Bile Specimens Research Articles

Prevalence of Salmonella Typhi in Bile of the Patients Undergoing Cholecystectomy at a Tertiary Care Hospital Karachi

Objective: To estimate the prevalence of salmonella typhi in bile specimens collected from patients undergoing cholecystectomy at a tertiary care Hospital Karachi. Methodology: The descriptive cross-sectional study, was conducted at Department of Surgical Unit 21, Jinnah Postgraduate Medical Center, Karachi, from January 2021 to June 2021. Patients with symptomatic Cholelithiasis like sudden pain in the upper right portion of your abdomen which radiate towards right shoulder, nausea, vomiting, undergo cholecystectomy of either gender male or female with age 14 to 60 years were included. After open cholecystectomy, bile was aspirated from the gallbladder at fundus in five ml syringe and long spinal needles was used in case of laparoscopic cholecystectomy to collect bile under vision of telescope. Specimens were sent to laboratory according to protocol for culture sensitivity. Results: A total of 171 patients who met the inclusion and exclusion criteria were included. Mean age of the patients was 49.39±8.78 years. 83 (48.5%) were male and 88 (51.5%) were female. Prevalence of salmonella typhi in bile specimens collected from patients undergoing cholecystectomy, was found 17.54%. Furthermore, the frequency of salmonella isolate was statically insignificant according to the patient’s gender, diabetes and obesity (p=>0.05), while it was statically significant according to age up to 40 years and residence in rural areas (p-<0.05. Conclusion: A significant proportion of patients were found to have Salmonella in bile from patients undergoing cholecystectomy. Therefore, close monitoring of patients must be done in all patients with regular follow-up.

Applicability of a Chemiluminescence Immunoassay to Screen Postmortem Bile Specimens and Its Agreement with Confirmation Analysis.

Bile has emerged as an alternative matrix for toxicological investigation of drugs in suspected forensic cases of overdose in adults and intoxications in children. Toxicological investigation consists in screening and, subsequently, confirming the result with specific techniques, such as liquid chromatography with tandem mass spectrometry (LC-MS/MS). As there is no screening test on the market to test postmortem bile specimens, the novelty of this study was in investigating the applicability of a chemiluminescence immunoassay, designed for other matrices and available on the market, on bile and validate its use, testing the agreement with LC-MS/MS analysis. Bile specimens were obtained from 25 forensic cases of suspected death from overdose and intoxication. Sample preparation for bile screening consists simply in centrifugation and dilution. Confirmation analysis allows simultaneous identification of 108 drugs and was validated on bile. Kappa analysis assessed a perfect agreement (0.81-1) between the assays for benzodiazepines, methadone, opiates, cocaine, oxycodone, cannabinoids, buprenorphine and pregabalin; a substantial agreement (0.41-0.6) was reported for barbiturates. No agreement was assessed for amphetamines, due to an abundance of putrefactive amines in postmortem specimens. In conclusion, this fast and easy immunoassay could be used for initial screening of bile specimens, identifying presence of drugs, except amphetamines, with reliability.

A Comparative Investigation of the Bile Microbiome in Patients with Choledocholithiasis and Cholecystolithiasis through Metagenomic Analysis.

While the precise triggers of gallstone formation remain incompletely understood, it is believed to arise from a complex interplay of genetic and environmental factors. The bile microbiome is being increasingly recognized as a possible contributor to the onset of gallstone disease. The primary objective of this study was to investigate distinctions in the microbial communities within bile specimens from patients with choledocholithiasis (common bile duct stones) and cholecystolithiasis (gallbladder stones). We employed massively parallel sequencing of the 16S rRNA gene to examine the microbial communities within bile samples obtained from 28 patients with choledocholithiasis (group DS) and cholecystolithiasis (group GS). The taxonomic composition of the bile microbial communities displayed significant disparities between the group DS and the group GS. Within the 16 prevalent genera, only Streptococcus, Ralstonia, Lactobacillus, and Enterococcus were predominantly found in the group GS. In contrast, the group DS displayed a more diverse range of genera. The alpha diversity of bile specimens was also notably lower in the group GS compared to the group DS (p = 0.041). Principal coordinate analysis unveiled distinct clustering of bile microbial communities depending on the location of the gallstone. Linear discriminant analysis effect size analysis, with a score threshold of >3 and the Kruskall-Wallis test (α < 0.05), recognized Bacilli and Lactobacillales as potential taxonomic markers for distinguishing patients with cholecystolithiasis limited to the gallbladder. Significant variations were found in the distribution and diversity of bile microbial communities between patients with choledocholithiasis and cholecystolithiasis. This observation suggests that alterations in the bile microbiome may contribute to the development of gallstones in these patients.

Microbiome Dysbiosis with Enterococcus Presence in the Upper Gastrointestinal Tract is A Risk Factor for Mortality in Patients Undergoing Surgery for Pancreatic Cancer.

