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23S rRNA base pair 2057-2611 determines ketolide susceptibility and fitness cost of the macrolide resistance mutation 2058A-->G.

ABSTRACT: The 23S rRNA A2058G alteration mediates macrolide, lincosamide, and streptogramin B resistance in the bacterial domain and determines the selectivity of macrolide antibiotics for eubacterial ribosomes, as opposed to eukaryotic ribosomes. However, this mutation is associated with a disparate resistance phenotype: It confers high-level resistance to ketolides in mycobacteria but only marginally affects ketolide susceptibility in streptococci. We used site-directed mutagenesis of nucleotides within domain V of 23S rRNA to study the molecular basis for this disparity. We show that mutational alteration of the polymorphic 2057-2611 base pair from A-U to G-C in isogenic mutants of Mycobacterium smegmatis significantly affects susceptibility to ketolides but does not influence susceptibility to other macrolide antibiotics. In addition, we provide evidence that the 2057-2611 polymorphism determines the fitness cost of the 23S rRNA A2058G resistance mutation. Supported by structural analysis, our results indicate that polymorphic nucleotides mediate the disparate phenotype of genotypically identical resistance mutations and provide an explanation for the large species differences in the epidemiology of defined drug resistance mutations.

SUBMITTER: Pfister P

PROVIDER: S-EPMC555689 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundSolithromycin, the fourth generation of ketolides, has been demonstrated potent antibacterial effect against commonly-isolated gram-positive strains. However, Staphylococcus aureus (S. aureus) strains with a higher solithromycin MIC have already been emerged, the mechanism of which is unknown.MethodsAntimicrobial susceptibility test was performed on 266 strains of S. aureus. The antibiotic resistance phenotype of erm-positive strain was determined by D-zone test. Spontaneous mutation frequency analysis was performed to compare the risk levels for solithromycin resistance among different strains. Efflux pumps and mutational analysis of ribosomal fragments as well as erm(B) gene domains were detected. Quantitative reverse transcription polymerase chain reaction was conducted to compare the transcriptional expression of the erm gene between the constitutive macrolide-lincosamide-streptogramin B (cMLSB)- and inducible MLSB (iMLSB)-phenotypes.ResultsIn the erm-positive S. aureus strains, the minimum inhibitory concentration (MIC)50/90 of solithromycin (2/>?16?mg/L) was significantly higher than that in the erm-negative strains (0.125/0.25?mg/L). Of note, the MIC50 value of the strains with iMLSB (0.25?mg/L) was significantly lower than that of the strains with cMLSB (4?mg/L). A comparison among strains demonstrated that the median mutational frequency in isolates with cMLSB (>?1.2?×?10-?4) was approximately >?57-fold and?>?3333-fold higher than that in iMLSB strains (2.1?×?10-?6) and in erythromycin-sensitive strains (3.6?×?10-?8), respectively. The differential antibiotic in vitro activity against strains between cMLSB and iMLSB could not be explained by efflux pump carriers or genetic mutations in the test genes. The expression of the erm genes in strains with cMLSB did not differ from that in strains with iMLSB.ConclusionsThe reduced susceptibility to solithromycin by S. aureus was associated with the cMLSB resistance phenotype mediated by erm.

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23S rRNA base pair 2057-2611 determines ketolide susceptibility and fitness cost of the macrolide resistance mutation 2058A-->G.

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