[Research advances of relationship between non-alcoholic fatty liver disease and biliary tract diseases].

Recent studies have found that non-alcoholic fatty liver disease(NAFLD) has great impact on the development of biliary tract diseases. Here in this review, we summarized the relationship between NAFLD and the occurrence and development, risk factors and severity of cholestasis, gallstones, intrahepatic cholangiocarcinoma, primary biliary cirrhosis and bile microbiota, so as to further illuminate the pathogenesis of NAFLD and biliary tract diseases, obtain better diagnostic and therapeutic outcomes on NAFLD and biliary tract diseases.

The dysregulation of biliary tract microflora is closely related to primary choledocholithiasis: a multicenter study.

Bile microecology changes play an important role in the occurrence and development of choledocholithiasis. At present, there is no clear report on the difference of bile microecology between asymptomatic patients with gallbladder polyps and choledocholithiasis. This study compared bile microecology between gallbladder polyp patients and patients with choledocholithiasis to identify risk factors for primary choledocholithiasis. This study was conducted in 3 hospitals in different regions of China. Bile samples from 26 patients with gallbladder polyps and 31 patients with choledocholithiasis were collected by laparoscopic cholecystectomy and endoscopic retrograde choledocholithiasis cholangiography (ERCP), respectively. The collected samples were used for 16S ribosomal RNA sequencing and liquid chromatography mass spectrometry analysis. The α-diversity of bile microecological colonies was similar between gallbladder polyp and choledocholithiasis, but the ß-diversity was different. Firmicutes, Proteobacteri, Bacteroidota and Actinobacteriota are the most common phyla in the gallbladder polyp group and choledocholithiasis group. However, compared with the gallbladder polyp patients, the abundance of Actinobacteriota has significantly lower in the choledocholithiasis group. At the genera level, the abundance of a variety of bacteria varies between the two groups, and Enterococcus was significantly elevated in choledocholithiasis group. In addition, bile biofilm formation-Pseudomonas aeruginosa was more metabolically active in the choledocholithiasis group, which was closely related to stone formation. The analysis of metabolites showed that a variety of metabolites decreased in the choledocholithiasis group, and the concentration of beta-muricholic acid decreased most significantly. For the first time, our study compared the bile of gallbladder polyp patients with patients with choledocholithiasis, and suggested that the change in the abundance of Actinobacteriota and Enterococcus were closely related to choledocholithiasis. The role of Pseudomonas aeruginosa biofilm in the formation of choledocholithiasis was discovered for the first time, and some prevention schemes for choledocholithiasis were discussed, which has important biological and medical significance.

Characteristics of bile microbiota in cholelithiasis, perihilar cholangiocarcinoma, distal cholangiocarcinoma, and pancreatic cancer.

Significant proof suggests an essential role played by the bile microbiota in biliary diseases. This study retrospectively analyzed the differences in biliary microbes among patients with perihilar cholangiocarcinoma (pCCA), distal cholangiocarcinoma (dCCA), pancreatic cancer (PC), and cholelithiasis (CH). Bile samples were obtained from 53 patients who underwent endoscopic retrograde cholangiopancreatography (ERCP), and the bile microbiota was analyzed through 16S rRNA gene analysis and next-generation sequencing. Based on the results of linear discriminant analysis effect size (LEfSe), the top three biomarkers for pCCA at the genus level were Pseudomonas, Sphingomonas, and Halomonas; for dCCA were Streptococcus, Prevotella, and Halomonas; and for PC were Pseudomonas, Chloroplast, and Acinetobacter. The top five genera in the pCCA, dCCA, and PC groups showed predictive values with areas under the receiver operating characteristic curves of 91.56%, 95.56%, and 96.59%, respectively. The PICRUSt2 analysis outcomes displayed the diversities of fifteen pathways between the CH and pCCA groups, 22 pathways between the CH and dCCA groups, and eighteen pathways between the CH and PC groups. As this pilot study identified specific microbial bile markers for patients with CH, pCCA, dCCA, and PC, the clinical implications are vast. Further study focusing on distinct bacterial populations in bile will help differentiate biliary diseases.

Screening of bacterial DNA in bile sampled from healthy dogs and dogs suffering from liver- or gallbladder-associated disease.

