High‐Throughput Assessment of Bacterial Growth Inhibition by Optical Density Measurements

The increasing incidence of antibiotic‐resistant bacterial infections both in hospitals and in the community intensifies the need for new antibacterial strategies and targets. Although high‐throughput screening against live bacteria allows rapid discovery of compounds with growth‐inhibitory activities, these efforts have failed to fill the pipeline with the anticipated antibacterial compounds because target identification is often onerous. Recently, a strategy was reported that employs a bacterial growth inhibition assay readout using optical density measurements on paired strains—both a wild‐type strain and a pathway‐null mutant—to find inhibitors of wild‐type bacterial growth that specifically target conditionally essential enzymes in the pathway of interest. Protocols are provided here for determining the robustness of an assay, screening in a high‐throughput format, and setting up dose‐response curves in paired Staphylococcus aureus strains. However, the protocols can be used to screen for growth‐inhibitory compounds in any bacterial strain of interest. Curr. Protoc. Chem. Biol . 2:195‐208 © 2010 by John Wiley & Sons, Inc.