Single-cell phenotypic heterogeneity shapes quorum signaling dynamics in Pseudomonas aeruginosa

Quorum sensing (QS) coordinates collective bacterial behavior, yet how individuality arises within a system built to unify action remains unclear. Using imaging-transcriptomics, we profile the QS response of Pseudomonas aeruginosa at single-cell resolution and uncover heterogeneity across all stages of QS. We find that most cells cooperate, but their contributions vary widely due to transcriptional noise. In contrast, expression of QS signal synthases, particularly in the Las and PQS systems, shows extreme cell-cell variability indicative of active differentiation. We show that cellular memory from prior growth cycles shapes QS dynamics by generating signaling-primed cells, yet it does not influence the de novo formation of hypersignaling cells. This differentiation is robust to exogenous autoinducers and conserved across diverse lab and clinical isolates. Together, our findings reveal a deliberately heterogeneous entry point embedded within an otherwise synchronizing program, shedding light on how individuality shapes cooperation and conflict in bacterial populations.