Comparative Transcriptomics and Metabolomics Uncover the Molecular Basis of Leaf Rust Resistance in Contrasting Germplasms.

(Trin.) Tzvel., a vital native forage grass in northern China for ecological restoration and livestock production, faces severe yield losses and grassland degradation due to rust ( spp.) infection. Current control strategies, reliant on chemical interventions, are limited by evolving resistance risks and environmental concerns, while rust-resistant breeding remains hindered by insufficient molecular insights. To address this, we systematically evaluated rust resistance in 24 germplasms from diverse geographic origins, identifying six highly resistant (HR) and five extremely susceptible (ES) genotypes. Integrating transcriptomics and metabolomics, we dissected molecular responses to infection, focusing on contrasting HR (Lc71) and ES (Lc5) germplasms at 48 h post-inoculation. Transcriptomic analysis revealed 1012 differentially expressed genes (DEGs: 247 upregulated, 765 downregulated), with enrichment in cell wall biosynthesis and photosynthesis pathways but suppression of flavonoid synthesis. Metabolomic profiling identified 287 differentially accumulated metabolites (DAMs: 133 upregulated, 188 downregulated), showing significant downregulation of pterocarpans and flavonoids in HR germplasms, alongside upregulated cutin synthesis-related metabolites. Multi-omics integration uncovered 79 co-enriched pathways, pinpointing critical regulatory networks: (1) In the nucleotide metabolism pathway, genes , , and exhibited negative cor-relations with metabolites Deoxycytidine and Cytosine. (2) In flavonoid biosynthesis, , , , , and were linked to naringenin and naringenin-7-O-glucoside accumulation. These candidate genes likely orchestrate rust resistance mechanisms in . Our findings advance the molecular understanding of rust resistance and provide actionable targets for breeding resilient germplasms.