Bile Microbiota in Patients with Pigment Common Bile Duct Stones.

BACKGROUND: Common bile duct (CBD) stone is one of the most prevalent gastroenterological diseases, but the role played by biliary microbiota in the pathogenesis of CBD stones remains obscure. The aim of this study was to investigate the characteristics of the biliary tract core microbiome and its potential association with the formation of pigment stones. METHODS: Twenty-eight patients with biliary obstruction of various causes were enrolled. Thirteen had new-onset pigment CBD stone. Of the remaining 15, four had benign biliary stricture, four had gallbladder cancer, three had pancreatic cancer, 3 had distal CBD cancer, and one had hepatocellular carcinoma. Endoscopic retrograde cholangiopancreatography was used to collect bile samples for DNA extraction, 16S ribosomal RNA gene sequencing, and bile microbiota composition analysis. RESULTS: Proteobacteria (61.7%), Firmicutes (25.1%), Bacteroidetes (5%), Fusobacteria (4.6%), and Actinobacteria (2.6%) were the most dominant phyla in the bile of the 28 study subjects. A comparison between new-onset choledocholithiasis and other causes of biliary obstruction (controls) showed Enterococcus was found to be significantly abundant in the CBD stone group at the genus level (linear discriminant analysis score = 4.38; P = 0.03). However, no other significant compositional difference was observed. CONCLUSION: This study demonstrates an abundance of microbiota in bile juice and presents a biliary microbiome composition similar to that of duodenum. The study also shows Enterococcus was significantly abundant in the bile juice of patients with a brown pigment stone than in controls, which suggests Enterococcus may play an important role in the development of pigment stones.

[MORPHOLOGICAL CHANGES OF THE LIVER IN OBTURATION JAUNDICE, CAUSED BY CHOLEDOCHOLITHIASIS, DEPENDING ON ITS DURATION].

Abstract The results of surgical treatment of 184 patients for obturation jaundice, caused by choledocholithiasis, were analyzed. Morphological changes of the liver were studied in 20 patients. There were three groups of patients delineated, depending on the obturation jaundice duration: up to 7 days, from 8 to 14 days, more than 15 days, and also a group of patients after the bile outflow restoration. The obturation jaundice occurrence in choledocholithiasis is accompanied by significant morphological changes in the liver, severity of which is enhancing while the obturation jaundice persistence increasing. While persistence of obturation jaundice through 8 days and more the connective tissue volume is enhancing, a relative volume of hepatocytes is reducing and a stromal-parenchymatous index is increasing. The bile outflow restoration secures significant reduction of intensity of alterative and inflammatory changes in hepatic parenchyma, as well as activation of reparative processes in the tissue. In cholangitis, caused by P. aeruginosa and E. coli, according to morphological investigations data, in the liver a diffuse purulent cholangitis on background of chronic changes in accordance to duration of the obturation jaundice persists.

[MICROBIAL LANDSCAPE OF BILE IN PATIENTS WITH AN ACUTE PURULENT CHOLANGITIS].

Bacteriological analysis was conducted in 136 patients with an acute purulent cholangitis (APCH). The APCH causes were: choledocholithiasis--in 40 (29.9%) patients, coexistence of a common biliary duct stricture and choledocholithiasis--in 39 (28.7%), compression of external biliary ducts by the oedematous pancreatic head in secondary pancreatitis--in 15 (11%), pericholedocheal lymphadenitis--in 3 (2.2%).

Diagnostic and post-treatment CT appearance of biopsy proven mixed Cryptococcus and Candida cholangitis.

