Comparative evaluation of two polymerase chain reactions targeting different genomic regions to detect Mycobacterium tuberculosis in sputum

Abstract

Purpose: Tuberculosis remains an important health problem all over the world, especially in resource poor settings like India. The Ziehl-Neelsen (ZN) staining of sputum smear is still the method of choice in the diagnosis of tuberculosis in spite of its low sensitivity and specificity. This paper evaluates comparison of two different polymerase chain reaction (PCR) assays with sputum smear findings to detect Mycobacterium tuberculosis. Materials and Methods: A total of 191 sputum samples were collected from 84 patients attending a tertiary care hospital, who were suspected of having pulmonary tuberculosis, were examined by PCR targeting two different genomic regions, namely, TRC4 by non-nested format and IS6110 insertion element by nested format in comparison to ZN staining of sputum smears. Results: Among the patients tested, 20.24% (Mid-p 95%CI: 31.5–52.4) were smear positive, 7.14% (Mid-p 95%CI: 2.94–14.26) were positive by TRC4 PCR and 41.67% (Mid-p 95%CI: 12.7–29.8) were positive by IS6110 nested PCR (nPCR). The median age of overall positive cases was 42 years. Among the nPCR positives, the median for age of rural and peri-urban community was 46 and 32 years, respectively. The kappa coefficient between smear findings and TRC4 PCR findings was 0.27 and an agreement of 0.83 was observed (Z = 2.99; one-tailed P = 0.001). TRC4 PCR picked two unique positives that were negative by smear and IS6110 nPCR. Conclusion: The non-nested TRC4 PCR showed inability for accurate detection of M. tuberculosis in sputum samples. The study concluded that the nPCR targeting IS6110 is superior and more sensitive than TRC4 PCR.