Recent retrospective studies suggest a role for distinct microbiota in the perioperative morbidity and mortality of pancreatic head resections. We aimed to prospectively investigate the microbial colonization of critical operative sites of pancreatic head resections to identify microbial stratification factors for surgical and long-term oncologic outcomes. Prospective biomarker study applying 16S rRNA sequencing and microbial culturing to samples collected from various sites of the GI tract and surgical sites of patients during pancreatic head resections at a German single high-volume pancreatic center. A total of 101 patients were included (38 non-cancer, 63 cancer patients [50 PDAC patients]) in the study. In a first data analysis series, 16S rRNA sequencing data were utilized from 96 patients to assess associations of microbiome profiles with clinical parameters and outcomes. In general, microbiome composition varied according to sampling site, cancer, age or preoperative ERCP intervention, notably for the bile microbiome. In the PDAC subcohort, compositional variance of the bile or periampullary microbiome was significantly associated with postoperative complications such as ICU admission; on a taxonomic level we observed Enterococcus spp. to be significantly more abundant in patients developing deep or organ-space surgical site infections (SSI). Elevated Enterococcus relative abundances in the upper GI tract, in turn, were associated with 6-months mortality rates. In a second step, we focused on microbiological cultures collected from bile aspirates during surgery and investigated associations with perioperative complications and long-term survival. Notably, Enterococcus spp. were among the most prevalent pathobiont isolates observed in cancer patient bile specimens that were associated with severe SSIs, and thereby elevated mortality rates up to 24 months. Clinically relevant postoperative pancreatic fistulas or severe SSI were found as other major variables determining short-term mortality in this cancer patient cohort. In the context of adverse microbiological factors, a preoperative ERCP was also observed to segregate long-term survival, and it appeared to interact with the presence of Enterococcus spp. as highest mortality rates were observed in PDAC patients with both preoperative ERCP and presence of E. faecalis in bile aspirates. The presence of Enterococcus spp. in bile ducts of PDAC patients undergoing pancreatic surgery represents a significant risk factor for perioperative infections and, thereby, elevated postoperative and long-term mortality. This finding supports previous data on the use of the antibiotic drug piperacillin-tazobactam as appropriate perioperative antibiotic prophylaxis for preventing adverse outcomes after pancreatoduodenectomy.

Variability of bile bacterial profiles and drug resistance in patients with choledocholithiasis combined with biliary tract infection: a retrospective study.

Biliary tract infection is a common complication of choledocholithiasis. This study aimed to analyse the distribution of pathogenic bacteria in bile cultures from patients with choledocholithiasis combined with biliary tract infection to guide clinical application of antimicrobials and reduce the emergence of drug resistance. A total of 880 patients were enrolled in this retrospective study from 30 March 2017 to 31 August 2022 at the Affiliated Hospital of Qingdao University in China. Bile specimens were extracted for microbiological culture under aseptic conditions using endoscopic retrograde cholangiopancreatography. Bacterial culture, strain identification, and antimicrobial susceptibility testing were conducted according to the standard protocol. Baseline data were retrieved from patient files. Overall, 90.34% (795/880) of bile samples showed positive microbiological results and 37.50% (330/880) demonstrated polymicrobial infections. Among the 795 bile specimens with positive culture results, 1,216 pathogenic bacteria were detected, with gram-negative bacilli accounting for 56.33%, gram-positive cocci for 41.86%, and fungi for 1.81%. The predominant gram-negative bacilli in the bile cultures were Escherichia coli (30.43%) and Klebsiella pneumoniae (13.98%), whereas the main gram-positive cocci were Enterococcus faecium (14.04%) and E. casseliflavus (4.28%). The annual trend analysis revealed a gradual decrease in the proportion of gram-negative bacilli and a gradual increase in the proportion of gram-positive cocci, with a concomitant decrease in the dominance of E. coli. Both E. faecium and E. coli showed high resistance to conventional antibiotics but high sensitivity to piperacillin/tazobactam, carbapenems, amikacin, and vancomycin. A significant change has occurred in the bile bacterial spectrum in patients with choledocholithiasis and biliary tract infection. The incidence of gram-positive cocci infections has increased annually, while that of gram-negative bacilli and E. coli infections has decreased. Antibiotic administration should be tailored based on the local bacterial profile.

A case of Vagococcus fluvialis isolated from the bile of a patient with calculous cholecystitis

