Roselle (Hibiscus sabdariffa L.) family Malvaceae is an important crop used in food, cosmetics and pharmaceutics industries. Roselle is cultivated mainly in Upper Egypt (Qena and Aswan governorates) producing 94% of total production. Root rot disease of roselle is one of the most important diseases that attack both seedlings and adult plants causing serious losses in crop productivity and quality. The main objective of the present study is to identify and characterize pathogens associated with root rot and wilt symptoms of roselle in Qena, Upper Egypt and evaluate their pathogenicity under greenhouse and field condition. Fusarium oxysporum, Macrophomina phaseolina, Fusarium solani, Fusarium equiseti and Fusarium semitectum were isolated from the natural root rot diseases in roselle. All isolated fungi were morphologically characterized and varied in their pathogenic potentialities. They could attack roselle plants causing damping-off and root rot/wilt diseases in different pathogenicity tests. The highest pathogenicity was caused by F. oxysporum and M. phaseolina followed by F. solani. The least pathogenic fungi were F. equiseti followed by F. semitectum. It obviously noted that Baladi roselle cultivar was more susceptible to infection with all tested fungi than Sobhia 17 under greenhouse and field conditions. This is the first report of fungal pathogens causing root rot and vascular wilt in roselle in Upper Egypt.