Cryptococcus neoformans and Candida are common opportunistic pathogens of human. There is very limited literature on Cryptococcus neoformans cholangitis or Candida cholangitis in immunocompetent patient while mixed Cryptococcus neoformans and Candida cholangitis has not been reported in the literature ever before. We hereby report the imaging findings of a case of mixed Cryptococcus neoformans and Candida cholangitis in an immunocompetent boy. The CT features included nodules in the cystic duct and common bile duct, dilatation of the intra- and extrahepatic bile ducts with mural thickening, irregular hypodense mass-like lesion extending along the bile ducts from the liver hilum to the periphery which was confirmed by the presence of enlarged and confluent lymph nodes. CT characteristics can contribute to timely diagnosis and treatment of this disease.

Characterization of the complex bacterial communities colonizing biliary stents reveals a host-dependent diversity.

This study provides a comprehensive survey of the spatial and temporal bacterial composition of biliary stent biofilms. The bacterial diversity, distribution and dynamics of 59 biliary and 4 pancreatic stent communities from 40 patients being treated at two different hospitals, which implant stents either simultaneously or consecutively, were characterized by single-strand conformation polymorphism (SSCP) analysis. Fifty-one phylotypes belonging to 5 bacterial phyla and 24 bacterial families were detected across 63 stents. This is a much broader diversity than previously detected through culture-dependent methods, particularly in regard to the diversity of obligate anaerobes. Stent bacterial diversity was patient-dependent and more similar when stents were implanted simultaneously rather than consecutively. Stent bacterial community composition differed between hospitals specifically because of the difference in abundance of Bifidobacteria. Co-colonization of Veillonella sp., Streptococcus anginosus and organisms closely related to Fusobacterium nucleatum revealed a potentially important attachment and survival strategy that has yet to be reported in biliary stents. This work reveals a more complete survey of the identities of bacterial species that form biofilms in biliary stents, their co-colonization patterns and the natural variation in species composition between different patients, hospitals and locations along the stent. Consideration of the community composition from individual patients will allow tailoring of prophylactic antibiotic treatments and thus will make the management of stent biofilms more effective.

Parásitos de fasciola hepática intracoledociano / Common bile duct fasciolasis: report of one case

La fasciolasis es una enfermedad parasitaria producida por el tremátodo digeneo Fasciola hepática. Durante la fase crónica de la enfermedad los parásitos pueden entran en los conductos biliares, y ser causa de ictericia obstructiva, colangitis, pancreatitis y otras complicaciones. Se presenta el caso clínico de una paciente intervenida por sospecha de colelitiasis y colédocolitiasis, a la cual durante la exploración coledociana intraoperatoria le fueron extraídos múltiples parásitos adultos de Fasciola hepática. Este hallazgo sólo fue aclarado posteriormente en la biopsia diferida. Se realiza una revisión breve del tema, métodos de diagnóstico, tratamiento médico y alternativas quirúrgicas de tratamiento.

Fasciolasis is a zoonotic disease caused by the digenean trematode Fasciola hepatica. During the chronic phase of the disease the parasites invade the main biliary ducts, causing obstructive jaundice, cholangitis, pancreatitis and other complications. We report a 75 years old female, operated due to the suspicion of a choledocholithiasis. During the exploration of the choledochus, adult F. hepatica parasites were extracted. The pathology report of the parasites confirmed the diagnosis.

Helicobacter species ribosomal DNA in the pancreas, stomach and duodenum of pancreatic cancer patients.

