Importance of maize late wilt caused by Harpophora maydis (Cephalosporium maydis Smra, Sabet & Hingorani) comes from the sudden and relatively rapid wilt of maize plants at the time of flowering and until shortly before maturity. So, the detection of the causal agent early before symptoms appearance may be useful for managing the disease. For this purpose, two maize plants were chosen randomly at 7, 21, 35, 49, 63, 77, 91 and 105 days after seeding in greenhouse and field infested soils, during 2016 and 2017 seasons. Fungal DNA was detected in emerged plant parts using the specific primer of H. maydis. The first detection of the fungal DNA was at 7d after seeding in roots and leaves of greenhouse and field seedlings. At the age of 21, 35, 49, 63 days the fungus was detected in all tested plant parts. With pathogen progression, from 77d to 105 d after sowing, clear amplified fungal DNA was found in samples taken from stalk-internodes, tassels, ear-shanks, cobs, kernels and leaves with the consistent of disease symptoms. But the unique band of H. maydis did not appear in the amplified DNA taken from mature kernels at the age of 105 days. As a conclusion of the obtained results, it is clearly pronounced that H. maydis colonized all maize plant parts emerged during the growing season. Also, polymerase chain reaction (PCR)-based assay was successfully employed to identify H. maydis directly from the mycelia that embedded in maize plant tissues, with no need to obtain pure fungal cultures for detection compared with traditional method. Thus, this protocol can be used for diagnosis of infected plants by late wilt causal agent.