Hepatitis E virus (HEV) infection is endemic in many developing countries and becomes of interest in the developed countries. Several animals are sources of HEV infection to humans. Recently, HEV was detected in the milk of cows in China, this data comes up with the probability of HEV transmission to humans via ingestion of contaminated milk. In Egypt, contaminated water and residing in rural communities are risk factors for HEV infection, while limited data is available on the zoonotic HEV transmission. Since pigs, wild boars, camels are not common in Egypt, we investigated if cows and/or cow milk represent a risk factor for HEV transmission in the Assiut governorate. Milk samples (n = 480), collected from Assiut city and 12 non-mixed dairy farms distributed in the rural communities, were tested for HEV markers such as anti-HEV IgG, HEV RNA, and HEV Ag. All milk samples collected from Assiut city (n = 220 …