Inability of L22 ribosomal protein alteration to increase macrolide MICs in the absence of efflux mechanism in Haemophilus influenzae HMC-S

J Antimicrob Chemother. 2004 Aug;54(2):393-400. doi: 10.1093/jac/dkh364. Epub 2004 Jul 8.


Background: Haemophilus influenzae HMC-C with high-level macrolide resistance after multi-step selection by clarithromycin reverted spontaneously and became hypersusceptible to macrolides.

Objective: Determination of macrolide resistance mechanism(s) in hypersusceptible and hyperresistant strains.

Methods: The presence of macrolide efflux in the strains was studied by radioactive erythromycin accumulation. Ribosomal mutations were investigated by sequencing. The possible role of acrAB clusters in macrolide resistance was studied by sequencing and expression analysis.

Results: The parent strain had no ribosomal alteration, but both high-level resistant and hypersusceptible strains had R88P mutations in ribosomal protein L22. Radioactive macrolide accumulation studies pointed to the presence of macrolide efflux in the high-level resistant and parent strains, but not in the hypersusceptible derivative. Transformation of hypersusceptible strains using total DNA from the parent strain restored the macrolide efflux system in the hypersusceptible strain, which was confirmed by MIC levels and radioactive erythromycin accumulation similar to that of the mutant resistant strain. Analysis of sequence and transcription of acrAB gene clusters showed no significant differences between resistant and hypersusceptible derivatives.

Conclusion: Mutation in ribosomal protein L22 alone does not confer high-level macrolide resistance unless efflux is present.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Bacterial Agents / pharmacology*
  • Blotting, Southern
  • Clarithromycin / pharmacology
  • DNA Primers
  • DNA, Bacterial / biosynthesis
  • DNA, Bacterial / genetics
  • Drug Resistance, Bacterial
  • Erythromycin / metabolism
  • Genes, Bacterial / genetics
  • Haemophilus influenzae / drug effects*
  • Haemophilus influenzae / metabolism*
  • Macrolides / pharmacology*
  • Microbial Sensitivity Tests
  • Multigene Family / genetics
  • Oligonucleotide Array Sequence Analysis
  • Protein Conformation
  • RNA, Bacterial / biosynthesis
  • RNA, Bacterial / genetics
  • Ribosomal Proteins / drug effects
  • Ribosomal Proteins / genetics
  • Ribosomal Proteins / metabolism*
  • Transformation, Bacterial


  • Anti-Bacterial Agents
  • DNA Primers
  • DNA, Bacterial
  • Macrolides
  • RNA, Bacterial
  • Ribosomal Proteins
  • Erythromycin
  • Clarithromycin