Alterations of the bile microbiome is associated with progression-free survival in pancreatic ductal adenocarcinoma patients.

BACKGROUND: Patients with pancreatic ductal adenocarcinoma (PDAC) display an altered oral, gastrointestinal, and intra-pancreatic microbiome compared to healthy individuals. However, knowledge regarding the bile microbiome and its potential impact on progression-free survival in PDACs remains limited. METHODS: Patients with PDAC (n = 45), including 20 matched pairs before and after surgery, and benign controls (n = 16) were included prospectively. The characteristics of the microbiomes of the total 81 bile were revealed by 16  S-rRNA gene sequencing. PDAC patients were divided into distinct groups based on tumor marker levels, disease staging, before and after surgery, as well as progression free survival (PFS) for further analysis. Disease diagnostic model was formulated utilizing the random forest algorithm. RESULTS: PDAC patients harbor a unique and diverse bile microbiome (PCoA, weighted Unifrac, p = 0.038), and the increasing microbial diversity is correlated with dysbiosis according to key microbes and microbial functions. Aliihoeflea emerged as the genus displaying the most significant alteration among two groups (p < 0.01). Significant differences were found in beta diversity of the bile microbiome between long-term PFS and short-term PFS groups (PCoA, weighted Unifrac, p = 0.005). Bacillota and Actinomycetota were identified as altered phylum between two groups associated with progression-free survival in all PDAC patients. Additionally, we identified three biomarkers as the most suitable set for the random forest model, which indicated a significantly elevated likelihood of disease occurrence in the PDAC group (p < 0.0001). The area under the receiver operating characteristic (ROC) curve reached 80.8% with a 95% confidence interval ranging from 55.0 to 100%. Due to the scarcity of bile samples, we were unable to conduct further external verification. CONCLUSION: PDAC is characterized by an altered microbiome of bile ducts. Biliary dysbiosis is linked with progression-free survival in all PDACs. This study revealed the alteration of the bile microbiome in PDACs and successfully developed a diagnostic model for PDAC.

Bilothorax: A Case Report and Systematic Literature Review of the Rare Entity.

Background: Bilothorax is defined as the presence of bile in the pleural space. It is a rare condition, and diagnosis is confirmed with a pleural fluid-to-serum bilirubin ratio of >1. Methods: The PubMed, Embase, Google Scholar, and CINAHL databases were searched using predetermined Boolean parameters. The systematic literature review was done per PRISMA guidelines. Retrospective studies, case series, case reports, and conference abstracts were included. The patients with reported pleural fluid analyses were pooled for fluid parameter data analysis. Results: Of 838 articles identified through the inclusion criteria and removing 105 duplicates, 732 articles were screened with abstracts, and 285 were screened for full article review. After this, 123 studies qualified for further detailed review, and of these, 115 were pooled for data analysis. The mean pleural fluid and serum bilirubin levels were 72 mg/dL and 61 mg/dL, respectively, with a mean pleural fluid-to-serum bilirubin ratio of 3.47. In most cases, the bilothorax was reported as a subacute or remote complication of hepatobiliary surgery or procedure, and traumatic injury to the chest or abdomen was the second most common cause. Tube thoracostomy was the main treatment modality (73.83%), followed by serial thoracentesis. Fifty-two patients (51.30%) had associated bronchopleural fistulas. The mortality was considerable, with 18/115 (15.65%) reported death. Most of the patients with mortality had advanced hepatobiliary cancer and were noted to die of complications not related to bilothorax. Conclusion: Bilothorax should be suspected in patients presenting with pleural effusion following surgical manipulation of hepatobiliary structures or a traumatic injury to the chest. This review is registered with CRD42023438426.

A Comprehensive Analysis of Fel Ursi and Its Common Adulterants Based on UHPLC-QTOF-MSE and Chemometrics.

