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Multi-omics analysis of Spongospora subterranea spore germination and interaction with its potato host

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Balotf, S

2022 , 'Multi-omics analysis of Spongospora subterranea spore germination and interaction with its potato host', PhD thesis, University of Tasmania.

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Abstract

The obligate biotroph Spongospora subterranea is an important pathogen of potato, the world’s third most valuable food crop for human consumption. Resting spores are essential for the spread and survival of S. subterranea and can remain viable in the soil in their dormant states for decades rendering fields unsuitable for planting. In the presence of potato roots, resting spores germinate and release zoospores that subsequently infect potato roots. A better understanding of the molecular basis of resting spore germination of S. subterranea and the regulatory principles underlying Spongospora-potato interactions could be important for developing novel disease interventions. However, as a soilborne and obligate biotroph pathogen, the application of omics techniques for the detailed study of the pre- and post-infection processes in S. subterranea has been problematic.
This thesis developed a method for the partial purification of S. subterranea resting spores utilizing Ludox\(^®\) gradient centrifugation. A series of preliminary experiments were then undertaken for S. subterranea protein preparation. Protein profiles between dormant and germination stimulant-treated resting spores were compared using label-free quantitative proteomics. A deep RNA sequencing approach was then employed to analyse S. subterranea resting spores' reprogramming during the transition to zoospores in an in vitro model. Further, to expand our understanding of S. subterranea biology during infection, the transcriptome and proteome of the pathogen during the invasion of roots of a susceptible and a resistant potato cultivar was characterized in planta. Later an integrated transcriptomic and proteomic dataset was employed to uncover these mechanisms underlying S. subterranea resistance in potato roots. In relation to this, the proteome and phosphoproteome of potato leaves were also profiled to explore potato resistance mechanisms to S. subterranea at the post-transcriptome levels.
Without affecting the viability of the spores, the Ludox\(^®\) purification protocol produced a semi- purified and concentrated suspension of S. subterranea resting spores and improved protein identification by approximately 40%. The transcriptome and proteome analysis of S. subterranea spore germination introduced several candidate genes and proteins related to the germination of the pathogen, including those belonging to transcription and translation, transport, energy metabolic processes, stress response, and DNA repair. The in planta analysis of S. subterranea transcriptome and proteome indicated that transportation, metabolic processes and cytoskeletal processes were induced in the resistant cultivar. Enzyme activity and nucleic acid metabolism were decreased in this cultivar, suggesting a probable influence of these processes in the virulence of S. subterranea. The transcriptomic and proteomic datasets of potato roots infected by S. subterranea uncovered the critical role of glutathione metabolism and lignin metabolic process in potato resistance to S. subterranea. In addition, this experiment also confirmed that the inositol phosphate pathway might be related to the susceptibility of potato plants to the root disease by S. subterranea. The proteome and phosphoproteome analysis of potato leaves after infection by S. subterranea showed that oxidoreductase activity, electron transfer, and photosynthesis were significant processes that differentially changed in the proteome of resistant cultivar. The phosphoproteomics results indicated increased activity of signal transduction and defence response functions in resistant cultivars but not in the susceptible cultivar. In contrast, the majority of increased phosphoproteins in the susceptible cultivar were related to transporter activity and subcellular localisation.
This thesis provides a comprehensive overview of the changes in transcriptome and proteome during the germination of S. subterranea resting spores and extends our knowledge on the spore germination in plasmodiophorids. In planta study and potato defence response analysis contributes to the significant progress in our understanding of the interaction between the obligate biotrophic pathogens and their host plants. More than that, the study increased the availability of omics data in such a complex interaction and can allow the exploitation of this knowledge for the benefit of agriculture.

