Isolates from lupin anthracnose in South Africa have previously been identified as Colletotrichum tortuosum, Colletotrichum gloeosporioides and Colletotrichum acutatum. Recently, lupin anthracnose isolates have been classified as a new species, Colletotrichum lupini, with two variants, Colletotrichum lupini var. setosum and Colletotrichum lupini var. lupini. A study was conducted to re-evaluate the classification of lupin anthracnose isolates from South Africa in light of this new classification. The South African isolates, Colletotrichum SHK2148, SHK1033 and SHK788, were compared morphologically with the two C. lupini extype cultures, and found to be similar to C. lupini var. setosum under all growth conditions but different from C. lupini var. lupini. No conidial differences were observed between isolates, although Colletotrichum SHK2148 produced setae, which have been reported for Colletotrichum lupini var. setosum. ITS and β-tubulin DNA sequences from Colletotrichum SHK2148, SHK1033 and SHK788 were compared with sequence data from C. lupini, C. gloeosporioides and C. acutatum isolates. Phylogenetic analysis supported the basic groupings from previous classifications and showed that isolates from lupin anthracnose from South Africa were most similar to C. lupini var. setosum.