title: Quantitation of mecA transcription in oxacillin-resistant Staphylococcus aureus clinical isolates.
abstract: The transcription of mecA, the gene required for oxacillin resistance in staphylococci, was quantified in a collection of 65 geographically and genetically diverse clinical and 8 defined laboratory Staphylococcus aureus isolates. mecA transcription was measured by real-time reverse transcription-PCR, confirmed by Northern blot analysis, and correlated with the presence and DNA sequence of the two mecA repressors, mecI and blaI. Isolates were first examined that contained mecI and/or blaI with wild-type sequence. BlaI provided significantly more repression of mecA transcription than did MecI, unrelated to blaI genetic location. Both together repressed mecA better than either one alone. In clinical isolates containing only wild-type mecI, mecA transcription repression was 10- to 25-fold less effective than that seen in previously studied constructs derived from strain N315. There was a difference in the mecI ribosomal binding site (RBS) between the clinical isolates (GGAA) and N315 (GGAG). The GGAA RBS was associated with 5.5- to 7.3-fold less mecA repression than GGAG in isogenic constructs. The values generated for wild-type repressors were compared to those in 26 isolates containing mecI mutations. mecA transcription appeared to be repressed only by BlaI in isolates with mecI nonsense and frameshift mutations. In contrast, mecI repression seemed to be partially or fully retained in many of the isolates with mecI and one isolate with blaI missense mutations, providing structure-function correlates with the site and type of mutation. We conclude that mecA repressor activity is highly variable in clinical S. aureus isolates due to mecI mutations, RBS polymorphisms, and unidentified genomic adaptations.
authors.affiliation: Department of Medicine, Medical College of Virginia Campus of Virginia Commonwealth University, Richmond, Virginia 23298-0049, USA.
authors.forename: Adriana E
authors.initials: AE
authors.lastname: Rosato
authors.forename: William A
authors.initials: WA
authors.lastname: Craig
authors.forename: Gordon L
authors.initials: GL
authors.lastname: Archer
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006602:Hexosyltransferases; D006801:Humans; D008969:Molecular Sequence Data; D002267:Muramoylpentapeptide Carboxypeptidase / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010068:Oxacillin / Q000494:pharmacology*; D010403:Penicillin Resistance / Q000235:genetics; D046915:Penicillin-Binding Proteins; D010406:Penicillins / Q000494:pharmacology; D010458:Peptidyl Transferases; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013203:Staphylococcal Infections / Q000382:microbiology*; D013211:Staphylococcus aureus / Q000187:drug effects* / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; C101640:MecI protein, Staphylococcus; D046915:Penicillin-Binding Proteins; D010406:Penicillins; D012097:Repressor Proteins; C101641:mecR1 protein, Staphylococcus aureus; D010458:Peptidyl Transferases; D006602:Hexosyltransferases; D002267:Muramoylpentapeptide Carboxypeptidase; D010068:Oxacillin