The endophytic and epiphytic microflora from walnut trees in South Tyrol (Northern Italy) was analyzed. A total of 16 trees from 8 sites were sampled at three dates during the warm season (May, July and September) in 2005, using plant material from three different tissue types (leaves, fruit and lignified twigs). Samples were either transferred directly onto nutrient medium or surface sterilized prior to plating to distinguish between epiphytic and endophytic growth. A total of 3,880 culturable isolates were obtained, the vast majority of the isolates (3,742) belonging to fungi (96.4%); only 138 (3.6%) were bacteria. Fungal isolates were classified into 30 genera, Alternaria, Penicillium, Phoma, Botryosphaeria, Fusarium, Cladosporium, Phyllosticta and Epicoccum being the most common taxa. Although epiphytic growth predominated (73.9% of the identified isolates), a significant number of endophytes (26.1%) were isolated as well. More isolates were obtained from leaves (45.3%) than from twigs (31.8%) and fruit (23.0%). Seasonal effects were not very distinct, but a tendency towards higher isolation success in September was observed.