A new method to determine postantibiotic effect and effects of subinhibitory antibiotic concentrations

It has been shown that bacteria in a postantibiotic (PA) phase exposed to subinhibitory concentrations (sub-MICs) of antibiotics show a long delay before regrowth. This effect has been named the PA sub-MIC effect (PA SME). In the present study, we have used a new method to demonstrate this phenomenon. A computerized incubator for bacteria, Bioscreen C (Lab Systems, Helsinki, Finland), which incubates the bacteria, measures growth continuously by vertical photometry, processes the data, and provides a printout of the results was used. With this method, one may easily test several antibiotics against different bacteria for PA effects (PAEs), PA SMEs, and SMEs. In this study, the effects of benzylpenicillin against beta-hemolytic streptococci and pneumococci were examined. The bacteria were exposed to 2, 10, or 50x MIC for 2 h, washed and diluted, incubated in the Bioscreen C incubator, and then exposed to 0.1 to 0.9x MIC. The regrowth was monitored for 20 h. The PAE was calculated as the difference in the time required for the exposed and unexposed bacteria to grow to a defined point (A50) on the absorbance curve. A50 was defined as 50% of the maximum absorbance for the control cultures. The PA SMEs were calculated as the difference in the time required for the reexposed cultures and the unexposed controls to reach A50. The PAEs ranged between 0.6 and 3.2 h and varied little with the concentration used for the induction of the PAEs. At 0.2x MIC, the PA SMEs were 2 to 3 h longer than the PAEs. Higher sub-MICs increased this delay before regrowth. Most cultures exposed to sub-MICs alone were only slightly affected compared with the controls.