Strengthen interactions among fungal and protistan taxa by increasing root biomass and soil nutrient in the topsoil than in the soil-rock mixing layer.

Soil depth plays a crucial role in shaping the interactions between soil microbes and nutrient availability. However, there is limited understanding of how bacterial, fungal, and protistan communities respond to different soil depths, particularly in the unique geological context and soil properties of karst regions. Organic matter, total nitrogen, and phosphorus, ammonium, nitrate, and plant root biomass, as well as bacterial and fungal abundances, bacterial and protistan diversity were higher in the 0-20 cm soil layer than those in the 20-40 cm and soil-rock mixing layers. In contrast, soil pH was higher in the 20-40 cm and soil-rock mixing layers than that in the 0-20 cm soil layer. The soil exchange of calcium, nitrate, and root biomass were identified as the primary factors regulating microbial assemblages across the depth transect. Moreover, co-occurrence network analysis revealed a greater degree of connectivity between protistan taxa and fungal taxa in the 0-20 cm soil layer than those in the 20-40 cm and soil-rock mixing layers. In contrast, the number of association links between protist-bacteria and bacteria-bacteria was higher in the soil-rock mixing layers compared to the 0-20 cm soil layer. Actinobacteria, Ascomycota, and unclassified protistan taxa were identified as keystones, displaying the highest number of connections with other microbial taxa. Collectively, these results suggested that the increased plant root biomass, coupled with sufficient available nutrient inputs in the upper 0-20 cm soil layer, facilitates strong interactions among fungal and protistan taxa, which play crucial roles in the topsoil. However, as nutrients become less available with increasing depth, competition among bacterial taxa and the predation between bacterial and protistan taxa intensify. Therefore, these findings indicate the interactions among keystone taxa at different soil depths has the potential to generate ecological implications during vegetation restoration in fragile ecosystems.