Carbon-Source Dependent Interplay of Copper and Manganese Ions Modulates the Morphology and Itaconic Acid Production in

The effects of the interplay of copper(II) and manganese(II) ions on growth, morphology and itaconic acid formation was investigated in a high-producing strain of (NRRL1960), using carbon sources metabolized either mainly via glycolysis (D-glucose, D-fructose) or primarily via the pentose phosphate shunt (D-xylose, L-arabinose). Limiting Mn concentration in the culture broth is indispensable to obtain high itaconic acid yields, while in the presence of higher Mn concentrations yield decreases and biomass formation is favored. However, this low yield in the presence of high Mn ion concentrations can be mitigated by increasing the Cu concentration in the medium when D-glucose or D-fructose is the growth substrate, whereas this effect was at best modest during growth on D-xylose or L-arabinose. displays a high tolerance to Cu which decreased when Mn availability became increasingly limiting. Under such conditions biomass formation on D-glucose or D-fructose could be sustained at concentrations up to 250 mg L Cu, while on D-xylose- or L-arabinose biomass formation was completely inhibited at 100 mg L. High (>75%) specific molar itaconic acid yields always coincided with an "overflow-associated" morphology, characterized by small compact pellets (<250 μm diameter) and short chains of "yeast-like" cells that exhibit increased diameters relative to the elongated cells in growing filamentous hyphae. At low concentrations (≤1 mg L) of Cu ions, manganese deficiency did not prevent filamentous growth. Mycelial- and cellular morphology progressively transformed into the typical overflow-associated one when external Cu concentrations increased, irrespective of the available Mn. Our results indicate that copper ions are relevant for overflow metabolism and should be considered when optimizing itaconic acid fermentation in s.