The effect of tannin on the gut microbiome of mice infected with Clostridioides difficile. Antimicrobial activity of tannic acid in vitro and its protective effect on mice against Clostridioides difficile

Background: Clostridioides difficile infection (CDI), recurrently reported as an urgent threat owing to its increased prevalence and mortality, has attracted significant attention. As the use of antibiotics to treat CDI has many limitations, such as high recurrence rate, the need to actively seek and develop other drugs that can effectively treat CDI with fewer side effects has become a key issue in CDI prevention and treatment. Objectives: This study aimed to evaluate the inhibitory effect of Galla Chinensis extract tannic acid (TA) against C. difficile in vitro and its therapeutic effect on CDI in vivo. Methods: Agar dilution assays were used to determine TA minimum inhibitory concentrations (MICs) against C. difficile. Sporulation test, ELISA, and Crystal Violet staining method were used to detect the effects of TA on sporulation, toxin production and biofilm formation, respectively. RT-qPCR was used to determine the effect of TA on the transcription of key sporulation gene spo0A and toxin-encoding genes tcdA and tcdB. CDI mice were treated with TA and observed for 30 days, and the survival rate, mean relative weight, clinical score, and serum levels of toxins A/B, IL-6 and IL-10 were measured. Fecal specimens of mice were used to count vegetative cells and spores, and the intestinal microbiota was analyzed by 16S rRNA sequencing. Results: Sub-MIC of TA significantly inhibited C. difficile sporulation, toxin production and biofilm formation in vitro. Treatment with 1/4MIC TA, reduced tcdA, tcdB and spo0A mRNA expression in C. difficile. When compared with the CDI control group, TA-treated mice lost less weight and presented significantly improved survival rate. TA significantly reduced the number of vegetative cells and spores, decreased serum TcdA level, and increased serum IL-10 level.