Experimental Design of the Pot Experiment

A pot experiment was conducted with sandy-loamy soil taken from the top horizon of agricultural soil located at Chenevières (Grand Est, France), which was previously spiked with Cd, Pb, Cu, and Zn and used for the cultivation of N. caerulescens as part of previous studies (Sterckeman et al., 2017, 2019). At the beginning of the present study, the Cd and Zn concentrations were 3.6 and 649 mg/kg, respectively. Some physico-chemical parameters of the soil are provided in Supplementary Table 1. The seeds of N. caerulescens originated from a metallicolous population from a mining site located in Ganges [Occitanie, France, Gonneau et al. (2014)]. The seeds were sown in a sterilized mix (v/v) of compost (75%) and sand (25%) and placed in a growth chamber. Seven weeks after germination, plants of similar developmental stages were selected for the pot experiment.

In the present study, we tested seven fungal endophytic strains (DBF60, DBF79, DBF81, DBF107, DBF108, DBF129, and DBF159) originating from the collection of strains isolated from N. caerulescens. They were selected based on their potential as PGP fungi, according to their description in the literature. The production of inoculum and the soil inoculation procedure are fully described in Yung et al. (2021b). As a control treatment (CTRL), we mixed one aliquot of soil with perlite containing fungus-free agar plugs. Five replicates were considered for each fungal treatment, and four were considered for the mock-inoculated treatment. Seedlings were transplanted into 40 plastic pots (8 treatments × 5 replicates) of 500 mL volume (8.6 cm in diameter) previously loaded with 400 g of the inoculated soils. The pots were then placed in a growth chamber with 16 h of light at 22°C and 8 h of darkness at 18°C, 70% relative humidity and a photon flux density of 200 μmol photons m^–2 s^–1 in the PAR range. Water was supplied every 2 or 3 days to 80% of the field capacity.