Mycobacterium tuberculosis inhibits gamma interferon (IFN-␥Mycobacterium tuberculosis is a facultative intracellular pathogen that resides and multiplies within human macrophages. Gamma interferon (IFN-␥), the predominant inducer of macrophage-mediated microbicidal functions (36), has been shown to be required for the prevention of progressive M. tuberculosis infection (13,19). In tuberculosis patients, IFN-␥ has been detected in CD4ϩ T cells and culture supernatants (8) as well as in infectious foci (7). Despite the availability of IFN-␥, the immune system is unable to eradicate the infection, indicating that M. tuberculosis selectively inhibits macrophage responsiveness to 61). This reduced response results in the inefficient induction of IFN-␥-inducible genes, such as major histocompatibility complex class II and others (24, 61).IFN-␥ binds to its cell surface receptor, IFN-␥R, which consists of two heterodimeric subunits, IFN-␥R1 (␣, ligand binding) and IFN-␥R2 (, signaling subunit) (6). The IFN-␥R is MATERIALS AND METHODSSubjects. Informed consent obtained from a cohort of 13 pulmonary tuberculosis (PTB) patients of the outpatient department of LRS Hospital of Tuberculosis and Respiratory Diseases and 16 laboratory personnel of the Biotechnology Department, AIIMS, New Delhi, India, were included in the study. All patients were administered standard antitubercular therapy (ATT). Six of them were available for the assessment of the level of expression of IFN-␥R1 at various time points prior to and after therapy. Scrutinizing patient clinical histories, physical examinations, and laboratory investigations ruled out the occurrence of concom-* Corresponding author. Mailing address:
BackgroundIndia has the highest estimated burden of tuberculosis in the world, accounting for 21% of all tuberculosis cases world-wide. However, due to lack of systematic analysis using multiple markers the available information on the genomic diversity of Mycobacterium tuberculosis in India is limited.Methodology/Principal FindingsThus, 65 M. tuberculosis isolates from New Delhi, India were analyzed by spoligotyping, MIRU-VNTR, large deletion PCR typing and single nucleotide polymorphism analysis (SNP). The Central Asian (CAS) 1 _DELHI sub-lineage was the most prevalent sub-lineage comprising 46.2% (n = 30) of all isolates, with shared-type (ST) 26 being the most dominant genotype comprising 24.6% (n = 16) of all isolates. Other sub-lineages observed were: East-African Indian (EAI)-5 (9.2%, n = 6), EAI6_BGD1 (6.2%, n = 4), EAI3_IND, CAS and T1 with 6.2% each (n = 4 each), Beijing (4.6%, n = 3), CAS2 (3.1%, n = 2), and X1 and X2 with 1 isolate each. Genotyping results from five isolates (7.7%) did not match any existing spoligopatterns, and one isolate, ST124, belonged to an undefined lineage. Twenty-six percent of the isolates belonged to the TbD1+ PGG1 genogroup. SNP analysis of the pncA gene revealed a CAS-lineage specific silent mutation, S65S, which was observed for all CAS-lineage isolates (except two ST26 isolates) and in 1 orphan. Mutations in the pncA gene, conferring resistance to pyrazinamide, were observed in 15.4% of all isolates. Collectively, mutations in the rpoB gene, the katG gene and in both rpoB and katG genes, conferring resistance to rifampicin and isoniazid, respectively, were more frequent in CAS1_DELHI isolates compared to non-CAS_DELHI isolates (OR: 3.1, CI95% [1.11, 8.70], P = 0.045). The increased frequency of drug-resistance could not be linked to the patients' history of previous anti-tuberculosis treatment (OR: 1.156, CI95% [0.40, 3.36], P = 0.79). Fifty-six percent of all new tuberculosis patients had mutations in either the katG gene or the rpoB gene, or in both katG and rpoB genes.ConclusionCAS1_DELHI isolates circulating in New Delhi, India have a high frequency of mutations in the rpoB and katG genes. A silent mutation (S65S) in the pncA gene can be used as a putative genetic marker for CAS-lineage isolates.
Nucleic acid amplification tests have improved tuberculosis diagnostics considerably. This study evaluates a new amplification test, the GenoType Mycobacteria Direct (GTMD) test, for detection of the Mycobacterium tuberculosis complex, Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium kansasii, and Mycobacterium malmoense directly in 61 sputum samples. Thirty (49.2%) samples were auramine smear positive, and 31 (50.8%) were smear negative. The GTMD results were compared to the Gen-Probe Amplified M. tuberculosis Direct (MTD) test results, using culturing and sequencing of the 16S rRNA gene as reference methods. The GTMD test could identify 28 of 29 samples containing the M. tuberculosis complex and was negative in a sputum sample containing M. intracellulare. The overall sensitivity and specificity results were 93.3% and 90.0% for the GTMD test, respectively, and 93.1% and 93.5% for the MTD test, respectively. The GTMD test is rapid and can be easily included in routine clinical laboratories for the direct detection of the M. tuberculosis complex in smear-positive sputum samples as an adjunct to microscopy and culture. Further studies are needed to evaluate the performance of the GTMD test for the detection of atypical mycobacteria.Worldwide, tuberculosis (TB) is a major cause of illness and death. WHO estimates that in 2006, 9.2 million new cases and 1.7 million deaths occurred from TB globally (25), and the incidence is increasing. The emergence of multidrug-resistant TB, and recently also extensively drug-resistant TB, and the human immunodeficiency virus-TB coinfection are further worsening the situation, and effort to accelerate progress in global TB control is needed. Important factors for TB control are increased case detection and treatment success rates (25). The slow growth of most pathogenic mycobacteria results in diagnosis and treatment delay and has stimulated the development of nucleic acid amplification (NAA) tests for identification of mycobacteria directly in clinical specimens. NAA tests provide test results within 1 day. In general, the specificity result for NAA tests ranges from 95% to 100% (1,12,16,23), but the sensitivity result, especially for acid-fast bacillus (AFB) smear-negative samples, varies greatly, from 33 to 96% (1,12,16,23). For AFB smear-positive respiratory specimens, the sensitivity level is approximately 95%.Two direct systems approved by the United States Food and Drug Administration (FDA) for detection of pulmonary TB are commercially available, as follows: the Amplicor Mycobacterium tuberculosis test (Roche Diagnostic Systems, Indianapolis, IN) and the Gen-Probe Amplified M. tuberculosis Direct test (MTD test; Gen-Probe, San Diego, CA). Both tests use the 16S rRNA gene as the target amplification gene. The 16S rRNA gene represents a stable property of microorganisms and is widely used as the target for identifying mycobacterium species. Several studies have confirmed an excellent test proficiency (sensitivity and specificity levels of more than 95%) in AF...
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