Development of a submerged-liquid sporulation medium for the johnsongrass bioherbicide Gloeocercospora sorghi

Submerged culture experiments were conducted in three phases to determine the optimal medium for rapidly producing conidia of the fungal bioherbicide Gloeocercospora sorghi. In phase I, 18 crude carbon sources were evaluated to determine which would support sporulation. Under the conditions tested, butter bean and lima bean brines (1.5-4.6 mS/cm) provided best conidiation. In phase II, a fractional-factorial design was utilized to screen 76 different medium adjuncts in combination with butter bean brine for improved sporulation. d-Mannitol and carboxymethylcellulose (CMC) were the only acceptable factors that resulted in a significant improvement. In phase III, a central composite design with response surface methodology was used to optimize concentrations of these critical factors. The model predicted optimal sporulation in a medium composed of 2.69 mS/cm butter bean brine +0.043 M d-mannitol +0.37% w/v CMC with an expected titer of 1.51x10(7) conidia/ml. Actual mean titer attained with the model-derived medium was 1.91x10(7) conidia/ml. Optimal sporulation occurred at 25.5 degrees C in this medium and conidia remained viable up to 2.71 days when stored at 12 degrees C. No significant difference was observed in virulence of conidia produced on agar vs washed conidia produced in the model-derived (liquid) medium.