Chemical ecology and physiology underlying Spongospora subterranea infection of potato : towards manipulating rhizosphere ecology, physiology, and inoculum load for sustainable management of potato powdery scab disease

Spongospora subterranea infection diminishes the quality and quantity of harvested potatoes wherever they are grown around the world. Disease control is largely ineffective, at least partially due to the abundance of the highly resistant and long-lived pathogen inoculum within cropping soils present as agglomerations of resting spores. Resting spore germination is induced by root exudate metabolites in a staggered manner over multiple years to release short-lived zoospores that facilitate infection of host plant roots. Premature germination of resting spores in absence of host plants has been suggested to reduce pathogen soil inoculum. Methods to synchronize germination of dormant spores that could be applied within a commercial field setting are however lacking. Movement of zoospores to host roots has been speculated to occur via chemical attraction processes (chemotaxis) but evidence for this has been lacking. Knowledge of zoospore attractants and/or repellents, and the communication channels that facilitate perception of these compounds may be useful for the development of novel disease control. This thesis developed a microcapillary assay to investigate the role of individual potato metabolites in the attraction or inhibition of S. subterranea zoospores. In relation to this, the effect of solution pH, pH buffer composition and compound concentration on zoospore chemotaxis were assessed. Zoospore attraction or inhibition towards a metabolite was determined by the number of zoospores that swam into a cavitated microcapillary containing the test compound. The potato metabolome was found to contain both zoospore attractants and inhibitors. Glutamine was the strongest attractant, whilst spermine was the strongest inhibitor. Strongly acidic and alkaline medium with pH < 5.3 and > 8.5 respectively were found inhibitory towards zoospore motility and chemotaxis. Total swimming distance was found to be the most important swimming parameter that determined zoospore chemotaxis. The role of Ca2\\(^+\\) signalling in transducing signals from these organic compounds across the cell membrane to stimulate or inhibit chemotaxis of the zoospores was also investigated. This study used a combination of glass slide assay, cavitated microcapillary assay, hydroponics set up and video microscopy to investigate the effect of Ca2\\(^+\\) antagonism on zoospore chemotaxis, attachment, root infection, and swimming patterns. Zoospore swimming patterns were altered, whilst chemotaxis, root attachment and zoosporangia root infection were constrained by inhibition of cytosolic Ca2\\(^+\\) fluxes with Ca2\\(^+\\) antagonist. LaCl3 and GdCl3 at concentrations ‚Äöv¢‚Ä¢ 50 ¬¨¬µM showed complete inhibition of zoospore chemotaxis, root attachment and zoosporangia root infection. High concentration of extracellular Ca2\\(^+\\) (‚Äöv¢‚Ä¢18 mM Ca(OH)\\(_2\\)) had similar chemotactic and motility impairments effects as the Ca2\\(^+\\) channel blockers. Helical swimming patterns observed in the control were always associated with high speed, acceleration, longer distance, high chemotaxis, and disease intensity. The further the deviation from this pattern with Ca2\\(^+\\) antagonist treatments the slower the speed, acceleration, distance travelled, and the lower the chemotaxis and disease. In a series of field and pot trials, the effects of pre-plant Fe-EDTA, fluazinam and their combination treatments on pathogen inoculum load and disease reduction were studied. Pathogen DNA load and S. subterranea disease intensities and prevalence in tomato and potato were tracked over time to ascertain treatment effect. Soil treatments appeared to have a beneficial effect on pathogen inoculum persistence and disease, with disease incidence diminished to a greater extent than detectable inoculum load. For example, fluazinam (4.5 kg/ha) applied four times a year reduced root infection, galling, powdery scab intensity and prevalence by over 70% but reduced inoculum load by only 11%. Similarly, Fe-EDTA had a positive, but marginal effect on disease control but was relatively ineffective at inoculum load reduction. This study provides key insights into the chemical ecology required for S. subterranea zoospore motility and chemoattraction and identifies potential novel breeding targets for combating Spongospora diseases. It also highlights the role of Ca2\\(^+\\) signalling in host:pathogen interactions and shows how disruption of Ca2\\(^+\\) signal transduction could control disease. Further studies into inoculum load reduction with more active and stable compounds could prove useful in the remediation of S. subterranea contaminated fields.