Living on the edge: emergence of spontaneous gac mutations in Pseudomonas protegens during swarming motility

Swarming motility is a flagella‐driven multicellular behaviour that allows bacteria to colonize new niches and escape competition. Here, we investigated the evolution of specific mutations in the GacS/GacA two‐component regulatory system in swarming colonies of Pseudomonas protegens Pf‐5. Experimental evolution assays showed that repeated rounds of swarming by wildtype Pf‐5 drives the accumulation of gacS/gacA spontaneous mutants on the swarming edge. These mutants cannot swarm on their own because they lack production of the biosurfactant orfamide A, but they do co‐swarm with orfamide‐producing wildtype Pf‐5. These co‐swarming assays further demonstrated that ΔgacA mutant cells indeed predominate on the edge and that initial ΔgacA:wildtype Pf‐5 ratios of at least 2:1 lead to a collapse of the swarming colony. Subsequent whole‐genome transcriptome analyses revealed that genes associated with motility, resource acquisition, chemotaxis and efflux were significantly upregulated in ΔgacA mutant on swarming medium. Moreover, transmission electron microscopy showed that ΔgacA mutant cells were longer and more flagellated than wildtype cells, which may explain their predominance on the swarming edge. We postulate that adaptive evolution through point mutations is a common feature of range‐expanding microbial populations and that the putative fitness benefits of these mutations during dispersal of bacteria into new territories are frequency‐dependent.