Assessment of linezolid resistance mechanisms among Staphylococcus epidermidis causing bacteraemia in Rome, Italy

Objectives To characterize linezolid resistance among blood cultured Staphylococcus epidermidis from patients at the Polyclinic Agostino Gemelli (2006–08). Isolates also showed elevated MICs of macrolide, lincosamide and streptogramin (MLS) compounds, which were investigated. Methods Ten S. epidermidis exhibiting linezolid MICs ≥ 4 mg/L were included. Isolates were screened for cfr mutations in 23S rRNA, L3, L4 and L22, and MLS genes by PCR/sequencing. Ribosomal proteins were compared with those from a linezolid-susceptible (MIC, 1 mg/L) clinical strain and ATCC 12228. cfr location was determined by Southern blot/hybridization. The cfr strain was submitted to plasmid curing. Epidemiology was assessed by PFGE and multilocus sequence typing (MLST). Results S. epidermidis displayed linezolid MICs of 4 or 8 mg/L, except for strain 4303A (MIC, 64 mg/L). These organisms and a linezolid-susceptible strain exhibited L3 Leu101Val compared with ATCC 12228. Isolates also showed L3 Phe147Leu and Ala157Arg, and L4 Asn158Ser. Strain 12375A possessed L4 Lys68Arg. Isolates were wild-type for 23S rRNA and L22. cfr was plasmid located in strain 4303A and the plasmid-cured strain exhibited a linezolid MIC (4 mg/L) similar to that for cfr-negative strains (4–8 mg/L). All organisms harboured erm(A) and msr(A), while vga(A) was detected in several isolates. All isolates were clonally related and ST-23. Conclusions L3 Phe147Leu and/or Ala157Arg appeared responsible for the elevated linezolid MIC, since adjacent alterations have been associated with resistance. L4 Asn158Ser has been reported in a linezolid-susceptible isolate and Lys68Arg detected here did not seem to provide an additive effect. Acquisition of cfr markedly increased (8- to 16-fold) the linezolid MICs. vga(A) was associated with higher MICs of quinupristin/dalfopristin and retapamulin.