Real time monitoring of the interactions between Pseudomonas syringae pv actinidiae and Actinidia species.

To elucidate the infection process and the colonization of plant tissues by P.syringae pv actinidiae, different strains of the bacterium were transformed with a stable and broad-host-range plasmid vector (pDSK-GFPuv) that strongly expresses the GFPuv protein. In comparison with the wild-type GFP, GFPuv produces 45-fold brighter green fluorescence in bacterial cells, while it retains the same excitation and emission maxima. In addition, GFPuv is a more soluble protein and does not produce, like wtGFP, nonfluorescent inclusion bodies. Finally, GFPuv has lower toxicity to bacteria than wtGFP. For these reasons, the use of GFPuv may potentially allow the direct observation of the bacterial colonization directly with naked eye under long-wavelength UV light (395 nm). The plasmid stability and the possible effect on P. syrinage actinidiae growth and virulence were tested both in vitro and in vivo. Successively, the use colonization of the plant tissue was monitored in planta on intact, viable plant tissues without any kind of staining of the specimens. The described methodology allows a non invasive observation of the plant-pathogen interaction both at the cell and whole plant level. Therefore, it may also be applied for investigating the influence of agricultural and phytosanitary practices on the host susceptibility and disease development