BackgroundChronic cholecystitis, characterized by persistent inflammation of the gallbladder, predominantly stems from the prolonged presence of gallstones. Calculous cholecystitis has demonstrated a consistent escalation in its incidence over time.Gallbladder stones have been recognized as a predisposing factor for the development of biliary tract infections.Concomitantly, there have been substantial shifts in the distribution and resistance profiles of pathogenic microorganisms responsible for biliary tract infections. The timely acquisition of bile samples for pathogen analysis is of paramount importance, given its critical role in guiding judicious clinical pharmacotherapy and enhancing patient prognosis.Case PresentationWe present a case involving a 66-year-old female patient who had previously undergone subtotal gastrectomy due to diffuse large B-cell lymphoma. The patient was admitted to our institution with complaints of abdominal pain. Subsequent diagnostic evaluation revealed concurrent choledocholithiasis and cholecystolithiasis. The patient underwent surgical cholecystectomy as the therapeutic approach. Histopathological examination of the excised gallbladder disclosed characteristic features indicative of chronic cholecystitis. Subsequent laboratory analysis of the patient’s bile specimen yielded Gram-positive cocci, subsequently identified through biochemical assays, mass spectrometry, and 16 S rRNA analysis as Vagococcus fluvialis. Further in vitro antimicrobial susceptibility testing using disk diffusion and microfluidic dilution showed that this strain exhibited inhibition zone diameters ranging from 12.0 to 32.0 mm in response to 26 antibiotics, including ampicillin, cefazolin, cefuroxime, cefotaxime, ceftriaxone, cefepime, ampicillin/sulbactam, piperacillin, ciprofloxacin, cefoperazone/sulbactam, imipenem, meropenem, piperacillin/tazobarb, penicillin, erythromycin, chloramphenicol, vancomycin, methotrexate/sulfamethoxazole, teicoplanin, linezolid, tigecycline, cefoxitin, ceftazidime, levofloxacin, minocycline and tobramycin. However, the inhibition zone diameters were 6.0 mm for amikacin, oxacillin, clindamycin, and tetracycline. The patient received ceftazidime anti-infective therapy both preoperatively and within 24 h postoperatively and was discharged successfully one week after surgery.ConclusionIn this study, we present the inaugural isolation and identification of Vagococcus fluvialis from bile specimens of patients afflicted with calculous cholecystitis. This novel finding lays a substantial experimental groundwork for guiding clinically rational antimicrobial therapy and advancing the exploration of relevant pathogenic mechanisms pertaining to Vagococcus fluvialis infections.

Biliary microbial patterns in primary sclerosing cholangitis are linked to poorer transplant-free survival.

Factors that determine individual disease course of patients with primary sclerosing cholangitis (PSC) are poorly understood. Although an association between gut microbes and disease outcome has been suggested, little is known about the role of microbes in the biliary tract. We analyzed microbial cultures from bile specimens obtained during routine endoscopic retrograde cholangiopancreatography (ERCP) and intraoperatively before liver transplantation in 114 patients with PSC in our tertiary academic center. The presence of bacterial and fungal species was correlated with clinical characteristics and outcome data. A total of 87 patients (76%) had positive bile culture results. The presence of concomitant inflammatory bowel disease (IBD) was associated with positive bile culture results in multivariate analysis (OR, 4.707; 95% CI, 1.688-13.128; p=0.003). Enterococcus spp. in the bile was associated with a more frequent occurrence of liver transplantation and/or death (OR, 2.778; 95% CI, 1.147-6.728; p=0.021) and recurrent (≥3) cholangitis episodes (OR, 2.839; 95% CI, 1.037-7.768; p=0.037). Biliary candidiasis was linked to a higher frequency of recurrent (≥3) cholangitis episodes (OR, 5.677; 95% CI, 1.940-16.616; p=0.001). Proton pump inhibitor intake conferred a clinical feature associated with biliary candidiasis in multivariate analysis (OR, 3.559; 95% CI, 1.275-9.937; p=0.016). Our data indicate that in patients with PSC, presence of Enterococcus spp. and Candida spp. in bile is associated with an adverse outcome. Concomitant IBD is linked to presence of microbes in bile, and proton pump inhibitor intake is a feature associated with biliary candidiasis in patients with PSC.

Microbiomic profiles of bile in patients with benign and malignant pancreaticobiliary disease.

The prognostic and pathophysiologic significance of the biliary microbiota in pancreaticobiliary malignancies is little understood. Our goal was to find malignancy-related microbiomic fingerprints in bile samples taken from patients with benign and malignant pancreaticobiliary diseases. Bile specimens were collected from consenting patients during routine endoscopic retrograde cholangiopancreatography. We used PowerViral RNA/DNA Isolation kit to extract DNA from bile specimens. The Illumina 16S Metagenomic Sequencing Library Preparation guide was used to amplify the bacterial 16S rRNA gene and create libraries. QIIME (Quantitative Insights Into Microbial Ecology), Bioconductor phyloseq, microbiomeSeq, and mixMC packages were used for post-sequencing analysis. Of 46 enrolled patients, 32 patients had pancreatic cancers, 6 had cholangiocarcinoma and 1 had gallbladder cancer. Rest of the patients had benign diseases including gallstones, and acute and chronic pancreatitis. We used multivariate approach in mixMC to classify Operational Taxonomic Units (OTUs). Doing this, we found a predominance of genera Dickeya (p = 0.00008), [Eubacterium] hallii group (p = 0.0004), Bacteroides (p = 0.0006), Faecalibacterium (p = 0.006), Escherichia-Shigella (p = 0.008), and Ruminococcus 1 (p = 0.008) in bile samples from pancreaticobiliary cancers as compared to benign diseases. Additionally, bile samples from patients with pancreatic cancer exhibited a predominance of genus Rothia (p = 0.008) as compared to those with cholangiocarcinoma, whereas bile samples from patients with cholangiocarcinoma exhibited a predominance of genera Akkermansia (p = 0.031) and Achromobacter (p = 0.031) as compared to those with pancreatic cancers. Both benign and malignant pancreaticobiliary diseases have distinct microbiomic fingerprints. The relative abundance of OTUs in bile samples varies between patients with benign and malignant pancreaticobiliary diseases, as well as between cholangiocarcinoma and pancreatic cancer. Our data suggest that either these OTUs play a role in carcinogenesis or that benign disease-specific microenvironmental changes differ from cancer-specific microenvironmental changes, resulting to a clear separation of OTU clusters. We need more research to confirm and expand on our findings.