BACKGROUND: As one of the four most valuable animal medicines, Fel Ursi, named Xiong Dan (XD) in China, has the effect of clearing heat, calming the liver, and brightening the eyes. However, due to the special source of XD and its high price, other animals' bile is often sold as XD or mixed with XD on the market, seriously affecting its clinical efficacy and consumers' rights and interests. In order to realize identification and adulteration analysis of XD, UHPLC-QTOF-MSE and multivariate statistical analysis were used to explore the differences in XD and six other animals' bile. METHODS: XD, pig gall (Zhu Dan, ZD), cow gall (Niu Dan, ND), rabbit gallbladder (Tu Dan, TD), duck gall (Yan Dan, YD), sheep gall (Yang Dan, YND), and chicken gall (Ji Dan, JD) were analyzed by UHPLC-QTOF-MSE, and the MS data, combined with multivariate analysis methods, were used to distinguish between them. Meanwhile, the potential chemical composition markers that contribute to their differences were further explored. RESULTS: The results showed that XD and six other animals' bile can be distinguished from each other obviously, with 27 ions with VIP > 1.0. We preliminarily identified 10 different bile acid-like components in XD and the other animals' bile with significant differences (p < 0.01) and VIP > 1.0, such as tauroursodeoxycholic acid, Glycohyodeoxycholic acid, and Glycodeoxycholic acid. CONCLUSIONS: The developed method was efficient and rapid in accurately distinguishing between XD and six other animals' bile. Based on the obtained chemical composition markers, it is beneficial to strengthen quality control for bile medicines.

Bile lakes in patients with biliary atresia who presented with jaundice-free native liver survival indicating the risk of subsequent liver transplantation due to various factors.

PURPOSE: The prognostic factors of subsequent liver transplantation (LT) in patients with biliary atresia (BA) who presented with jaundice-free native liver survival were investigated. METHODS: This study retrospectively reviewed patients who underwent portoenterostomy (PE) for BA. Patients with jaundice-free native liver survival at 1 year postoperatively were divided into the autologous liver survivor and liver transplant recipient groups. Peri- and postoperative data were compared between the two groups. RESULTS: Among 97 patients with BA, 29 who received LT within 1 year after PE were excluded from the analysis. Further, 48 patients currently living with native liver and 20 who received LT after 1 year postoperatively were compared. Bile lake (BL) was the strongest risk factor of LT. The risk score was 2.38 ∗ B L s c o r e + 0.00466 ∗ T B A , and the area under the receiver operating characteristic curve was 0.83. Patients with BL and those without significantly differed in terms of the native liver survival rate. Patients with BL who presented with not only cholangitis but also gastrointestinal hemorrhage and hepatopulmonary syndrome received LT. CONCLUSION: BL can cause different pathologies. Moreover, it is an evident risk factor of subsequent LT in patients with BA who are living with native liver at 1 year after PE.

Treatment Strategies for Bile Leak Following Pediatric Liver Transplantation.

There are no standard management protocols for the treatment of bile leak (BL) after liver transplantation. The objective of this study is to describe treatment options for BL after pediatric LT. METHODS: Retrospective analysis (January 2010-March 2023). VARIABLES STUDIED: preoperative data, status at diagnosis, and postoperative outcome. Four groups: observation (n = 9), percutaneous transhepatic cholangiography (PTC, n = 38), ERCP (2), and surgery (n = 27). RESULTS: Nine hundred and thirty-one pediatric liver transplantation (859 LDLT and 72 DDT); 78 (8.3%) patients had BL, all in LDLT. The median (IQR) peritoneal bilirubin (PB) level and fluid-to-serum bilirubin ratio (FSBR) at diagnosis was 14.40 mg/dL (8.5-29), and 10.7 (4.1-23.7). Patients who required surgery for treatment underwent the procedure earlier, at a median of 14 days (IQR: 7-19) versus 22 days for PTC (IQR: 15-27, p = 0.002). PB and FSBR were significantly lower in the observation group. In 11 cases, conservative management had resolution of the BL in an average time of 35 days, and 38 patients underwent PTC in a median time of 22 days (15-27). Twenty-seven (34.6%) patients were reoperated as initial treatment for BL in a median time of 17 days (1-108 days); 25 (33%) patients evolved with biliary stricture, 5 (18.5%) after surgery, and 20 (52.6%) after PTC (p = 0.01). CONCLUSION: Patients with BL who were observed presented significantly lower levels of PB and FSBR versus those who underwent PTC or surgery. Patients treated with PTC presented higher rates of biliary stricture during the follow-up.

