Evaluation of loop mediated isothermal amplification for diagnosis of Mycobacterium tuberculosis complex in clinical samples

Abstract

Tuberculosis (TB) remains an important global public health problem. The lack of rapid and accurate diagnostic testing is an important impediment to global tuberculosis control. Loop mediated isothermal amplification (LAMP) is a rapid method for nucleic acid amplification. In this study, we assessed the performance of an in-house LAMP assay for the detection of tuberculosis. Six oligonucleotide primers specific for Mycobacterium tuberculosis complex were designed corresponding to IS6110 gene sequence. Optimization of LAMP reaction was performed. A total of 133 clinical sputum samples and 80 bacterial cultures were studied by LAMP method. Sensitivity of this assay for detection of genomic DNA was 5 fg. This assay successfully detected M. tuberculosis complex not only in the bacterial cultures but also in the clinical sputum samples from patients with TB. The sensitivity of LAMP in culturepositive samples was 100% (60/60) and the specificity in culture-negative samples was 95.9% (70/73, 95% confidence interval 91.3 to 98.7%). Thus, LAMP is a rapid, highly sensitive and specific DNA amplification technique for early diagnosis of TB.

Key words: Loop mediated isothermal amplification (LAMP), polymerase chain reaction (PCR), IS6110 gene, Mycobacterium tuberculosis, diagnosis.