Colletotrichum species are important plant pathogens, causing anthracnose in virtually every crop grown throughout the world. However, little is known about the species that infect watermelon. A total of 526 strains were isolated from diseased watermelon samples of eight major watermelon growing provinces in China. Phylogenetic analyses using seven loci (ITS, gadph, chs-1, his3, act, tub2, and gs) coupled with morphology of 146 representative isolates showed that they belonged to 12 known species of Colletotrichum, including C. aenigma, C. chlorophyti, C. fructicola, C. jiangxiense, C. karstii, C. magnum, C. nymphaeae, C. nigrum, C. orbiculare, C. plurivorum, C. sojae, and C. truncatum and three new species, here described as C. citrulli, C. kaifengense, and C. qilinense. Colletotrichum orbiculare was the dominant species. Pathogenicity tests revealed that all isolates of the species described above were pathogenic, with C. magnum and C. kaifengense being the most aggressive to leaves and fruits, respectively. This is the first report of C. aenigma, C. chlorophyti, C. fructicola, C. jiangxiense, C. nymphaeae, C. nigrum, C. plurivorum, and C. sojae on watermelon. These findings shed light on the Colletotrichum spp. involved in watermelon anthracnose and provide useful information for implementing effective control of watermelon anthracnose in China.