Grapevine phyllosphere pan-metagenomics reveals pan-microbiome structure, diversity, and functional roles in downy mildew resistance

Grapevines are among the most economically important fruit crops, and microbiome profoundly influence their health, yield, and quality. However, mechanistic insights into microbiome-orchestrated grapevine biology remain limited. Here, we conduct large-scale pan-metagenomic and pan-metatranscriptomic analyses of the phyllosphere microbiome from 107 grapevine accessions spanning 34 Vitis species. We show that the grapevine core microbiome is dominated by phylum Bacillota and Pseudomonadota. Leveraging HiFi long-read sequencing technology, we assembled 19 high-quality metagenome-assembled genomes (MAGs) from the grapevine pan-microbiome, representing the first MAG reconstruction in plant-associated microbial communities using HiFi long reads. These MAGs encode genes associated with antibiotic resistance, secondary metabolism, and carbohydrate-active enzymes (CAZymes), which could potentially influence grapevine biology. During downy mildew (DM) infection, DM-resistant grapevines exhibit significantly higher microbial network complexity than susceptible counterparts. Among the key taxa contributing to this complexity, Bacillota emerged as the dominant phylum with displaying strong abundance correlations with phylum Euglenozoa and Cyanobacteriota, and an isolated Bacillota species from the grapevine leaves, Bacillus cereus, demonstrated potent biocontrol activity against DM infection. Pan-metatranscriptomic analysis further revealed significant upregulation of eukaryotic microbial genes involved in primary and secondary metabolism. This pan-metagenomic study offers unprecedented insights into the complex structure, diversity and functional roles of grapevine phyllosphere microbiome, and presents valuable genomic and microbial resources for microbiome research and engineering to enhance viticulture productivity and quality.