Tissue Microbiome Profiling Identifies an Enrichment of Specific Enteric Bacteria in Opisthorchis viverrini Associated Cholangiocarcinoma

• Stenotrophomonas, implicated in bile duct infections, is enriched in tumor tissues of non-fluke related cholangiocarcinoma.
• O. viverrini infection may alter composition of bile duct tissue microbiomes.
• Enteric bacteria with metabolic outputs linked to carcinogenesis are enriched in O. viverrini associated tissue microbiomes.The link between microbiota and cancer of the gastrointestinal (GI) tract has been extensively studied. However, the role of tissue microbiome in cholangiocarcinoma (CCA), cancer of the bile duct (an organ connected to the GI tract), is largely unknown. In this study, we detected intriguing compositional differences in the tissue microbiomes of liver fluke related and non-related CCA. Taken together, our data suggests a connection between parasitic infections, tissue microbiome alterations, tissue micro-environment changes and CCA development.

Cholangiocarcinoma (CCA) is the primary cancer of the bile duct system. The role of bile duct tissue microbiomes in CCA tumorigenesis is unestablished. To address this, sixty primary CCA tumors and matched normals, from both liver fluke (Opisthorchis viverrini) associated (OVa, n = 28) and non-O. viverrini associated (non-OVa, n = 32) cancers, were profiled using high-throughput 16S rRNA sequencing. A distinct, tissue-specific microbiome dominated by the bacterial families Dietziaceae, Pseudomonadaceae and Oxalobacteraceae was observed in bile duct tissues. Systemic perturbation of the microbiome was noted in tumor and paired normal samples (vs non-cancer normals) for several bacterial families with a significant increase in Stenotrophomonas species distinguishing tumors vs paired normals. Comparison of parasite associated (OVa) vs non-associated (non-OVa) groups identified enrichment for specific enteric bacteria (Bifidobacteriaceae, Enterobacteriaceae and Enterococcaceae). One of the enriched families, Bifidobacteriaceae, was found to be dominant in the O. viverrini microbiome, providing a mechanistic link to the parasite. Functional analysis and comparison of CCA microbiomes revealed higher potential for producing bile acids and ammonia in OVa tissues, linking the altered microbiota to carcinogenesis. These results define how the unique microbial communities resident in the bile duct, parasitic infections and the tissue microenvironment can influence each other, and contribute to cancer.