The increasingly evident involvement of microbes in basic host function has led to a more holistic perspective of plants to be considered. Here, plants and their associated microbiotas interact with each other as holobionts in performing various biological functions in natural ecosystems and crop fields as a single ecological unit. The extent to which microbial components of the holobiont contribute to host health, however, is not fully understood, especially in natural environments. Here, I applied the use of peat-based gnotobiotic growth systems to generate plants grown with and without exposure to microbiota to characterize the role of microbiota on the development of plant immunocompetence.In my first chapter, I review the current understanding on the interplay between microbiota and plant health and immunity. I begin with background on microbial detection and response in plants. I then discuss plant-associated microbiota and provide examples of how perturbation to microbiota homeostasis, such as during dysbiosis for example, can be associated with positive and negative impacts on host health. Host factors regulating microbiota homeostasis in plants are discussed. Lastly, I describe tools and approaches that can be used to the study of plant-microbiota interactions and highlight recent findings involving the modulation of plant immune responses by plant microbiotas.In my second chapter, I highlight the contributions I made to the improvement of two peat-based gnotobiotic plant growth systems for plant microbiome research recently developed in Dr. Sheng Yang He's laboratory: the FlowPot and GnotoPot systems. I adapted the FlowPot system to use a field soil, highlighting its versatility, and optimized several abiotic conditions associated with plant growth in GnotoPots. Additionally, I used 16S rRNA gene amplicon sequencing to characterize the colonization of a natural, soil-derived microbial community and several preparations of a synthetic bacterial community, demonstrating the use of GnotoPots for colonization studies. We expect both systems to be useful tools for the research community to address a wide variety of questions related to plant-microbiota interactions.In my third chapter, I implement the optimizations made to peat-based gnotobiotic growth systems described in the previous chapter to characterize the role of basal microbiota colonization on plant immunocompetence. I found that compared to plants colonized by a soil-derived microbiota, axenic plants grown without exposure to a microbiota lacked robust age-dependent immunity. Axenic plants were defective in several aspects of pattern-triggered immunity including flg22-induced production of reactive oxygen species, signaling through MAPK pathways, and induction of defense-related genes and hypersusceptible to disease a bacterial foliar pathogen. Additionally, I found that a synthetic microbiota composed of culturable leaf endosphere bacteria was able to restore immunocompetence similar to plants inoculated with a soil-derived community in a growth substrate-dependent manner. These results demonstrate a role of microbiota in immunocompetence and age-dependent immunity, which was previously thought to be an intrinsic trait of plants.
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