Possibility of Cell Block Specimens from Overnight-Stored Bile for Next-Generation Sequencing of Cholangiocarcinoma.

The identification of anticancer therapies using next-generation sequencing (NGS) is necessary for the treatment of cholangiocarcinoma. NGS can be easily performed when cell blocks (CB) are obtained from bile stored overnight. We compared NGS results of paired CB and surgically resected specimens (SRS) from the same cholangiocarcinoma cases. Of the prospectively collected 64 bile CBs from 2018 to 2023, NGS was performed for three cases of cholangiocarcinoma that could be compared with the SRS results. The median numbers of DNA and RNA reads were 95,077,806 [CB] vs. 93,161,788 [SRS] and 22,101,328 [CB] vs. 24,806,180 [SRS], respectively. We evaluated 588 genes and found that almost all genetic alterations were attributed to single-nucleotide variants, insertions/deletions, and multi-nucleotide variants. The coverage rate of variants in SRS by those found in CB was 97.9-99.2%, and the coverage rate of SRS genes by CB genes was 99.6-99.7%. The NGS results of CB fully covered the variants and genetic alterations observed in paired SRS samples. As bile CB is easy to prepare in general hospitals, our results suggest the potential use of bile CB as a novel method for NGS-based evaluation of cholangiocarcinoma.

Metabolite identification and excretion of pinocembrin-7-O-ß-D-glucoside in rats by UHPLC/MS.

Pinocembrin-7-O-ß-D-glucoside (PCBG) isolated from Penthorum chinense Pursh was proven to display a wide range of pharmacological effects including hepatoprotection, anti-hepatoma and antifungal activities, etc. The research aims to qualitatively analyze the metabolites of PCBG in rat plasma, urine, bile and feces, and further perform the excretion study of PCBG and its major metabolite pinocembrin (PCB). Fifteen rats were divided into three groups (n=5 for each group) for blood, bile, urine and feces collection, respectively. After PCBG suspension was intragastrically administered to rats at 50 mg/kg, biological samples were collected and processed. The metabolites in each matrix were detected by UHPLC-Q-Exactive-MS/MS. A total of 111 metabolites were observed in plasma, urine, bile and feces, which include hydroxylated, sulfated and glucuronized metabolites, etc. In addition, an UHPLC-MS/MS method was established and applied for the excretion quantification of PCBG and PCB in rat urine, bile, and feces samples. Studies on excretion have shown that PCBG is mainly excreted through feces. The cumulative excretion rates of PCBG and PCB in rat urine, bile and feces were (4.5±2.4)%, (0.2±0.1)% and (18.4±10.5)%, respectively. After hydrolysis by ß-glucuronidase/sulfatase, the excretion rates of PCB in urine and bile were (5.7±2.8)% and (8.9±4.2)%. This study contributes to preclinical research on PCBG and explains its pharmacological effects.

Viability assessment of the liver during ex-situ machine perfusion prior to transplantation.

