Imaging the Human Immunological Synapse.

The purpose of the method is to generate an immunological synapse (IS), an example of cell-to-cell conjugation formed by an antigen-presenting cell (APC) and an effector helper T lymphocyte (Th) cell, and to record the images corresponding to the first stages of the IS formation and the subsequent trafficking events (occurring both in the APC and in the Th cell). These events will eventually lead to polarized secretion at the IS. In this protocol, Jurkat cells challenged with Staphylococcus enterotoxin E (SEE)-pulsed Raji cells as a cell synapse model was used, because of the closeness of this experimental system to the biological reality (Th cell-APC synaptic conjugates).

Bedaquiline and linezolid MIC distributions and epidemiological cut-off values for Mycobacterium tuberculosis in the Latin American region.

Objectives: To describe the distributions of bedaquiline and linezolid MIC values for the Mycobacterium tuberculosis WT population and to define the corresponding epidemiological cut-offs (ECOFFs) in three Latin American countries.

Methods: MICs of bedaquiline and linezolid were determined by the resazurin microtitre assay (REMA). In phase 1, interlaboratory reproducibility was assessed using a panel of 10 fully susceptible M.

String test: A new tool for tuberculosis diagnosis and drug-resistance detection in children.

Background: There is a critical need to improve the diagnostic accuracy of tuberculosis (TB) in children. Several techniques have been developed to improve the quality of sputum samples; however, these procedures are very unpleasant and invasive and require hospitalization and trained personnel. This study aims to explore the potential use of a new and noninvasive tool, "string test," for TB diagnosis in children and in adults not able to render sputum samples and at risk of developing multidrug-resistant TB (MDR-TB).

Antimicrobial activity against Mycobacterium tuberculosis under in vitro lipid-rich dormancy conditions.

Although tuberculosis treatment is dependent on drug-susceptibility testing (DST) and molecular drug-resistance detection, treatment failure and relapse remain a challenge. This could be partially due to the emergence of antibiotic-tolerant dormant mycobacteria, where host lipids have been shown to play an important role. This study evaluated the susceptibility of Mycobacterium tuberculosis to two antibiotic combinations - rifampicin, moxifloxacin, amikacin and metronidazole (RIF-MXF-AMK-MTZ), and rifampicin, moxifloxacin, amikacin and pretomanid (RIF-MXF-AMK-PA) - in a lipid-rich dormancy model.

Performance of a highly successful outbreak strain of Mycobacterium tuberculosis in a multifaceted approach to bacterial fitness assessment.

Determining bacterial fitness represents a major challenge and no single parameter can accurately predict the ability of a certain pathogen to succeed. The M strain of Mycobacterium tuberculosis managed to spread and establish in the community and caused the largest multidrug-resistant tuberculosis outbreak in Latin America. We have previously shown that the M strain can manipulate the host immune response, but we still have no direct evidence, other than epidemiology, that can account for the enhanced fitness of the M strain.

The transcriptome of Mycobacterium tuberculosis in a lipid-rich dormancy model through RNAseq analysis.

Tuberculosis (TB) is currently the number one killer among infectious diseases worldwide. Lipids are abundant molecules during the infectious cycle of Mycobacterium tuberculosis (Mtb) and studies better mimicking its actual metabolic state during pathogenesis are needed. Though most studies have focused on the mycobacterial lipid metabolism under standard culture conditions, little is known about the transcriptome of Mtb in a lipid environment.

In vitro activity of bedaquiline against rapidly growing nontuberculous mycobacteria.

Bedaquiline (BDQ) has been proven to be effective in the treatment of multidrug-resistant tuberculosis. We hypothesized that BDQ could be a potential agent to treat nontuberculous mycobacterial (NTM) infection. The objective of this study was to evaluate the in vitro activity of BDQ against rapidly growing mycobacteria by assessing the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) against 18 NTM strains.

Optimizing of a protein extraction method for Mycobacterium tuberculosis proteome analysis using mass spectrometry.

A critical step in proteomic analyses comprises the implementation of a reliable cell lysis method with high yields of qualitative proteins. In Mycobacteria, the protein extraction step is often hampered by the thick waxy cell wall which is rich in mycolic acids. Harsh disruption techniques to release proteins from the cells are thus required.

