Abstract:[Background] Longyangxia valley is one of the most important lake wetlands in the source region of the Yellow River, demonstrating great potential in water regulation. [Objective] To explore the soil fungal diversity and screen biocontrol isolates from Longyangxia valley, enrich the fungal resources of lake wetlands in alpine regions, and provide basic materials for subsequent development of biocontrol agents. [Methods] Ten soil samples were collected from the shoreside of Longyangxia reservoir in Longyangxia valley, and fungi were isolated by the dilution coating method. The isolates were identified by morphological observation and methods of molecular biology, and the biocontrol isolates were screened by the plate confrontation method. [Results] One hundred and four fungal strains belonging to 23 genera were isolated, including Preussia, Trichoderma, Alternaria, Cladosporium, Bipolaris, Pseudogymnoascus, and Penicillium. The isolates belonging to Preussia and Trichoderma were the most, with the number of 21 and 19 and the isolation frequency of 20.19% and 18.27%, respectively. The isolates of Botryotrichum, Westerdykella, Leptosphaeria, Arthrinium, Scolecobasidium, Sporothrix, Tetracladium, and Aphanoascus were the fewest, and all these genera had only one isolate, with the isolation frequency of 0.96%. Nineteen isolates of Trichoderma were screened out from the 104 fungal isolates, and these isolates had great biocontrol effects on eight fungal pathogens isolated from the main crops of Qinghai Province, with the inhibition rates between 51.6%–73.5%. Ten Preussia isolates, 4 Pseudogymnoascus isolates, 3 Cladosporium isolates, 2 Bipolaris isolates, and 1 isolate each of Arthrinium, Leptosphaeria, Westerdykella, Penicillium, and Emericellopsis also exerted strong inhibitory effects on the eight pathogens, with the largest inhibition zone ≥10 mm. [Conclusion] The soil of Longyangxia valley harbors abundant fungal resources and demonstrates high potential of exploitation and utilization.