The complete genome sequence of the Arabidopsis and tomato pathogen Pseudomonas syringae pv. tomato DC3000

Abstract

We report the complete genome sequence of the model bacterial pathogen Pseudomonas syringae pathovar tomato DC3000 (DC3000), which is pathogenic on tomato and Arabidopsis thaliana. The DC3000 genome (6.5 megabases) contains a circular chromosome and two plasmids, which collectively encode 5,763 ORFs. We identified 298 established and putative virulence genes, including several clusters of genes encoding 31 confirmed and 19 predicted type III secretion system effector proteins. Many of the virulence genes were members of paralogous families and also were proximal to mobile elements, which collectively comprise 7% of the DC3000 genome. The bacterium possesses a large repertoire of transporters for the acquisition of nutrients, particularly sugars, as well as genes implicated in attachment to plant surfaces. Over 12% of the genes are dedicated to regulation, which may reflect the need for rapid adaptation to the diverse environments encountered during epiphytic growth and pathogenesis. Comparative analyses confirmed a high degree of similarity with two sequenced pseudomonads, Pseudomonas putida and Pseudomonas aeruginosa, yet revealed 1,159 genes unique to DC3000, of which 811 lack a known function.

Fig. 1.

Features of the DC3000 genome. (A) The chromosome, with the outermost circle depicting ORFs on the positive strand and the second circle depicting ORFs on the negative strand. The ORFs are color-coded based on the major grouping of role categories as follows: amino acid biosynthesis (salmon); biosynthesis of cofactors, prosthetic groups, and carriers (light blue); cell envelope (light green); cellular processes (red); central intermediary metabolism (brown); DNA metabolism (yellow); energy metabolism (green); fatty acid and phospholipid metabolism (purple); protein fate/synthesis (pink); purines, pyrimidines, nucleosides, and nucleotides (orange); regulatory functions (navy blue); transcription (gray); transport and binding proteins (teal); and unknown function, conserved hypothetical, and hypothetical (black). The third circle represents the ORFs with homologs present in all three Pseudomonas spp. (E < 10^-5). The fourth circle depicts the ORFs that are unique to DC3000 compared with P. aeruginosa and P. putida (E ≥ 10^-5). The fifth circle represents mobile genetic elements, including IS elements (red), phage-related genes (light green), and plasmid-related functions (blue). The sixth circle depicts ORFs associated with virulence, which are color coded as follows: adhesins and other cell surface-associated factors (dark green), miscellaneous virulence factors (yellow), TTSS effectors, candidates, and helpers (red), TTSS regulatory and secretory proteins (brown), and low molecular weight diffusible factors (blue). The seventh circle is the GC skew, and the eighth circle represents atypical nucleotides. (B) The plasmids with the outermost circle representing ORFs on the positive strand and the second circle depicting ORFs on the negative strand, with the same color coding as A. The third circle represents ORFs shared by the two plasmids (E < 10^-5). The fourth circle depicts the ORFs with homologs present in all three Pseudomonas spp. (E < 10^-5) in navy blue and the ORFs that are unique to DC3000 compared with P. aeruginosa and P. putida (E ≥ 10^-5) in green. The fifth circle represents mobile genetic elements (color-coded as in A). The sixth circle depicts ORFs associated with virulence (color-coded as in A).

Fig. 2.

Diagrammatic overview of P. syringae-plant interactions, with virulence factors color-coded as in Fig. 1. Host-pathogen factors associated with defense and disease are separately grouped on the left and right of the diagram, respectively. The central pathogenic process is the injection of multiple effector proteins into plant cells by the TTSS, which is depicted as the brown structure traversing the bacterial inner and outer membranes, plant cell wall, and plasma membrane. The effectors may suppress defenses and promote nutrient and water accumulation in the apoplast unless any one of them is detected by a resistance (R) gene-encoded sentinel, in which case strong defenses associated with the hypersensitive response (HR) are triggered. Motility favors bacterial epiphytic survival and entry into leaves, but flagellin is also recognized by means of the FLS2 receptor-like kinase as an elicitor of general defenses (62), which are not as strong as HR-associated defenses and may be suppressed by Hrp effectors. Exclusion or tolerance of reactive oxygen species (ROS) and other antimicrobials is likely promoted by extracellular polysaccharides (EPS), scavengers, and ABC exporters. Favoring virulence are type IV pili (Tfp) and possibly adhesins, coronatine synthesis and regulation, plant cell-wall-degrading enzymes (CWDE) and other variously secreted proteins, iron-scavenging siderophores (Sid), indoleacetic acid (IAA), and probably numerous ABC transporters and other nutrient uptake systems.

Fig. 3.

Detailed view of one region of the DC3000 genome that is enriched in virulence factors and mobile genetic elements. Only virulence-related genes, “hrp box” promoters, and mobile genetic elements are shown. Features are color-coded as in Fig. 1 (circle 6), with the exception that a lighter blue is used for the nonribosomal peptide synthase genes, and the mobile genetic elements are white. The genome display was generated by using artemis (www.sanger.ac.uk/Software/Artemis/) to view the chromosome (AE016853) with the aid of input files for a variety of virulence-related features that are available at http://pseudomonas-syringae.org and http://monod.cornell.edu.