Overexpression of the Plant Manganese Superoxide Dismutase Gene GeSOD7 Enhances Cold Stress Tolerance in Armillaria mellea

Background: Fungal symbionts are critical for host plant survival but are vulnerable to abiotic stresses such as low temperature, which limits their agricultural utility. Armillaria mellea is an essential fungal partner for the cultivation of the valuable medicinal orchid Gastrodia elata (G. elata). Enhancing its cold tolerance is a key step toward stabilizing G. elata production. Methods: Based on G. elata transcriptome data, a manganese superoxide dismutase gene (GeSOD7) was identified and heterologously expressed in Escherichia coli for enzymatic characterization. The gene was then overexpressed in A. mellea via Agrobacterium tumefaciens-mediated transformation. Transgenic and wild-type strains were subjected to cold stress (13 °C for 45 days), after which physiological, biochemical, and molecular responses were analyzed. Results: Recombinant GeSOD7 showed optimal activity at pH 6.0 and 60 °C, with inhibition under high concentrations of metal ions, especially Mn2+ and Cu2+. Overexpression of GeSOD7 in A. mellea significantly improved hyphal growth and fresh weight under cold stress. Transgenic strains exhibited higher activities of catalase and glutathione peroxidase, increased accumulation of glutathione and proline, and reduced levels of hydrogen peroxide and malondialdehyde. Expression of genes involved in glutathione synthesis and peroxide detoxification was coordinately upregulated. Conclusions: This study demonstrates that heterologous expression of a plant-derived Mn-SOD can effectively enhance the cold stress tolerance of a symbiotic fungus by boosting its enzymatic and non-enzymatic antioxidant systems. These findings provide a novel genetic strategy for improving stress resilience in agriculturally important fungi and contribute to the sustainable cultivation of G. elata.