Effect of storage and changes in bacterial growth phase and antibiotic concentrations on antimicrobial tolerance in Staphylococcus aureus

Forty clinical isolates of Staphylococcus aureus were tested for tolerance to oxacillin and cephalothin by broth dilution susceptibility tests and killing curves. Most experiments were carried out with stationary-phase inocula, but nine tolerant isolates were retested with log-phase inocula. All 40 isolates were retested in killing curves at double the antibiotic concentrations used in initial tests. Isolates were retested for tolerance to oxacillin after storage at -70°C for 1 year. In broth dilution tests, 23 of 40 (57.5%) and 20 of 40 (50%) isolates were tolerant to oxacillin and cephalothin, respectively. By killing curves, 25 of 40 (62.5%) and 22 of 40 (55%) isolates were tolerant to oxacillin and cephalothin, respectively. When nine tolerant isolates were retested with log-phase inocula, none manifested tolerance. Only 25 to 30% of the isolates were tolerant in killing curves performed with oxacillin and cephalothin at concentrations double those used in initial tests. After storage at -70°C for 1 year, only two-thirds of the isolates remained tolerant. In isolates that remained tolerant, the degree of tolerance diminished to about 25% of that observed in initial tests.