PURPOSE OF REVIEW: In an attempt to reduce waiting list mortality in liver transplantation, less-than-ideal quality donor livers from extended criteria donors are increasingly accepted. Predicting the outcome of these organs remains a challenge. Machine perfusion provides the unique possibility to assess donor liver viability pretransplantation and predict postreperfusion organ function. RECENT FINDINGS: Assessing liver viability during hypothermic machine perfusion remains challenging, as the liver is not metabolically active. Nevertheless, the levels of flavin mononucleotide, transaminases, lactate dehydrogenase, glucose and pH in the perfusate have proven to be predictors of liver viability. During normothermic machine perfusion, the liver is metabolically active and in addition to the perfusate levels of pH, transaminases, glucose and lactate, the production of bile is a crucial criterion for hepatocyte viability. Cholangiocyte viability can be determined by analyzing bile composition. The differences between perfusate and bile levels of pH, bicarbonate and glucose are good predictors of freedom from ischemic cholangiopathy. SUMMARY: Although consensus is lacking regarding precise cut-off values during machine perfusion, there is general consensus on the importance of evaluating both hepatocyte and cholangiocyte compartments. The challenge is to reach consensus for increased organ utilization, while at the same time pushing the boundaries by expanding the possibilities for viability testing.

Genipin improves obesity through promoting bile secretion and changing bile acids composition in diet-induced obese rats.

OBJECTIVES: Bile acids (BAs), as signaling molecules to regulate metabolism, have received considerable attention. Genipin is an iridoid compound extracted from Fructus Gradeniae, which has been shown to relieve adiposity and metabolic syndrome. Here, we investigated the mechanism of genipin counteracting obesity and its relationship with BAs signals in diet-induced obese (DIO) rats. METHODS: The DIO rats were received intraperitoneal injections of genipin for 10 days. The body weight, visceral fat, lipid metabolism in the liver, thermogenic genes expressions in brown fat, BAs metabolism and signals, and key enzymes for BAs synthesis were determined. KEY FINDINGS: Genipin inhibited fat synthesis and promoted lipolysis in the liver, and upregulated thermogenic gene expressions in brown adipose tissue of DIO rats. Genipin increased bile flow rate and upregulated the expressions of aquaporin 8 and the transporters of BAs in liver. Furthermore, genipin changed BAs composition by promoting alternative pathways and inhibiting classical pathways for BAs synthesis and upregulated the expressions of bile acid receptors synchronously. CONCLUSIONS: These results suggest that genipin ameliorate obesity through BAs-mediated signaling pathways.

Sampling time for different matrices in stress assessment of farmed Atlantic salmon post-smolt.

The sustainability of commercial aquaculture production depends critically on prioritizing fish welfare management. Besides monitoring welfare parameters such as fish behaviour and water quality, fish stress level can also provide a reliable measure of the welfare status of farmed fish. Cortisol and 5 of its metabolites (5ß-THF, cortisone, 5ß-DHE, 5ß-THE, ß-cortolone) were previously identified by the authors as suitable stress biomarkers of farmed Atlantic salmon. Based on this knowledge, the present study aimed to investigate the time-related dynamics of these metabolites in plasma, skin mucus, bile and faeces over a 72 h- period. The objective was to determine the optimal sampling time for each matrix and to understand the clearance pathway of these metabolites following stress. An experiment was carried out using a total of 90 Atlantic salmon with an average weight of 438 (±132) g. The average sea temperature was 6.9 °C during the experimental period. A control group of 10 fish was first collected before the remaining 80 fish were submitted to a stress of netting and subsequent relocation into two separate cages. From each of these two stress groups, 10 fish were sampled at 1 h, 2 h, 4 h, 6 h and 12 h, 24 h, 48 h, 72 h after the stress event respectively. The concentrations of cortisol and its metabolites were measured at each of the sampling timepoint. The results demonstrated that plasma cortisol metabolites reached the highest concentration 4 h after stress and remained elevated despite the slight decrease for the remaining timepoints. The peak level was observed at 12 h post-stress in skin mucus and 24 h in bile and faeces. The findings suggest that these timepoints are the optimal for sampling Atlantic salmon post-smolt following stressful events in acute stress studies. Furthermore, the results reveal that analysing cortisol and its metabolites, both in free and conjugated forms, rather than free cortisol provides greater flexibility as their concentrations are less affected by sampling procedure. This study confirms the appropriateness of skin mucus and faeces as less-invasive sample matrices for fish stress evaluation and provides a basis for further developing low invasive tools for monitoring the welfare of farmed salmonid.

