Compound Bacillus agents inhibit Cytospora mali-induced valsa canker in Hanfu apple trees through multiple pathways

Abstract Background Apple Valsa canker caused by Cytospora mali (Cma) has emerged as a devastating fungal disease threatening apple production worldwide, particularly in cold-resistant cultivars like Hanfu apple. Current control strategies predominantly rely on chemical fungicides, which pose environmental risks and may lead to pathogen resistance. There is an urgent need to develop sustainable biocontrol alternatives. Result Through systematic screening of 123 Bacillus isolates from Xinjiang wild fruit forest soils, we identified three elite strains with superior biocontrol potential: B. subtilis Bsu181, B. licheniformis Bli279, and B. amyloliquefaciens Bam190. The developed Bacillus compound agent, comprising Bsu181, Bli279 and Bam190 in equal proportions, demonstrated comprehensive disease control through multiple synergistic mechanisms: (1) Directly inhibiting Cma growth (75.56–97.78% inhibition rates) and causing mycelial deformation; (2) Enhancing host resistance by elevating antioxidant enzymes (POD: 1505.7-2036.48 U/g·min; PPO: 163.51-201.52 U/g·min) and upregulating defense genes PR1 (4.70-fold); (3) Promoting tree growth through increased stomatal opening (0.414 μm) and photosynthetic efficiency; and (4) Restructuring rhizosphere microbiomes by reducing pathogenic fungi ( Penicillium , Cladophialophora ) while enriching beneficial microbes ( Chaetomium , Mortierella , Sphingomonas ). Conclusion The Bacillus compound agent achieved exceptional disease control efficacy of 74.8–81.9% while simultaneously enhancing tree growth and health. These findings demonstrate that this carefully formulated microbial consortium can serve as an effective and environmentally friendly alternative to conventional fungicides for managing apple Valsa canker. The multiple synergistic mechanisms of action, combining direct pathogen inhibition with plant immunity induction and microbiome modulation, provide durable protection against Cma infection while supporting sustainable apple production systems.