The diversity and biological activities of the endophytic fungi associated with the Brazilian medicinal plant Stryphnodendron adstringens were studied. A total of 320 fungal isolates were obtained, and 66 phylotypes comprising 25 genera were identified. The fungal community of S. adstringens displayed high richness, diversity and low dominance indices. The most abundant phylotypes were closely related to Diaporthe phaseolorum, Guignardia camelliae, and Preussia pseudominima. Sixteen fungal extracts displayed biological activities when screened against bacteria, fungi, cancer cell lines, and amastigote forms of Leishmania amazonensis. The extract of phylotype Nigrospora cf. oryzae exhibited a selective antifungal activity and inhibited the growth of Candida albicans and Cladosporium sphaerospermum. The extracts of Diaporthe cf. phaseolorum and Xylaria sp. phylotypes displayed anticancer activities. Our results indicate that the endophytes associate with this medicinal plant may be a source for novel drugs.