AIM: To determine whether gastric and enteric Helicobacter species are associated with pancreatic cancer. METHODS: Patients with exocrine pancreatic cancer (n = 40), neuroendocrine cancer (n = 14), multiple endocrine neoplasia type 1 (n = 8), and chronic pancreatitis (n = 5) were studied. Other benign pancreatic diseases (n = 10) and specimens of normal pancreas (n = 7) were included as controls. Pancreatic tissue specimens were analyzed by Helicobacter-specific PCR-assay and products were characterized by denaturing gradient electrophoresis and DNA-sequencing. From a subset of the pancreatic cancer patients, gastric and/or duodenal tissue as well as gallbladder and ductus choledochus tissue were analyzed. Gallbladder and choledochus samples were included as controls. Stomach and duodenum samples were investigated to analyze whether a gastric helicobacter might disseminate to the pancreas in pancreatic cancer patients. Pancreatic specimens were analyzed by Bacteroides-specific PCR for detecting the translocation of indigenous gut microbes to the diseased pancreas. RESULTS: Helicobacter DNA was detected in pancreas (tumor and/or surrounding tissue) of 75% of patients with exocrine cancer, 57% of patients with neuroendocrine cancer, 38% of patients with multiple endocrine neoplasia, and 60% of patients with chronic pancreatitis. All samples from other benign pancreatic diseases and normal pancreas were negative. Thirty-three percent of the patients were helicobacter-positive in gastroduodenal specimens. Surprisingly, H. bilis was identified in 60% of the positive gastroduodenal samples. All gallbladder and ductus choledochus specimens were negative for helicobacter. Bacteroides PCR-assay was negative for all pancreatic samples. CONCLUSION: Helicobacter DNA commonly detected in pancreatic cancer suggests a possible role of the emerging pathogens in the development of chronic pancreatitis and pancreatic cancer.

Loss of the major duodenal papilla results in brown pigment biliary stone formation in pdx1 null mice.

BACKGROUND & AIMS: Pdx1 plays a pivotal role in pancreas organogenesis and specification of some types of cells in the duodenum and antral stomach. However, its expression is not restricted to pancreas, duodenum, and antral stomach but is also found in the common bile duct during embryogenesis. This study aimed to elucidate the role of Pdx1 in the development of the common bile duct, major duodenal papilla, and duodenum. METHODS: Expression pattern of pdx1 during embryogenesis and the morphology of the common bile duct, major duodenal papilla, and duodenum in pdx1 null mice were analyzed. RESULTS: The major duodenal papilla, peribiliary glands, and mucin-producing cells in the common bile duct were not formed in pdx1 null mice. Pdx1 null mice had shorter periampullary duodenal villi than wild-type mice at postnatal stages associated with reduced cell proliferation and increased apoptosis of the duodenal epithelial cells. Loss of the major duodenal papilla allowed duodeno-biliary reflux and bile infection, resulting in the formation of brown pigment biliary stones in pdx1 null mice, and antibiotics treatment significantly reduced the incidence of biliary stone formation. CONCLUSIONS: Pdx1 is required for proper development of the major duodenal papilla, peribiliary glands, and mucin-producing cells in the common bile duct and for maintenance of the periampullary duodenal epithelial cells during perinatal period. Bile infection because of loss of the major duodenal papilla plays a significant role in the formation of brown pigment biliary stones in pdx1 null mice.

[Bile duct infections as a late complication after endoscopic sphincterotomy]. / Zakazenia dróg zólciowych jako pózne powiklanie sfinkterotomii endoskopowej.