Dual-energy CT in acute cholecystitis- features predicting culture-positive bile and outcome.

PURPOSE: Low mono-energetic CT has been shown to improve visualization of acute abdominal inflammatory processes. We aimed to determine its utility in patients with acute cholecystitis and potential added value in clinical decision making. METHODS: Sixty-seven consecutive patients with radiological signs of cholecystitis on contrast-enhanced dual-layer CT imaging were retrospectively identified over a four-year period (2/17-8/21). A ranked Likert scale was created for imaging findings present in acute cholecystitis, including gallbladder mucosal integrity and enhancement and pericholecystic liver parenchymal enhancement. These rankings were correlated with laboratory data, followed by sensitivity, specificity, and odds-ratios calculations. RESULTS: Mucosal integrity and pericholecystic liver enhancement were better seen on low-energetic images by unanimous consensus. Presence of pericholecystic liver enhancement and poorer mucosal wall integrity correlated with positive bile cultures (sensitivity: 93.8 % and 96.9 %, specificity: 37.5 and 50.0 %; odds-ratio: 9.0[1.1-68.1 95 %CI] and 31.0 [2.7-350.7 95 %CI], p = 0.017 and p ≤ 0.001) in patients undergoing cholecystostomy (n = 40/67). Moreover, binary regression modeling showed that the strongest predictor variable for bile culture positivity was the score for pericholecystic liver enhancement (Exp(B) = 0.6, P = 0.022). By contrast, other laboratory markers and other imaging findings (such as GB wall thickness) showed lower sensitivities (76-82 %), specificities (16-21 %) and odds ratios (0.2-4.4) for the prediction of infected bile. CONCLUSIONS: Pericholecystic liver enhancement and gallbladder wall integrity are better visualized on low-DECT images. These findings also potentially predict bile culture positivity in patients with cholecystitis, which may influence clinical management including the need for intervention.

Bile promotes Lactobacillus johnsonii N6.2 extracellular vesicle production with conserved immunomodulatory properties.

Recently, Lactobacillus johnsonii N6.2-derived extracellular vesicles (EVs) were shown to reduce apoptosis in human beta cell lines and stimulate insulin secretion in human islets. Our goal was to identify a physiologically relevant environmental condition that induces a hypervesiculation phenotype in L. johnsonii N6.2 and to evaluate if transcriptional changes are involved in this process. Culturing this strain in the presence of 0.2% bovine bile, which mimics a stressor encountered by the bacterium in the small intestine, resulted in approximately a 100-fold increase in EVs relative to cells grown in media without bile. Whole transcriptome analysis of cells grown with bile revealed upregulation of several peptidoglycan hydrolases as well as several genes involved in fatty acid utilization. These results suggest that the hypervesiculation phenotype may be the result of increased cell wall turnover combined with increased accumulation of phospholipids, in agreement with our previous proteomic and lipidomics results. Additionally, EVs isolated from L. johnsonii N6.2 grown in presence of bile maintained their immunomodulatory properties in host-derived ßlox5 pancreatic and THP-1 macrophage cell lines. Our findings suggest that in L. johnsonii N6.2 vesiculogenesis is significantly impacted by the expression of cell wall modifying enzymes and proteins utilized for exogenous fatty acid uptake that are regulated at the transcriptional level. Furthermore, this data suggests that vesiculogenesis could be stimulated in vivo using small molecules thereby maximizing the beneficial interactions between bacteria and their hosts.

Unique bile acid profiles in the bile ducts of patients with primary sclerosing cholangitis.

