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Date palm diseases are rising as an important concern during the last decades. The fungal pathogens of date palm are considered as the most serious problem causing significant reductions of growth, development, and production of date palm. The present study was aimed at the characterization of two Nigrospora species, isolated from different cultivars of date palm, based on morphological, molecular and pathological characteristics. In the current study, the identity of both Nigrospora species have been revealed to be as N. oryzae and N. sphaerica on the basis of their morphological characteristics and molecular analysis of the Internal Transcribed Spacer (ITS) region. Results showed that both pathogens were found to be true pathogens on different date palm cultivars. Compared to N. Sphaerica, N. oryzae was more aggressive on the following cultivars: Al-Sayer, Hillawi, Zahdi, Leloy and Kantar. After 30 days post-inoculation, the overall average lesion diameter was 1.85 cm in response to the artificial infection with N. oryzae, whereas infection with N. sphaerica produced 1.42 cm lesions. Al-Sayer cv. was the most susceptible, among the tested cultivars to both Nigrospora species; the lesion diameter was 2.50 cm, in contrast with cv. Leloy, 1.10 cm, which showed the lowest level of susceptibility. The extracellular enzymatic activity of both pathogens revealed that N. oryzae surpassed N. sphaerica in the production of cellulase and protease enzymes; whereas, lipase enzyme activity was absent in both fungi. The high enzymatic activity and virulence of N. oryzae on different date palm cultivars were approved in contrast with the species of N …