Mild Constitutive Immune Activation Promotes a Pathogen-Depleted Rhizosphere Microbiome in Maize

Constitutive activation of plant immune responses can enhance disease resistance but often incurs fitness costs and may alter assembly of the rhizosphere microbiome. However, how the intensity of constitutive immune activation influences rhizosphere microbial communities remains poorly understood. Using nine pairs of maize near-isogenic lines and hybrids differing in the presence or absence of the auto-active NLR allele Rp1-D21, we investigated how a genetically modulated gradient of constitutive hypersensitive re-sponse (HR) intensity, ranging from mild to severe, influences rhizosphere microbiome assembly. Root transcriptome profiling confirmed activation of canonical defense pathways, including oxidative stress and pathogenesis-related responses, providing mechanistic support for constitutive immune activation. Mild-HR plants represented a distinct rhizosphere microbiome state characterized by maintenance of overall microbial diversity together with the greatest abundance and diversity of taxa previously associat-ed with disease suppression and the lowest abundance and diversity of putative pathogens. In contrast, wild-type plants harbored the greatest putative pathogen diversity and abundance, whereas moderate and severe HR were associated with reduced microbial diversity and increasing enrichment of opportunistic necrotrophic and saprophytic taxa. Differential abundance analysis using ANCOM-BC2 supported these community-level patterns. Collectively, these findings suggest that maintaining a mild constitutive defense state may promote assembly of a functionally favorable rhizosphere microbiome while limiting pathogen colonization, providing new insights into how host immune signaling shapes rhizosphere microbiome or-ganization and highlighting a potential strategy for engineering favorable rhizosphere microbiomes.