Colletotrichum spp. are known to occur on and cause diseases of Proteaceae, but their identities are confused and poorly understood. The aim of the present study thus was to identify accurately the Colletotrichum spp. associated with diseases of cultivated Proteaceae. Colletotrichum spp. associated with proteaceous hosts growing in various parts of the world were identified based on morphology, sequence data of the internal transcribed spacer region (ITS-1, ITS-2), the 5.8S gene, and partial sequences of the β-tubulin gene. Four species of Colletotrichum were found to be associated with Proteaceae. Colletotrichum gloeosporioides, a cosmopolitan species known to occur on numerous hosts, was isolated from Protea cynaroides cultivated in South Africa and Zimbabwe, and from a Leucospermum sp. in Portugal. A recently described species, C. boninense was associated with Zimbabwean and Australian Proteaceae but also occurred on a Eucalyptus sp. in South Africa. This represents a major geographical and host extension for the species and a description of the African strains is provided. Colletotrichum crassipes was represented by a single isolate obtained from a Dryandra plant in Madeira. Colletotrichum acutatum was isolated from Protea and Leucadendron in South Africa as well as from other hosts occurring elsewhere. A pathologically distinct population of this species was found to occur on Hakea in South Africa. This population is described as C. acutatum f. sp. hakeae, and its relationship with other strains of C. acutatum is discussed. Contrary to earlier literature reports linking C. gloeosporioides to anthracnose of Proteaceae, the present study has shown that several distinct species of Colletotrichum are associated with different diseases of this crop, which has serious implications for quarantine and disease control practices.