Rapid differentiation of Mycobacterium tuberculosis and Mycobacterium leprae from sputum by polymerase chain reaction.

Differentiation of M tuberculosis and M leprae by polymerase chain reaction (PCR), when acid-fast bacilli (AFB) were present in sputum from patients at Anandaban hospital, was carried out. Thirty sputum samples microscopy positive for AFB were collected and were subjected to culture. Bacterial DNA was extracted and PCR was performed using primers specific for Mycobacterium tuberculosis and Mycobacterium leprae DNA. Twenty samples were from patients with clinical TB and 10 from patients with clinical leprosy. Fifteen of the TB samples were positive in both TB PCR and culture, among the reminders four were TB PCR negative and one was positive for TB PCR. All TB samples were negative for leprosy PCR. Of the leprosy samples, five were TB PCR and culture positive, and negative for leprosy PCR. The remaining five samples were negative for both TB PCR and culture but positive in leprosy PCR. Five often clinical leprosy samples were positive for tuberculosis. This indicates that AFB in the sputum of leprosy patients might be M. tuberculosis or M. leprae. Thus PCR can be used for rapid differentiation of M. tuberculosis and M. leprae present in sputum where AFB microscopy is inconclusive.

Reverse line probe assay for the rapid detection of rifampicin resistance in Mycobacterium leprae.

Mutations in the rpoB gene of 40 biopsy isolates of Mycobacterium leprae were analyzed by reverse hybridization-based line probe assay after PCR, and nine distinct single-nucleotide substitutions were found. Among them, a 3-nucleotide substitution was found in two, and 2-nucleotide substitutions were found in seven isolates. This is a new finding of multiple mutations in a single point of the rpoB gene for rifampicin resistance. This investigation demonstrates that the pattern of mutations in the rpoB gene for rifampicin resistance in Nepal involves more variety.