title: A novel MATE family efflux pump contributes to the reduced susceptibility of laboratory-derived Staphylococcus aureus mutants to tigecycline.
abstract: Tigecycline, an expanded-broad-spectrum glycylcycline antibiotic is not affected by the classical tetracycline resistance determinants found in Staphylococcus aureus. The in vitro selection of mutants with reduced susceptibility to tigecycline was evaluated for two methicillin-resistant S. aureus strains by serial passage in increasing concentrations of tigecycline. Both strains showed a stepwise elevation in tigecycline MIC over a period of 16 days, resulting in an increase in tigecycline MIC of 16- and 32-fold for N315 and Mu3, respectively. Transcriptional profiling revealed that both mutants exhibited over 100-fold increased expression of a gene cluster, mepRAB (multidrug export protein), encoding a MarR-like transcriptional regulator (mepR), a novel MATE family efflux pump (mepA), and a hypothetical protein of unknown function (mepB). Sequencing of the mepR gene in the mutant strains identified changes that presumably inactivated the MepR protein, which suggested that MepR functions as a repressor of mepA. Overexpression of mepA in a wild-type background caused a decrease in susceptibility to tigecycline and other substrates for MATE-type efflux pumps, although it was not sufficient to confer high-level resistance to tigecycline. Complementation of the mepR defect by overexpressing a wild-type mepR gene reduced mepA transcription and lowered the tigecycline MIC in the mutants. Transcription of tet(M) also increased by over 40-fold in the Mu3 mutant. This was attributed to a deletion in the promoter region of the gene that removed a stem-loop responsible for transcriptional attenuation. However, overexpression of the tet(M) transcript in a tigecycline-susceptible strain was not enough to significantly increase the MIC of tigecycline. These results suggest that the overexpression of mepA but not tet(M) may contribute to decreased susceptibility of tigecycline in S. aureus.
authors.affiliation: Wyeth Research, 401 N. Middletown Rd., Pearl River, NY 10965, USA. mcaleef@wyeth.com
authors.forename: Fionnuala
authors.initials: F
authors.lastname: McAleese
authors.forename: Peter
authors.initials: P
authors.lastname: Petersen
authors.forename: Alexey
authors.initials: A
authors.lastname: Ruzin
authors.forename: Paul M
authors.initials: PM
authors.lastname: Dunman
authors.forename: Ellen
authors.initials: E
authors.lastname: Murphy
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Projan
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Bradford
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D015342:DNA Probes; D004269:DNA, Bacterial / Q000235:genetics; D024881:Drug Resistance, Bacterial; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D005798:Genes, Bacterial / Q000235:genetics; D008826:Microbial Sensitivity Tests; D008911:Minocycline / Q000031:analogs & derivatives* / Q000494:pharmacology*; D009154:Mutation / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D010957:Plasmids / Q000235:genetics; D012329:RNA, Bacterial / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013211:Staphylococcus aureus / Q000187:drug effects*; D000078304:Tigecycline
chemical_list: D000900:Anti-Bacterial Agents; D015342:DNA Probes; D004269:DNA, Bacterial; D012329:RNA, Bacterial; D000078304:Tigecycline; D008911:Minocycline