Bile detection of squamous cell carcinoma antigen (SCCA) in extrahepatic cholangiocarcinoma.

Cholangiocarcinoma (CCA) is a rare biliary tract tumor with poor prognosis that often is challenging to diagnose and the majority of patients present with advanced stage. Squamous cell carcinoma antigen 1 (SCCA1) overexpression has been found in different tumors associated with poor prognosis and chemoresistance. To assess the presence and possible prognostic role of SCCA1/2 isoforms in bile and serum of patients with CCA. Forty seven surgical patients (36 with CCA and 11 with benign diseases) were prospectively included in the study. Serum and bile specimens were collected at the time of surgery and free and IgM-complexed SCCA was quantified by ELISA (Xeptagen, srl). Free or IgM linked SCCA was rarely found in serum, while SCCA was detectable in bile samples of patients with CCA, especially in those with extrahepatic form (43% vs 17%, p=0.008), but not in controls. Despite similar tumor stage, these positive patients presented a trend toward a higher percentage of portal invasion (27% vs 15%) and of tumor recurrence than negative cases (62% vs 40%), although the difference was not statistically significant. These preliminary results indicate that bile testing for SCCA is a specific marker of extrahepatic CCA, with potential prognostic value.

Heroin-Related Fatalities in Jeddah, Saudi Arabia, between 2008 and 2018

To date, epidemiological studies have not evaluated heroin-related deaths in the Middle East and North African regions, especially Saudi Arabia. All heroin-related postmortem cases reported at the Jeddah Poison Control Center (JPCC) over a 10-year period (21 January 2008 to 31 July 2018) were reviewed. In addition, liquid chromatography electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) was utilized to determine the 6-monoacetylmorphine (6-MAM), 6-acetylcodeine (6-AC), morphine (MOR), and codeine contents in unhydrolyzed postmortem specimens. Ninety-seven heroin-related deaths were assessed in this study, and they represented 2% of the total postmortem cases at the JPCC (median age, 38; 98% male). In the blood, urine, vitreous humor, and bile samples, the median morphine concentrations were 280 ng/mL, 1400 ng/mL, 90 ng/mL, and 2200 ng/mL, respectively; 6-MAM was detected in 60%, 100%, 99%, and 59% of the samples, respectively; and 6-AC was detected in 24%, 68%, 50%, and 30% of the samples, respectively. The highest number of deaths (33% of total cases) was observed in the 21-30 age group. In addition, 61% of cases were classified as "rapid deaths," while 24% were classified as "delayed deaths." The majority (76%) of deaths were accidental; 7% were from suicide; 5% were from homicide; and 11% were undetermined. This is the first epidemiological study to investigate heroin-related fatalities in Saudi Arabia and the Middle East and North African region. The rate of heroin-related deaths in Jeddah remained stable but increased slightly at the end of the study period. Most patients were heroin-dependent abusers and from the middle-aged group. The availability of urine, vitreous humor, and bile specimens provided valuable information regarding the opioids that were administered and the survival time following heroin injection.

Microbial spectrum and drug resistance of pathogens cultured from gallbladder bile specimens of patients with cholelithiasis: A single-center retrospective study

