Neurospora intermedia for single-cell protein production: Enhancement by medium optimization and atmospheric and room...

Neurospora intermedia can efficiently convert low-cost substrates into single-cell protein; however, systematic strategies for achieving high-density cultivation remain poorly understood. In this study, we developed a high-protein-producing mutant, T154, derived from Neurospora intermedia FF171 via atmospheric and room temperature plasma (ARTP) mutagenesis, which exhibited stable genetic traits. Medium optimization using molasses as the carbon source and a Box-Behnken design resulted in high cell density (∼60 g/L dry cell weight) and increased protein yield from 18.0 to 28.9 g/L. Notably, the T154 strain (obtained after 120-s ARTP exposure) achieved a final yield of 35.1 g/L, representing a significant 22% improvement over medium optimization alone. Furthermore, meat analogs formulated with T154 biomass exhibited low water loss (35%) and enhanced textural properties (e.g., chewiness: 290.5 g). Overall, this study establishes an effective strategy for the high-density cultivation of Neurospora intermedia and highlights its promising potential as a sustainable alternative protein source.