No-tillage, N fertilization and cover cropping affect physical, chemical and biological qualities of soil. We investigated the effect of 15-yr of tillage systems, N fertilization and cover crops on soil organic matter, aggregation, bulk density and on microbial community in the sandy loam soil of central Italy. The soil in no-tillage (NT) system had 50% more organic matter and 3 folds higher aggregate stability than the soil in conventional tillage (CT) system. The NT system significantly increased soil moisture and decreased bulk density in upper 10 cm layer than in CT system but the effect was not significant in the soil below 10 cm. Similarly, the NT system significantly increased number and diversity of micro-arthropods groups than CT system, thereby improving soil biological quality measured as QBS (Qualita biological del suolo). N fertilization and use of cover crops significantly increased soil organic matter and aggregate stability under the NT system but not under the CT system. Use of cover crops, particularly hairy vetch, also improved soil moisture content, bulk density and QBS value of soil compared with no cover crops. Our results on major physical, chemical and biological properties of soil suggests that combination of NT, cover crops and moderate rate of N fertilization could be a good strategy for enhancing long-term soil quality in the sandy loam soil of the Mediterranean region.