BURDEN OF MYCOBACTERIUM-TUBERCULOSIS IN SPUTUM SAMPLES CAN BE RELIABLY DETERMINED USING A QUANTITATIVE, NONRADIOACTIVE POLYMERASE CHAIN-REACTION ASSAY

Objective: To develop a rapid assay for quantitation of Mycobacterium tuberculosis in sputum samples using the competitive polymerase chain reaction (PCR) and a colorimetric microtiter well detection format. De sign: The assay relies on the co-amplification of a 419 base pair (bp) pab fragment of M. tuberculosis together with a target template (pab/ tet) made by splicing a fragment of tet excised from pbr322 between th e 5' and 3' ends of the pub fragment to create a 380 bp hybrid templat e amplified with the same primers but readily distinguishable using pr obes specific for either pub or tet. Results: We demonstrate a good co rrelation between the results obtained using this assay and the result s of quantitative culture. Conclusion: This assay provides quantitativ e information regarding M. tuberculosis burden in samples containing b etween 10(3) and 10(8) colony forming units/milliliter (CFU/ml).