Phosphorus stress and spatial confinement lower the quorum-sensing activation threshold for phenazine production in Pseudomonas synxantha

Bacteria often coordinate collective behaviors such as biofilm formation and secondary metabolite production through quorum sensing (QS), a regulatory system traditionally linked to high cell density. However, in environments such as soil, where microbial populations are spatially fragmented, sparse, nutrient-limited, and subject to mass transport, the mechanisms that enable QS-dependent processes remain incompletely understood. Here, we investigate the regulation of a secreted redox-active metabolite, phenazine-1-carboxylic acid (PCA), in Pseudomonas synxantha 2-79, a model rhizobacterium, under phosphorus (P) limitation, a persistent stress in many soils. Using a combination of microscopy and molecular genetic approaches, we show that P limitation sensitizes the QS activation threshold by an order of magnitude, enabling phenazine induction at relatively low population densities compared with P-replete conditions. This induction is abolished in QS-deficient mutants and restored by the addition of exogenous acyl-homoserine lactone (AHL), demonstrating that QS remains essential, but its threshold becomes environmentally tuned. Under P limitation, spatial confinement and pore saturation levels further shape the timing and location of induction, illustrating how physical structure and nutrient stress can modulate bacterial activities. Moreover, P stress confers both collaborative and competitive advantages, enabling P. synxantha to undergo low-cell-density AHL cross-induction with related Pseudomonas spp. and to suppress other rhizobacteria. Lastly, on plant roots, phenazine biosynthetic genes are more strongly induced under P limitation. These findings illustrate how the nutrient status of an environment can modulate the onset of QS, enabling quorum-regulated behaviors to activate at lower thresholds.