UNLABELLED: Nowadays endoscopic sphincterotomy (SE) is considered to be a "gold standard" in the treatment of residual choledocholithiasis and biliary pancreatitis. Nevertheless patients after invasive procedures concerning bile ducts with destruction of Oddi's sphincter often have increased succeptibility to chronic infections of bile ducts. AIM: Assessment of the influence of SE on bacterial colonization, development of main bile duct chronic inflammation and recurrent choledocholithiasis. MATERIAL AND METHODS: We included to the study 50 patients with main bile duct lithiasis, who underwent endoscopic cholangiopancreatography (ERCP). Among them there were 32 women in age from 45 to 87 (mean 69.7 +/- 12.3) and 18 men in age from 39 to 84 (mean 59.8 +/- 11.56). The studied group was divided into 3 subgroups: In I (control) group were included 10 patients diagnosed with ERCP techniques because of jaundice of unknown origin. Those patients did't have ERCP or operative revision of bile ducts before, but all of them underwent cholecystectomy in the past. Each II and III group consisted of 20 patients, who underwent SE before 12-24 and 25-36 months earlier respectively. Sterile uodenoscope was inserted to the region of major duodenal papilla, and then a sterile brush was used to collect material to cytological smear (brushing) from main bile duct. In all groups 2 ml of bile were taken by a catheter to a syringe. On the basis of radiograms from ERCP findings, the width of main bile duct was assessed. In all patients blood cell count, aspartate and alanine aminotransferases, gammaglutamyltranspeptidase, alkaline phosphatase, amylase, bilirubin, C-reactive protein and procalcitonine levels were measured. RESULTS: In our study all 40 patients with previously performed endoscopic sphincterotomy (SE) had bacterial colonization in bile ducts, mostly with Gramm negative pathogens. The most virulent pathogens vere cultured from bile duct swabs from patients after two or more revisions of common bile duct, and after a longer time from sphincterotomy. Those patients had multibacterial bile duct infections (Escherichia coli + Pseudomonas aeruginosa; P. aeruginosa + Enterococcus faecalis; E. faecalis + Enterobacter cloacae + Candida albicans; P. aeruginosa + E. cloacae). E. coil and E. faecalis were most frequently cultured (20 and 16 cases respectively). It was observed that with length of time from endoscopic sphincterotomy, the width of main bile duct and also amount of cultured pathogens and their virulency grow. Elevated transaminases activity and serum procalcitonine levels were observed in studied groups. 9 patients after previous sphincterotomy had recurrent lithiasis. CONCLUSIONS: Sphincterotomy contributes to colonization of main bile duct with pathogenic bacteria. With length of time from endoscopic sphincterotomy, and amount of performed endoscopic sphincterotomies the amount of cultured pathogens, their virulency and drug resistance grow. Chronic infection of bile ducts after endoscopic sphincterotomy causes higher risk of recurrent lithiasis.

Microbiology of choledochal bile in patients with choledocholithiasis admitted to a tertiary hospital.

AIM: The present study was designed to investigate the microbiology of choledochal bile of patients with cholangitis and choledocholithiasis. METHODS: We identified and determined the antimicrobial susceptibility of bacteria isolated in the bile of patients with cholangitis and choledocholithiasis diagnosed by endoscopic retrograde cholangiopancreatography (ERCP). RESULTS: Nineteen (82.6%) of 23 patients with choledocholithiasis had positive bile cultures. A single agent was detected in 11 patients (57.9%), while a mixed growth, with pathogens ranging from two to three species, were seen in eight patients (42.1%). Patients with clinical manifestations of cholangitis had significantly higher counts of colonies per mL of bile (> 105 cfu/mL). The predominant Gram-negative aerobic bacteria isolated were Escherichia coli (9, 31.0%), Klebsiella pneumoniae (5, 17.2%), Enterobacter cloacae (2, 6.9%), Pantoea agglomerans (1, 3.4%), and Pseudomonas aeruginosa (1, 3.4%). The predominant Gram-positive bacteria were Enterococcus faecalis (5, 17.2%) and Streptococcus sp. (5, 17.2%). Bacteroides fragilis was isolated in one patient with mixed growth. All Gram-positive bacteria isolated in bile were sensitive to ampicillin, and all Gram-negative bacteria isolated were sensitive to gentamicin with a minimum inhibitory concentration (CIM90) ranging from 0.5 to 1.0- micro g/mL. Gram-negative bacteria were also sensitive to imipenem, fluorquinolones, second and third generation cephalosporins. Although all five isolates of E. faecalis were sensitive to ampicillin, two of five (40%) E. faecalis isolates demonstrated high levels of resistance to gentamicin. CONCLUSION: E. coli, K. pneumoniae, E. faecalis and Streptoccocus sp. were the most common bacteria isolated in the bile of patients with cholangitis and choledocholithiasis, which were sensitive to a simple therapeutic regimen, such as the combination of ampicilin and gentamicin.

Bacterial translocation from the biliary tract to blood and lymph in rats with obstructive jaundice.

