Evaluation of Nested Polymerase Chain Reaction for Rapid Diagnosis of Clinically Suspected Tuberculous Pleurisy

Abstract

Background: Early diagnosis of tuberculosis is important in its control. The conventional techniques like smear microscopy and culture suffer from low sensitivity for diagnosis of extra-pulmonary tuberculosis like Pleural Tuberculosis (PTB) due to paucibacillary nature of the fluid. Polymerase Chain Reaction (PCR) is presently seen as a promising alternative to conventional techniques. In this study we have evaluated IS6110 sequence based nested PCR (nPCR) for the detection of Mycobacterium tuberculosis (MTB) DNA directly from clinical samples. The results of PCR were compared with the results of conventional methods like smear, culture and Adenosine Deaminase (ADA) activity. Material and Methods: A total of 50 pleural fluid samples from the patients with history suggestive of tuberculosis were taken. All the samples were processed for Ziehl-Neelsan (ZN) staining for Acid Fast Bacilli (AFB), culture ADA activity and PCR with primers targeting 123bp fragment of IS6110 of MTB complex. Results: A significant difference was seen in the sensitivities of conventional methods and PCR (p<0.05). Out of these 50 samples 3 were positive by smear, culture was positive in 5 samples, 21 samples showed high ADA activity and 29 were positive by PCR with overall 100% sensitivity of PCR using culture on LJ media as gold standard. Conclusions: The combined analysis of nPCR, ADA activity and other lab investigations can be very useful in the rapid diagnosis in cases of PTB.

Keywords

• pleural tuberculosis
• mycobacterium tuberculosis
• nested polymerase chain reaction
• is6110 sequence