title: A cfr-positive clinical staphylococcal isolate from India with multiple mechanisms of linezolid-resistance.
abstract: OBJECTIVE
  Linezolid, a member of the oxazolidinone class of antibiotics, has been an effective therapeutic option to treat severe infections caused by multidrug resistant Gram positive bacteria. Emergence of linezolid resistant clinical strains is a serious issue in the healthcare settings worldwide. We report here the molecular characterization of a linezolid resistant clinical isolate of Staphylococcus haemolyticus from India.
  
  
  METHODS
  The species of the clinical isolate was identified by 16S rRNA gene sequencing. The minimum inhibitory concentrations (MICs) of linezolid, clindamycin, chloramphenicol and oxacillin were determined by E-test method. To elucidate the mechanism of linezolid-resistance, presence of cfr gene (chloramphenicol florfenicol resistance) and mutations in 23S rRNA and ribosomal proteins (L3, L4 and L22) were investigated. Staphylococcal Cassette Chromosome mec (SCCmec) typing was performed by multiplex PCR.
  
  
  RESULTS
  The study documented a rare clinical S. haemolyticus strain with three independent mechanisms of linezolid-resistance. The strain carried cfr gene, the only known transmissible mechanism of linezolid-resistance. The strain also possessed resistance-conferring mutations such as G 2576 T in domain V of 23S rRNA gene and Met 156 Thr in L3 ribosomal protein. The other ribosomal proteins (L4 and L22) did not exhibit mutations accountable for linezolid-resistance. Restriction digestion by NheI revealed that all the alleles of 23S rRNA gene were mutated. The isolate showed elevated MIC values (>256 μg ml -[1] of linezolid, clindamycin, chloramphenicol and oxacillin. Methicillin resistance was conferred by type I SCCmec element. The strain also harboured lsa(B) gene which encodes an ABC transporter that can efflux clindamycin.
  
  
  CONCLUSIONS
  The present study reports the first clinical strain from India with transmissible and multiple mechanisms of linezolid-resistance. Judicious use of linezolid in clinical practice and proper surveillance of cfr-positive strains are of utmost importance to safeguard the efficacy of linezolid.
authors.forename: Vineeth
authors.initials: V
authors.lastname: Rajan
authors.forename: Vijay Gowdara Shankarappa
authors.initials: VG
authors.lastname: Kumar
authors.affiliation: Department of Studies in Microbiology, University of Mysore, Mysore, India.
authors.forename: Shubha
authors.initials: S
authors.lastname: Gopal
mesh_terms: D000081:Acetamides; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D000069349:Linezolid; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D023303:Oxazolidinones; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012338:RNA, Ribosomal, 23S / Q000235:genetics; D017422:Sequence Analysis, DNA; D041162:Staphylococcus haemolyticus / Q000235:genetics*
chemical_list: D000081:Acetamides; D001426:Bacterial Proteins; D017931:DNA Primers; D023303:Oxazolidinones; D012336:RNA, Ribosomal, 16S; D012338:RNA, Ribosomal, 23S; D000069349:Linezolid