Harnessing Aspergillus and host M genes for sustainable phyllosphere microbiome engineering

Abstract

The phyllosphere microbiota plays a crucial role in maintaining plant health and enhancing disease resistance [1-5]. Recent studies have indicated that plants actively shape their microbial communities prior to stress and selectively recruit beneficial microbes in response to pathogen infection, thereby establishing an extended layer of plant immunity [6]. Despite accumulating evidence highlighting the critical role of metabo-lites as mediators in plant-microbe interactions [7], the mechanisms by which microbiota-derived signals promote host resistance remain poorly understood. In a recent study published in Nature Microbiology, Fan et al. identified 2,4-Di-tert-butylphenol (2,4-DTBP), a small molecule produced by the phyllosphere-associated fungus Aspergillus, which enhances resistance of rice to the fungal pathogen Rhizoctonia solani [8]. By sterilizing the leaves of three representative rice cultivars with varying resistance to R. solani, researchers demonstrated a loss of cultivar-specific resistance, underscoring the essential role of the native phyllo-sphere microbiota in disease registance. To identify key microbes, they performed phyllosphere microbi-ome sequencing in the absence of pathogen inoculation and discovered a positive correlation between Aspergillus abundance and resistance levels. Researchers subsequently hypothesized that specific Aspergillus metabolites underlie this protective effect. This hypothesis was supported when metabolites derived from Aspergillus cvjetkovicii were shown to suppress R. solani infection. Through metabolomic and bioassay-guided approaches, they identified 2,4-DTBP as the active metabolite responsible for R. solani suppression. Notably, this inhibitory effect extends beyond rice, providing protection to other crops, such as cucumber, maize, soybean, and tomato, even under field conditions , highlighting its broad-spectrum potential. Subsequent transcriptomic analyses of R. solani treated with 2,4-DTBP revealed significant downregu-lation of R. solani AMT1, encoding an ammonium transporter. The overexpression of RsAMT1 promoted hyphal growth, sclerotia formation, and pathogenic-ity, indicating that AMT1 positively regulates R. solani pathogenicity. 2,4-DTBP was found to reduce the accumulation of reactive oxygen species (ROS) in R. solani. Conversely, H 2 O 2 treatment upregulated AMT1 expression in R. solani as well as in another fungal pathogen, Fusarium fujikuroi. These findings collectively suggest that A. cvjetkovicii-derived 2,4-DTBP suppresses RsAMT1 expression by decreasing ROS levels, thereby dampening its pathogenicity. The composition and structure of the plant microbi-ome are shaped by the plant genotype, developmental stage, and nutrient uptake, reflecting plant adaptations to various environments to impact traits such as disease resistance beyond innate immunity [9, 10]. This adaptive capacity has been attributed to micro-biome-shaping (M) genes, denoting the genetic basis by which plants manipulate and reshape their associated microbiome [11, 12]. However, the specific genes