Maize (Zea mays L.) is one of the most important staple crops in developing countries. Fungal spoilage on maize resulted in grain deterioration and mycotoxin contamination. In Ethiopia, maize production is prone to losses due to mold contamination. The purpose of this study was to isolate and enumerate fungi and determine the contributing factors to fungal spoilage in Maize. A total of 72 maize samples were collected based on incremental sampling method from the sampling sites. Samples were properly labled with sample history and the moisture content was measured upon collection. Fungal isolation was done on Potato Dextrose Agar (PDA) amended with 0.01% chloramphenicol for 5-7 days at 25. Fungal colonies were purified by sub-cultured on PDA. Morphological and microscopic characterization of the isolates was done based on fungal identification manual. From the total 1080 maize grains analyzed, 613 fungal isolates were recorded with 54.1% mean percent of infection. Of these, 448 isolates belongs to the genus Fusarium followed by 52 Penicillium, 46 Muchor and Rhizopus, 33 Aspergillus and 34 others. Samples obtained from Dano district showed higher mean percent of fungal infection (68.15%) followed by Adami Tulu (60.37%), Bako Tibe (50%) and Arsi Negelle (37.78%). Maize stored in plastic bag showed higher fungal percent of infection (62.5%) followed by gotera (51.1%) and fertilizer bag (49.63%). Grains collected from warehouse were more infected (58.88%) followed by store (55%), open market (54.28%) and household (52.78%). Overall, the prevalence of fungal infection in the study sites was high. Awareness creation to farmers, and capacity building training to experts and agricultural extension workers about pre and postharvest handling, cumulative integrated management, monitoring, and precautionary measures are recommended. Keywords: Fungal Spoilage, Maize, Mold, Mycoflora, Mycotoxin