Date of Award

Fall 10-1-2021

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Forestry and Environmental Studies

First Advisor

Brodersen, Craig

Abstract

Xylella fastidiosa (Xf) is a xylem-dwelling bacterium that causes Pierce’s disease (PD) in grapevines (Vitis vinifera L.), and disease in a range of other ecologically and economically important woody plants. To successfully colonize the xylem network, Xf cells accumulate on the vessel walls and form a biofilm. The biofilm contains cell wall-degrading enzymes, allowing the bacteria to breach the intervessel pit membranes. Thus, Xf can move from one vessel to another and colonize the xylem network. Degraded intervessel pit membranes and the production of tyloses in response to the presence of Xf likely contribute to significant declines in both hydraulic conductivity and resistance to drought-induced embolism spread. Indeed, Xf-infected grapevines typically display a range of symptoms that are often associated with water transport dysfunction.Despite the consensus that PD susceptibility is associated with Xf multiplication and systemic spread within the xylem network, there significant gaps in our understanding of the relationships between xylem structure and function that allow for Xf establishment and colonization still remain. For instance, although Xf can breach pit membranes to move from one vessel to another, the consequences of the breakdown of pit membranes within the context of embolism spread and hydraulic conductivity, and the subsequent implications for whole-plant physiological decline, remain inconclusive. Furthermore, the physical structure of the xylem network, i.e. the spatial distribution of xylem connections that might facilitate the spread of Xf, are largely unknown because of the complex, three-dimensional nature of the network. In my dissertation research, I explored the roles of the xylem structure and function related to the mechanisms of PD resistance. Throughout my work, I applied a holistic approach, coupling anatomical and physiological measurements across different grapevine genotypes with different levels of PD resistance -- from non-cultivated North American species to commercial European vinifera cultivars and their hybrids. In the first chapter, I tested the hypothesis where if the 3D structure of the xylem network connectivity plays a significant role in Xf spread, then PD resistant grapevine genotypes should have fewer total connections in the lateral and radial directions, which thereby limits the total number of pathways. Given that the Xf spread is essentially dependent on the intervessel connections, comparing the number and orientation of connections was a logical step in the fundamental understanding of this host-pathogen relationship. The chapter concludes, however, that there was limited evidence to support this hypothesis, and network connectivity does not appear to be strongly correlated with PD resistance and Xf spread. While network connectivity in the radial and lateral directions is somewhat variable within the genus Vitis, no clear trends emerged linking connectivity with resistance to PD. In the second chapter, I investigated the consequences of the extracellular cell wall-degrading enzyme released by Xf on pit membrane integrity and the downstream effects on water transport. The enzymatic breakdown of the pit membranes was relatively small, less than 10% of the pit aperture area, but enough to weaken pit membrane resistance to air-seeding by introducing pores into the membrane. Not only would larger pore diameters facilitate Xf movement, but they would subsequently increase the vulnerability of those vessels to drought-induced embolism spread. These factors would significantly affect the water transport capacity of infected grapevines and put them at greater risk to the effects of drought. In the third chapter, my objective was to determine the key physiological mechanisms that lead to mortality in the Xf infection process. This chapter reveals the mechanistic cascade of events that occur after Xf inoculation, with a coordinated decline in hydraulic conductivity, photosynthesis, and starch storage in PD susceptible grapevine genotypes. The results support the theory that hydraulic failure and carbon starvation underlie plant mortality resulting from PD. My dissertation explored the roles of the xylem structure and function on the PD mechanisms of resistance. Collectively, this work (1) identifies the variability in 3D xylem network traits in six different Vitis genotypes, representing the most complete analysis of its type for any plant group; (2) reveals that in young shoots the axial pathway appears to be the most important in determining the long-distance movement and systemic spread of Xf in the xylem network, (3) and provides a more robust, mechanistic understanding of the timing and sequence of events from initial Xf inoculation to ultimate death, as well as the variability in this mortality sequence in resistant and susceptible genotypes. As we do not have an effective remedy against the Xf bacterium, a more accurate understanding of how some grapevines resist to the infection process is one piece of this very important puzzle.

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