Lasiodiplodia species cause dieback, shoot blight, fruit rot, and stem-end rot of woody plants. Recently, studies on taxonomic revision of Lasiodiplodia species have been conducted globally and have led to more clarification of the species diversity of this genus. However, only three species of the genus have been reported in Japan. To evaluate the species diversity of Lasiodiplodia in Japan, we re-examined the taxonomical positions of 30 isolates based on their morphological and cultural characteristics and phylogenetic relationships. Phylogenetic analyses were performed using a matrix comprising ITS, tef1, tub2, and rpb2 sequences, and the results indicate that the examined Japanese isolates belong to six species: Lasiodiplodia brasiliensis, L. hormozganensis, L. pseudotheobromae, L. thailandica, L. theobromae, and Lasiodiplodia sp. Moreover, three of these species were newly added to the Japanese mycoflora.