Exploring tuberculosis by molecular tests on DNA isolated from smear microscopy slides.

Tuberculosis (TB) is an infectious disease of global public health importance caused by Mycobacterium tuberculosis complex. The disease has worsened with the emergence of multidrug-resistant (MDR)-TB strains. The timely diagnosis and treatment of TB remains a key public health priority, and laboratories have a critical role in the rapid and accurate detection of TB and drug resistance.

"Genetic regulation of Mycobacterium tuberculosis in a lipid-rich environment".

Tuberculosis (TB) remains as one of the leading causes of morbidity and mortality among infectious diseases worldwide. Although lipids (mainly fatty acids and cholesterol) have been reported to play an important role during active and latent infection of M. tuberculosis, there are other molecular aspects of bacterial response to those substrates that are not fully understood, involving gene regulation background.

The potential role of trimethoprim-sulfamethoxazole in the treatment of drug-resistant tuberculosis.

Tuberculosis (TB) remains a serious public health threat worsened by emerging drug resistance. Mycobacterium tuberculosis has become resistant not only to front-line drugs but also to second-line antimicrobials directed at drug-resistant TB. Renewed efforts are devoted for the development of new antibiotics active against TB.

Resazurin microtiter assay for isoniazid, rifampicin, ethambutol and streptomycin resistance detection in Mycobacterium tuberculosis: Updated meta-analysis.

Aims: The present meta-analysis aims to assess the evidence regarding the diagnostic accuracy and performance characteristics of the colorimetric redox indicator (CRI) assay with a special emphasis on the use of the resazurin microtiter assay (REMA) for determination of primary anti-tuberculosis drug resistance.

Subject And Methods: By updating previous literature searches in Medline PubMed, ISI Web, Web of Science and Google academic databases of the REMA test for determination of primary anti-tuberculosis drug resistance, this meta-analysis includes 14 studies for isoniazid (INH); 15 studies for rifampicin (RIF); 6 studies for streptomycin (STR); and 5 studies for ethambutol (EMB). SROC curve analysis was performed for meta-analysis and diagnostic accuracy was summarized.

Performance of Four Transport and Storage Systems for Molecular Detection of Multidrug-Resistant Tuberculosis.

Background: Detection of drug-resistant tuberculosis is essential for the control of the disease but it is often hampered by the limitation of transport and storage of samples from remote locations to the reference laboratory. We performed a retrospective field study to evaluate the performance of four supports enabling the transport and storage of samples to be used for molecular detection of drug resistance using the GenoType MTBDRplus.

Methods: Two hundred Mycobacterium tuberculosis strains were selected and spotted on slides, FTA cards, GenoCards, and in ethanol.

Mycobacterium saopaulense sp. nov., a rapidly growing mycobacterium closely related to members of the Mycobacterium chelonae--Mycobacterium abscessus group.

Five isolates of non-pigmented, rapidly growing mycobacteria were isolated from three patients and,in an earlier study, from zebrafish. Phenotypic and molecular tests confirmed that these isolates belong to the Mycobacterium chelonae-Mycobacterium abscessus group, but they could not be confidently assigned to any known species of this group. Phenotypic analysis and biochemical tests were not helpful for distinguishing these isolates from other members of the M.

Sensitivity Pattern of Second Line Anti-Tuberculosis Drugs against Clinical Isolates of Multidrug Resistant Mycobacterium tuberculosis.

Objective: To determine the current sensitivity pattern of second line anti-tuberculosis drugs against clinical isolates of Multidrug Resistant Mycobacterium tuberculosis (MDR-TB).

Study Design: A cross-sectional study.

Place And Duration Of Study: Department of Microbiology, Armed Forces Institute of Pathology (AFIP), Rawalpindi, from November 2011 to April 2013.

Mycobacterium franklinii sp. nov., a species closely related to members of the Mycobacterium chelonae-Mycobacterium abscessus group.

