A study of the regulating gene of femA from methicillin-resistant Staphylococcus aureus clinical isolates.
The regulating gene of femA was studied in methicillin-resistant Staphylococcus aureus (MRSA). High-level MRSA, low-level MRSA and methicillin-sensitive S. aureus (MSSA) were identified by agar diffusion. Beta-lactamases were detected by nitrocephin and the presence of the mecA gene was determined by polymerase chain reaction (PCR). Only isolates that were both beta-lactamase-negative and mecA-positive were used. The femA gene and its 250 base pair (bp) upstream sequence were amplified by PCR and expression was determined by real-time fluorescent quantitative PCR. The 250 bp upstream sequence was labelled by BrightStar Psoralen-Biotin and detected by electrophoretic mobility shift assay (EMSA). Expression levels of femA in MSSA, low-level MRSA and high-level MRSA were 3.53 x 10(-3)% - 29.91%, 5.54 x 10(-3)% - 3.1 x 10(2)% and 13.88 - 5.50 x 10(4)%, respectively. EMSA detected a signal shift in 57 high-level MRSA isolates but not in four low-level MRSA and four MSSA strains. Expression of femA in high-level MRSA (non-beta-lactamase-producing) was higher than in low-level MRSA and MSSA. The femA regulating gene probably lies in the 250 bp upstream sequence in MRSA and high-level expression is essential for high-level methicillin resistance.
Li
X
X
Department of Infectious Diseases, West China Hospital, Sichuan University, Chengdu, China.
Xiong
Y
Y
Fan
X
X
Zhong
Z
Z
Feng
P
P
Tang
H
H
Zhou
T
T
D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D004587:Electrophoresis, Agar Gel; D024202:Electrophoretic Mobility Shift Assay; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D016106:Methicillin Resistance / Q000235:genetics*; D008826:Microbial Sensitivity Tests; D008957:Models, Genetic; D008969:Molecular Sequence Data; D046915:Penicillin-Binding Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D013211:Staphylococcus aureus / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics* / Q000302:isolation & purification; D001618:beta-Lactamases / Q000378:metabolism
D001426:Bacterial Proteins; C096686:FemA protein, Bacteria; D046915:Penicillin-Binding Proteins; C487789:mecA protein, Staphylococcus aureus; D001618:beta-Lactamases