Feedback experiment: phase 1

The soil collected from each of the five replicate fields of the four field types was mixed with sterilized background soil in a proportion of 1:6, based on dry mass, to avoid confounding effects of differences in soil physics and chemistry among fields. The background soil was collected from a former agricultural field where in the previous year the nutrient-rich topsoil had been removed, sieved with a 1 cm mesh size sieve, homogenized and sterilized by γ-irradiation (≥25 kGy). This dosage is known to effectively kill all soil biota (McNamara et al. 2003). Pots with a volume of 4 L (21 cm top diameter, 18 cm tall) were filled with the moist soil mixture (equivalent of 5.70 kg dry mass per pot). To obtain sufficient amounts of conditioned soil, for each treatment replicate in phase 1 we filled three pots with soil, whereas after conditioning the soil of the three pots was mixed and used as a substrate in phase 2. This procedure enabled us to condition ample amounts of soil while keeping the true replicates (the five fields per field type) independent. The pots were planted with either seedlings typical to original or to degraded fen meadows. In spite of seedling limitation we ensured that at least one seedling of each plant species was planted in one of the three pots, making sure that all the replicates 1 were planted with the same number of individuals of a species, as well as the replicates 2 and 3, creating similar plant configurations among the replicate pots of the fields. This resulted in 40 soil treatments (2 conditioning plant mixtures × 4 field types × 5 replicate fields).

In order to mimic moist fen meadow conditions, the pots were placed in individual containers with a water level set to 12 cm below the soil surface. Three times per week, the water table was reset using demineralized water and weed seedlings were removed. The pots were placed randomly on trolleys, which were rotated once every week to minimize effects of differences in microclimate in the greenhouse. The plants were grown for 20 weeks under controlled greenhouse conditions at 21/16 °C with a 16/8 h light regime. If needed, the plants were provided with extra light to ensure a minimum photosynthetic photon flux rate of 200 µmol m⁻² s⁻¹.

At harvest, the plants were clipped at ground level. The clipped shoot material was separated by species, dried at 70 °C for at least 48 h and weighed. Soil samples were taken for analysis of nematodes, ergosterol (an indicator of fungal biomass other than arbuscular mycorrhizal fungi (AMF)), neutral lipid fatty acids (NLFA; a measure of AMF) and abiotic soil characteristics [see Supporting Information—Appendix S1]. The remaining roots were clipped in 1–2 cm pieces, mixed with the remaining root zone and bulk soil and stored at 4 °C for 5–6 weeks until the start of phase 2 of the experiment.