DETECTION OF MYCOBACTERIUM-TUBERCULOSIS IN SPUTUM SAMPLES BY POLYMERASE CHAIN-REACTION USING A SIMPLIFIED PROCEDURE

A repetitive sequence of Mycobacterium tuberculosis DNA was amplified by polymerase chain reaction (PCR), from sputum samples, for the diagn osis of pulmonary tuberculosis. The method of heating the sample in a boiling water bath to break down the bacterial cell wall and to releas e the DNA was compared with that of enzymatic lysis of bacteria and th en phenol-chloroform extraction of DNA. Heating the sample was the bet ter method with a sensitivity of approximately 10 microorganisms. A to tal of 78 sputum specimens prepared by heating were examined by PCR, a nd the results were compared with the results of acid-fast stained sme ars, cultures, and clinical data. M. tuberculosis was detected by PCR in all smear- and culture-positive and smear-negative, culture-positiv e cases. Additionally, PCR was capable of detecting four of nine cases which were smear and culture negative but clinically suspected of tub erculosis. DNA amplification by PCR is a sensitive and specific method for the diagnosis of tuberculosis, and with this simplified DNA isola tion procedure it can be used in routine clinical practice.