Bacterial infection is an important cause of cholelithiasis or gallstones and interferes with its treatment. There is no consensus on bile microbial culture profiles in previous studies, and identified microbial spectrum and drug resistance is helpful for targeted preventive and therapeutic drugs in the perioperative period. To analyze the bile microbial spectrum of patients with cholelithiasis and the drug susceptibility patterns in order to establish an empirical antibiotic treatment for cholelithiasis-associated infection. A retrospective single-center study was conducted on patients diagnosed with cholelithiasis between May 2013 and December 2018. This study included 185 patients, of whom 163 (88.1%) were diagnosed with gallstones and 22 (11.9%) were diagnosed with gallstones and common bile duct stones (CBDSs). Bile culture in 38 cases (20.5%) was positive. The presence of CBDSs (OR = 5.4, 95%CI: 1.3-21.9, P = 0.03) and longer operation time (> 80 min) (OR = 4.3, 95%CI: 1.4-13.1, P = 0.01) were identified as independent risk factors for positive bile culture. Gram-negative bacteria were detected in 28 positive bile specimens, and Escherichia coli (E. coli) (19/28) and Klebsiella pneumoniae (5/28) were the most frequently identified species. Gram-positive bacteria were present in 10 specimens. The resistance rate to cephalosporin in E. coli was above 42% and varied across generations. All the isolated E. coli strains were sensitive to carbapenems, with the exception of one imipenem-resistant strain. K. pneumoniae showed a similar resistance spectrum to E. coli. Enterococcus spp. was largely sensitive to glycopeptides and penicillin, except for a few strains of E. faecium. The presence of common bile duct stones and longer operation time were identified as independent risk factors for positive bile culture in patients with cholelithiasis. The most commonly detected bacterium was E. coli. The combination of β-lactam antibiotics and β-lactamase inhibitors prescribed perioperatively appears to be effective against bile pathogens and is recommended. Additionally, regular monitoring of emerging resistance patterns is required in the future.

Effect of sphincter of Oddi dysfunction on the abundance of biliary microbiota (biliary microecology) in patients with common bile duct stones.

Biliary calculi, a common benign disease of the gastrointestinal tract, are affected by multiple factors, including diet, lifestyle, living environment, and personal and genetic background. Its occurrence is believed to be related to a change in biliary microbiota. Approximately 10%-20% of symptomatic patients with cholecystolithiasis have choledocholithiasis, resulting in infection, abdominal pain, jaundice, and biliary pancreatitis. This study aimed to determine whether a dysfunction in the sphincter of Oddi, which controls the outflow of bile and separates the bile duct from the intestine, leads to a change in biliary microbiota and the occurrence of biliary calculi. Forty patients with cholecystolithiasis and choledocholithiasis were prospectively recruited. Bile specimens were obtained, and biliary pressure was measured during and after surgery. The collected specimens were analyzed with 16S rRNA gene to characterize the biliary microbiota. The risk factors of common bile duct calculi were analyzed numerically combined with the pressure in the sphincter of Oddi. Different biliary microbiota were found in all cases. Patients with sphincter of Oddi dysfunction had significantly increased biliary microbiota as well as significantly higher level of systemic inflammation than patients with normal sphincter of Oddi. The systemic inflammatory response of patients with sphincter of Oddi dysfunction is more severe, and their microbial community significantly differs from that of patients with normal sphincter of Oddi, which makes biliary tract infection more likely; furthermore, the biliary tract of patients with sphincter of Oddi dysfunction has more gallstone-related bacterial communities.

THE IMPACT OF BILIARY DRAINAGE MODE ON BACTERIOBILIA OCCURRENCE IN PATIENTS WITH HILAR MALIGNANT OBSTRUCTION

The purpose of this study was comparative assessment of the frequency of bacterial colonization of the bile in patients with hilar malignant biliary obstruction after the palliative biliary decompression using different methodological approaches. Methods. 50 patients with proximal mechanical jaundice of tumor origin aged of ~ 62 years (25 males and 25 females), who were on steady-state treatment in Main military clinical hospital (Kyiv, Ukraine) were recruited in this prospective study. All patients underwent cholangiostomy using percutaneous transhepatic (PTBD) and external-internal suprapapillary (EISBD) approaches. Bile specimens were taken right after the biliary drainage. Identification of bacterial isolates was performed using standard cultural and biochemical methods. Results. The incidence of cholangitis was almost twice lower in EISBD group (n=26) than in PTBD group (n=24): 25.6% vs 49.1%. The rates of bacteriobilia did not differ significantly in patients from different groups: 23.1% in EISBD group and 25.0% in PTBD group. However, the frequency of biliary bacterial colonization coupled with cholangitis was also 2 times lower in EISBD group in comparison with patients underwent PTBD: 7.7% vs 16.7%. Escherichia coli predominated in bile specimens from patients with bacteriobilia associated with cholangitis in both groups. Conclusions. The use of EISBD for palliative biliary decompression in patients with proximal mechanical jaundice of tumor origin is associated with lower risk of bacterial colonization of the bile as compared to PTBD approach, and as a result with less risk of the development of infectious complications

Chlopyrifos fatal intoxication: homicide or suicide?

