Abstract: Pine wilt disease is currently a serious threat to global forest ecosystems and a highly contagious pest affecting pine trees. The screening of effective biocontrol strains is an urgent issue that the world is facing. From March 2023 to February 2024, this study monitored the nematode-trapping fungi (NTF) resources in five healthy Yunnan pine trees selected from the Cangshan National Nature Reserve in Dali, Yunnan. Bark samples were collected, and NTF were cultured, isolated, and purified in the laboratory. Species identification was conducted using morphological and molecular techniques, and the spatiotemporal dynamics of NTF communities were analyzed across different heights of the tree trunk: upper (80 cm < height ≤ 200 cm), middle (20 cm < height ≤ 80 cm), and lower (0 cm ≤ height ≤ 20 cm). The results revealed the presence of NTF in the pine bark, with fungi of the genus Dactylellina being the dominant group, and Dactylellina ellipspspora as the dominant species (5.48%). The greatest number of NTF species and strains was detected in the lower part of the tree and during August. A spatiotemporal variation in the number of detected NTF species and strains was observed. Indigenous NTF in pine trees can serve as ideal candidates for screening efficient biocontrol strains against pine wilt disease.