BACKGROUND: The disruption of the hepatocyte tight junctions observed in biliary obstruction suggests altered permeability of the blood-bile barrier. In this study the role of biliary obstruction and increased biliary pressure on the translocation of bacteria from biliary tract to bloodstream and lymphatic system were evaluated. MATERIALS AND METHODS: Rats underwent distal bile duct ligation (BDL, n = 33) for two weeks or a sham celiotomy (n = 21). Seventeen of the 33 BDL rats underwent subsequent biliary decompression by a choledochojejunostomy (CJ). Two weeks after the final operation, a laparotomy was performed again and the CBD, the thoracic duct, and the caval vein were canulated. Next, a suspension containing 10(8) Escherichia coli/ml was retrogradely infused in the CBD for 5 min at 5 or 20 cm H2O above the secretory biliary pressure. RESULTS: A higher biliary infusion pressure resulted in a significant increase of cfu E.coli per milliliter of blood in all the three groups (Sham, BDL, CJ). BDL rats showed significantly more bacterial translocation to the bloodstream than the shams. After biliary decompression, translocation normalized to the control levels. At 5 cm H2O infusion pressure only one lymph culture was positive (CJ group). At 20 cm H2O overpressure, nine lymph cultures were E.coli positive (P = 0.03). These were found mainly in groups with a nonobstructed bile duct (Sham and CJ 40% vs BDL 10%). CONCLUSION: Translocation of bacteria from biliary tract to bloodstream increased at higher intrabiliary pressures. Longstanding bile duct obstruction was an independent determinant for cholangiovenous reflux. Bacterial translocation to the lymphatic system did not parallel translocation to the bloodstream, although in the nonobstructed biliary tract, increased bacterial translocation to the lymphatic system was pressure related.

Mucus production of choledochal epithelial cells due to bile stagnation in the rat.

Mucus secretion is increased by various causative agents of inflammation in the mucosa. Although mucus production of the choledochal epithelial cells (except goblet cells) has never been noted in the rat in normal physiological conditions, it is seen as an inflammatory reaction in animals with choledochoenterostomy, which brings bacteria and active digestive enzymes into the common bile duct through regurgitation of intestinal contents. It is known that stagnant bile alone can cause inflammation in the mucosa of the gallbladder in patients with aseptic acalculous cholecystitis. In this study, aseptic bile stagnation was caused by choledochal dilation made by detaching the common bile duct from the hepatoduodenal ligament in five rats, and histological changes of the choledochal epithelium were observed by light and transmission electron microscopy 17 months after the operative procedure to determine whether stagnant bile could cause mucus production in choledochal epithelial cells. Mucus production was noted in two rats by light microscopy and in all the rats by transmission electron microscopy, and so it was demonstrated that stagnant bile could give rise to this phenomenon in the rat choledochal epithelium.

A bacteriologic and scanning electron microscope study after implantation of foreign bodies in the biliary tract in rats.

BACKGROUND: Bacterial adherence to the stent surfaces, concomitant colonization, and possible stent blockage are the main complications after the use of biliary stents. The present study was assigned to investigate bacteriologic and morphologic changes in the biliary tract after the implantation of biliary drain materials. METHODS: Rubber and silicone pieces with a surface area of 1 cm2 were implanted into the biliary tract in rats after temporary obstruction of the common bile duct by the use of a mini-occluder. The animals were killed at 4, 8 and 14 weeks, respectively, after implantation, and the implants were retrieved, cultured, and examined by scanning electron microscopy (SEM). Bacterial culture and SEM were also performed on tissue samples obtained from the mucosal surface of the biliary tract. RESULTS: Bacterial colonization and biofilm formation were found on the surfaces of the implanted materials and on the mucosal surface of the biliary tract in animals with implants but not on the biliary tract mucosa in rats without implants. CONCLUSION: Foreign bodies implanted in the biliary tract facilitate bacterial adherence not only to the surface of the implants but also to the mucosal surface in the biliary tract.