To document a fatal case where the organophosphorus compound chlorpyrifos is the cause of death, but the manner of exposure (voluntary or surreptitious) remains unclear. Chlorpyrifos is an organophosphorus insecticide used against a wide range of pests in commercially important crops and also for ornamental garden plants. In addition to being a particularly harmful substance at an environmental level, it also has a dangerous profile for humans. Voluntary suicidal intoxication generally occurs by oral ingestion and gastrointestinal absorption; systemic effects occur within minutes of inhalation when exposed to vapors or aerosols, but are delayed in the case of gastrointestinal absorption. A case of human intoxication is discussed: a 77 y.o. man was admitted at the emergency department due to a sudden alteration of the sensory, with tremors in the upper limbs and poor contactability. His GCS was 10, and he exhibited neurological symptoms of unclear etiology, with myoclonus, diffuse fasciculations, strong asthenia, hyponatremia. Family members did not refer any suspect ingestion nor peculiar circumstances. Only nine days after hospitalization, upon the release of liquid faeces mixed with sand, the toxicological evaluation of biological samples was undertaken thus disclosing the presence of chlorpyrifos. Death occurred after 30 days of coma. A judiciary autopsy was requested by the prosecutor office. Blood, stool, hair were collected during hospitalization. At the autopsy, blood from femoral vein, vitreous, urine, bile, liver, brain, fat and lung tissues were collected. A specific screening and confirmation method for pesticides was performed by GC-MS/MS at the Laboratory of Forensic Toxicology after liquid/liquid extraction with dichloromethane. Chlorpyrifos metabolites (chlorpyrifos oxon, 3,5,6-trichloro-2-pyridinol (TCPy), diethyl thiophosphate, diethylphosphate and glucuronide and sulfate conjugates of TCPy) were also investigated. In vivo: chlorpyrifos levels in blood samples were 893 ng/mL (admittance), 319 ng/mL (2 days later), 92 ng/mL (10 days later), 26 ng/mL (20 days later), and negative (<1 ng/mL) 22 days later. Metabolites were also detected but not quantified due to the lack of reference standards. In the 0-1 cm proximal section of 3 cm long white hair, the parent molecule was absent while O,O,O-triethylthiophosphate was identified. The solid material found in faeces with the appearance of sand was identified as sepiolite. Post-mortem: chlorpyrifos was detected at low levels (<1 ng/g) in fat and brain specimens; nor chlorpyrifos nor its metabolites were found in blood, urine, vitreous, bile, liver and lung specimens. A sealed bottle of “universal insecticide” found near his garden proved to be an emulsion containing chlorpyrifos, with different co-formulants but without any solid material such as sepiolite. The way of ingestion was deemed to be oral comparing the case blood levels with those reported for different ways of exposure. The doubt arose about the intentionality of the ingestion. The sepiolite found in faeces suggested the use of a solid formulation of chlorpyriphos, whereas finding the sealed bottle of liquid formulation seemed to be purportedly confusing. In insecticides containing chlorpyrifos found on the market, solid silica can be used as formulants or co-formulants, and the resulting products are described as free of significant odor, which would make them not excessively disgusting to ingest or to be administered orally as such or mixed with foods/drinks. The cause of death was acute chlorpyrifos intoxication, associated with prolonged coma, multi-organ failure and terminal cardiac arrest. There were no circumstantial and documentary elements useful for defining the dynamics of the intoxicating event as a suicide or a homicide. Voluntary suicide with organophosphorus compounds represents a common occurrence in Asian countries and in Eastern European countries but not in Italy.

Microbiota analysis with next-generation 16S rDNA gene sequencing in recurrent common bile duct stones.

BackgroundEndoscopic retrograde cholangiopancreatography (ERCP) is the main remedy for gallstones, but the postoperative recurrence rate is high. Recent research has indicated that the biliary microbiome takes part in the pathogenesis of cholelithiasis. However, it is not yet known whether biliary microbiome dysbiosis is relevant to recurrent cholelithiasis.MethodsThus, we investigated the bacterial communities of the biliary microbiomes of patients with recurrent common bile duct (CBD) stones and analyzed the relationship between recurrent CBD stones and biliary microbiota. The bile specimens of 5 patients with recurrent CBD stones (FF) and 45 patients with primary CBD stones (YF) were collected during the ERCP process. The microbiota was analyzed using 16S ribosomal DNA (rDNA) high-throughput sequencing. We also identified the link between recurrent CBD stones and biliary microbiota.ResultsOur results showed that at the phylum level, proteobacteria and firmicutes were the main two genera groups, and proteobacteria was high in FF patients. Additionally, synergistetes were high, but Bacteroidetes and actinobacteria were low in FF patients. The microbiomes in the bile of the YF patients were more evenly distributed than those in the bile of the FF patients. We also discovered that FF patients had decreased microbial bile diversity. At the genus level, klebsiella dominated in the FF patients, while Escherichia-shigella dominated in the YF patients. Additionally, klebsiella was higher in the FF patients than the YF patients.ConclusionsThe observed differences in the genera between the recurrent CBD stone FF patients and the YF patients provide novel insights into the link between biliary microbiota changes and recurrent CBD stones.

A Retrospective Study on Bile Culture and Antibiotic Susceptibility Patterns of Patients with Biliary Tract Infections.