BACKGROUND: The relationship between primary sclerosing cholangitis (PSC) and biliary bile acids (BAs) remains unclear. Although a few studies have compared PSC biliary BAs with other diseases, they did not exclude the influence of cholestasis, which affects the composition of BAs. We compared biliary BAs and microbiota among patients with PSC, controls without cholestasis, and controls with cholestasis, based on the hypothesis that alterations in BAs underlie the pathophysiology of PSC. METHODS: Bile samples were obtained using endoscopic retrograde cholangiopancreatography from patients with PSC (n = 14), non-hepato-pancreato-biliary patients without cholestasis (n = 15), and patients with cholestasis (n = 13). RESULTS: The BA profiles showed that patients with PSC and cholestasis controls had significantly lower secondary BAs than non-cholestasis controls, as expected, whereas the ratio of cholic acid/chenodeoxycholic acid in patients with PSC was significantly lower despite cholestasis, and the ratio of (cholic acid + deoxycholic acid)/(chenodeoxycholic acid + lithocholic acid) in patients with PSC was significantly lower than that in the controls with or without cholestasis. The BA ratio in the bile of patients with PSC showed a similar trend in the serum. Moreover, there were correlations between the alteration of BAs and clinical data that differed from those of the cholestasis controls. Biliary microbiota did not differ among the groups. CONCLUSIONS: Patients with PSC showed characteristic biliary and serum BA compositions that were different from those in other groups. These findings suggest that the BA synthesis system in patients with PSC differs from that in controls and patients with other cholestatic diseases. Our approach to assessing BAs provides insights into the pathophysiology of PSC.

Bile Spillage as a Prognostic Factor for Gall Bladder Cancer: A Systematic Review and Meta-Analysis.

INTRODUCTION: Biliary spillage (BS) is a common complication following initial cholecystectomy for gall bladder cancer (GBC). Few studies have explored the importance of BS as a long-term prognostic factor. We perform a meta-analysis of the association between BS and survival in GBC. METHODS: A systematic literature search was performed in February 2023. Studies evaluating the incidence of BS and its association with long-term outcomes in patients undergoing initial laparoscopic or open cholecystectomy for either incidental or resectable GBC were included. Overall survival (OS), disease-free survival (DFS), and rate of peritoneal carcinomatosis (RPC) were the primary end points. Forest plot analyses were used to calculate the pooled hazard ratios (HRs) of OS, DFS, and RPC. Metaregression was used to evaluate study-level association between BS and perioperative risk factors. RESULTS: Of 181 published articles, 11 met inclusion criteria with a sample size of 1116 patients. The rate of BS ranged between 9% and 67%. On pooled analysis, BS was associated with worse OS (HR = 1.68, 95% confidence interval [CI] = 1.32-2.14), DFS (pooled HR= 2.19, 95% CI = 1.30-3.68), and higher RPC (odds ratio = 9.37, 95% CI = 3.49-25.2). The rate of BS was not associated with higher T stage, lymph node metastasis, higher grade, positive margin status, reresection, or conversion rates. CONCLUSIONS: Our meta-analysis shows that BS is a predictor of higher peritoneal recurrence and poor survival in GBC. BS was not associated with tumor characteristics or conversion rates. Further research is needed to identify other potential risk factors for BS and investigate the ideal treatment schedule to improve survival.

Kidney ischemia/reperfusion injury causes cholangiocytes primary cilia disruption and abnormal bile secretion.

