Dissertation
Plant-nematode interactions and the application of RNA interference for controlling root-knot nematodes
Doctor of Philosophy (PhD), Washington State University
01/2015
Handle:
https://hdl.handle.net/2376/5496
Abstract
Plant parasitic nematodes are significant pests in major agricultural systems. Sedentary endoparasites like root knot nematodes (RKN; Meloidogyne sp.) obtain plant-derived nutrients from feeding sites formed in host roots thereby supporting nematode development. The formation of feeding sites is hypothesized to be mediated by nematode effectors, proteinaceous secretions from nematode esophageal gland cells.
An inherent challenge in studying most plant-nematode interactions is the difficulty in directly observing nematodes within plant roots. Traditional microscopy techniques are unusable because developing nematodes are surrounded by layers of root cells. In this study, a novel, nondestructive technique was developed to observe the progression of nematode pathogenesis in planta. Pratylenchus penetrans, Heterodera schachtii and Meloidogyne chitwoodi were fluorescently labeled with the lipid specific stain PKH26 and inoculated onto Arabidopsis thaliana seedlings growing in microscopy rhizosphere chambers. The migration patterns and morphology of live nematodes then were observed using confocal microscopy during the parasitic life cycles.
Host-nematode interactions were studied at the molecular level by characterizing the highly conserved RKN effector, 16D10. Overexpression of 16D10 in A. thaliana increased the transcript level of VND7, a xylem development marker, and metaxylem root cell numbers, thereby enhancing susceptibility of A. thaliana to M. incognita. Modifying xylem development by 16D10 possibly facilitates M. incognita feeding site formation. An ortholog of 16D10, Mc16D10L, was cloned from M. chitwoodi. Plant-mediated 16D10 RNA interference (RNAi) silenced Mc16D10L and significantly reduced M. chitwoodi race 1 reproduction by up to 71% in A. thaliana and potato (Solanum tuberosum cvs Russet Burbank and Désirée) plants. Introducing 16D10 RNAi into potato breeding line PA99N82-4 also decreased reproduction of M. chitwoodi pathotype Roza by 50%; this pathotype breaks RMc1(blb), a resistant gene of PA99N82-4. The RNAi effect of Mc16D10L was transmitted to M. chitwoodi offspring, and significantly reduced pathogenicity of nematode offspring on non-RNAi plants. The potato RNAi line, D21 further proved resistant to M. incognita, M. javanica, M. arenaria and M. hapla. Plant-mediated 16D10 RNAi offers a promising new tool for molecular breeding against RKN in potato.
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Details
- Title
- Plant-nematode interactions and the application of RNA interference for controlling root-knot nematodes
- Creators
- Phuong T.Y Dinh
- Contributors
- Debra A. Inglis (Advisor)Charles R. Brown (Committee Member)Lori M. Carris (Committee Member)Axel A. Elling (Committee Member)Kiwamu Tanaka (Committee Member)
- Awarding Institution
- Washington State University
- Academic Unit
- Plant Pathology, Department of
- Theses and Dissertations
- Doctor of Philosophy (PhD), Washington State University
- Number of pages
- 275
- Identifiers
- 99900581439701842
- Language
- English
- Resource Type
- Dissertation