Alley cropping agroforestry systems are complex agroecosystems highlighted for their positive effects on soil quality. However, the potential spatial heterogeneity of soil quality created by tree rows at the plot scale has seldom been studied. The aim of this study was to evaluate soil quality at the plot scale, under tree rows and along transects perpendicular to the tree row and to compare alley cropping systems with monocropping systems. This study was performed on an alley cropping system that combined hybrid walnut trees (21 years old) and peas. Topsoil was sampled at tree rows between 1 and 2, 2 and 4 and 4 and 6.5 m from the tree row in the alley cropping system, as well as in a neighbouring monocropping plot. Physical, chemical and microbiological indicators of soil quality were measured. Tree row implantation induced spatial heterogeneity in the chemical indicators, microbial biomass, activities and community structure at the alley cropping plot scale. Alley cropping not only improved microbiological soil quality indicators within the tree rows but also in the interrows when compared to a monocropping system. These indicators were then integrated into one soil quality index (SQI) built through a statistical approach. The soil quality index was calculated for the monocropping plot and for each position within the alley cropping plot. After 21 years of agroforestry practice, tree rows and permanent grass cover improved the SQI until 2 m in the interrow. Weighted SQIs were calculated relative to the surface area of each location for the entire alley cropping plot (i.e., tree row + interrow positions) and for the entire alley cropping interrow (i.e., removing the tree row surface area). The weighted SQI of the entire alley cropping plot significantly increased compared with that of the monocropping plot.