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Ag/TiO₂ wood-based nanocomposites were prepared by the methods of ultrasound impregnation and vacuum impregnation. The as-prepared samples were characterized by field emission scanning electron microscopy (FESEM), energy-dispersive spectroscopy (EDS), Fourier transform infrared spectroscopy (FTIR), mercury intrusion porosimetry (MIP), and water contact angles (WCAs). The anti-mold properties of the Ag/TiO₂ wood-based nanocomposites were improved by 14 times compared to those of the original wood. The nano-Ag/TiO₂, which was impregnated in the tracheid and attached to the cell walls, was able to form a two-stage rough structure and reduce the number of hydroxyl functional groups on the wood surfaces. The resulting decline of wood hydrophobic and equilibrium moisture content (EMC) destroyed the moisture environment necessary for mold survival. Ag/TiO₂ was deposited in the wood pores, which reduced the number and volume of pores and blocked the path of mold infection. Thus, the anti-mold properties of the Ag/TiO₂ wood-based nanocomposite were improved by cutting off the water source and blocking the mold infection path. This study reveals the anti-mold mechanism of Ag/TiO₂ wood-based nanocomposites and provides a feasible pathway for wood-based nanocomposites with anti-mold functions.