Molecular and evolutionary basis of O-antigenic polysaccharide-driven phage sensitivity in environmental pseudomonads

Pseudomonas protegens CHA0, a bacterial strain able to suppress plant pathogens as well as efficiently kill lepidopteran pest insects, has been studied as a biocontrol agent to prevent ensuing agricultural damage. However, the success of this method is dependent on efficient plant colonization by the bacterial inoculant, while it faces competition from the resident microbiota as well as predators such as bacteriophages. One of these naturally occurring phages, Phi GP100, was found to drastically reduce the abundance of CHA0 once inoculated into plant microcosms, resulting in the loss of plant protection effect against a phytopathogen. Here, we investigated the molecular determinants implicated in the interaction between CHA0 and the phage Phi GP100 using a high-density transposon-sequencing approach. We show that lipopolysaccharide cell surface decorations, specifically the longer OBC3-type O-antigenic polysaccharide (O-PS, O-antigen) of the two dominant O-PS of CHA0, are essential for the attachment and infection of Phi GP100. Moreover, when exploring the distribution of the OBC3 cluster in bacterial genomes, we identified several parts of this gene cluster that are conserved in phylogenetically distant bacteria. Through heterologous complementation, we integrated an OBC3-type gene copy from a phylogenetically distant bacterium and were able to restore the phage sensitivity of a CHA0 mutant which lacked the ability to form long O-PS. Finally, we evidence that the OBC3 gene cluster of CHA0 displays a high genomic plasticity and likely underwent several horizontal acquisitions and genomic rearrangements. Collectively, this study underlines the complexity of phage-bacteria interactions and the multifunctional aspect of bacterial cell surface decorations.