<p dir="ltr">Plant secondary metabolites (PSMs) are specialized compounds produced in response to a range of insect herbivores and microbes, making them important in shaping tri-trophic interactions. However, despite being well-studied in the context of plant-insect coevolution, it is unclear how PSMs impact microbial communities associated with plants and the insect herbivores that feed on them. The overarching goal of this dissertation was to better understand how variation in plant defensive responses, particularly expression of PSMs, influences the composition and functional potential of microbial communities associated with plant tissues (roots and leaves) and insect herbivores. Monarchs (<i>Danaus plexippus</i>) and their milkweed hosts (<i>Asclepias spp.)</i> are well-studied for mechanisms of plant defense and insect counter defense, but little is known about the role of associated microbial communities in this iconic system. To address this knowledge gap, a combination of metabarcoding and metagenomics was used to characterize the taxonomic composition and functional gene profiles of bacterial communities associated with plant tissues (i.e., phyllosphere and rhizosphere) and monarch caterpillars fed on multiple milkweed species (<i>A. curassavica</i>, <i>A. syriaca</i>, and <i>A. tuberosa</i>). Findings show the composition of phyllosphere, rhizosphere, and monarch microbiomes vary across milkweed species in terms of diversity and relative abundance of bacterial taxa. Furthermore, phyllosphere and rhizosphere microbiomes were shown to have distinct functional gene profiles and presence of potential PSM metabolism genes that also varied across milkweed species. Rhizosphere microbiomes had a greater overall capacity for PSM metabolism compared to the phyllosphere, having more genes, and associated metabolic pathways involved in degradation or detoxification of known classes of PSMs. However, plant associated microbiomes were not generally affected by monarch feeding, evidenced by few changes in taxonomic composition or abundance of genes predicted to be involved in PSM metabolism. Interestingly, monarch microbiomes shared >90% of their taxa with their host plants, but there was little evidence of PSM metabolism genes present in functional gene profiles. Overall, this dissertation lays the foundation for understanding how PSMs shape all the microbial communities associated with monarchs and their milkweed hosts. Findings suggest plant defensive responses affect the assembly, functional potential and ultimately the evolution of plant and insect microbiomes.</p>