Green manuring improved tobacco performance by modulating soil microbial species pool and microbial communities across soil-root continuum

Green manure-tobacco rotation represents a promising strategy for modulating root-associated microbiome, offering significant potential to enhance crop productivity and support sustainable tobacco cultivation in Southwest China. Here, pot experiments including treatments of smooth vetch (SV), ryegrass (R), and radish (RD) as green manures rotated with tobacco were established. The spatiotemporal dynamics of soil microbial species pool and root-associated microbiomes as well as their functional profiles and contributions to tobacco growth were investigated. Results showed that green manures effectively captured soil available nutrients at their growth stage, then SV significantly increased soil inorganic nitrogen, R and RD subsequently increased dissolved organic nitrogen across later tobacco growth. Furthermore, SV significantly increased tobacco shoot and root biomass respectively by 4.4% and 36.1%, and increased tobacco shoot nitrogen content by 0.25%, compared to fallow. Host selection index (HSI) for bacteria and fungi is 2.23 and 3.84 in fallow, while 1.99–2.32 and 1.8–2.79 in green manuring treatments, indicating green manure can mitigate host selection intensity and facilitate the colonization of soil microbiota to root (with the exception of SV bacteria, HSI=2.32). The gradually enriched microbes across soil- root continuum of green manuring treatments dominated by Rhizobiaceae , Sphingomonadaceae and Sordariomycetes , Dothideomycetes, respectively, suggesting that green manuring increased colonization of growth promoting taxa (e.g., diazotrophs and saprotrophs) in . Functional predictions highlighted that the enriched taxa of SV favored nitrogen cycling, R and RD enhanced saprotrophs, showing that green manure differentially facilitated various functional enriched taxa. These findings collectively demonstrate that the SV could be the suitable rotation practice which simultaneously enhance nutrient availability and microbiome-mediated benefits across different green manure treatments, providing insights for targeted soil microbiome management in sustainable agriculture.