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.

Fluconazole-resistant Candida glabrata is an emerging pathogen that causes fungemia. Polymyxin B, a last-resort antibiotic used to treat multidrug-resistant Gram-negative bacterial infections, has been found to possess in vitro fungicidal activity and showed synergy with fluconazole against a single strain of C. glabrata. Since both agents may be used simultaneously in intensive care unit (ICU) patients, this study was performed to test for possible synergy of this combination against 35 C. glabrata blood isolates, using 2 methods: a time-kill assay and an experimental MIC-MIC Etest method. Thirty-five genetically unique C. glabrata bloodstream isolates were collected from 2009 to 2011, identified using an API 20C system, and genotyped by repetitive sequence-based PCR (rep-PCR). MICs were determined by Etest and broth microdilution methods. Synergy testing was performed using a modified bacterial Etest synergy method and time-kill assay, with final results read at 24 h. The Etest method showed synergy against 19/35 (54%) isolates; the time-kill assay showed synergy against 21/35 (60%) isolates. Isolates not showing drug synergy had an indifferent status. Concordance between methods was 60%. In vitro synergy of polymyxin B and fluconazole against the majority of C. glabrata isolates was demonstrated by both methods. The bacterial Etest synergy method adapted well when used with C. glabrata. Etest was easier to perform than time-kill assay and may be found to be an acceptable alternative to time-kill assay with antifungals. Candida glabrata has been a rising cause of candidemia in the past few years, second only to Candida albicans (1). According to the Centers for Disease Control and Prevention (CDC), candidemia is the fourth most common hospital-acquired infection in the United States (2). C. glabrata has been known to develop resistance to the azole family of antifungals, the most inexpensive and easily accessible medications to treat candidemia. Further data from the CDC have shown that 7% of Candida species have become fluconazole resistant (2). This development of resistance is most likely due to the routine use of azoles in treatment or prophylaxis. Other treatment options for Candida infections include amphotericin B and the echinocandin family (to which resistance by C. glabrata has occurred) (1). A U.S. surveillance study evaluating Candida isolates from San Francisco, CA, Atlanta, GA, Baltimore, MD, and Connecticut (from 1998 to 2010) determined that C. glabrata isolates are able to adapt to new environments to increase their fitness and antifungal resistance (3). Another surveillance study, evaluating developing resistance in fluconazolesusceptible isolates collected from 2008 to 2011, showed that among 2,329 Candida isolates, the prevalence of azole resistance was unchanged (7%), 32 of the isolates were echinocandin resistant, and 8 C. glabrata isolates were resistant to both fluconazole and echinocandins (4).Polymyxin B is a last-resort antibiotic used to treat multidrug-resistant Gram-negative bacte...

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