Endopathogenic lifestyle of Pseudomonas savastanoi pv. savastanoi in olive knots.
Bottom Line: The endophytic phase of Pseudomonas savastanoi pv. savastanoi in olive stems and the structural and ultrastructural histogenesis of olive knots have been studied.Hypertrophy of the stem tissue was concomitant with the formation of bacterial aggregates, microcolonies and multilayer biofilms, over the cell surfaces and the interior of plasmolysed cells facing the air-tissue interface of internal opened fissures, and was followed by invasion of the outer layers of the hypertrophied tissue.This is the first real-time monitoring of P. savastanoi disease development and the first illustrated description of the ultrastructure of P. savastanoi-induced knots.
Affiliation: Departamento de Biología Celular, Genética y Fisiología, Universidad de Málaga, Campus de Teatinos s/n, E-29071, Málaga, Spain.Show MeSH
Related in: MedlinePlus
Mentions: In order to determine whether knots induced by P. savastanoi pv. savastanoi NCPPB 3335‐GFP on in vitro olive plants exhibited similar histological structures to those previously reported for older olive plants, semi‐thin and fixed‐frozen transverse sections of knots collected at different times after pathogen inoculation (for details see Fig. 4) were stained with toluidine blue or methylene blue‐picrofuchsin, respectively, and visualized by light microscopy. As previously reported for olive knots developed under natural conditions, a panoramic view of knot sections stained with toluidine blue clearly showed two structurally different regions: a vascular cylinder similar to that of non‐infected stems, composed of the pith parenchyma, xylem, cambium, phloem and epidermis; and expanding from this area, a hypertrophied parenchymatic tissue (Fig. 4A). Hypertrophied tissue was predominantly made up of disorganized cells and showed internal open fissures surrounded by plasmolysed cells and clusters of primary cell walls stained in intense dark blue (Fig. 4A and B). A closer view of this area also showed the existence of longitudinal xylem vessels running out of the stem vascular system towards the hypertrophied knot tissue (Fig. 4B). In addition, magnification of the knot outer layers allowed visualization of both hypertrophic and hyperplasic cells, which frequently showed irregular thickenings of the primary wall. Hyperplasic cells were clearly distinguishable by the formation of a slim middle lamella separating two nuclei (Fig. 4C). Knot sections stained with methylene blue‐picrofuchsin showed newly formed bundles of spiral xylem vessels inside the hypertrophied area (Fig. 4D and E). Interestingly, a connection between the primary vascular cylinder and newly formed spiral vessels was clearly observed in longitudinal sections of knots (Fig. 4F and G). Identical results were obtained from sections of knots collected from olive plants inoculated with the wild‐type strain NCPPB 3335 (data not shown).
Affiliation: Departamento de Biología Celular, Genética y Fisiología, Universidad de Málaga, Campus de Teatinos s/n, E-29071, Málaga, Spain.