Plant root-associated fungal communities play a pivotal role in enhancing plant growth, nutrient absorption, disease resistance and environmental stress adaptation. Despite their importance, the assembly processes of these communities remain inadequately explored. In this study, we utilizzzed high-throughput sequencing, co-occurrence network analysis and null models to examine the diversity, composition, interaction patterns and assembly mechanisms of the root-associated fungal communities of Mussaenda pubescens, a drought-tolerant shrub that thrives in stressful environments and is widely used for Chinese medicine. Our findings revealed pronounced regional and ecological niche-based variations in the diversity and assembly of total fungi and essential functional guilds, including saprotrophs, symbiotrophs and plant pathogens. Significantly, the fungal diversity of plant pathogens decreased with elevation, whereas total fungi, saprotrophs and symbiotrophs were minimally affected. Stochastic processes, such as dispersal limitation, played a significant role in fungal assembly. Furthermore, soil physicochemical properties, climatic conditions and spatial variables emerged as critical determinants of fungal community structure. This study enriches our understanding of the dynamics governing root-associated fungal community assemblies and underscores the factors essential for sustaining fungal diversity.