Endophytic fungi significantly influence plant health, growth, and ecological interactions, yet comprehensive insights into their diversity within medicinal plants remain limited. This study provides the first comprehensive analysis of alpha and beta diversity of fungal endophytic communities residing within Wrightia tinctoria (Roxb.) R.Br., examining variations across different plant parts (leaf, stem, and bark), seasons, and geographic locations. A total of 3929 fungal isolates representing 32 morphotypes, primarily from the phylum Ascomycota, were isolated from 6075 tissue segments. Notably, Fusarium cassiae, Neocosmospora magnoliae, Xylaria rohrensis, and Pestalotiopsis papuana were globally reported as endophytes for the first time. Colonization frequency varied significantly with maximum colonization observed at location 1 (80.88%), specifically in leaf tissues (84.64%) and during the monsoon season (80.91%). Analyses of alpha and beta diversity revealed marked variations in locations, plant parts, and seasons. Beta diversity analyses further illustrated both unique and overlapping fungal communities across different conditions, supported by non-metric multidimensional scaling and hierarchical clustering based on Bray-Curtis dissimilarity. Principal component analysis indicated that the first two components explained 40.4% of the observed diversity variations, primarily influenced by location, plant part, and seasonal dynamics. The study concludes that fungal endophytic diversity within W. tinctoria is significantly structured by ecological factors such as plant tissue type, seasonal variation, and geographic location, emphasizing the complexity and specificity of plant-endophyte interactions.