial species are known to activate nucleotide-binding oligomerization domain-containing protein 2 (NOD2) innate immune receptors by as yet uncharacterized ligands, which are presumed to be PG fragments (muramyl di- and tripeptides).

Succinate dehydrogenase is the regulator of respiration in Mycobacterium tuberculosis.

In chronic infection, Mycobacterium tuberculosis bacilli are thought to enter a metabolic program that provides sufficient energy for maintenance of the protonmotive force, but is insufficient to meet the demands of cellular growth. We sought to understand this metabolic downshift genetically by targeting succinate dehydrogenase, the enzyme which couples the growth processes controlled by the TCA cycle with the energy production resulting from the electron transport chain. M.

The p60 and NamA autolysins from Listeria monocytogenes contribute to host colonization and induction of protective memory.

Inducing long-term protective memory CD8(+) T-cells is a desirable goal for vaccines against intracellular pathogens. However, the mechanisms of differentiation of CD8(+) T-cells into long-lived memory cells capable of mediating protection of immunized hosts remain incompletely understood. We have developed an experimental system using mice immunized with wild type (WT) or mutants of the intracellular bacterium Listeria monocytogenes (Lm) that either do or do not develop protective memory CD8(+) T-cells.

Phosphorylation of KasB regulates virulence and acid-fastness in Mycobacterium tuberculosis.

Mycobacterium tuberculosis bacilli display two signature features: acid-fast staining and the capacity to induce long-term latent infections in humans. However, the mechanisms governing these two important processes remain largely unknown. Ser/Thr phosphorylation has recently emerged as an important regulatory mechanism allowing mycobacteria to adapt their cell wall structure/composition in response to their environment.

Role for Mycobacterium tuberculosis membrane vesicles in iron acquisition.

Mycobacterium tuberculosis releases membrane vesicles packed with molecules that can modulate the immune response. Because environmental conditions often influence the production and content of bacterial vesicles, this study examined M. tuberculosis microvesicles released under iron limitation, a common condition faced by pathogens inside the host.

Identification of a small molecule with activity against drug-resistant and persistent tuberculosis.

A cell-based phenotypic screen for inhibitors of biofilm formation in mycobacteria identified the small molecule TCA1, which has bactericidal activity against both drug-susceptible and -resistant Mycobacterium tuberculosis (Mtb) and sterilizes Mtb in vitro combined with rifampicin or isoniazid. In addition, TCA1 has bactericidal activity against nonreplicating Mtb in vitro and is efficacious in acute and chronic Mtb infection mouse models both alone and combined with rifampicin or isoniazid. Transcriptional analysis revealed that TCA1 down-regulates genes known to be involved in Mtb persistence.

Mycobacterium tuberculosis is extraordinarily sensitive to killing by a vitamin C-induced Fenton reaction.

Drugs that kill tuberculosis more quickly could shorten chemotherapy significantly. In Escherichia coli, a common mechanism of cell death by bactericidal antibiotics involves the generation of highly reactive hydroxyl radicals via the Fenton reaction. Here we show that vitamin C, a compound known to drive the Fenton reaction, sterilizes cultures of drug-susceptible and drug-resistant Mycobacterium tuberculosis, the causative agent of tuberculosis.

Keto-mycolic acid-dependent pellicle formation confers tolerance to drug-sensitive Mycobacterium tuberculosis.

ABSTRACT The chronic nature of tuberculosis (TB), its requirement of long duration of treatment, its ability to evade immune intervention, and its propensity to relapse after drug treatment is discontinued are reminiscent of other chronic, biofilm-associated bacterial diseases. Historically, Mycobacterium tuberculosis was grown as a pellicle, a biofilm-like structure, at the liquid-air interface in a variety of synthetic media. Notably, the most widely administered human vaccine, BCG, is grown as a pellicle for vaccine production.