Evolutionary genomics of Pseudomonas syringae plasmids

Plasmids are accessory DNA molecules that have been shown to enhance the fitness and adaptability of bacterial pathogens. To understand the processes driving plasmid evolution, I utilized a comparative genomics approach to examine the plasmids of the "Group 5" strains of the phytopathogenic bacterium Pseudomonas syringae. The Group 5 strains include three pathogens of Raphanus sativus (radish) and a single pathogen of Coriandrum sativum (cilantro/coriander), providing two host plants and an appropriate phylogenetic structure to assess the evolution of plasmids and their impact on host-specific virulence. To initiate comparative analyses, the five-plasmid complement of the radish pathogen P. syringae pv. maculicola ES4326 was sequenced and analyzed, revealing a predominant role for insertion sequence (IS) elements in driving plasmid evolution. This analysis also led to the identification of a new miniature transposable element that appeared to be predominantly virulence-associated and plasmid localized. I examined the role of IS elements further by comparing the three pPMA4326B-like plasmids from the radish pathogens. This analysis not only reinforced the importance of IS elements in plasmid evolution, but also revealed that this plasmid has undergone gradual deletion of the gene cluster responsible for horizontal transfer. I explored the possibility that this switch to vertical transmission was due to the presence of two virulence-associated type III secreted effectors (T3SEs), which may significantly enhance fitness and maintain the plasmid stably. One of the T3SEs, hopW1-2 confers a significant growth advantage on radish, but reduces the ability of the cilantro pathogen to grow in cilantro, suggesting the pPMA4326B-like plasmid may have once been present in the cilantro lineage but was subsequently lost. Although only one T3SE confers a measurable growth advantage, the two pPMA4326B-borne T3SEs share high sequence identity, suggesting they were derived from the same ancestral sequence. This led to the identification of a new evolutionary process, terminal reassortment, which involves the shuffling of existing T3SEs leading to the formation of gene fusions that results in chimeric T3SEs with new virulence function. As labile genomes, plasmids are sources and sinks for genetic variation, and have the capacity to significantly impact the evolution of their bacterial hosts.