Single and dual mutations at positions 2058, 2503 and 2504 of 23S rRNA and their relationship to resistance to antibiotics that target the large ribosomal subunit.

OBJECTIVES To study mutations at positions A2058, A2503 and U2504 (Escherichia coli numbering) of 23S rRNA and their relationship to resistance to antibiotics that target the large ribosomal subunit. METHODS Single and dual mutations at positions 2058, 2503 and 2504 of 23S rRNA were introduced into a Mycobacterium smegmatis strain with a single functional rRNA operon. MICs of macrolide, pleuromutilin, phenicol, lincosamide and oxazolidinone antibiotics were determined for the engineered mutants. The doubling times of the mutant strains were measured to investigate how the introduced mutations affected growth rate. RESULTS Single mutations A2058G, A2503U and U2504G and double mutations A2058G-A2503U and A2058G-U2504G were successfully introduced. The A2058G mutation resulted in various levels of resistance to macrolides and clindamycin. The A2503U and U2504G mutations conferred resistance to valnemulin, chloramphenicol, florfenicol and linezolid. In addition, the A2503U mutant showed reduced susceptibility to the 16-membered macrolides tylosin, spiramycin and josamycin, and the U2504G mutant exhibited decreased susceptibility to spiramycin and josamycin. Moreover, the dual mutations A2058G-A2503U and A2058G-U2504G had co-effects on resistance to 16-membered macrolides. CONCLUSIONS 23S rRNA mutations A2058G, A2503U and U2504G play key roles in resistance to clinically useful antibiotics that target the large ribosomal subunit. Furthermore, the double mutations A2058G-A2503U and A2058G-U2504G have combined effects on resistance to 16-membered macrolides.