Metagenomic sequencing of bile from gallstone patients to identify different microbial community patterns and novel biliary bacteria.

Hongzhang Shen,Lei Li,Ying Chen,Peisong Xu,Fei Meng,Jianfeng Yang,Fuqiang Ye,Xiaochen Bo,Ming Ni,Xiaofeng Zhang,Lu Xie,Zhen Li

doi:10.1038/srep17450

Abstract

Despite the high worldwide prevalence of gallstone disease, the role of the biliary microbiota in gallstone pathogenesis remains obscure. Next-generation sequencing offers advantages for systematically understanding the human microbiota; however, there have been few such investigations of the biliary microbiome. Here, we performed whole-metagenome shotgun (WMS) sequencing and 16S rRNA sequencing on bile samples from 15 Chinese patients with gallstone disease. Microbial communities of most individuals were clustered into two types, according to the relative enrichment of different intestinal bacterial species. In the bile samples, oral cavity/respiratory tract inhabitants were more prevalent than intestinal inhabitants and existed in both community types. Unexpectedly, the two types were not associated with fever status or surgical history, and many bacteria were patient-specific. We identified 13 novel biliary bacteria based on WMS sequencing, as well as genes encoding putative proteins related to gallstone formation and bile resistance (e.g., β-glucuronidase and multidrug efflux pumps). Bile samples from gallstone patients had reduced microbial diversity compared to healthy faecal samples. Patient samples were enriched in pathways related to oxidative stress and flagellar assembly, whereas carbohydrate metabolic pathways showed varying behaviours. As the first biliary WMS survey, our study reveals the complexity and specificity of biliary microecology.

Highlights

De novo formation of brown pigment stones (BPSs) in bile ducts is generally thought to be related to biliary bacteria/parasite infection and partial biliary tract obstruction[2,9]
We found that oral cavity and respiratory tract inhabitants were more prevalent than intestinal inhabitants in the bile samples
The 11 intestinal inhabitants were basically distributed in Cluster I, which was highly enriched in E. coli

Summary

Introduction

De novo formation of BPSs in bile ducts is generally thought to be related to biliary bacteria/parasite infection and partial biliary tract obstruction[2,9]. Many bacteria, such as Escherichia coli, Klebsiella pneumoniae, Enterococcus faecium, Enterobacter cloacae and Pseudomonas aeruginosa, have been identified in bile or gallstone samples through cultivation or polymerase chain reaction (PCR). Wu et al performed bacterial 16S rRNA amplicon sequencing (referred to as ‘16S sequencing’) on gallbladder bile, gallstones and faeces from cholesterol gallstone patients[16] They detected many gut bacterial operational taxonomic units (OTUs) in the biliary tract and found that faecal samples from gallstone patients exhibited microbiota dysbiosis. We investigated genes possibly related to gallstone formation and bile resistance, and the metabolic network of the biliary microbiota was reconstructed

Methods

Results

Discussion

Conclusion