Experimental foreign-body infection in the biliary tract in rats.

BACKGROUND: Biomaterials used for biliary drainage may potentially result in biomaterial-associated infections. METHODS: Foreign-body infection in the biliary tract was investigated in rats. Rubber drain pieces were implanted in the biliary tract in rats for 1-4 weeks, followed by challenges with 10(2) to 10(4) colony-forming units (cfu) Escherichia coli injected into the common bile duct. The rate of infection was calculated, the bacterial growth in the biliary tract was observed over 72 h after challenges, and the opsonic activity in bile and in sera was assessed. RESULTS: In the group with drain material, inocula as small as 10(2) cfu produced persisting infection in the common bile duct in 90% of animals, whereas the same number of E. coli infected only 30% of rats in the control and sham implantation groups (p < 0.01, chi-square test). Complement-mediated opsonic activity in bile of animals with implanted drain pieces gradually decreased with time, whereas opsonic activity in sera from the same animals remained unchanged. CONCLUSIONS: Implants in the biliary tract impair local host defense, resulting in an increased susceptibility to microbial infection.

Sensitivity to new generation of antibiotics in biliary surgery.

The treatment of septic complications of cholecystectomy patients requires special attention. In a prospective study of 1009 consecutive cholecystectomy patients (including all acute and elective patients, excluding cases of malignancy), we routinely took cultures from the cystic duct and the gallbladder, and checked the strain for sensitivity. Positive cultures were found in 31.4% of the series, the most frequent bacteria being E. coli (50.3%). The aminoglycosides and the second and third generations of cephaloridins were found more specific for the strains that were cultured. While the new cephaloridins are the first choice for prophylaxis, the combination of aminoglycosides with ampicillin is to be preferred from the medical and cost effective standpoint while the aim is therapeutic.

Bile secretory immunoglobulin A in biliary infection and cholelithiasis.

Bile samples from 71 patients with cholelithiasis and a control group of 10 subjects without hepatobiliary diseases were cultured for bacteria and measured for secretory immunoglobulin A (SIgA) using enzyme immunoassay specific for SIgA. The results of bile bacterial culture were all positive in patients with primary bile duct pigment stones, and significantly lower bile SIgA levels were observed than in normal controls (P less than 0.005). It was also shown that the constitutent ratios of SIgA to total bile immunoglobulin and the bile-serum ratio of SIgA were markedly lower in these patients than in normal controls (P less than 0.001, P less than 0.001). In patients with cholecystolithiasis, bile SIgA concentrations of patients with biliary infections were remarkably lower than those of patients without biliary infection (P less than 0.01) and those of normal controls (P less than 0.01). These results suggest a close relationship between biliary tract infection and low concentrations of bile SIgA.

[Correlativity of plasma endotoxin, plasma fibronectin and common bile duct pressure in severe acute cholangitis].

On the basis of common bile duct pressure measurement (CBDP) in 18 patients of severe acute cholangitis, plasma endotoxin (ET) was determined by modified synthetic chromogenic limulus amebocyte lysate assay and plasma fibronectin (FN) was detected with Laurell's rocket immunoelectrophoresis. CBDP was 2.23 +/- 0.49 KPa in patient group. There was striking positive correlation between ET and CBDP. Preoperative ET was 202.73 +/- 88.57 ng/L in patient group which was much higher than 15.47 +/- 7.38 ng/L in the control (P less than 0.001). Preoperative FN was 141.77 +/- 82.37 mg/L in the patient group which was lower than 317.21 +/- 12.57 mg/L in the control (p less than 0.001). Statistical differences could be noticed in postoperative ET and FN between the survivor and the dead. The study suggested that plasma ET levels are greatly influenced by pressure gradient of bile duct, dynamic observations of ET and FN levels are helpful to monitor disease course and predict prognosis.