title: Mutation landscape of acquired cross-resistance to glycopeptide and β-lactam antibiotics in Enterococcus faecium.
abstract: Bypass of the d,d-transpeptidase activity of penicillin-binding proteins by an l,d-transpeptidase (Ldtfm) results in resistance to ampicillin and glycopeptides in Enterococcus faecium M9, a mutant obtained by nine consecutive selection steps. Resistance requires activation of a cryptic locus for production of the essential tetrapeptide-containing substrate of Ldtfm and impaired activity of protein phosphatase StpA. Here, whole-genome sequencing revealed a high mutation rate for the entire selection procedure (79 mutations in 900 generations). Acquisition of a mutation in the mismatch repair gene mutL had little impact on the frequency of rifampin-resistant mutants although the mutation spectrum of M9 was typical of impaired MutL with high transversion to transition (40/11) and substitution to deletion (51/28) ratios. M9 did not mainly accumulate neutral mutations since base substitutions occurred more frequently in coding sequences than expected (χ(2) = 5.0; P < 0.05) and silent mutations were underrepresented (χ(2) = 5.72; P < 0.02). None of the mutations directly affected recognition of the tetrapeptide substrate of Ldtfm by peptidoglycan synthesis enzymes. Instead, mutations appear to remodel regulatory circuits involving two-component regulatory systems and sugar metabolism. The high number of mutations required for activation of the l,d-transpeptidase pathway may strongly limit emergence of cross-resistance to ampicillin and glycopeptides by this mechanism.
authors.affiliation: INSERM, UMR_S 1138, Centre de Recherche des Cordeliers, Equipe 12, Paris, France Sorbonne Universités, UPMC Université Paris 06, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France Université Paris Descartes, Sorbonne Paris Cité, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Sacco
authors.affiliation: INSERM, UMR_S 1138, Centre de Recherche des Cordeliers, Equipe 12, Paris, France Sorbonne Universités, UPMC Université Paris 06, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France Université Paris Descartes, Sorbonne Paris Cité, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France.
authors.forename: Mélanie
authors.initials: M
authors.lastname: Cortes
authors.affiliation: INSERM, UMR_S 1138, Centre de Recherche des Cordeliers, Equipe 12, Paris, France Sorbonne Universités, UPMC Université Paris 06, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France Université Paris Descartes, Sorbonne Paris Cité, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Josseaume
authors.affiliation: Institut Pasteur, Genomic Platform, Paris, France.
authors.forename: Christiane
authors.initials: C
authors.lastname: Bouchier
authors.affiliation: INSERM, UMR_S 1138, Centre de Recherche des Cordeliers, Equipe 12, Paris, France Sorbonne Universités, UPMC Université Paris 06, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France Université Paris Descartes, Sorbonne Paris Cité, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Dubée
authors.affiliation: INSERM, UMR_S 1138, Centre de Recherche des Cordeliers, Equipe 12, Paris, France Sorbonne Universités, UPMC Université Paris 06, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France Université Paris Descartes, Sorbonne Paris Cité, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France.
authors.forename: Jean-Emmanuel
authors.initials: JE
authors.lastname: Hugonnet
authors.affiliation: INSERM, UMR_S 1138, Centre de Recherche des Cordeliers, Equipe 12, Paris, France Sorbonne Universités, UPMC Université Paris 06, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France Université Paris Descartes, Sorbonne Paris Cité, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France Assistance Publique-Hôpitaux de Paris, Hôpital Européen Georges Pompidou, Paris, France.
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Mainardi
authors.affiliation: Rhode Island Hospital, Brown University, Providence, Rhode Island, USA.
authors.forename: Louis B
authors.initials: LB
authors.lastname: Rice
authors.affiliation: INSERM, UMR_S 1138, Centre de Recherche des Cordeliers, Equipe 12, Paris, France Sorbonne Universités, UPMC Université Paris 06, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France Université Paris Descartes, Sorbonne Paris Cité, UMR_S 1138, Centre de Recherche des Cordeliers, Paris, France michel.arthur@crc.jussieu.fr.
authors.forename: Michel
authors.initials: M
authors.lastname: Arthur
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics; D016984:Enterococcus faecium / Q000187:drug effects* / Q000235:genetics; D006020:Glycopeptides / Q000494:pharmacology*; D009154:Mutation; D047090:beta-Lactams / Q000494:pharmacology*
chemical_list: D000900:Anti-Bacterial Agents; D006020:Glycopeptides; D047090:beta-Lactams