BACKGROUND: Seed-borne diseases have seriously affected the sustainability of sorghum cultivation in China as the demand for organic products in the winemaking industry has limited the use of chemical fungicides. RESULTS: This study conducted a comprehensive analysis of fungal diversity in sorghum seeds from three major growing regions in Guizhou Province. Using a combination of traditional tissue separation and high-throughput sequencing, we identified Colletotrichum, Fusarium, Cladosporium, and Alternaria as dominant fungi. Pathogenicity tests revealed that strains GD202206, GD202219, and GD202242 were pathogenic and were identified as C. sublineola through morphological and multi-locus phylogeny analysis (ITS、CAPDH、ACT、CHS-1 and TUB2). 16 fungicides for seed priming experiments with sorghum seeds, the results indicated that priming with KHCO₃ significantly enhanced germination of the sorghum seeds, with both indoor and outdoor emergence rates notably higher. Analysis of the fungal changes before and after KHCO₃ priming revealed a significant reduction in the abundance of the Colletotrichum genus. Additionally, KHCO₃ altered the community structure of fungi within the sorghum seeds, reducing population richness. Inter-generic relationships were rebalanced, with antagonism decreased and synergy increased following KHCO₃ treatment. Non-target metabolomic analysis indicated that KHCO₃ enhances sorghum seed germination via the phenylalanine and flavonoid pathways and exhibits antifungal properties through the cyanoamino acid metabolic pathway. CONCLUSION: This study identified C. sublineola as the primary pathogenic fungus carried by sorghum seeds. KHCO₃ treatment has a dual effect on sorghum seeds: on one hand, it suppresses pathogen transmission by reducing the abundance of the Colletotrichum genus
on the other hand, it promotes germination and seedling emergence, thereby enhancing both germination and emergence rates.