Pyrazinamide Is Active against Mycobacterium tuberculosis Cultures at Neutral pH and Low Temperature.

For the past decades, an acidic pH has been used to render Mycobacterium tuberculosis susceptible to pyrazinamide for in vitro testing. Here, we show that at the standard breakpoint concentration and reduced culture temperatures, pyrazinamide (PZA) is active against tuberculosis (TB) at neutral pH. This finding should help unravel the mechanism of action of PZA and allow drug susceptibility testing (DST) methods to be optimized.

Cytokine Kinetics in the First Week of Tuberculosis Therapy as a Tool to Confirm a Clinical Diagnosis and Guide Therapy.

Background: Many patients treated for tuberculosis (TB) in low and middle income countries are treated based on clinical suspicion without bacteriological confirmation. This is often due to lack of rapid simple accurate diagnostics and low healthcare provider confidence in the predictive value of current tests. We previously reported in an animal TB model that levels of host markers rapidly change in response to treatment initiation.

Pyrazinamide resistance in Mycobacterium tuberculosis fails to bite?

In contrast to most other antimycobacterial drugs where--particularly in multidrug-resistant (MDR) strains--a limited number of resistance mutations dominate, pyrazinamide (PZA) resistance associated mutations remain highly diverse with limited clustering. This apparent lack of evolutionary selection for successful PZA resistance mechanisms deserves attention. A clear understanding of the epidemiology of PZA resistance acquisition and spread would be expected to result in important insights into how PZA might be better exploited in treatment regimens to minimize the amplification of Mycobacterium tuberculosis (MTB) drug resistance.

Evaluation of a microcolony growth monitoring method for the rapid determination of ethambutol resistance in Mycobacterium tuberculosis.

Background: Due to the increasing prevalence of Mycobacterium tuberculosis strains resistant to one or more antibiotics, there is a need for new quantitative culture methods both for drug susceptibility testing and for validation of mutations putatively associated with drug resistance. We previously developed a (myco) bacterial culture method, in which multiple growing microcolonies are monitored individually. Transfer of the growing microcolonies to selective medium allows the effect on the growth rate of each individual colony to be determined.

Early specific host response associated with starting effective tuberculosis treatment in an infection controlled placebo controlled mouse study.

Recently we proposed exploring the potential of treatment stimulated testing as diagnostic method for tuberculosis (TB). An infection controlled placebo controlled mouse study was performed to investigate whether serum cytokine levels changed measurably during the early phase of TB chemotherapy. Serum was collected prior to and during the first 3 weeks of isoniazid (INH) and rifampicin (RIF) chemotherapy, and levels of 23 selected cytokines/chemokines were measured using a liquid bead array.

Simplified automated image analysis for detection and phenotyping of Mycobacterium tuberculosis on porous supports by monitoring growing microcolonies.

Background: Even with the advent of nucleic acid (NA) amplification technologies the culture of mycobacteria for diagnostic and other applications remains of critical importance. Notably microscopic observed drug susceptibility testing (MODS), as opposed to traditional culture on solid media or automated liquid culture, has shown potential to both speed up and increase the provision of mycobacterial culture in high burden settings.

Methods: Here we explore the growth of Mycobacterial tuberculosis microcolonies, imaged by automated digital microscopy, cultured on a porous aluminium oxide (PAO) supports.

Structure-function relationship of the human antimicrobial peptide LL-37 and LL-37 fragments in the modulation of TLR responses.

Cathelicidins are effector molecules of the innate host defense system that establish an antimicrobial barrier at epithelial interfaces. The human cathelicidin LL-37, in addition to its antimicrobial activity, also exhibits immunomodulatory effects, such as inhibition of pro-inflammatory responses to bacterial LPS in human monocytic cells. In this report, we demonstrate that LL-37 almost completely prevents the pro-inflammatory cytokine release by human peripheral blood mononuclear cells (PBMCs) following stimulation with Toll-like receptor (TLR)4 and TLR2/1 agonists while leaving TLR2/6, TLR5, TLR7 and TLR8 responses unchanged.

The human cathelicidin peptide LL-37 and truncated variants induce segregation of lipids and proteins in the plasma membrane of Candida albicans.

The human cathelicidin peptide LL-37 and several truncated variants differ in their capability to transmigrate over the plasma membrane of Candida albicans. We investigated whether retention at the cell perimeter or membrane transmigration affects their membrane-disrupting activities and candidacidal properties. Using fluorescein-labeled peptides, we demonstrate that LL-37 and its C-terminally truncated peptide LL-31 remain permanently associated with the perimeter of the cell.

Candidacidal effects of two antimicrobial peptides: histatin 5 causes small membrane defects, but LL-37 causes massive disruption of the cell membrane.

The effects of antimicrobial peptides on artificial membranes have been well-documented; however, reports on the ultrastructural effects on the membranes of micro-organisms are relatively scarce. We compared the effects of histatin 5 and LL-37, two antimicrobial peptides present in human saliva, on the functional and morphological properties of the Candida albicans cell membrane. Fluorescence microscopy and immunogold transmission electron microscopy revealed that LL-37 remained associated with the cell wall and cell membrane, whereas histatin 5 transmigrated over the membrane and accumulated intracellularly.

Reactive oxygen species play no role in the candidacidal activity of the salivary antimicrobial peptide histatin 5.

The mechanism of action of antimicrobial peptides is still a matter of debate. The formation of ROS (reactive oxygen species) has been suggested to be the crucial step in the fungicidal mechanism of a number of antimicrobial peptides, including histatin 5 and lactoferrin-derived peptides. In the present study we have investigated the effects of histatin 5 and of a more amphipathic synthetic derivative, dhvar4, on the generation of ROS in the yeast Candida albicans, using dihydroethidium as an indicator for ROS.

Interactions of histatin 5 and histatin 5-derived peptides with liposome membranes: surface effects, translocation and permeabilization.

A number of cationic antimicrobial peptides, among which are histatin 5 and the derived peptides dhvar4 and dhvar5, enter their target cells and interact with internal organelles. There still are questions about the mechanisms by which antimicrobial peptides translocate across the membrane. We used a liposome model to study membrane binding, translocation and membrane-perturbing capacities of histatin 5, dhvar4 and dhvar5.