Soils are key drivers for forest health and productivity. Beyond physical and chemical properties that impact water retention, pH and mineral composition, the biotic factors are being acknowledged as important components to ecosystem functioning. In the soils a high diversity of life forms can be found, including the microbiome, e.g., bacteria and fungi. Generally, they have an impact on plant growth and health and on ecosystem stability. As sustainable forests are becoming a major priority, it is important to evaluate the impact of monocultures on soil quality and microbial diversity and composition. In this study, we aimed to understand the soil microbiome diversity and composition, and their implications on soil functionality among different forest plantations, Eucalyptus globulus, Pinus pinea, and Quercus suber, growing under similar edaphoclimatic conditions in a site located in Abrantes, Portugal. Using Oxford-Nanopore Next-Generation Sequencing we assessed the microbial community structure of soils from these three plantations. This approach was complemented with functional predictions and physicochemical soil analysis. Extracellular enzymatic activities included phosphatase, dehydrogenase, N-acetylglucosaminidase and cellulase evaluation. Results show that overall soil microbial alpha diversity metrics did not differ among eucalypts, stone pine and cork oak plantations. However, microbial compositional patterns clearly diverged, indicating that microbiome assemblages depend on tree species. Bacterial communities appeared more closely related to soil physicochemical parameters, and shared common genera across plantations. Fungal communities displayed higher specificity to tree species and were more strongly associated with organic matter inputs and litter characteristics. These patterns determined the multifunctionality of the microbiome of each tree plantation, highlighting the role of plant–soil associations in structuring belowground communities. This work enhanced our knowledge on soil functions and soil microbial communities from different tree plantations, contributing towards more sustainable and site-adapted forest management strategies.

Acknowledgments: This work was funded by Fundação para a Ciência e Tecnologia (FCT) through CE3C - Centre for Ecology, Evolution and Environmental Changes unit funding (UIDB/00329/2025; https://doi.org/10.54499/UID/00329/2025).