title: The functional vanGCd cluster of Clostridium difficile does not confer vancomycin resistance.
abstract: vanGCd, a cryptic gene cluster highly homologous to the vanG gene cluster of Enterococcus faecalis is largely spread in Clostridium difficile. Since emergence of vancomycin resistance would have dramatic clinical consequences, we have evaluated the capacity of the vanGCd cluster to confer resistance. We showed that expression of vanGCd is inducible by vancomycin and that VanGCd , VanXYCd and VanTCd are functional, exhibiting D-Ala : D-Ser ligase, D,D-dipeptidase and D-Ser racemase activities respectively. In other bacteria, these enzymes are sufficient to promote vancomycin resistance. Trans-complementation of C. difficile with the vanC resistance operon of Enterococcus gallinarum faintly impacted the MIC of vancomycin, but did not promote vancomycin resistance in C. difficile. Sublethal concentration of vancomycin led to production of UDP-MurNAc-pentapeptide[D-Ser], suggesting that the vanGCd gene cluster is able to modify the peptidoglycan precursors. Our results indicated amidation of UDP-MurNAc-tetrapeptide, UDP-MurNAc-pentapeptide[D-Ala] and UDP-MurNAc-pentapeptide[D-Ser]. This modification is passed on the mature peptidoglycan where a muropeptide Tetra-Tetra is amidated on the meso-diaminopimelic acid. Taken together, our results suggest that the vanGCd gene cluster is functional and is prevented from promoting vancomycin resistance in C. difficile.
authors.affiliation: EA4043, Faculté de Pharmacie, Université Paris Sud, Châtenay-Malabry, France.
authors.forename: Fariza
authors.initials: F
authors.lastname: Ammam
authors.forename: Djalal
authors.initials: D
authors.lastname: Meziane-Cherif
authors.forename: Dominique
authors.initials: D
authors.lastname: Mengin-Lecreulx
authors.forename: Didier
authors.initials: D
authors.lastname: Blanot
authors.forename: Delphine
authors.initials: D
authors.lastname: Patin
authors.forename: Ivo G
authors.initials: IG
authors.lastname: Boneca
authors.forename: Patrice
authors.initials: P
authors.lastname: Courvalin
authors.forename: Thierry
authors.initials: T
authors.lastname: Lambert
authors.forename: Thomas
authors.initials: T
authors.lastname: Candela
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001419:Bacteria; D001426:Bacterial Proteins / Q000235:genetics; D016360:Clostridioides difficile / Q000187:drug effects* / Q000235:genetics*; D004352:Drug Resistance, Microbial / Q000235:genetics; D016983:Enterococcus; D013293:Enterococcus faecalis; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D008826:Microbial Sensitivity Tests; D005810:Multigene Family; D010453:Peptide Synthases / Q000235:genetics; D014640:Vancomycin / Q000494:pharmacology*; D020713:Vancomycin Resistance
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; C417270:VanG protein, Enterococcus faecalis; D014640:Vancomycin; D010453:Peptide Synthases; C108395:VanC protein, Enterococcus