Evolution of land plants began and has since occurred, in concert with complex communities of microorganisms, giving rise to a vast spectrum of plant-microbe relationships. Over the past decade, plant molecular biologists and microbial ecologists have worked together to identify drivers of microbiome composition that inspire hypotheses about microbiome functional potential, but many fall short of offering empirical evidence of microbiome-mediated influence on host phenotypes. Herein, I introduce a new suite of tools to explore microbiome function and report that many facets of plant immunocompetence are microbiome-dependent.Chapter One summarizes the current understanding of plant innate immunity and notable progress of plant microbiome research, including: (1) detection and response to microbe-associated molecular patterns, (2) hormone signaling during biotic interactions, (3) technology for exploration of plant microbiome ecology, (4) factors that influence microbiome community structure, and (5) a review of relevant model systems and gnotobiotic growth platforms. Chapter Two describes the development of a novel “FlowPot” growth system: a peat-based platform conducive to axenic (microbe-free), gnotobiotic (defined microbiota), and holoxenic (undefined, complex microbiota) Arabidopsis thaliana growth. This system provides the ability to maintain control of abiotic parameters and exogenous microbiota, thus providing a valuable platform for discovery for plant microbiome research. The FlowPot system and offers a substantial improvement over alternative growth systems regarding plant health, tractability to greenhouse conditions, and maintenance of bacterial alpha diversity upon inoculation with soil-derived microbiota. An implementation of the growth system is detailed in Chapter Three, featuring a comparative analysis of the axenic vs. holoxenic Arabidopsis transcriptome, metabolome, and immunocompetence. Axenic Arabidopsis has a reduced level of defense- and immunity-associated gene expression and the defense hormone salicylic acid (SA). We report that axenic Arabidopsis is compromised in defense against the foliar pathogen Pseudomonas syringae pv. tomato DC3000 (Pst). Immune elicitation experiments revealed that axenic Arabidopsis is also compromised in the ability to recognize and/or mount normal defense responses to the microbe-associated molecular pattern flg22. Axenic susceptibility to Pst is partially explained by defective innate immunity. Finally, we report the identity of differentially abundant metabolites and transcripts in axenic and holoxenic Arabidopsis that may be involved in microbiome-influenced host phenotypes. Collectively, research described in this dissertation provides new tools and a discovery platform to empirically characterize the function of plant microbiota, as well as detailed characterization of axenic phenotypes and axenic immune deficiency.
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