Article

Improved diagnosis of central nervous system tuberculosis by MPB64-Target PCR

Neurosurgery.
Brazilian Journal of Microbiology (Impact Factor: 0.59). 06/2008; 39(2):209-213. DOI: 10.1590/S1517-83822008000200002
Source: PubMed

ABSTRACT

Central nervous system (CNS) tuberculosis is a serious clinical problem, the treatment of which is sometimes hampered by delayed diagnosis. Clearly, prompt laboratory diagnosis is of vital importance as the spectrum of disease is wide and abnormalities of the cerebrospinal fluid (CSF) are incredibly variable. Since delayed hypersensitivity is the underlying immune response, bacterial load is very low. The conventional bacteriological methods rarely detect Mycobacterium tuberculosis in CSF and are of limited use in diagnosis of tuberculous meningitis (TBM). This double blind study was, therefore, directed to the molecular analysis of CNS tuberculosis by an in-house-developed PCR targeted for amplification of a 240bp nucleotide sequence coding for MPB64 protein specific for Mycobacterium tuberculosis. Based on the clinical criteria, 47 patients with CNS tuberculosis and a control group of 10 patients having non-tubercular lesions of the CNS were included in the study. Analyses were done in three groups; one group consisting of 27 patients of TBM, a second group of 20 patients with intracranial tuberculomas and a third group of 10 patients having nontubercular lesions of the CNS acted as control. There were no false positive results by PCR and the specificity worked out to be 100%. In the three study groups, routine CSF analysis (cells and chemistry), CSF for AFB smear and culture were negative in all cases. PCR was positive for 21/27 patients (77.7% sensitivity) of the first group of TBM patients, 6/20 patients (30% sensitivity) of the second group with intracranial tuberculomas were positive by PCR and none was PCR-positive (100% specificity) in the third group. Thus, PCR was found to be more sensitive than any other conventional method in the diagnosis of clinically suspected tubercular meningitis.

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    • "We have chosen IS6110 primers because of multiple copy numbers (6–24) in the Mycobacterium genome.[11] MPB64 primer has shown good sensitivity for diagnosis of CNS TB.[13] To the best of our knowledge, this is one of the first few studies in which role of multiplex PCR using IS6110 and MPB64 for early diagnosis of GITB has been evaluated. "
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    ABSTRACT: Rapid and specific diagnosis of gastrointestinal tuberculosis (GITB) is of utmost importance. To evaluate Multiplex PCR (MPCR) using MPB64 and IS6110 primers specific for M. tuberculosis for rapid diagnosis of GITB. MPCR was performed on colonoscopy biopsy specimens on 11 GITB confirmed (culture/AFB/histopathology was positive), 29 GITB suspected and 30 Non GITB (control group) patients. MPB64 PCR had sensitivity and specificity of 90% and 100% for confirmed GITB cases. In 29 clinically diagnosed but unconfirmed GITB cases, MPCR was positive in 72.41%. MPCR was negative in all control group patients. The overall sensitivity and specificity of microscopy, culture, histopathology and MPCR was 5%, 2% 20% and 77.5% and 100%, 100%, 100% and 100% respectively. MPCR has good sensitivity and specificity in diagnosing gastrointestinal tuberculosis.
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    • "The most extensively used molecular epidemiology technique is Restriction Fragment Length Polymorphism (RFLP) typing, which uses the insertion sequence IS6110 to differentiate clinical isolates (5,37). Polymerase Chain Reaction (PCR) is the most sensitive method in the diagnosis of clinically suspected tuberculosis (1,8,25). New typing methods based on the PCR, such as spoligotyping (18), and mycobacterial interspersed repetitive units (MIRU) typing have also been described (34). "
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    Full-text · Article · Apr 2009 · Brazilian Journal of Microbiology
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    [Show abstract] [Hide abstract]
    ABSTRACT: Designing newer drugs, vaccines, and diagnostic techniques is dependent on better understanding of M. tuberculosis virulence mechanism. In this study the prevalence of pcaA gene was determined in M. tuberculosis strains typed by spoligotyping. The associated risk factors among patients with different nationalities residing in Iran were also determined. The isolated M. tuberculosis strains have been characterized by performing susceptibility tests against four first-line antituberculosis drugs and were then subjected to spoligotyping characterization. PCR was used for detection of pcaA gene and its nucleotide sequence was also determined. Spoligotyping of M. tuberculosis strains resulted in 140 different patterns. One hundred twenty two (87.1%) of these spoligotype isolates were unique and reported for the first time. The remaining18 (12.8%) spoligotype patterns were previously reported from other geographical regions of the world. Haarlem family was most prevalent than other genotype. Antibiotic resistances were higher in those isolated from the Iranian patients. The pcaA gene was detected in M. tuberculosis clinical isolates but not in saprophyte strains such as M. kansasi. The results showed that, spread of M. tuberculosis strains belonging to the Beijing family among Iranian patients has to be considered seriously. This study confirmed the widespread existence of pcaA gene in almost all the clinical isolates. It is also important to undertake studies to identify which factors are the most significant to consider in tuberculosis control program.
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