A one-step DNA PCR-based method for the detection of Mycobacterium tuberculosis complex grown on Lowenstein-Jensen media.

A simple, rapid, and sensitive direct colony polymerase chain reaction (PCR) method to detect Mycobacterium tuberculosis complex grown on Lowenstein-Jensen media is described. M. tuberculosis is killed by treating it for 2 hours with 70% ethanol. Whole Mycobacterium cells inactivated by ethanol are added to a PCR mix that is designed to amplify the IS6110 insertion sequence. All 44 isolates tested were positive by this method. Our results show that PCR can be performed directly on bacterial colonies without the need for DNA extraction before PCR. Moreover, inactivation of M. tuberculosis before DNA amplification reduces the potential exposure of workers to viable M. tuberculosis. The exclusion of DNA extraction and inactivation of colonies before PCR provide a safe and low-cost preparatory technique for PCR reaction compared with expensive conventional extraction protocols that are based on chemical and enzymatic lysis, especially for countries with limited resources.