Denitrifying bacteria respond to and shape microscale gradients within particulate matrices.

Heterotrophic denitrification enables facultative anaerobes to continue growing even when limited by oxygen (O2) availability. Particles in particular provide physical matrices characterized by reduced O2 permeability even in well-oxygenated bulk conditions, creating microenvironments where microbial denitrifiers may proliferate. Whereas numerical particle models generally describe denitrification as a function of radius, here we provide evidence for heterogeneity of intraparticle denitrification activity due to local interactions within and among microcolonies. Pseudomonas aeruginosa cells and microcolonies act to metabolically shade each other, fostering anaerobic processes just microns from O2-saturated bulk water. Even within well-oxygenated fluid, suboxia and denitrification reproducibly developed and migrated along sharp 10 to 100 µm gradients, driven by the balance of oxidant diffusion and local respiration. Moreover, metabolic differentiation among densely packed cells is dictated by the diffusional supply of O2, leading to distinct bimodality in the distribution of nitrate and nitrite reductase expression. The initial seeding density controls the speed at which anoxia develops, and even particles seeded with few bacteria remain capable of becoming anoxic. Our empirical results capture the dynamics of denitrifier gene expression in direct association with O2 concentrations over microscale physical matrices, providing observations of the co-occurrence and spatial arrangement of aerobic and anaerobic processes.