Although the biliary system is generally aseptic, gallbladder microbiota has been reported in humans and some animals apart from dogs. We screened and analyzed the bacterial deoxyribonucleic acid in canine gallbladders using bile sampled from 7 healthy dogs and 52 dogs with liver- or gallbladder-associated disease. PCR screening detected bacteria in 17.3% of diseased dogs (9/52) and none in healthy dogs. Microbiota analysis of PCR-positive samples showed that the microbial diversity differed between liver- and gallbladder-associated disease groups. Thus, a specific bacterial community appears to occur at a certain frequency in the bile of diseased dogs.

ABO-Incompatible Liver Transplantation under the Desensitization Protocol with Rituximab: Effect on Biliary Microbiota and Metabolites.

Background: ABO-incompatible liver transplantation (ABOi LT) under the desensitization protocol with rituximab had excellent survival outcomes comparable to those of ABO-compatible liver transplantation (ABOc LT). In this work, we explored the effect of ABOi LT on recipients from the perspective of biliary microbiota and metabonomics. Methods: Liver transplant (LT) recipients treated at our center were enrolled in the study. In total, 6 ABOi LT recipients and 12 ABOc LT recipients were enrolled, and we collected their bile five times (during LT and at 2 days, 1 week, 2 weeks and 1 month after LT). The collected samples were used for 16S ribosomal RNA sequencing and liquid chromatography mass spectrometry analysis. Results: We obtained 90 bile samples. Whether in group ABOi LT or ABOc LT, the most common phyla in all of the samples were Firmicutes, Proteobacteria, Bacteroidetes and Actinobacteria. The most common genera were Lactobacillus, Weissella, Klebsiella, Pantoea and Lactococcus. There was no significant difference in the diversity between the two groups at 1 week, 2 weeks and 1 month after LT. However, the biggest disparities between the ABOi LT recipients and ABOc LT recipients were observed 2 days after LT, including increased biodiversity with a higher ACE, Chao1, OBS and Shannon index (p < 0.05), and more Staphylococcus in ABOi LT and binary−Jaccard dissimilarity, which indicated varying ß-diversity (p = 0.046). These differences were not observed at 1 week, 2 weeks and 1 month after LT. The principal coordinate analysis (PCoA) revealed that the composition of the bile microbiota did not change significantly within 1 month after LT by longitudinal comparison. In an analysis of the bile components, the metabolites were not significantly different every time. However, four enrichment KEGG pathways were observed among the groups. Conclusion: These findings suggest that ABOi LT under the desensitization protocol with rituximab did not significantly affect the biliary microbiota and metabolites of recipients.

Dysbiosis of the Fecal and Biliary Microbiota in Biliary Tract Cancer.

Characteristic bile duct and gut microbiota have been identified in patients with chronic biliary tract disease. This study aimed to characterize the fecal and bile microbiota in biliary tract cancer (BTC) patients and their relationship. Patients with BTC (n = 30) and benign biliary disease (BBD) without cholangitis (n = 11) were included. Ten healthy, age-matched subjects were also recruited for fecal microbiota comparison. The fecal and bile duct microbiotas were analyzed by sequencing the 16S rRNA gene V3-V4 region. Live bacteria were obtained in the bile from three BTC patients by culture, and metagenomics-based identification was performed. Linear discriminant analysis effect size showed a higher Enterobacteriaceae abundance and a lower Clostridia abundance, including that of Faecalibacterium and Coprococcus, in the BTC patients than in the other subjects. Ten of 17 operational taxonomic units (OTUs) assigned to Enterobacteriaceae in the bile were matched with the OTUs found in the BTC subject fecal samples. Furthermore, a bile-isolated strain possessed the carcinogenic bacterial colipolyketide synthase-encoding gene. Enterobacteriaceae was enriched in the BTC feces, and more than half of Enterobacteriaceae in the bile matched that in the feces at the OTU level. Our data suggests that fecal microbiota dysbiosis may contribute to BTC onset.

Bile microbiota: new insights into biliary complications in liver transplant recipients.

