Culture of extracellular protease-producing and non-producing bacterial strains for antimicrobial activity assay

To determine the specificity of the microbial proteases in triggering the respiratory proteins to produce bactericidal ROS, we compared the level of bacterial clearance of protease-producing and non-producing strains of typical Gram-negative and Gram-positive bacteria. For Gram-negative bacteria, we chose Pseudomonas aeruginosa strain PAO-Iglewski which produces PAE (the major extracellular protease virulence factor), and the PAE-knockout mutant PAO-B1A1 which does not produce PAE (1). For Gram-positive bacteria, we cultured Staphylococcus aureus PC1839 strain which produces active V8-protease, and AK3 strain which is a V8 protease-mutant (2). The genes controlling PAE in P. aeruginosa and the V8-protease in S. aureus are under the control of quorum-sensing signals. Therefore the culture conditions which facilitate the production of active extracellular proteases are utilized.