Abstract: Streptomyces sp. TH053, an actinomycete strain with antifungal activity isolated from the sediment of Taihu Lake, was preliminarily identified, and its taxonomic position was determined by 16S rRNA gene sequence analysis. By bioassay-guided fractionation, the major antifungal constituents in the concentrated extract of the fermentation broth were isolated and purified by silica gel chromatography, gel column chromatography, and preparative HPLC, and their structures were elucidated by MS and 1D/2D NMR. The antifungal activities of the obtained compounds were verified by the filter paper disk method. Three compounds were obtained and identified as mycotrienin I (1), mycotrienin II (2), and ansafurantrienin A (3). Compounds 1 and 2 exhibited notable antifungal activity against Colletotrichum camelliae, C. siamense, C. fructicola and C. gloeosporioides. At 50 μg per filter paper, compound 2 showed inhibition rates of 42.00%, 65.78%, 60.78%, and 59.33% against the four anthracnose pathogens, respectively, comparable to those of the positive control fungicide tebuconazole (48.78%, 66.17%, 61.50%, and 58.67%). On day 4, compound 2 at 100 mg/L provided control efficacies of 100% and 60.70% against C. gloeosporioides on mango (Mangifera indica L.) and C. fructicola on grape (Vitis vinifera L.), respectively, effectively protecting mango and grape fruits from anthracnose. These results provide a reference for the practical application of such compounds in the control of anthracnose pathogens in crops.