In vitro biofilm formation by a beneficial bacterium partially predicts in planta protection against rhizosphere pathogens

Plant roots form associations with beneficial and pathogenic soil microorganisms. Although members of the rhizosphere microbiome can protect against pathogens, the mechanisms are poorly understood. We hypothesized that the ability to form a biofilm on the root surface is necessary for the exclusion of pathogens; however, it is not known if the same biofilm formation components required in vitro are necessary in vivo. Pseudomonas brassicacearum WCS365 is a beneficial strain that is phylogenetically closely related to an opportunistic pathogen Pseudomonas sp. N2C3 and confers protection against N2C3 in the rhizosphere. We used this plant-mutualist-pathogen model to screen collections of P. brassicacearum WCS365 increased attachment mutants (iam) and surface attachment defective (sad) transposon insertion mutants that form increased or decreased biofilm on abiotic surfaces, respectively. We found that whereas the P. brassicacearum WCS365 mutants had altered biofilm formation in vitro, only a subset of these mutants lost protection against N2C3. Non-protective mutants those involved in large adhesion protein (LapA) biosynthesis, flagellar synthesis and function, and O-antigen biosynthesis. We found that the inability of P. brassicacearum WCS365 mutants to grow in planta, and the inability to suppress pathogen growth, both partially contributed to loss of plant protection. We did not find a correlation between the extent of biofilm formed in vitro and pathogen protection in planta indicating that biofilm formation on abiotic surfaces may not fully predict pathogen exclusion in planta. Collectively, our work provides insights into mechanisms of biofilm formation and host colonization that shape the outcomes of host-microbe-pathogen interactions.