BACKGROUND: Acute kidney injury (AKI) causes distant liver injury, to date, which causes poor outcomes of patients with AKI. Many studies have been performed to overcome AKI-associated liver injury. However, those studies have mainly focused on hepatocytes, and AKI-induced liver injury still remains a clinical problem. Here, we investigated the implication of cholangiocytes and their primary cilia which are critical in final bile secretion. Cholangiocyte, a lining cell of bile ducts, are the only liver epithelial cell containing primary cilium (a microtubule-based cell surface signal-sensing organelle). METHODS: Cystathione γ-lyase (CSE, a transsulfuration enzyme) deficient and wild-type mice were subjected to kidney ischemia followed by reperfusion (KIR). Some mice were administered with N-acetyl-cysteine (NAC). RESULTS: KIR damaged hepatocytes and cholagiocytes, disrupted cholangiocytes primary cilia, released the disrupted ciliary fragments into the bile, and caused abnormal bile secretion. Glutathione (GSH) and H2S levels in the livers were significantly reduced by KIR, resulting in increased the ratio oxidized GSH to total GSH, and oxidation of tissue and bile. CSE and cystathione ß-synthase (CBS) expression were lowered in the liver after KIR. NAC administration increased total GSH and H2S levels in the liver and attenuated KIR-induced liver injuries. In contrast, Cse deletion caused the reduction of total GSH levels and worsened KIR-induced liver injuries, including primary cilia damage and abnormal bile secretion. CONCLUSIONS: These results indicate that KIR causes cholangiocyte damage, cholangiocytes primary cilia disruption, and abnormal bile secretion through reduced antioxidative ability of the liver.

Amanitin determination in bile samples by UHPLC-MS: LR-MS and HR-MS analytical performance.

Consumption of misidentified foraged mushrooms containing bicyclic amanitin octapeptides is a worldwide public health and veterinary problem, being considered one of the deadliest accidental human and canine food ingestion due to acute liver failure (ALF). Reversal of advanced ALF and complete clinical recovery can be achieved following definitive removal of accumulated amatoxin laden bile from the gallbladder. An accurate means of quantifying amanitin content in aspirated bile is, therefore, urgently needed. Building on our prior work validating a method to detect and quantify amanitin in hepatic autopsy tissue, the development of an accurate method of measuring α- and ß-amanitin in aspirated gallbladder bile was performed to evaluate the efficiency of this emergency procedure applied as a clinical treatment for intoxicated patients. A solid-phase extraction (SPE) procedure was optimized followed by detection based on ultra-high performance liquid chromatography coupled with mass spectrometry (UHPLC-MS). Low resolution mass spectrometry (LRMS) was compared with high resolution (HRMS) by the validation of UHPLC-MS/MS (triple quadrupole MS) and UHPLC-ToF-MS (time-of-flight MS). Both methods were able to detect amatoxins in bile with limits of detection and quantification ranging from 2.71 to 3.46 µg.kg-1, and 8.36-9.03 µg.kg-1 for α-amanitin and, 0.32-1.69 µg.kg-1 and 0.55-5.62 µg.kg-1 for ß-amanitin, respectively. Validation was completed with the evaluation of linearity, specificity, robustness, recovery, and precision following the ICH guidelines and CIR 808/2021. The validated methods were finally applied to bile samples obtained 48-96 hours + post-ingestion from 4 amatoxin poisoning patients who underwent gallbladder drainage procedures in Vietnam, Canada, and California. Gallbladder bile from patients with amatoxin mushroom poisoning contained significant amanitin content, even when aspirated several days post-ingestion, thus confirming the important role of enterohepatic circulation in amatoxin hepatotoxicity. This work represents a high and unique analytical throughput in amanitin poisoning allowing to efficiently respond to this fatal health problem.

Pharmacokinetics and residues of florfenicol in Nile tilapia (Oreochromis niloticus) post-oral gavage.

In the study on Oreochromis niloticus, singular oral gavage of florfenicol (FFC) at 15 mg/kg biomass/day was conducted, mimicking approved aquaculture dosing. Samples of plasma, bile, muscle, intestine, skin, liver, kidney, gill, and brain tissues were collected at 0, 2, 3, 4, 6, 8, 12, 16, 24, 32, 48, 64, 96, and 128 hours (h) after oral gavage. LC-MS/MS analysis revealed FFC concentrations peaked at 12.15 µg/mL in plasma and 77.92 µg/mL in bile, both at 24 hours. Elimination half-lives were 28.17 h (plasma) and 26.88 h (bile). The residues of FFC ranked muscle>intestine>skin>liver>kidney>gill. In contrast, the residues of florfenicol amine (FFA) ranked kidney>skin>liver>muscle>gill>intestine>brain, particularly notable in tropical summer conditions. The minimum inhibitory concentration of FFC was elucidated against several bacterial pathogens revealing its superior efficacy. Results highlight bile's crucial role in FFC elimination. Further investigation, especially during winter when fish susceptibility to infections rises, is warranted.

