Distinct types of multicellular aggregates in Pseudomonas aeruginosa liquid cultures

ABSTRACT Pseudomonas aeruginosa forms suspended multicellular aggregates when cultured in liquid media. Such aggregates may be important in disease, and/or as a pathway to biofilm formation. The polysaccharide Psl and extracellular DNA (eDNA) have both been implicated in aggregation, but previous results depend strongly on the experimental conditions. Here we develop a quantitative microscopy-based method for assessing changes in the size distribution of suspended aggregates over time in growing cultures. For exponentially growing cultures of P. aeruginosa PAO1, we find that aggregation is mediated by cell-associated Psl, rather than by either eDNA or se-creted Psl. These aggregates arise de novo within the culture via a growth process that involves both collisions and clonal growth. They are “non-cheatable” since Psl non-producing cells do not aggregate with producers. In contrast, we find that stationary phase (overnight) cultures contain a different type of multicelullar aggregate, in which both eDNA and Psl mediate cohesion. Our findings suggest that the physical and bi-ological properties of multicellular aggregates may be very different in early-stage vs late-stage bacterial cultures.