Role of multiplex polymerase chain reaction using IS6110 and Protein b for the diagnosis of extra-pulmonary tuberculosis: North India

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Publisher: Medknow Publications and Media Pvt. Ltd.
Document Type: Report
Length: 2,297 words
Lexile Measure: 1550L

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Byline: Kusum. Sharma, Suma. Appannanavar, Manish. Modi, Malkit. Singh, Aman. Sharma, Subhash. Varma

Background: Prompt and accurate diagnosis of extra-pulmonary tuberculosis (TB) is highly challenging. Current conventional techniques lack sensitivity and are time-consuming. Here, we report our experience with multiplex polymerase chain reaction (MPCR) using two targets namely IS6110 and protein antigen b in the diagnosis of extra-pulmonary TB. Materials and Methods: A total of 150 patients of extra-pulmonary TB visiting tertiary care center in north India between September 2008 and December 2009 were included in the study. Sixty-six biopsy samples and 84 were body fluids from these patients were subjected for microscopy (Ziehl-Neelsen), culture on LJ medium and for Multiplex PCR using IS6110 and Protein b antigen. Results: Smear positivity was noted in 11 samples (7.33%), and LJ culture yielded Mycobacterium tuberculosis in 8 biopsies and 9 body fluids with overall positivity of 11.3%. The multi-targeted PCR could detect M. tuberculosis in a total of 112 samples. Of 112 positive samples, only Protein b band was detected in 7 samples and only IS6110 was detected in 5 samples. Overall Protein b, PCR could detect 71.33% of the cases, whereas IS6110 was positive in 66.6% of the cases. Overall the sensitivities of microscopy, culture, IS6110 PCR, Protein b PCR and MPCR were 7.33%, 11.3%, 66.67%, 71.3% and 74.6%, respectively. Thus by using more than two targets the sensitivity increased from 66.67% of IS6110 to 74.6% in MPCR. Conclusion: Multiplex polymerase chain reaction using IS6110 and Protein b antigen is a highly sensitive and specific tool in the diagnosis of pauci-bacillary conditions like extra-pulmonary TB.

INTRODUCTION

Extra-pulmonary tuberculosis (TB) accounts for around 20-50% of all TB cases. [sup][1] Prompt and accurate diagnosis is essential for better management of this group of pauci-bacillary patients. Currently, the diagnosis of extra-pulmonary TB relies on clinical features, imaging findings and conventional microbiological tools like microscopy and culture. Microscopy is quite insensitive especially because of the pauci-bacillary nature of extra-pulmonary TB. Although culture is a gold standard, it has a slow turnaround time of 6-8 weeks. Thus, there is a need for rapid, sensitive and specific tests for the diagnosis of extra-pulmonary TB. Nucleic acid based tests have proven beneficial in the diagnosis of TB. The polymerase chain reaction is the most common nucleic acid amplification test. [sup][2] A large number of sequences of mycobacterial genome like IS6110, IS986, 65 kDa and 38 kDa antigens have been used as targets in polymerase chain reaction (PCR) test. [sup][3],[4] Most of the PCR-based studies have shown single target, out of which IS6110 is the most commonly used target because it has multiple copies (16-20). [sup][5] However, in the Indian subcontinent, Chauhan et al . have shown that 10-40% of the Mycobacterial strains either have low copies or no copy of IS6110. [sup][6] More reliable report may be obtained by using more than one target. Another important target is protein antigen b (38 kDa protein), which is specific for Mycobacterium tuberculosis complex and has shown good clinical utility in...

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Gale Document Number: GALE|A410607023