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Mango (Mangifera indica L.) is an economically significant fruit crop in provinces of southern China including Hainan, Yunnan, Sichuan, Guizhou, Guangdong and Fujian. The objective of this study was to examine the diversity of Colletotrichum species infecting mango cultivars in major growing areas in China, using morphological and molecular techniques together with pathogenicity tests on detached leaves and fruits. Over 200 Colletotrichum isolates were obtained across all mango orchards investigated, and 128 of them were selected for sequencing and analyses of actin (ACT), chitin synthase (CHS-1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), the internal transcribed spacer (ITS) region, β-tubulin (TUB2) genomic regions. Our results showed that the most common fungal isolates associated with mango in southern China involved 13 species: Colletotrichum asianum, C. cliviicola, C. cordylinicola, C. endophytica, C. fructicola, C. gigasporum, C. gloeosporioides, C. karstii, C. liaoningense, C. musae, C. scovillei, C. siamense and C. tropicale. The dominant species were C. asianum and C. siamense each accounting for 30%, and C. fructicola for 25%. Only C. asianum, C. fructicola, C. scovillei and C. siamense have previously been reported on mango, while the other nine Colletotrichum species listed above were first reports associated with mango in China. From this study, five Colletotrichum species, namely C. cordylinicola, C. endophytica, C. gigasporum, C. liaoningense and C. musae were the first report on mango worldwide. Pathogenicity tests revealed that all 13 species caused symptoms on artificially wounded mango fruit and leaves (cv. Tainong). There was no obvious relationship between aggressiveness and the geographic origin of the isolates. These findings will help in mango disease management and future disease resistance breeding.