Two isolates from water, D16Q19 and D16R27, were shown to be highly similar in their 16S rRNA, 16S-23S internal transcribed spacer (ITS), hsp65 and rpoB gene sequences to 'Mycobacterium franklinii' DSM 45524, described in 2011 but with the name not validly published. They are all nonpigmented rapid growers and are related phenotypically and genetically to the Mycobacterium chelonae-Mycobacterium abscessus group. Extensive characterization by phenotypic analysis, biochemical tests, drug susceptibility testing, PCR restriction enzyme analysis of the hsp65 gene and ITS, DNA sequencing of housekeeping genes and DNA-DNA hybridization demonstrated that 'M.

Evaluation of the Speed-Oligo Mycobacteria assay for the identification of nontuberculous mycobacteria.

Nontuberculous mycobacteria (NTM) causing human infectious disease have become increasingly common. Rapid and accurate identification to the species level is, therefore, critical. The Speed-Oligo Mycobacteria assay is an oligochromatographic method that was made available recently for the identification and differentiation of mycobacteria.

Classical and new assays for detecting drug resistance in tuberculosis.

Tuberculosis is a public health concern worldwide. Particularly worrying is the emergence of severe forms of drug resistance, such as extensively drug resistant and totally drug resistant tuberculosis, with few treatment options for the afflicted patients. To avoid further spread of drug resistance, its early detection is extremely important.

Drug Resistance Mechanisms in Mycobacterium tuberculosis.

Tuberculosis (TB) is a serious public health problem worldwide. Its situation is worsened by the presence of multidrug resistant (MDR) strains of Mycobacterium tuberculosis, the causative agent of the disease. In recent years, even more serious forms of drug resistance have been reported.

TMC207 becomes bedaquiline, a new anti-TB drug.

TB still represents a serious public health problem. The latest reports estimate an incidence of 8.7 million cases in 2011 and 1.

Prospective multicentre evaluation of the direct nitrate reductase assay for the rapid detection of extensively drug-resistant tuberculosis.

Objectives: To perform a multicentre study evaluating the performance of the direct nitrate reductase assay (NRA) for the detection of multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis in sputum samples.

Methods: The study was conducted in six laboratories performing tuberculosis diagnosis that were located in six different countries. The NRA was performed directly on sputum samples in parallel with the reference method used at each site.

Tuberculosis-spoligo-rifampin-isoniazid typing: an all-in-one assay technique for surveillance and control of multidrug-resistant tuberculosis on Luminex devices.

As a follow-up of the "spoligoriftyping" development, we present here an extension of this technique which includes the detection of isoniazid resistance-associated mutations in a new 59-plex assay, i.e., tuberculosis-spoligo-rifampin-isoniazid typing (TB-SPRINT), running on microbead-based multiplexed systems.

Tuberculosis clinical trial update and the current anti-tuberculosis drug portfolio.

Tuberculosis (TB), an ongoing public health threat, is worsened by the emergence of drug resistance. With an estimated 630000 cases per year of multidrug resistant (MDR)-TB, and 9% of those being extensively drug resistant (XDR)-TB, there is an urgent need for new and more effective anti-TB drugs. New TB treatment regimens should be able to shorten the duration of therapy that currently takes at least six months.

Genotypes of Mycobacterium tuberculosis in patients at risk of drug resistance in Bolivia.

Bolivia ranks among the 10 Latin American countries with the highest rates of tuberculosis (TB) and multidrug resistant (MDR) TB. In view of this, and of the lacking information on the population structure of Mycobacterium tuberculosis in the country, we explored genotype associations with drug resistance and clustering by analyzing isolates collected in 2010 from 100 consecutive TB patients at risk of drug resistance in seven of the nine departments in which Bolivia is divided. Fourteen isolates were MDR, 29 had other drug resistance profiles, and 57 were pansusceptible.

Field evaluation of the direct detection of multidrug resistant Mycobacterium tuberculosis by nitrate reductase assay on 7H11 agar.

In this study we evaluated the performance of colorimetric nitrate reductase assay (NRA) on Middlebrook 7H11 agar instead of Lowenstein-Jensen medium for detection of isoniazid (INH) and rifampin (RIF) resistance directly on 114 smear positive sputum specimens and compared the results with direct proportion method on LJ medium. The results of both methods were in 100% agreement for detection of RIF resistance while agreement for INH was 96.4%.