Aim This study aimed to provide profiles of microorganisms isolated from bile and antibiotic susceptibility patterns of biliary tract infections (BTIs) in our center. Methods A total of 277 patients diagnosed with BTIs at the Second Affiliated Hospital of Harbin Medical University from 2011 to 2018 were included in this study. Medical records were reviewed to obtain clinical and demographic data. Bile specimens were prepared through endoscopic retrograde cholangiopancreatography (ERCP), percutaneous transhepatic cholangiodrainage (PTCD), and percutaneous transhepatic gallbladder drainage (PTGD) under aseptic conditions. In those with positive bile culture results, blood cultures were concurrently conducted. The concordance of the results between bile culture and blood culture were also analysed. Results Two hundred and sixty-seven bile cultures were positive, while 280 strains of micro-organisms were isolated. Among these, 76.8% were Gram-negative, 22.5% were Gram-positive and 0.7% were fungi. The most common microorganisms were Escherichia coli, Klebsiella pneumoniae, and Enterococcus faecalis. Gram-negative bacteria we tested were highly sensitive to ertapenem, imipenem, tigecycline, and amikacin. Gram-positive bacteria we tested were highly sensitive to tigecycline, teicoplanin, linezolid, vancomycin, and chloramphenicol. For the 44 patients with positive bile cultures, a blood culture was also performed. Among them, 29 cases yielded positive blood culture results. Among those cases with positive blood culture, 48.3% showed complete agreement with bile culture, 3.4% showed partial agreement, and 48.3% showed disagreement. The most common microorganisms in blood culture were the same as in bile culture. Additionally, the proportion of Staphylococcus epidermidis was significantly higher in blood culture (P < 0.05). Conclusion Our study provided a comprehensive analysis of the bacteria distribution and drug resistance profiles in patients with BTIs in northern China. Further studies should be conducted to validate our findings.

Quantitative Metabolomics of Tissue, Perfusate, and Bile from Rat Livers Subjected to Normothermic Machine Perfusion.

Machine perfusion (MP) allows the maintenance of liver cells in a metabolically active state ex vivo and can potentially revert metabolic perturbations caused by donor warm ischemia, procurement, and static cold storage (SCS). The present preclinical research investigated the metabolic outcome of the MP procedure by analyzing rat liver tissue, bile, and perfusate samples by means of high-field (600 MHz) nuclear magnetic resonance (NMR) spectroscopy. An established rat model of normothermic MP (NMP) was used. Experiments were carried out with the addition of an oxygen carrier (OxC) to the perfusion fluid (OxC-NMP, n = 5) or without (h-NMP, n = 5). Bile and perfusate samples were collected throughout the procedure, while biopsies were only taken at the end of NMP. Two additional groups were: (1) Native, in which tissue or bile specimens were collected from rats in resting conditions; and (2) SCS, in which biopsies were taken from cold-stored livers. Generally, NMP groups showed a distinctive metabolomic signature in all the analyzed biological matrices. In particular, many of the differentially expressed metabolites were involved in mitochondrial biochemical pathways. Succinate, acetate, 3-hydroxybutyrate, creatine, and O-phosphocholine were deeply modulated in ex vivo perfused livers compared to both the Native and SCS groups. These novel results demonstrate a broad modulation of mitochondrial metabolism during NMP that exceeds energy production and redox balance maintenance.

Comparison of Drug Resistance of Bile Isolated Bacteria in Nanjing from 2017 to 2020

Background: Analyzed pathogenic bacteria isolated from bile samples of patients with biliary tract infection and their drug resistance in the first Affiliated Hospital of Nanjing Medical University from 2017 to 2020, so as to provide evidence for the treatment of biliary tract infection. Methods: Clinical strains isolated from bile specimens of patients suspected of biliary infection in the First Affiliated Hospital of Nanjing Medical University from 2017 to 2020 were collected, The drug susceptibility criteria are based on the standards published by the National Standardization Committee of the US Clinical Laboratories. WHONET 5.6 software was used to analyze the distribution of pathogens and drug resistance. Results: From January 2017 to December 2020, a total of 2006 strains of bile pathogenic bacteria were cultured and identified. There were 1253 cases of gram-negative bacilli, 638 cases of gram-positive cocci, 98 cases of fungi, and 17 cases of Gram-positive bacilli. The top five pathogens of Gram negative bacilli were Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, Pseudomonas aeruginosa, Acinetobacter baumannii. The top five pathogens of gram positive cocci were Enterococcus faecium, Enterococcus faecalis, Enterococcus gallinarum, Enterococcus faecalis and Staphylococcus epidermidis. the resistance rate of Escherichia coli to ampicillin, ampicillin / sulbactam, cefotetan, ceftazidime, ceftriaxone,cefepime, itronam, compound trimethoprim decreased, the resistance rate of Klebsiella pneumoniae to gentamicin, levofloxacin decreased, the resistance rate of Enterobacter cloacae to ciprofloxacin, levofloxacin decreased. the resistance rate of Pseudomonas aeruginosa to ceftazidime, cefepime, gentamicin decreased. the resistance rate of Acinetobacter baumannii to ceftazidime, ciprofloxacin decreased. the resistance rate of Enterococcus faecium to penicillin G, high concentration streptomycin, erythromycin decreased. Conclusions: Enterogenic pathogens were the main pathogens in China. Such as Escherichia coli, Klebsiella pneumoniae, Enterococcus faecalis and Enterococcus faecalis; Nosocomial infection pathogens. For example, Pseudomonas aeruginosa and Acinetobacter also need to be paid attention to. Clinicians should improve their awareness of microbiological examination and provide more reliable pathogenic evidence for the selection of clinical antibiotic.

