Pinus cembroides is among the pine species best adapted to arid and semi-arid ecosystems in the Americas, and its potential distribution is projected to expand under climate change. However, the success of this expansion will depend on belowground processes, particularly the role of soil fungal communities, which in subtropical forests are key for nutrient cycling and plant resilience to environmental stress. Yet their vertical stratification and responses to forest management remain poorly understood, especially in semi-arid systems. Here, we characterized fungal communities from mineral soil and litter associated with P. cembroides across a forest management gradient (mature forests, disturbed stands, and pine plantations) at the southern limit of the species' distribution. We evaluated the influence of climatic, edaphic, vegetation structure, and microbial activity variables (soil moisture, precipitation, pH, tree density, vegetation cover, temperature and extracellular enzyme activity) on fungal community composition. We found strong vertical stratification between litter and mineral soil. When both substrates were analyzed together as an integrated soil profile, forest condition had no significant effect on alpha diversity; however, substrate-specific analyses revealed higher richness in mineral soil of mature forests compared to disturbed and plantation sites, while litter communities showed no significant differences among conditions. Litter communities were characterized by saprotrophic and endophytic fungi, whereas mineral soil communities were enriched in ectomycorrhizal and other root-associated taxa. Distance-based redundancy analysis indicated that fungal community composition was primarily associated with moisture content, precipitation, pH, tree density, and carbon-degrading enzyme activity. These results highlight the importance of substrate differentiation and environmental gradients in shaping fungal communities in semi-arid pine forests, and provide a baseline for understanding how management and climate change influence soil fungal diversity and ecosystem functioning.