Detection of a New cfr -Like Gene, cfr (B), in Enterococcus faecium Isolates Recovered from Human Specimens in the United States as Part of the SENTRY Antimicrobial Surveillance Program

Two linezolid-resistant Enterococcus faecium isolates (MICs, 8 μg/ml) from unique patients of a medical center in New Orleans were included in this study. Isolates were initially investigated for the presence of mutations in the V domain of 23S rRNA genes and L3, L4, and L22 ribosomal proteins, as well as cfr . Isolates were subjected to pulsed-field gel electrophoresis (just one band difference), and one representative strain was submitted to whole-genome sequencing. Gene location was also determined by hybridization, and cfr genes were cloned and expressed in a Staphylococcus aureus background. The two isolates had one out of six 23S rRNA alleles mutated (G2576T), had wild-type L3, L4, and L22 sequences, and were positive for a cfr -like gene. The sequence of the protein encoded by the cfr -like gene was most similar (99.7%) to that found in Peptoclostridium difficile , which shared only 74.9% amino acid identity with the proteins encoded by genes previously identified in staphylococci and non- faecium enterococci and was, therefore, denominated Cfr(B). When expressed in S. aureus , the protein conferred a resistance profile similar to that of Cfr. Two copies of cfr (B) were chromosomally located and embedded in a Tn 6218 similar to the cfr -carrying transposon described in P. difficile. This study reports the first detection of cfr genes in E. faecium clinical isolates in the United States and characterization of a new cfr variant, cfr (B). cfr (B) has been observed in mobile genetic elements in E. faecium and P. difficile , suggesting potential for dissemination. However, further analysis is necessary to access the resistance levels conferred by cfr (B) when expressed in enterococci.