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1057 UNDERESTIMATION OF S. AUREUS TOLERANCE

Recent work indicates that antibiotic tolerance among S. aureus strains may be of clinical importance. The percentage of strains found to be tolerant varies greatly in published reports. In addition, the percentage of the bacteria within the tolerant strains has also differed greatly (from 0.1% to nearly 50%). The different techniques used by individual investigators may account for these discrepancies. We report here that a major cause of underestimating tolerance is the carryover of antibiotic from the tubes used to determine the minimal inhibitory concentration (MIC) to the blood agar subculture plates that are used to identify and count the surviving organisms.We consider a strain to be tolerant if more than 0.1% of its bacterial population survives 18 hours in 32 times the concentration of the MIC and is able to grow when subcultured onto a blood agar plate. It has been assumed that spreading the incubated antibiotic-containing broth culture onto agar plates removed the inhibitory effect of the drug. However, when we tested 15 S. aureus strains by the standard method only 5 qualified as tolerant. In contrast, when the inhibitory effect of the antibiotic was removed by twentyfold dilution before plating, 5 additional strains were found to be tolerant. Also, in all 15 strains tested a greater number of bacteria survived at 32 times the MIC than occurred with the standard method.