1. The enhancement of biodiversity in meadow grassland, an environmental aim of European agricultural policy, requires definition of appropriate management regimes and the rate at which they enhance biodiversity and change ecosystem properties. We describe vegetation changes in a 10-year trial on mesotrophic grassland that was previously agriculturally improved, plus change in the soil microbial community and fertility, important factors that influence biodiversity. 2. Management treatments were three hay-cut dates, plus two mineral fertilizer, two seed addition and two farmyard manure (FYM) applications. Treatment combinations included the traditional management regime (21 July hay-cut date, no mineral fertilizer, autumn grazing with cattle, spring grazing with sheep), modern variants of this (14 June hay-cut date, mineral fertilizer) and exceptional historic variants (1 September hay-cut date). 3. Sowing seed increased species richness and, in the absence of fertilizer and FYM, produced a plant community similar to Geranium sylvaticum-Anthoxanthum odouratum grassland. The greatest cover of sown species was found in seeded treatments, cut for hay on 21 July, in the absence of mineral fertilizer. The target plant community (MG3b grassland) was most rapidly achieved with a 21 July hay cut. Initial decrease in Ellenberg fertility scores only persisted in the 21 July and 1 September cut dates when mineral fertilizer was absent. Soil phosphate was lowest in the joint absence of mineral fertilizer and FYM. 4. There were few treatment effects on the soil microflora. Bacterial biomass was reduced when FYM was applied with the 14 June cut date, but increased when FYM was applied with the 1 September cut date. Fungal biomass decreased when mineral fertilizer was applied. 5. Increased species richness, primarily through an increase in legumes, stress-tolerant and stress-tolerant ruderal plant strategists, was associated with an increase in soil fungi and the abundance of fungi relative to bacteria. All these were associated with seed addition to unfertilized plots cut on 21 July, in the absence of FYM, indicating a functional role for individual species. 6. Synthesis and applications. The enhancement of biodiversity in meadow grassland is a long-term (> 10-year) secondary succession, most rapidly achieved in the absence of mineral fertilizer by cutting for hay in mid-July and autumn grazing with cattle. The sowing of key functional species, i.e. legumes and Rhinanthus minor, was important in facilitating the staged colonization of other sown species.