DETECTION OF A GENETIC-LOCUS ENCODING RESISTANCE TO RIFAMPIN IN MYCOBACTERIAL CULTURES AND IN CLINICAL SPECIMENS

The polymerase chain reaction (PCR) and automated DNA sequencing were used to detect a genetic locus, rpoB, associated with rifampin resista nce in Mycobacterium tuberculosis (TB) in clinical isolates and direct ly in clinical specimens. Primers derived from the sequence of a TB rp oB gene fragment were used to amplify DNA from bacterial and mycobacte rial isolates. An rpoB-specific PCR product was obtained for five of f ive TB, seven of eight other mycobacterial species, Nocardia sp., Cory nebacterium sp., Streptomyces sp., Actinomyces sp., and Rhodococcus sp ., but not for 15 isolates (eight genera) representing usual bacterial flora. Sequence comparison of the amplified rpoB region revealed the occurrence of TB-specific ''signature nucleotides'' at three positions . PCR yielded amplification products for seven of 16 clinical specimen s. Five of the seven contained TB-specific DNA, as well as sequences t hat predicted rifampin susceptibility in accord with agar dilution res ults. None of ten specimens that were culture negative for TB yielded TB-specific PCR products. These results with a limited number of clini cal specimens demonstrate the feasibility of direct detection by PCX o f rifampin-resistant TB in clinical specimens. Such testing may serve as a rapid surrogate test for multidrug-resistant TB in laboratories w ith PCR and automated sequencing capability.