ABSTRACT
The conventional techniques like the Acid-Fast Bacilli (AFB) staining and smear microscopy employed for the diagnosis of tuberculosis (TB) have a significant disadvantage of poor sensitivity and the gold standard diagnostic method employing culture is time-consuming. This study was conducted to isolate and identify Mycobacterium tuberculosis (MTB) from the sputum sample and to evaluate the efficacy of PCR as a modern diagnostic tool, for diagnosis of tuberculosis, especially in the smear-negative cases. The multiplex polymerase chain reaction (MPCR) offers a great promise as a diagnostic tool for TB owing to the rapid analysis, significant sensitivity, and specificity. Amplified nucleic acid hybridization assays such as MPCR have shown promising results. The aim of this study was to perform MPCR analysis using MTB genes namely PPE41, MPT53, LPQH, ESAT-6, and CFP-10, and analyze its efficiency in the diagnosis of TB. 252 sputum samples were collected from TB Hospital and they were subjected to Ziehl-Neelsen, culture analysis employing Lowenstein-Jensen (L-J) 224medium, and Multiplex PCR (MPCR) for the specific detection of mycobacterium DNA. The MPCR was performed for targeting genes of MTB. Of 252 cases, 102 samples showed positive for AFB, 124 samples were positive for culture method and 196 samples showed positive for MPCR. The overall results showed a sensitivity as well as specificity of the MPCR assay as 97.7% and 100%, respectively. Whereas the sensitivity and specificity in the case of culture was 89.2% and 98.1% and with AFB it was 63.3% and 92%, respectively. Analysis of the positive and negative predictive values of MPCR was found to be 91.1% and 99.3% when compared to the culture (78.2% and 87.1%) and AFB (71.2% and 82%), respectively. MPCR analysis performed by employing specified MTB primers was able to detect more TB positive cases when compared to the conventional AFB and L-J culture method for the detection of MTB. MPCR was found to be a more rapid, reliable, more sensitive, and highly specific diagnostic tool for the purpose of detection as well as the differentiation of MTB patients and this can serve as the potential diagnostic tool for the analysis of sputum samples.
