Microbial Composition Study in Cholangiocarcinoma: Stage-Specific Diversity Analysis in Bile and Tumor Microbiome

Cholangiocarcinoma (CCA) is an aggressive malignancy of the biliary tract with poor prognosis and limited early diagnostic tools. Microbial dysbiosis has been implicated in carcinogenesis, yet microbial signatures across disease stages and anatomical sites in CCA remain poorly defined. This study aimed to characterize the microbial diversity in bile and tumor tissue of CCA patients and evaluate association with disease stage.

Microbiome profiles were analyzed from 36 subjects using 16Sr RNA gene sequencing data from the Sequence Read Archive (SRA). Alpha diversity (Chao1 index) and beta diversity (Bray-Curtis dissimilarity) were assessed across bile and intratumoral samples. Linear discriminant analysis effect size (LEfSe) was employed to identify stage-specific microbial taxa.

Alpha diversity did not differ significantly between disease stages (ANOVA, F = 0.385, p = 0.816), suggesting stable microbial richness throughout progression. However, beta diversity analysis (PERMANOVA, R² = 0.279, p = 0.013) revealed distinct clustering by sample type and tumor stage. LEfSe identified enrichment of Fusobacterium, Escherichia-Shigella, and Enterococcus in advanced-stage tumor samples, while Lactobacillus and Streptococcus were more abundant in early-stage bile samples.

This study demonstrates distinct microbial composition patterns across disease stages and anatomical sites in CCA. Enrichment of specific pathogenic taxa in advanced stages supports a role for microbiome alterations in CCA pathogenesis. Microbial markers may serve as potential tools for staging and prognosis, warranting further functional and prospective validation studies.