Elucidating the effects of long-term cultivation on pore structure and infiltration characteristics is essential for understanding soil degradation mechanism and improving cropland sustainability. In this study, we evaluated a number of indicators regarding soil properties, pore structure, and water infiltration during long-term cultivation and the relationship of basic properties and pore structure with soil infiltration in northeast Mollisol region of China. The studied cropland involved four cultivation durations (20, 40, 60, and 100 years) and one forest (FR) as the control. X-ray computed tomography (CT) and mini disk infiltrometer were applied to assess the effects of long-term cultivation on soil pore structure and infiltration characteristics at soil depths of 0–100 cm. The pore properties including soil porosity (TP), pore number (PN), pore size distribution, mean shape factor (MSF), fractal dimension (FA), anisotropy (DA), and Euler number (EN), and infiltration properties including mean (IR), initial (IIR), stable infiltration rate (SIR), and saturated hydraulic conductivity (Ks) were planned to be measured in this study. The results showed that soil organic carbon (SOC) in cropland was significantly lower than that in FR, while bulk density (BD) and mean weight diameter (MWD) exhibited an opposite trend. Long-term cultivation altered the soil pore size and shape distribution, and decreased the macroporosity (>500 μm) and elongated porosity, which suggested that long-term cultivation results in a simpler pore network. FR took a longer time to reach a stable state in infiltration curve over time, while cropland had lower IR, IIR, SIR and Ks, indicating negative effects of cultivation on water infiltration. > 500 μm porosity and elongated porosity were positively correlated with infiltration characteristics (P<0.01). The infiltration parameters were mainly affected by soil pore properties (MSF, PN, and DA) and basic properties (SOC, MWD, and BD), highlighting the importance of pore morphology in the infiltration process. Our results provide new insights into the evolution of soil structure and properties and explanations for water infiltration dynamics under long-term cultivation from the microscale.