Sensitive and specific detection of Mycobacterium tuberculosis from clinical samples by amplification of a repeated sequence of the REP13E12 family

Abstract

Objectives: To design and implement of PCR method for sensitivity and specific detection of M. tuberculosis oriented to opportune and correctly diagnosis of tuberculosis by the application on clinical specimens. Material an methods: Specific oligonucleotides for the amplification of a fragment of 318pb after a multiple alignment of sequences REP13E12 were designed. The PCR method was standarize and also the analytical and diagnostic sensibility and specifity was evaluate; using clinical samples with smear or culture diagnosis positive and negative. Results: The REP13E12-PCR detected up to 100 fg of genomic DNA. It was specific for the detection of complex M. tuberculosis and able to distinguish them of non-tuberculous mycobacteria. This preliminary evaluation provided 100% of sensitivity and specificity in sputum samples of patients with TB diagnosis. Conclusions: The REP13E12 amplification by PCR is an alternative method for the rapid diagnosis of patients with TB, especially for cases which difficult or unclear diagnosis by the gold standard technique.