Understanding the complex relationships among soil quality, crop productivity, and management practices would help to develop more sustainable agricultural production systems. In this study, we investigated the combined effects of crop rotations and fertilization treatments on soil quality and crop yield in a long term (about 50 years) field experiment. Crop rotations included continuous corn (Zea mays L.), a 2 year corn-wheat (Triticum aestivum L.) rotation, and a 9 year corn-wheat-corn-wheat-corn-wheat-alfalfa-alfalfa-alfalfa (Medicago sativa L.) rotation. Fertilization treatments included control, mineral fertilization with urea and triple superphosphate, and amendment with cattle manure. Crop rotations and fertilization treatments were combined in a factorial experimental design with two replications for each rotation and six replications for each fertilization treatment. The continuous corn and the corn-wheat rotations had negative effects on the main soil quality indicators considered (carbon (C) and nitrogen (N) pools, microbial biomass and activity). On the contrary, the 9 year rotation had positive effects on soil organic carbon (+24%) and total nitrogen (+23%) but resulted in impoverished available P (−5%). Positive effects on soil microbial biomass (+37% of microbial biomass C and +23% of microbial biomass N) and activity (+19%) were also observed. Soil amendment with manure built up soil organic carbon (+13%), increased nutrient content (+31% of extractable C and +19% of extractable N), including that of available P (+47%), and stimulated microbial growth (+34%) and activity (+8%). As compared to manure, mineral fertilization increased the soil nutrient content to a lesser extent. This study showed that the combined use of rotations, including legume forage crops, and soil amendment with manure may help preserve soil quality and crop productivity in the long term.