This study investigates the relationship between gut microbiota and disease resistance in piglets, focusing on the impact of Porcine Epidemic Diarrhea Virus (PEDV) infection. Single-cell transcriptomics, 16S amplicon sequencing, metagenomics, and untargeted metabolomics were used to explore the gut microbiota's role in PEDV infection. Key findings include: 1. **Gut Microbiota Changes After PEDV Infection**: PEDV infection significantly alters the gut microbiota of piglets, with Landrace pigs losing their resistance more quickly compared to Min pigs. Transplanting the fecal microbiota of Min pigs to Landrace pigs improved their infection status. 2. **Role of Specific Bacteria**: *Lactobacillus reuteri* and *Lactobacillus amylovorus* were identified as playing an anti-infective role in the gut microbiota. Metabolomic screening revealed that deoxycholic acid (DCA) and lithocholic acid (LCA), secondary bile acids, were significantly correlated with these bacteria, but only LCA exhibited protective effects in animal models. 3. **LCA's Protective Mechanism**: LCA supplementation altered the distribution of intestinal T-cell populations, increasing the number of CD8+ cytotoxic T lymphocytes (CTLs). In vitro and in vivo experiments showed that LCA increased SLA-I expression in porcine intestinal epithelial cells via FXR receptors, recruiting CD8+ CTLs to exert antiviral effects. 4. **Conclusion**: The diversity of gut microbiota influences disease development, and manipulating *Lactobacillus reuteri* and *Lactobacillus amylovorus*, as well as LCA, represents a promising strategy to improve PEDV infection in piglets. Keywords: Infection resistance, Gut microbiota, Macrogenomic, Metabolomic, T cell responseThis study investigates the relationship between gut microbiota and disease resistance in piglets, focusing on the impact of Porcine Epidemic Diarrhea Virus (PEDV) infection. Single-cell transcriptomics, 16S amplicon sequencing, metagenomics, and untargeted metabolomics were used to explore the gut microbiota's role in PEDV infection. Key findings include: 1. **Gut Microbiota Changes After PEDV Infection**: PEDV infection significantly alters the gut microbiota of piglets, with Landrace pigs losing their resistance more quickly compared to Min pigs. Transplanting the fecal microbiota of Min pigs to Landrace pigs improved their infection status. 2. **Role of Specific Bacteria**: *Lactobacillus reuteri* and *Lactobacillus amylovorus* were identified as playing an anti-infective role in the gut microbiota. Metabolomic screening revealed that deoxycholic acid (DCA) and lithocholic acid (LCA), secondary bile acids, were significantly correlated with these bacteria, but only LCA exhibited protective effects in animal models. 3. **LCA's Protective Mechanism**: LCA supplementation altered the distribution of intestinal T-cell populations, increasing the number of CD8+ cytotoxic T lymphocytes (CTLs). In vitro and in vivo experiments showed that LCA increased SLA-I expression in porcine intestinal epithelial cells via FXR receptors, recruiting CD8+ CTLs to exert antiviral effects. 4. **Conclusion**: The diversity of gut microbiota influences disease development, and manipulating *Lactobacillus reuteri* and *Lactobacillus amylovorus*, as well as LCA, represents a promising strategy to improve PEDV infection in piglets. Keywords: Infection resistance, Gut microbiota, Macrogenomic, Metabolomic, T cell response