Abstract:Abstract: Objective: To screen a high trehalose-producing bacterium from Volvariella volvacea cultivation substrate and optimize its fermentation conditions, thereby providing a microbial resource and technical basis for the large-scale production of trehalose. Methods: Bacterial strains were isolated and purified from Volvariella volvacea cultivation substrate. A high trehalose-producing strain was screened using trehalose content in the fermentation broth as the evaluation index. The selected strain was identified based on morphological characteristics, physiological and biochemical properties, and 16S rRNA gene sequence analysis. Single-factor experiments were conducted to optimize the fermentation conditions, including carbon source, nitrogen source, metal salt type and concentration, inoculum size, cultivation time, cultivation temperature, and initial medium pH. Results: A high trehalose-producing Bacillus sp. was isolated from Volvariella volvacea cultivation substrate. The optimal fermentation conditions were as follows: glucose as the carbon source, yeast extract as the nitrogen source, and sodium chloride as the metal salt, with concentrations of 2 mg/mL glucose, 1.5 mg/mL yeast extract, and 1 mg/mL sodium chloride, respectively; inoculum size of 4%, cultivation time of 24 h, cultivation temperature of 29 °C, and initial medium pH of 7. Under these optimized conditions, the trehalose concentration in the fermentation broth increased from 6.234 mg/mL to 17.623 mg/mL, representing an increase of 182.6% compared with that before optimization. Conclusion: The screened Bacillus sp. exhibited a high trehalose production capacity under the optimized culture conditions and shows considerable potential for further development as a microbial resource for large-scale trehalose production.