Objective: To identify the species and abundance of microbes in the bile of patients with biliary injury after liver transplantation. To explore the potential microbial involvement, we evaluated the differences in biliary microbiota in patients with and without biliary injury after liver transplantation. Methods: Bile was collected by percutaneous transhepatic cholangiography drainage from 5 liver transplant recipients with biliary injury (case group) and from the T-tube in 4 liver transplant recipients without biliary injury (control group). 16S rRNA sequencing was performed on the bile samples. Results: The biliary bacterial phyla in patients after liver transplantation were Firmicutes, Proteobacteria, Actinobacteria, Bacteroidetes, and Acidobacteria in order of abundance from highest to lowest. There were differences in genus Prevotella (P = 0.03), Porphyromonas (P = 0.02), and Fusobacterium (P = 0.02) between the two groups, which were significantly abundant in the case group. There was no significant difference in the diversity analysis between the two groups (P > 0.05). The terpenoid backbone biosynthesis pathway and the histidine degradation pathway were significantly enriched in the bile samples in the case group. Compared with the control group, the pyruvate ferredoxin oxidoreductase pathway was depleted in the samples from the case group. Conclusion: This is the first bile microbiota report using high-throughput 16S rRNA gene sequencing of bile material in liver transplant recipients. There were significant differences in bacterial abundance between patients with and without biliary tract injury after liver transplantation but no significant differences in the diversity. KEGG analysis showed that there were differences in metabolic pathways between the two groups.