Adaptive laboratory evolution of Naematelia aurantialba under high temperature for efficient production of exopolysaccharide.

Liquid fermentation could revolutionize mushroom polysaccharide production, but the low temperature constraint hampers the process. This study implemented adaptive laboratory evolution (ALE) to enhance the thermotolerance of Naematelia aurantialba strains and increase expolysaccharide production. After 75 ALE cycles at 30 °C, the adaptive strain surpassed the wild-type strain by 5 °C. In a 7.5 L fermentor at 30 °C, the ALE strain yielded 17 % more exopolysaccharide than the wild type strain at 25 °C. Although the exopolysaccharide synthesized by both strains shares a consistent monosaccharide composition, infrared spectrum, and glycosidic bond composition, the ALE strain's exopolysaccharide has a larger molecular weight. Furthermore, the ALE strain's exopolysaccharide exhibits superior cryoprotection performance compared to that produced by the original strain. The adapted strain demonstrated lower ROS levels and increased activity of antioxidant enzymes, indicating improved performance. Fatty acid profiling and transcriptomics revealed reconfiguration of carbohydrate metabolism, amino acid metabolism, and membrane lipid synthesis in thermophilic strains, maintaining cellular homeostasis and productivity. This study provides efficient strains and fermentation methods for high-temperature mushroom polysaccharide production, reducing energy consumption and costs.