Cassava (Manihot esculenta Crantz), is a staple crop widely cultivated by small farmers in developing countries. However, despite the low required level of technology required for its management, it can be affected by several diseases, with anthracnose as the main threat. There is little information about the main species of Colletotrichum that infect cassava in Brazil. Thus, the objective of this work was to study the diversity, prevalence and virulence of Colletotrichum species that cause anthracnose in cassava leaves in northern Brazil. Twenty municipalities of Pará and Tocantins states were selected and leaves with symptoms were collected in those locations. Pure cultures were isolated in the laboratory. Species were identified using phylogenetic analyses of multiple loci and their pathogenicity, aggressivity and virulence levels were assessed. Our results found a greatest diversity of Colletotrichum associated with anthracnose in cassava plants of the “Formosa” cultivar on Tocantins and Pará states. We determined the presence of Colletotrichum chrysophilum, C. truncatum, C. siamense, C. fructicola, C. plurivorum, C. musicola and C. karsti; with C. chrysophilum as the most aggressive and virulent. Our findings provide accurate identifications of species of Colletotrichum causing anthracnose in cassava crops and provide important information for a more effective control of this disease and the adoption of anthracnose management strategies. These include the development of cultivars with resistance genes to the different pathotypes prevalent in the areas of cultivation, which will potentially reduce production costs.