Penicillin-Binding Proteins and Associated Protein Mutations Confer Oxacillin/Cefoxitin Tolerance in Borderline Oxacillin-ResistantStaphylococcus aureus

Among clinical isolates ofStaphylococcus aureus, borderline oxacillin-resistantS. aureus(BORSA), which is mildly resistant to oxacillin (OXA) without harboring themecAormecCgene, is considered a risk factor for further resistance against multiple antibiotics. In this study, BORSA isolates and their derivatives were characterized through antibiotic susceptibility testing and mutation analysis of the genes encoding penicillin-binding proteins (PBPs) and their related proteins, including the promoter region. Eight BORSA isolates were confirmed to harbor theblaZ gene, and hyperproduction ofblaZ-encoded penicillinase was predicted based on the minimum inhibitory concentrations (MICs). Of these, four derivative strains that were spontaneously selected based on viability on media containing high concentrations of OXA showed higher MICs than the parent isolates. The minimum bactericidal concentrations, MIC ratios, and TDtest results identified many strains with cefoxitin tolerance. Sequencing ofpbp1,pbp2,pbp3,pbp4,gdpP, andyjbH, and the promoter ofpbp4revealed mutations in BORSA isolates and derivatives, despite their absence in parent isolates, suggesting that mutations in PBPs confer OXA/cefoxitin tolerance in BORSA strains.