Challenging the Paradigm: Non-Canonical Exoprotease Cheating in Clinical Pseudomonas aeruginosa Isolates

Pseudomonas aeruginosa is a key model for studying social behaviors in bacteria, such as the exploitation of the public good exoprotease by social cheaters. The current paradigm holds that continuous culture of exoprotease producing individuals with protein as the sole carbon source selects for exoprotease non producers mutants with an impaired quorum-sensing regulator, LasR, which controls exoprotease expression. However, recent studies with clinical and environmental strains reveal that some isolates lacking functional LasR still produce exoproteases under the control of another quorum-sensing regulator, RhlR. Under these conditions, exoprotease-non-producers are selected, often leading to population collapses. Here, we extended this study to two clinical strains, AUS 411 and AUS 531, isolated from cystic fibrosis patients and harboring functional LasR. Surprisingly, in AUS 411, exoprotease-non-producers appeared from the first growth passages, but most lost exoprotease production only transiently, with stable non-producers isolated only in late passages. In contrast, AUS 531 slowly selected stable non-producers with limited cheating ability, which neither accumulated to high proportions nor caused population collapses. Contrary to the paradigm, these non-producers had no inactivating mutations in lasRyet were more fit than laboratory-derived lasR deletion mutants in both casein and casamino acid media. Our findings illustrate that social behavior in clinical isolates can differ markedly from that in reference strains, suggesting some P. aeruginosa strains evolve quorum-sensing networks with robust resistance to exploitation.