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Studies on the biological activity of chitosans and their mode of action in plant protection

Sowohl konventionelle als auch ökologische Landwirtschaft werden noch immer vom Einsatz umweltschädlicher und oft giftiger Pestizide bestimmt. Chitosane sind Zuckermoleküle, die einerseits aktiv die Resistenz von Pflanzen gegenüber Krankheiten erhöhen, andererseits Pflanzenschädlinge direkt angreifen können. Da Chitosane unter anderem aus Krabbenschalen oder Insektenpanzern gewonnen werden können, sind sie eine erneuerbare und nachhaltige Ressource und stellen eine attraktive Alternative zum chemisch geprägten Pflanzenschutz dar. In dieser Arbeit wurde die Eignung von Chitosan als Pflanzenschutzmittel überprüft, indem es als Fungizid gegen den Weizenschädling Fusarium graminearum und als Resistenzinduktor auf Kartoffelpflanzen getestet wurde. Dabei hat sich einerseits gezeigt, dass Chitosane das Wachstum von Fusarium graminearum verhindert, andererseits wurden verschiedene Abwehrmaßnahmen in der Kartoffelpflanze aktiviert, die auch auf genetischer Ebene nachgewiesen werden konnten.

Both conventional and organic agriculture are still largely dependent on the usage of ecologically harmful and toxic pesticides. Chitosans are sugars, which can induce the resistance of plants against diseases and prevent growth and spread of phytopathogens at the same time. Chitosans display a sustainable and renewable resource as they are attained from exoskeletons of crabs or insect cuticles. This thesis shows the antifungal activity of chitosans against the wheat pathogen Fusarium graminearum as well as its resistance-inducing potential on potato plants. It was observed that chitosans inhibit the growth of Fusarium graminearum and trigger defense mechanisms in potato plants that could also be determined via transcriptome expression analysis.

Titel: Studies on the biological activity of chitosans and their mode of action in plant protection
Verfasser: Lemke, Philipp GND
Gutachter: Moerschbacher, Bruno Maria GND
Organisation: FB 13: Biologie
Dokumenttyp: Dissertation/Habilitation
Medientyp: Text
Erscheinungsdatum: 2019
Publikation in MIAMI: 24.10.2019
Datum der letzten Änderung: 24.10.2019
Schlagwörter: Chitosan; Pflanzenschutz; Fungizid; oxidative burst; RNA-seq; synergistisch; Kartoffel
Chitosan; plant protection; fungicide; oxidative burst; RNA-seq; synergistic; potato
Fachgebiete: Biowissenschaften; Biologie
Lizenz: CC BY 4.0
Sprache: Englisch
Format: PDF-Dokument
URN: urn:nbn:de:hbz:6-73139723274
Permalink: https://nbn-resolving.org/urn:nbn:de:hbz:6-73139723274
Onlinezugriff:
Inhalt:
Table of content ......................................................................................................................... I
Abbreviations .......................................................................................................................... IV
List of tables .......................................................................................................................... VII
List of figures ....................................................................................................................... VIII
Abstract .................................................................................................................................... X
1. Introduction ...................................................................................................................... 1
1.1. The global challenge to feed the world ....................................................................... 1
1.2. Potato – “a dainty dish even for Spaniards” ................................................................ 3
1.3. From classic crop protection to modern crop improvement ........................................ 5
1.4. Chitosan – the jack of all trades ................................................................................... 9
1.5. What’s it all for? ........................................................................................................ 17
2. Material and methods .................................................................................................... 20
2.1. Chitosan preparation and characterization................................................................. 20
2.1.1. Chitosan supply sources ..................................................................................... 20
2.1.2. Preparation of chitosan solutions ....................................................................... 20
2.1.3. Molecular weight analysis .................................................................................. 20
2.1.4. Fraction of acetylation determination................................................................. 21
2.1.5. Separation of oligomer and polymer fractions of chitosans ............................... 21
2.1.6. Enzymatical de-N-acetylation of chitin oligomers ............................................. 21
2.1.7. Chemical re-N-acetylation of chitosans ............................................................. 22
2.2. Antifungal activity of chitosan .................................................................................. 23
2.2.1. Cultivation and induction of conidia production of Fusarium graminearum .... 23
2.2.2. Antifungal activity .............................................................................................. 24
2.3. Eliciting activity of chitosan ...................................................................................... 24
2.3.1. Cultivation of potato plants ................................................................................ 24
2.3.2. Preparation of leaf discs ..................................................................................... 25
2.3.3. Oxidative burst assay ......................................................................................... 25
2.4. Test for synergistic activity ....................................................................................... 26
2.5. Statistics ..................................................................................................................... 26
2.6. Gene expression studies ............................................................................................. 27
2.6.1. Plant treatment .................................................................................................... 27
2.6.2. Pathogen treatment ............................................................................................. 27
2.6.3. RNA extraction .................................................................................................. 27
2.6.4. cDNA synthesis .................................................................................................. 28
2.6.5. Primer design ...................................................................................................... 28
2.6.6. Primer efficiencies .............................................................................................. 33
2.6.7. Real-time PCR .................................................................................................... 33
2.6.8. RNA-seq ............................................................................................................. 34
3. Results ............................................................................................................................. 38
3.1. Antifungal activity of chitosan against Fusarium graminearum ............................... 38
3.1.1. Antifungal activity of COS and paCOS ............................................................. 38
3.1.2. FA-dependent antifungal activity ........................................................................ 39
3.1.3. DP-dependent antifungal activity ....................................................................... 40
3.1.4. Antifungal activity of chitosan combinations .................................................... 44
3.2. Eliciting activity of chitosan ...................................................................................... 51
3.2.1 Eliciting activity of COS and paCOS ................................................................. 51
3.2.2 FA-dependent eliciting activity ........................................................................... 52
3.2.3 DP-dependent eliciting activity .......................................................................... 53
3.2.4 Eliciting activity of chitosan combinations ........................................................ 58
3.3. Chitosan-induced gene expression ............................................................................ 62
3.3.1. Gene expression profile of F. graminearum in response to chitosan ................. 62
3.3.2. Transcriptome analysis of chitosan-treated potato leaves .................................. 66
4. Discussion ........................................................................................................................ 75
4.1. Antifungal activity of chitosan .................................................................................. 75
4.1.1. Chitosan as conidium germination and mycelium growth inhibitor .................. 76
4.1.2. Gene expression changes in response to chitosan .............................................. 81
4.1.3. Chitosan as new big seller on the fungicide market? ......................................... 84
4.2. Eliciting activity of chitosan ...................................................................................... 85
4.2.1. Chitosan-induced oxidative bursts in plants ....................................................... 86
4.2.2. Chitosan-induced gene expression ..................................................................... 91
4.2.3. Chitosan-triggered plant resistance – more than just a simple counterattack .. 101
4.3. Biologically active chitosan in plant protection – the bottom line .......................... 102
5. Summary and outlook .................................................................................................. 106
6. References ..................................................................................................................... 109
Acknowledgements ............................................................................................................... 142
Curriculum vitae .................................................................................................................... 143