Simin Zhang, Jianyu Wei, Jili Zhang, Minghui Chen, Yingying Zhang, Yixia Cai, Wei Wang
• Tobacco soil-borne diseases were well controlled by combined application of organic fertilizer, biochar and microbes. • Soil quality was improved and potassium content affected enzyme activities strongly. • Fungal abundance negatively correlated with disease incidence, but positively with Lanceotoxin A. • A “soil-microbiome-metabolite” cascade underlies the defense against soil-borne diseases. The application of green additives is an effective strategy to enhance soil health and control tobacco diseases. In this study, microbiome and metabolomics techniques were employed to investigate the mechanisms by which the combined application (SP) of organic bran fertilizer, biochar, and microbial inoculants suppresses tobacco soil-borne diseases. The results showed that SP significantly decreased the incidence of bacterial blight, black shank, and root rot by 66.58 %, 76.12 %, and 64.37 %, respectively. Soil pH, the contents of organic matter and total potassium were improved obviously, and the activities of catalase, polyphenol oxidase and invertase activities were enhanced, too. The abundance of beneficial bacterial genera, such as Sphingomonas and Bacillus , and fungal genera, including Pseudeurotium and Rhizophagus , increased significantly, whereas pathogenic genera such as Gemmatimonas and Fusarium were significantly suppressed. Bacterial functional groups associated with “peptidases and inhibitors” and fungal saprotroph-symbiotroph groups were distinctly enhanced. A total of 925 differential metabolites were identified in tobacco roots, stems, and rhizosphere soil. Lanceotoxin A increased by 119.96-fold in rhizosphere soil, with key metabolic pathways, including glucosinolates and alkaloids, activated, promoting the biosynthesis of disease-resistant metabolites such as gingerol and 12-KETE. Additionally, plant resistance was supported by Albizziin as well as by Rhizophagus and Bacillus . Structural equation modeling revealed that fungal abundance was negatively correlated with disease incidence but positively correlated with Lanceotoxin A. Soil nutrients accounted for a substantial proportion of the variation in enzyme activity and fungal community composition. Overall, SP provides a feasible and effective approach to controlling soil-borne diseases and enhancing tobacco yields through improved soil quality, regulated microbial communities, and activated host metabolic pathways.