Degradation experiment and metabolite identification.

Cells of strain DY-1 were cultured in R2A medium for approximately 72 h at 30°C and then collected by centrifugation at 6,000 × g for 5 min. The cell pellets were washed twice with sterilized MSM, adjusted to an optical density at 600 nm (OD₆₀₀) of approximately 1.5, and used as the inoculant. The standard degradation assay was performed in MSM containing 0.15 mM trifloxystrobin (pH 7.0) at 30°C with shaking (180 rpm). The initial inoculum of strain DY-1 cells was set at 1.15 × 10⁷ CFU · ml⁻¹. The concentration of trifloxystrobin was determined by HPLC, and the metabolites were identified by HPLC-MS/MS. The effect of pH and temperature on the degradation of trifloxystrobin by strain DY-1 was investigated by the standard degradation assay at different pHs (pH 4.0 to 10.0, in increments of 1 pH unit) and incubation temperatures (15°C to 55°C, in increments of 5°C). The degradation of strobilurin fungicides, including azoxystrobin, picoxystrobin, and pyraclostrobin, by strain DY-1 was also studied using the standard degradation assay method. Each treatment was performed in triplicate, and the means and standard errors were calculated. The uninoculated medium was used as a control, and the change in the tested substrate concentration in the control was negligible.