Filamentous fungi in soil branch hyphae through pores, creating an interconnected fiber network, which is known as mycelium. Fungal mycelium can cross-link and entangle soil particles, which alters soil pore structures. Fungi can also secret hydrophobic compounds, changing the water wettability of soils. These fungal traits can affect the hydraulic properties of soils. This study investigated the effect of fungi on soil water retention and hydraulic conductivity of the Ottawa 50/70 sand treated by a saprotrophic fungus, Trichoderma virens (commonly existing in soil). The soil water retention curve (SWRC) and hydraulic conductivity tests were performed on fungi-treated and untreated Ottawa 50/70 sand. Water repellency of fungi-treated sand was also assessed by measuring contact angles (between the water droplet and fungi on sand specimen) and water drop penetration time. The results of SWRC tests showed an approximate 6-fold increase of air entry suction in the fungi-treated sand, indicating the fungal treatment improved water retention capability. The increased air entry suction was attributed to the change of pore geometry due to mycelium network. A 2-fold reduction in hydraulic conductivity was observed in the fungi-treated sand when growing fungi for 10 days. The hydraulic conductivity reduction was attributed to the enhanced discontinuity of fluid channel by cross-linking and entangling mycelium network. Also, strong hydrophobicity of mycelium layer on the specimen surface contributed to the reduction of hydraulic conductivity. Scanning electron microscopy (SEM) imaging was conducted to assess the morphologies of sand matrix treated by fungi.