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Modeling Pseudomonas aeruginosa pathogenesis in plant hosts

Nature Protocols volume 4, pages 117–124 (2009)Cite this article

Abstract

A pathogenic model in which both the pathogen and its host are amenable to genetic manipulation can greatly facilitate the understanding of bacterial pathogenesis. Plants are genetically tractable and can be used as experimental models for human microbial pathogenesis. We present protocols for both lettuce and Arabidopsis leaf infection models using the opportunistic human bacterial pathogen, Pseudomonas aeruginosa. The lettuce model allows for high-throughput qualitative analysis of virulence and is suitable for screening large numbers of bacterial strains, whereas the Arabidopsis model provides a quantitative approach and permits the tracking of bacterial cell proliferation in planta. The lettuce model takes ∼24 h including bacterial growth using store-bought lettuce, and the Arabidopsis model takes 4–6 weeks to grow the plants and a similar time as with lettuce to infect the plants. Both models are monitored for up to 5 d post-infection. These methodologies can and have been used to identify novel and critical P. aeruginosa pathogenicity agents, as virulence factors are often conserved across phylogeny.

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Figure 1
Figure 2
Figure 3
Figure 4: The amount of cells in the inoculum can impact disease progression in Arabidopsis plants.
Figure 5: Inoculate the underside of each leaf by gently pressing the syringe against your thumb.
Figure 6
Figure 7: When inoculating lettuce midribs, avoid penetrating the middle space of the plant.
Figure 8: Infected lettuce leaves show severe rotting (black coloration) of the entire midrib when infected with the wild-type strain PA14.

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Acknowledgements

This work was partially supported by the research grant R01AI063433. M.S. was supported by the Shriners Research Fellowship no. 8506. Thank you to Jenifer Bush for growing and maintaining our Arabidopsis plants.

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Authors and Affiliations

  1. Department of Surgery at Massachusetts General Hospital, 50 Blossom St., Boston, 02114, Massachusetts, USA

    Melissa Starkey & Laurence G Rahme

  2. Department of Surgery at Harvard Medical School, 200 Longwood Ave., Boston, 02115, MA, USA

    Melissa Starkey & Laurence G Rahme

  3. Shriners Burns Institute, 51 Blossom St., Boston, 02114, Massachusetts, USA

    Melissa Starkey & Laurence G Rahme

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Correspondence to Laurence G Rahme.

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Starkey, M., Rahme, L. Modeling Pseudomonas aeruginosa pathogenesis in plant hosts. Nat Protoc 4, 117–124 (2009). https://doi.org/10.1038/nprot.2008.224

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