Modulation of rhizosphere microbiota by <i>Bacillus subtilis</i> R31 enhances long‐term suppression of banana Fusarium wilt

Continuous cropping of bananas leads to Fusarium wilt, affecting crop health, yet employing biocontrol bacteria to modulate rhizosphere microbial communities may offer effective disease suppression. This study revealed the secondary succession characteristics of rhizosphere microbiota in banana fields after more than 5-year continuous cropping and assessed the suppressive effects induced by biocontrol strain <i>Bacillus subtilis</i> R31. Through high-throughput sequencing, we observed a convergent enrichment of core bacterial genera <i>Burkholderia</i>-<i>Dyella</i> and <i>Arthrobacter</i>-<i>Ralstonia</i> in naturally suppressive and R31-treated suppressive soils, indicating broad-spectrum disease-suppressive soil traits induced by R31. R31 significantly enhanced associations between <i>Streptomyces</i> and the rhizosphere core community while weakening <i>Burkholderia</i>'s linkage with membrane transport and energy metabolism pathways; moreover, it strengthened positive correlations between <i>Rhizobium</i> and terpenoid and polyketide metabolism (<i>r</i> = 0.65, <i>p</i> < 0.01). Culture-dependent assays showed that among 46 isolates from root and rhizosphere, those with high activities of indole-3-acetic acid, protease, cellulase, chitinase, and <i>β</i>-1,3-glucanase exhibited pronounced antagonistic activities against <i>Fusarium oxysporum</i>. Although R31 was undetectable in the rhizosphere in the second year, its modulation of rhizosphere function persisted, displaying a "legacy effect" consistent with the priority effects theory, whereby early colonizing beneficial communities resist subsequent pathogen invasions. R31 induced functional bacteria colonization in nascent banana root hair tissues, establishing new microbiome patterns that contributed to long-term disease suppression. Pot experiments further indicated that endophytic bacteria within roots exhibited stronger Fusarium wilt control than rhizosphere isolates. However, single-strain treatments frequently led to sheath rot co-infection, suggesting that synergistic actions of multiple strains might be more effective for disease suppression. This study highlighted R31's potential to sustainably modulate the rhizosphere microbiome, enhance enzyme activity, and promote beneficial bacteria colonization, laying the groundwork for constructing efficient synthetic bacterial communities for biocontrol.