Potato dextrose agar medium pda

Pre-cultures of strains MACI46 (NRRL 66708), MACI219 (NRRL 66709), MACI254 (NRRL 66710) and MACI264 (NRRL 66711) incubated in the dark at 27 °C for seven days. For metabolite profile characterization, isolates were cultured in four different media: MEA, CYA, YES agar, and Potato Dextrose Agar medium (PDA) (Sigma-Aldrich, Saint Quentin Fallavier, France). For each medium, three biological replicates were inoculated centrally with 10 µL of calibrated spore suspensions (10⁵ spores/mL) prepared from seven-day cultures on 7.5 cm Petri dishes. The samples were incubated in the dark at 27 °C for seven days.
To perform extrolite extractions, culture media were macerated and placed in 50 mL sterilized tubes, and thereafter 35 mL of ethyl acetate was added to each sample. Samples were agitated 48 h in an orbital incubator at 170 rpm at room temperature. Ethyl acetate was filtered through a Whatman 1PS phase separator (GE Healthcare Life Sciences, Vélizy-Villacoublay, France) and evaporated at 60 °C until dry. Samples were then dissolved in 400 µL of methanol. To eliminate possible impurities, each sample was filtered through a 0.45 µm disk filter (ThermoFisher Scientific, Illkirch, France) [81 (link)].

Wheat cultivar Alpowa, susceptible to soilborne fungal pathogen R. solani AG8, was used in this study. All wheat seeds were derived from the same seed source to reduce plant variation. Seeds were treated with 5% sodium hypochlorite for 3 min for surface disinfestation and rinsed three times with sterilized double-distilled water (ddH₂O) before germination. All wheat plants were grown in a greenhouse in 16/8 h light/darkness at 16°C. Arabidopsis thaliana Col-0 accession (hereafter Arabidopsis) was used to evaluate the influence of bacteria on root growth. The soil used in this study was collected from the Washington State University Dryland Research Station near Lind (47°0’N, 118°34’W), WA, United States, as described previously (Yin et al., 2021 (link)). The fungal pathogen R. solani AG8 was used and grown in potato dextrose agar medium (PDA, Sigma-Aldrich, St. Louise, MO) at 25°C. The inoculum of R. solani AG8 was prepared with twice-autoclaved millet seeds, as described previously (Yin et al., 2021 (link)). The bacterial strains isolated from the wheat rhizosphere (Yin et al., 2013 (link), 2021 (link)) were grown on 1/4 x tryptic soy agar/1 x tryptic soy broth (TSA/TSB, BD, Franklin Lakes, NJ) at 25°C.

Monilinia fructicola (G. Winter) honey strain MeCV-2 used in this study belongs to the IVIA CTP collection of postharvest pathogens. This strain was isolated from a rotten peach fruit from a stone fruit packinghouse in Carlet (Valencia, Spain) and, after isolation and identification, it was selected among other isolates due to its aggressiveness and uniform behavior. The strain is deposited and available with the accession number CECT 21161 at the Spanish Type Culture Collection (CECT, University of Valencia, Valencia Spain). Before the experiment, the isolate was grown on potato dextrose agar medium (PDA, Sigma-Aldrich Chemie, Steinheim, Germany) in Petri dishes at 25 °C for 7–14 days. High-density conidial suspensions of spores were prepared in Tween 80 (0.05% w/v; Panreac-Química S.A., Barcelona, Catalonia, Spain) in sterile water. After passing through two layers of cheesecloth, the density of the suspension was measured with a hemacytometer and diluted with sterile water to obtain an exact inoculum density of 1 × 10³ spores/mL [6 (link)].