Most rifampicin-resistant strains have been associated with mutations in an 81-bp rifampicin resistance-determining region (RRDR) in the Mycobacterium tuberculosis gene rpoB. However, when targeting this region alone, rifampicin-resistant strains with mutations outside the RRDR would not be detected. In this study, among fifty-one rifampicin-resistant clinical isolates analyzing by sequencing 1681-bp-long DNA fragments containing the RRDR, 47 isolates contained mutations within the RRDR, three isolates have mutation both within and outside of RRDR while only one isolate had single missense mutation (Arg548His) located outside the RRDR. The drug susceptibility test of recombinant Mycobacterium smegmatis and M. tuberculosis carrying mutated rpoB (Arg548His) showed an increased minimum inhibitory concentration (MIC) for rifampicin, compared to control strains. Modelling the Arg548His mutant RpoB-DNA complex revealed that the His548 side chain formed a more stable hydrogen-bond structure than Arg548, reducing the flexibility of the rifampicin-resistant cluster II region of RpoB, suggesting that the RpoB Arg548His mutant does not effectively interact with rifampicin and resulting in bacterial resistance to the drug. This is the first report on the relationship between the mutation of codon 548 of RpoB and rifampicin resistance in tuberculosis. The novel mutational profile of the rpoB gene described here will contribute to the comprehensive understanding of rifampicin resistance patterns and to the development of a useful tool for simple and rapid drug susceptibility tests.
Date:
2015-03
Relation:
Antimicrobial Agents and Chemotherapy. 2015 Mar;59(3):1542-1548.
