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

Abstract

Two linezolid-resistant Enterococcus faecium isolates (MICs, 8 g/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 Tn6218 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.

Presence of the optrA Gene in Methicillin-Resistant Staphylococcus sciuri of Porcine Origin

“…The cfr gene was first discovered in a bovine Staphylococcus sciuri strain and was later reported in various Staphylococcus species (10)(11)(12). Recently, a cfr homologue, cfr(B), was discovered in Clostridium difficile and Enterococcus faecium (13,14). Besides resistance to oxazolidinones, the cfr gene also mediates cross-resistance to phenicols, lincosamides, pleuromutilins, and streptogramin A antibiotics (15).…”

Five-Year Summary of In Vitro Activity and Resistance Mechanisms of Linezolid against Clinically Important Gram-Positive Cocci in the United States from the LEADER Surveillance Program (2011 to 2015)

“…The clinical data have been supported by the LEADER Surveillance Program, established in 2004 to detect emerging antimicrobial resistance among Gram-positive cocci (GPC) in the United States (5). For 12 consecutive years, this program has provided yearly information regarding linezolid resistance mechanisms, including the identification of new and emerging mechanisms (6)(7)(8)(9)(10)(11).…”

“…Resistance development appeared early in clinical use in staphylococci and enterococci through ribosomal mutation in the 23S rRNA (6,13). Subsequently, an rRNA methyltransferase was identified that conferred resistance to linezolid and other antimicrobial agents (7,9,14,15; see https://doi-org .ezproxy.library.tufts.edu/10.1093/jac/dkx023). This cfr rRNA methyltransferase has the potential to mobilize, and, although it has been detected among an expanding number of GPC, it is still not the dominant mechanism of linezolid resistance among clinical GPC (5).…”

Nationwide Surveillance of Novel Oxazolidinone Resistance Gene optrA in Enterococcus Isolates in China from 2004 to 2014

“…Besides these mutations, the cfr gene, the first reported transferable oxazolidinone resistance gene, encodes a methyltransferase that modifies the 23S rRNA at position A2503 that mediates the resistance not only to linezolid but also to phenicols, lincosamides, pleuromutilins, and streptogramin A. Moreover, a cfr variant gene, designated cfr(B) (5,6), and the second transferable oxazolidinone resistance gene optrA have been identified (7).…”

“…MICs were determined by the 2-fold agar dilution method according to the recommendations of the CLSI, and the reference strain E. faecalis ATCC 29212 served as a quality control (12). Moreover, all of the optrA-positive isolates were screened for cfr and cfr(B), and isolates with linezolid resistance (MIC of Ն8 mg/ liter) were also screened for the presence of the 23S rRNA mutation and ribosomal protein (L3/L4) mutation as previously described (5,7,13).…”