Isolation, identification and characteristics of <i>Clostridium butyricum</i> CB-a together with its high-density fermentation

Clostridium butyricum is a strictly anaerobic probiotic widely used in food, medicine, and agriculture. However, its industrial application is limited by low fermentation yields and cultivation difficulties. In this study, a novel strain, C. butyricum CB-a, was isolated from free-range chicken feces and exhibited notable probiotic traits. It maintained 25.0% and 32.5% survival rates after 2-h exposure to simulated gastric and intestinal juices, respectively, and over 42.0% survival in 0.2%–0.5% bile salt solutions. Moreover, it showed clear antimicrobial activity against Escherichia coli and Staphylococcus aureus. To enhance biomass production, one-factor and orthogonal optimization experiments were conducted. Glucose was selected as the optimal carbon source, and soy peptone, yeast extract, and peptone as key nitrogen sources. The optimized medium, combined with suitable pH (6.0), inoculum ratio (3%), and temperature (34 °C), enabled a viable cell count of 4.56 × 10⁹ CFU/mL after 24 h. Although this value is slightly below the conventional high-density threshold, it represents a substantial increase compared to previous C. butyricum reports. This study provides a practical and economical strategy for cultivating C. butyricum with industrial potential and lays the groundwork for its future probiotic application.