Cultivation and Genomic Characterization of the Bile Bacterial Species From Cholecystitis Patients.

The microbes in human bile are closely related to gallbladder health and other potential disorders. Although the bile microbial community has been investigated by recent studies using amplicon or metagenomic sequencing technologies, the genomic information of the microbial species resident in bile is rarely reported. Herein, we isolated 138 bacterial colonies from the fresh bile specimens of four cholecystitis patients using a culturome approach and genomically characterized 35 non-redundant strains using whole-genome shotgun sequencing. The bile bacterial isolates spanned 3 classes, 6 orders, 10 families, and 14 genera, of which the members of Enterococcus, Escherichia–Shigella, Lysinibacillus, and Enterobacter frequently appeared. Genomic analysis identified three species, including Providencia sp. D135, Psychrobacter sp. D093, and Vibrio sp. D074, which are not represented in existing reference genome databases. Based on the genome data, the functional capacity between bile and gut isolates was compared. The bile strains encoded 5,488 KEGG orthologs, of which 4.9% were specific to the gut strains, including the enzymes involved in biofilm formation, two-component systems, and quorum-sensing pathways. A total of 472 antibiotic resistance genes (ARGs) were identified from the bile genomes including multidrug resistance proteins (42.6%), fluoroquinolone resistance proteins (12.3%), aminoglycoside resistance proteins (9.1%), and β-lactamase (7.2%). Moreover, in vitro experiments showed that some bile bacteria have the capabilities for bile salt deconjugation or biotransformation (of primary bile acids into secondary bile acids). Although the physiological or pathological significance of these bacteria needs further exploration, our works expanded knowledge about the genome, diversity, and function of human bile bacteria.

Demographic, Chemical, and Helicobacter pylori Positivity Assessment in Different Types of Gallstones and the Bile in a Random Sample of Cholecystectomied Iranian Patients with Cholelithiasis.

Background The occurrence of stones in the gallbladder or common bile duct and the symptoms and complications they cause is called gallstone disease. The symptoms of gallstone disease range from mild, nonspecific symptoms to a severe right quadrant abdominal pain. Characteristics of gallstone types in an Iranian population have not been well studied before and there are very limited studies on the demographic pattern of stone types in our country, so this study is one of the first studies on its kind on the epidemiology of gallstone types in Iran. As information on chemical components of the stone will help in the management and prevention of gallstones, in this study, we aimed to do chemical component analysis of gallstones including cholesterol, bilirubin, and calcium. Given the conflicting reports about the relationship between H. pylori infections and gallstone formation, this study aimed to investigate the relationship between H. pylori positivity in the bile specimen of Iranian patients with cholelithiasis and formation and type of stone. Methods This prospective study reviewed a total of 196 patients who underwent cholecystectomy for symptomatic cholelithiasis at Shahid Beheshti Training and Research Hospital affiliated to the Yasuj University of Medical Sciences between September 2015 and May 2018. Chemical analysis of gallstone components performed using the colorimetry method. Microbiological analysis for H. pylori was done using the OnSite H. pylori Ag Rapid Test on the bile sample. For the validation test of bile, the H. pylori Rapid Stool Ag Test on stool was used, and Cohen's Kappa statistical analysis was done next. Results There were significant associations between the stone types and age, chemical composition of the stones such as calcium, cholesterol, and bilirubin levels, and also H. pylori positivity and cholesterol and bilirubin levels; however, no significant association was found between the stone types and sex, H. pylori positivity and age, sex, stone types, and calcium level. The main bile and validity tests were matched to the substantial agreement according to Cohen's Kappa analysis. The most common drugs used were proton pump inhibitors, nonsteroidal anti-inflammatory drugs, antihypertensive drugs, and oral contraceptives. Conclusions This study suggested that the chemical composition of the stones could predict the presence of bacteria, there is no correlation between H. pylori and gallstone formation, and some of the drugs could be predisposing factors for gallstones. This work provides an objective basis for further research into gallbladder stone formation; meanwhile, it has great significance in the treatment and prevention of gallbladder stones. Trial registration. The project was found to be in accordance to the ethical principles and the national norms and standards for conducting research in Iran with the approval ID IR.YUMS.REC.1399.147 and date 2020.09.23, and this project is the result of a residency dissertation to obtain the specialty in general surgery, which has been registered with the research project number 960159 in the Vice Chancellor for Research and Technology Development of the Yasuj University of Medical Sciences, Yasuj, Iran, URL: https://ethics.research.ac.ir/EthicsProposalViewEn.php?id=160634.