Nontuberculous mycobacterial infections in Indian AIDS patients detected by a novel set of ESAT-6 polymerase chain reaction primers.

Nontuberculous mycobacteria are often underdiagnosed due to lack of proper diagnostic facilities. To overcome this, we created a rapid PCR method for the species-specific diagnosis of Mycobacterium tuberculosis and its differentiation from other mycobacteria. A set of PCR primers targeting the gene encoding for early-secreted antigen-6 (ESAT-6) of the M. tuberculosis complex was designed and standardized on mycobacterial standard strains and on 75 recent isolates from AIDS patients and 70 isolates from HIV-negative patients seen at the hospital of the All India Institute of Medical Sciences, New Delhi, India. All 145 fresh mycobacterial isolates were identified using phenotypic methods and 16S rRNA PCR followed by sequencing of hypervariable region A. The ESAT-6 PCR detected all of the M. tuberculosis strains correctly (100% sensitivity), but none of the nontuberculous Mycobacterium spp. gave positive results (100% specific). Most nontuberculous mycobacteria were identified in patients with AIDS (24%) followed by those with tuberculous lymphadenitis (12.5%) and those with pulmonary tuberculosis whose treatment had failed (4.3%). The most common nontuberculous mycobacterial species isolated from AIDS patients was M. avium (6.6%), followed by M. fortuitum (5.7%), M. intracellulare and M. terrae (2.6% each). M. celatum, M. duvalii, M. austroafricanum, M. phlei and M. flavescence were also isolated from one patient each. The combination of genus-specific PCR primers with the novel ESAT-6 primer set could provide accurate and rapid diagnosis of mycobacteriosis.