Peng Yang

Molasses is an abundant and low-cost fermentation feedstock, but its direct utilization is often constrained by its complex composition and inhibitory components. In this study, a molasses-utilizing yeast strain, M0, was isolated from tea rhizosphere soil and identified as Pichia ethanolica. M0 grew directly in untreated molasses diluted only with water, and adaptive laboratory evolution (ALE) was performed under stepwise increasing molasses concentrations. After 70 transfers, the evolved population reached an OD₅₆₀ of 26.41, representing a 73.1% increase relative to the parental strain, and a superior isolate was designated M70. M70 showed lower residual sucrose after 48 h in 100 g/L molasses medium (16.02 vs. 21.11 g/L) and improved growth under high-molasses, -temperature, and -NaCl conditions, with additional growth advantages under selected ethanol and oxidative-stress conditions. Genome resequencing identified five small candidate variants and one interchromosomal duplicative insertion, including alterations involving RIB1, SOL3, and a CET1-like/REX2-like region. Transcriptomic and metabolomic analyses identified 179 differentially expressed genes and 169 differentially accumulated metabolites. Integrated analysis highlighted coordinated changes in D-amino acid, arginine and proline, beta-alanine, and glyoxylate and dicarboxylate metabolism. These findings indicate multilevel genomic, transcriptional, and metabolic remodeling associated with improved molasses adaptation in P. ethanolica.

Corynebacterium glutamicum has been widely used for bulk and fine chemicals fermentation these years. In this study, we developed a defined medium for this bacteria based on the widely used CGXII minimal medium. We evaluated the effects of different components in CGXII on cell growth of C. glutamicum ATCC 13032 and improved the medium through single-factor experiment and central composite design (CCD). Urea, K₂HPO₄ and MgSO₄ were found to be significant factors. 7 out of the total 15 components were modified. (NH₄)₂SO₄, KH₂PO₄, and protocatechuic acid were eliminated. Amounts of urea and MgSO₄ were increased, and concentrations of biotin and glucose were reduced. The resulting R2 medium was proved to be more suitable for cell growth, plasmid amplification and protein production than the original recipe. Remarkably, cell biomass accumulation in R2 increased by 54.36% than CGXII. Transcriptome analysis revealed alteration of carbon metabolism, cation transport and energy synthesis, which might be beneficial for cell growth in R2. Considering the high nitrogen content and availability of urea, the new medium is simplified and cost effective, which holds attractive potential for future study.