A Study of the Regulating Gene of femA from Methicillin-resistant Staphylococcus aureus Clinical Isolates

Abstract

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. β-Lactamases were detected by nitrocephin and the presence of the mecA gene was determined by polymerase chain reaction (PCR). Only isolates that were both β-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 × 10⁻³%–29.91%, 5.54 × 10⁻³%–3.1 × 10²% and 13.88–5.50 × 10⁴%, 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-β-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.