Item Type:	Thesis - PhD
Authors/Creators:	Balotf, S
Keywords:	Spongospora subterranea, resting spore, potato, powdery scab, transcriptomics, proteomics, phosphoproteomics
DOI / ID Number:	https://doi.org/10.25959/100.00047588
Copyright Information:	Copyright 2022 the author
Additional Information:	Chapter 2 appears to be the equivalent of a post-print version of an article published as: Balotf, S., Wilson, R., Tegg, R. S., Nichols, D. S., Calum W. R., 2022. Shotgun proteomics as a powerful tool for the study of the proteomes of plants, their pathogens, and plant-pathogen interactions, Proteomes10(1), 5. © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution 4.0 International (CC BY 4.0) license (https://creativecommons.org/licenses/by/4.0/). Chapter 3 appears to be the equivalent of a post-print version of an article published as: Balotf, S., Wilson, R., Tegg, R. S., Nichols, D. S., Wilson, C. R., 2020. Optimisation of sporosori purification and protein extraction techniques for the biotrophic protozoan plant pathogen Spongospora subterranea, Molecules 25(14), 3109. © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution 4.0 International (CC BY 4.0) license (https://creativecommons.org/licenses/by/4.0/). Chapter 4 appears to be the equivalent of the peer reviewed version of the following article: Balotf, S., Wilson, R., Tegg, R .S., Nichols, D. S., Wilson, C. R., 2021. Quantitative proteomics provides an insight into germination-related proteins in the obligate biotrophic plant pathogen Spongospora subterranea, Environmental microbiology reports 13(4), 521-532, which has been published in final form at https://doi.org/10.1111/1758-2229.12955. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions. This article may not be enhanced, enriched or otherwise transformed into a derivative work, without express permission from Wiley or by statutory rights under applicable legislation. Copyright notices must not be removed, obscured or modified. The article must be linked to Wiley’s version of record on Wiley Online Library and any embedding, framing or otherwise making available the article or pages thereof by third parties from platforms, services and websites other than Wiley Online Library must be prohibited. Chapter 5 appears to be the equivalent of a post-print version of an article published as: Balotf, S., Tegg, R. S., Nichols, D. S., Wilson, C. R., 2021. Spore germination of the obligate biotroph Spongospora subterranea: transcriptome analysis reveals germination associated genes, Frontiers in microbiology 12, 1557. Copyright © 2021 Balotf, Tegg, Nichols and Wilson. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International (CC BY 4.0) license (https://creativecommons.org/licenses/by/4.0/). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Chapter 6 appears to be the equivalent of a post-print version of an article published as: Balotf, S., Tegg, R. S., Nichols, D. S., Wilson, C. R., 2021. In planta transcriptome and proteome profiles of Spongospora subterranea in resistant and susceptible host environments illuminates regulatory principles underlying host-pathogen interaction, Biology 10(9), 840. © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution 4.0 International (CC BY 4.0) license (https://creativecommons.org/licenses/by/4.0/). Chapter 7 appears to be the equivalent of a pre-print version of an article published as: Balotf, S., Wilson, R., Nichols, D. S., Tegg, R. S., Wilson, C. R., 2022. Multi-omics reveals mechanisms of resistance to potato root infection by Spongospora subterranea, Scientific reports, 12, 10804. © The Author(s) 2022. This article is licensed under a Creative Commons Attribution 4.0 International (CC BY 4.0) License, (https://creativecommons.org/licenses/by/4.0/) which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. Chapter 8 appears to be the equivalent of a post-print version of an article published as: Balotf, S., Wilson, C. R., Tegg, R. S., Nichols, D. S., Wilson, R., 2022. Large-scale protein and phosphoprotein profiling to explore potato resistance mechanisms to Spongospora subterranea infection. Frontiers in plant science 13, 872901. Copyright © 2022 Balotf, Wilson, Tegg, Nichols and Wilson. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International (CC BY 4.0) license (https://creativecommons.org/licenses/by/4.0/). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
Related URLs:	Chapter 2 (published version) Chapter 3 (published version) Chapter 4 (published version) Chapter 5 (published version) Chapter 6 (published version) Chapter 7 (published version) Chapter 8 (published version)
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