12-h photoperiod: Columbia View orchard soil was passed through a 4-mm sieve and 100 g samples were placed into 487 mL microcosms (glass jars). Jar lids were perforated to make a 1 cm diameter hole into which a rubber septum was inserted. The experiment consisted of two treatments; ASD amended with rice bran (RB) and a no-carbon input ASD control (ASD-NA), seven time points; day 0, 1, 2, 3, 7, 11, and 15, with four samples per treatment at each incubation duration point. In total, across all treatments and time points, 56 samples analyzed. For the purposes of this study, RB soil amendment treatment was not included as pathogen suppression (e.g., F. oxysporum f. sp. fragariae) was not realized under such conditions (Figure 1). In addition, RB soil incorporation does not yield an anaerobic environment (Mazzola et al., 2018), nor result in amplification of target microbial populations commonly associated with ASD-induced disease suppression, such as Clostridium spp. For the ASD treatment, 1.57 g of RB (equivalent to 20 Mg per ha per 15 cm depth in the field) was homogeneously mixed into soil. Estimated nutrient concentration of RB was C = 47.8%, N = 2.49%, P = 1.53%, K = 1.60%, S = 0.18%, C:N ratio = 19:1, pH = 6.2 (Soiltest Farm Consultants, Inc., Moses Lake, WA, United States). For both treatments, 25 mL of distilled water was added to saturate the soil pore spaces. The headspace volume over the microcosms was 409 mL. Jars were incubated in an environmental growth chamber for 15 days at 24/18°C with 12-h photoperiod. Photosynthetically active radiation (PAR, 400–700 nm) just above the jars was 226 μmol m–2 sec–2. Immediately after initiating the experiment, headspace volatiles were extracted, and a representative soil sample was collected using a #3 cork borer (7 cores from each microcosm). Soil samples were placed in a chilled sample cup, followed by immediate immersion in liquid nitrogen and stored at −80°C. This procedure was followed for soil sampling at all incubation duration points. Frozen soil cores were cryogenically milled to a fine powder and stored again at −80°C before use for metabolite analysis. For day 0 and day 1 time points, headspace O2 and CO2 composition was analyzed every 4 h and once for the remaining incubation duration points as described below (https://www.frontiersin.org/articles/10.3389/fmicb.2019.02365/full).
12-h photoperiod: Columbia View orchard soil was passed through a 4-mm sieve and 100 g samples were placed into 487 mL microcosms (glass jars). Jar lids were perforated to make a 1 cm diameter hole into which a rubber septum was inserted. The experiment consisted of two treatments; ASD amended with rice bran (RB) and a no-carbon input ASD control (ASD-NA), seven time points; day 0, 1, 2, 3, 7, 11, and 15, with four samples per treatment at each incubation duration point. In total, across all treatments and time points, 56 samples analyzed. For the purposes of this study, RB soil amendment treatment was not included as pathogen suppression (e.g., F. oxysporum f. sp. fragariae) was not realized under such conditions (Figure 1). In addition, RB soil incorporation does not yield an anaerobic environment (Mazzola et al., 2018), nor result in amplification of target microbial populations commonly associated with ASD-induced disease suppression, such as Clostridium spp. For the ASD treatment, 1.57 g of RB (equivalent to 20 Mg per ha per 15 cm depth in the field) was homogeneously mixed into soil. Estimated nutrient concentration of RB was C = 47.8%, N = 2.49%, P = 1.53%, K = 1.60%, S = 0.18%, C:N ratio = 19:1, pH = 6.2 (Soiltest Farm Consultants, Inc., Moses Lake, WA, United States). For both treatments, 25 mL of distilled water was added to saturate the soil pore spaces. The headspace volume over the microcosms was 409 mL. Jars were incubated in an environmental growth chamber for 15 days at 24/18°C with 12-h photoperiod. Photosynthetically active radiation (PAR, 400–700 nm) just above the jars was 226 μmol m–2 sec–2. Immediately after initiating the experiment, headspace volatiles were extracted, and a representative soil sample was collected using a #3 cork borer (7 cores from each microcosm). Soil samples were placed in a chilled sample cup, followed by immediate immersion in liquid nitrogen and stored at −80°C. This procedure was followed for soil sampling at all incubation duration points. Frozen soil cores were cryogenically milled to a fine powder and stored again at −80°C before use for metabolite analysis. For day 0 and day 1 time points, headspace O2 and CO2 composition was analyzed every 4 h and once for the remaining incubation duration points as described below (https://www.frontiersin.org/articles/10.3389/fmicb.2019.02365/full).