Role of albumin in the metabolism and excretion of ochratoxin A.

Ochratoxin A (OTA) is known to be strongly bound to serum albumin, but it remains unknown how albumin affects its metabolism and kinetics. To close this gap, we used a mouse model, where heterozygous albumin deletion reduces serum albumin to concentrations similar to hypoalbuminemic patients and completely eliminates albumin by a homozygous knockout. OTA and its potential metabolites (OTα, 4-OH-OTA, 7'-OH-OTA, OTHQ, OP-OTA, OTB-GSH, OTB-NAC, OTB) were time-dependently analyzed in plasma, bile, and urine by LC-MS/MS and were compared to previously published hepatotoxicity and nephrotoxicity data. Homozygous albumin deletion strongly accelerated plasma clearance as well as biliary and urinary excretion of the parent compound and its hydroxylation products. Decreasing albumin in mice by the heterozygous and even more by the homozygous knockout leads to an increase in the parent compound in urine which corresponded to increased nephrotoxicity. The role of albumin in OTA-induced hepatotoxicity is more complex, since heterozygous but not homozygous nor wild-type mice showed a strong biliary increase in the toxic open lactone OP-OTA. Correspondingly, OTA-induced hepatotoxicity was higher in heterozygous than in wild-type and homozygous animals. We present evidence that albumin-mediated retention of OTA in hepatocytes is required for formation of the toxic OP-OTA, while complete albumin elimination leads to rapid biliary clearance of OTA from hepatocytes with less formation of OP-OTA. In conclusion, albumin has a strong influence on metabolism and toxicity of OTA. In hypoalbuminemia, the parent OTA is associated with increased nephrotoxicity and the open lactone with increased hepatotoxicity.

Relationship between Intraoperative Bile Culture Outcomes and Subsequent Postoperative Infectious Complications: A Retrospective Cohort Study.

The presence of positive bile culture during intraoperative procedures has been associated with elevated morbidity and mortality rates in hepatobiliopancreatic surgeries, contributing to increased healthcare expenditures. However, the precise impact of bactobilia on the development of postoperative complications remains uncertain due to existing disparities in the published literature. In this retrospective cohort study, we assessed 137 patients who underwent major hepatobiliopancreatic surgery to examine the relationship between intraoperative bile culture outcomes and subsequent postoperative infectious complications. Among patients with bactobilia, a significant 35.1% exhibited systemic or local infectious complications, whereas only 11.1% of those with negative culture results experienced any infectious complications (p = 0.002). Similarly, a notable difference was observed in the incidence of surgical site infections, with 24.3% in the bactobilia group compared to 7.9% in the negative culture group (p = 0.01). A total of 74 monomicrobial cultures with microbiological growth were isolated, predominantly featuring Gram-negative microorganisms, primarily Enterobacteriaceae in 49 cultures. Escherichia coli was identified in 37.8% of positive cultures, while Klebsiella pneumoniae was evident in 21.6%. Gram-positive microorganisms were present in 10 cultures, with Enterococcus emerging as the prevailing species. The logistic regression model identified a positive bile culture as an independent factor significantly associated with infection development (OR: 2.26; 95% confidence interval: 1.23-11; p = 0.02). Considering the limitations of the study, these findings underscore the critical importance of conducting bile cultures during the intraoperative phase to enable vigilant monitoring and prompt management of infectious complications.