Effects of Fermented Compound Chinese Herbal Feed on Gut Microbiota, Immune Response, and Disease Resistance in Chinese Soft-Shelled Turtle (Pelodiscus sinensis)

In this study, Chinese medicinal herbs were evaluated as potential antibiotic substitutes for Chinese soft-shelled turtle (Pelodiscus sinensis). Forty-five herbs were initially screened for antibacterial activity against Salmonella enteritidis, Escherichia coli, and Shigella flexneri. Nine herbs exhibiting broad-spectrum inhibitory effects were selected and subjected to microbial fermentation, after which their antibacterial activities were reassessed and applied as dietary supplements in feeding trials. The results showed that fermentation altered the antibacterial activities of several herbs and enhanced their overall functional performance. Dietary supplementation with fermented Chinese herbal medicine did not adversely affect feed utilization but significantly improved hematological parameters, liver and kidney function indicators, antioxidant capacity, and nonspecific immune responses. Furthermore, turtles fed fermented herbal diets exhibited higher survival rates following bacterial challenge. Intestinal microbiota analysis based on 16S rRNA gene sequencing indicated that fermented herbal supplementation modulated microbial community structure by reducing potential pathogens and increasing beneficial bacterial taxa associated with intestinal health. These findings suggest that microbial fermentation effectively enhances the biological efficacy of Chinese medicinal herbs. Fermented herbal feed additives represent a promising green alternative to antibiotics for soft-shelled turtle aquaculture. The global ban on prophylactic antibiotics drives the need for safe, effective feed alternatives. Microbial fermentation of Chinese herbs (FCM) is proposed to enhance efficacy and detoxification, but its comprehensive effects in aquaculture require deeper investigation. This study evaluated compound unfermented (CM) and fermented (FCM) Chinese herbal supplements on the Chinese soft-shelled turtle (Pelodiscus sinensis). Initial screening showed fermentation generally enhanced the antibacterial activity of the herbs against common enteric pathogens (S. enteritidis, E. coli, S. flexneri). Results indicated that the FCM diet significantly improved physiological status, leading to higher red blood cell counts, better liver/kidney function (reduced ALT/AST, UREA), and stronger immune/antioxidant responses (increased Lysozyme and T-AOC) compared to CM or control diets. Critically, the FCM group achieved the highest survival rates across all single and combined pathogen challenges, demonstrating superior protective efficacy. Furthermore, FCM effectively modulated the gut microbiota, enriching beneficial fermentative bacteria. In conclusion, microbial fermentation significantly amplifies the health-promoting and protective benefits of Chinese herbal supplements in soft-shelled turtles, positioning FCM as a promising green alternative for disease control in aquaculture.