BACKGROUND: Biliary complications represent a major problem associated with liver transplantation. This report represents the first study to use high-throughput 16S ribosomal RNA (rRNA) gene sequencing to assess bile microbiota within bile samples of liver transplant recipients with biliary complications. Our goal in this report was to identify the species and abundance of microbes and examine the potential for microbial involvement of bile in liver transplantation patients with biliary complications. METHODS: Liver transplant recipients treated at our center over the period from September 2015 to June 2017 were enrolled in the study. Patients satisfying the inclusion criteria were divided into two groups, control (N=13) and symptom (N=10). Sequencing of 16s rDNA was then performed on bile samples from both groups. RESULTS: The main bacterial phyla of bile samples in the symptom group included Proteobacteria (55.19%), Firmicutes (32.36%), Actinobacteria (10.24%) and Bacteroidetes (1.23%) and the main bacterial genera were Pseudomonas (23.31%), Klebsiella (18.42%), Lactococcus (9.61%), Rhodococcus (9.59%) and Rhizobium (5.08%). Proteobacteria and Staphylococcus were enriched in the symptom group (P<0.05), whereas Firmicutes (P<0.05) and Enterococcus (P<0.01) were enriched in the control group. Pathways involved as determined with use of the Kyoto Encyclopedia of Genes and Genomes (KEGG), revealed that metabolism pathways of glyoxylate and dicarboxylate, porphyrin and chlorophyll, arginine and proline, glycine, serine and threonine, as well as the bacterial secretion system were all enriched in bile samples from the symptom group (P<0.05). CONCLUSIONS: Clear differences exist in microbial species distribution in bile samples from the symptom versus control group. The species and pathways enriched in bile samples within the symptom group may be involved in the pathogenesis of biliary complication after liver transplantation.

The human gallbladder microbiome is related to the physiological state and the biliary metabolic profile.

BACKGROUND: The microbial populations of the human intestinal tract and their relationship to specific diseases have been extensively studied during the last decade. However, the characterization of the human bile microbiota as a whole has been hampered by difficulties in accessing biological samples and the lack of adequate methodologies to assess molecular studies. Although a few reports have described the biliary microbiota in some hepatobiliary diseases, the bile microbiota of healthy individuals has not been described. With this in mind, the goal of the present study was to generate fundamental knowledge on the composition and activity of the human bile microbiota, as well as establishing its potential relationship with human bile-related disorders. RESULTS: Human bile samples from the gallbladder of individuals from a control group, without any record of hepatobiliary disorder, were obtained from liver donors during liver transplantation surgery. A bile DNA extraction method was optimized together with a quantitative PCR (qPCR) assay for determining the bacterial load. This allows the selection of samples to perform functional metagenomic analysis. Bile samples from the gallbladder of individuals suffering from lithiasis were collected during gallbladder resection and the microbial profiles assessed, using a 16S rRNA gene-based sequencing analysis, and compared with those of the control group. Additionally, the metabolic profile of the samples was analyzed by nuclear magnetic resonance (NMR). We detected, for the first time, bacterial communities in gallbladder samples of individuals without any hepatobiliary pathology. In the biliary microecosystem, the main bacterial phyla were represented by Firmicutes, Bacteroidetes, Actinobacteria, and Proteobacteria. Significant differences in the relative abundance of different taxa of both groups were found. Sequences belonging to the family Propionibacteriaceae were more abundant in bile samples from control subjects; meanwhile, in patients with cholelithiasis members of the families Bacteroidaceae, Prevotellaceae, Porphyromonadaceae, and Veillonellaceae were more frequently detected. Furthermore, the metabolomics analysis showed that the two study groups have different metabolic profiles. CONCLUSIONS: Our results indicate that the gallbladder of human individuals, without diagnosed hepatobiliary pathology, harbors a microbial ecosystem that is described for the first time in this study. Its bacterial representatives and metabolites are different from those detected in people suffering from cholelithiasis. In this regard, since liver donors have been subjected to the specific conditions of the hospital's intensive care unit, including an antibiotic treatment, we must be cautious in stating that their bile samples contain a physiologically normal biliary microbiome. In any case, our results open up new possibilities to discover bacterial functions in a microbial ecosystem that has not previously been explored.

Research advances of relationship between non-alcoholic fatty liver disease and biliary tract diseases / 中华肝脏病杂志

Recent studies have found that non-alcoholic fatty liver disease(NAFLD) has great impact on the development of biliary tract diseases. Here in this review, we summarized the relationship between NAFLD and the occurrence and development, risk factors and severity of cholestasis, gallstones, intrahepatic cholangiocarcinoma, primary biliary cirrhosis and bile microbiota, so as to further illuminate the pathogenesis of NAFLD and biliary tract diseases, obtain better diagnostic and therapeutic outcomes on NAFLD and biliary tract diseases.