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Characterization of the Human Gallbladder Microbiome

Periodic Reporting for period 1 - GallBiome (Characterization of the Human Gallbladder Microbiome)

Reporting period: 2016-05-01 to 2018-04-30

Cholecystitis, or inflammation of the gallbladder, occurs due to gallstone production or blockages in the cystic ducts and causes severe pain (biliary colic) and fever, often requiring surgery. When bile flow is interrupted, a number of secondary problems such as fat-soluble vitamin malabsorption and liver damage can occur. Once the gallbladder is surgically removed, patients often encounter complications in the form of acute infections or chronic bowel difficulties that can last months or years, making gallbladder disease one of the most common and costly of all digestive diseases. Post-cholecystectomy Syndrome (PCS) arises from alterations in bile flow due to loss of the reservoir function of the gallbladder, and affects approximately 20-40% of patients. Although the exact reasons have not yet been elucidated, these complications are examples of a much wider problem – unknown effects of gallbladder removal on the human body’s biggest organ, its microbiome. Without the gallbladder, the body lacks a way of regulating the flow of bile into the intestines. It is expected that the increased bile flow leads to downstream changes in the intestinal population, but no such studies have been performed in humans.

Cholecystectomies rank number one for the most frequently performed major surgery. The National Health and Nutrition Examination Survey in the US estimated that 6.3 million men and 14.2 million women aged 20-74 yrs suffer gallbladder disease. The prevalence is 20 million cases among Europeans, whereby the incidence of gallstones is 1 million new cases/yr. In the US, direct and indirect costs of gallbladder disease represented a consumption of $6.2 billion in 2012, increasing by more than 20% over the last three decades. In 2013 England reported >12,000 cases of cholelithiasis, making it one of the primary reasons for hospital visits relating to gastrointestinal issues. Treatment is governed by non-specific diagnoses and usually includes pharmacologic or surgical approaches. As these number increase, considerable burden is placed on the healthcare system and taxpayer funds. However, the degree of research in this topic does not reflect demand, and much remains to be known about the epidemiology of gallbladder disease.

The MSCA-funded GallBiome project aimed to define the core gallbladder microbiome. It was hypothesized that measurable differences would be seen among the dominant phyla in healthy compared to diseased gallbladders and that removal of the gallbladder would lead to gut dysbiosis. To investigate these questions the fellow sought to achieve the following objectives: 1) mapping community changes linked to gallbladder disease; 2) quantifying changes in bacterial communities from gallbladder mucosa, bile, and stool as a function of time and patient status, 3) evaluations of gallbladder removal on the gut population (3) correlations among population profiles with metabolite profiles to glean microbial functions, and 4) the isolation and characterization of previously uncultivated anaerobic bacteria. Ultimately, this project seeks to understand the role of bacteria in gallbladder disease and will form the basis for establishing relationships between gallbladder microbiota and gut microbiota to benefit human health.
"The Human GallBiome project is still currently on-going, and the fellow is repeating tasks using a Metabolic Syndrome pig model, whereby gallbladders, bile, and faecal samples have been processed for microbiological and metabolomic analysis. This additional investigation has revealed significant differences in gut taxa (among other parameters) of pigs fed high-fat diet vs. control pigs on normal chow, and metabolites present in both bile and gallbladder mucosae separated distinctly when visualized with heatmaps and PCoA plots.

The dissemination strategy included a number activities such as seminars, hands-on anaerobic workshops, training and technology transfer, and presentations at conferences, symposia and Postdoctoral workshops. Accordingly, information about the project and/or researcher was disseminated in the following ways: 1) featured as a Research Highlight in the APC Microbiome Institute’s publication, 'Gut Reaction' 2) article titled ""Bacteria and Bugs in Guts and Gallbladders"" appeared in the ‘Researchers’ Lives’ column of the Irish Times newspaper 3) progress presented at APC Marie Curie Festival – “Characterisation of the Human Gallbladder Microbiome” 4) fellow profiled in TResearch publication for special edition “Teagasc Women in STEM” 5) presented “GallBiome: Tips and Tricks from a Funded Fellow” discussing progress and funding at 'The Proposal Writing Workshop for MSCA Individual Fellowships', 6) delivered a talk at The Chinese Nutrition Science Congress in Beijing on “The Human Microbiome, at the Interface of Nutritional Health and Disease” 7) poster abstract “The Human GallBiome, At The Interface of Hepatic Health and Disease” published in the conference proceedings of The International Symposium of Probiotics and Prebiotics (ISAPP). Manuscripts are in preparation."
The information generated by this work is expected to define better options for educating patients, health care professionals and industry about the sources and problems associated with bacterial dysbiosis related to gallbladder surgery. Analyses of biopsies and bile has 1) presented unique DNA based community sequencing surveys for establishing healthy/diseased gallbladder microbiome profiles 2) revealed metabolic activities of the microbes involved, and 3) provided a road-map for the recovery of novel bacterial. Furthermore, data from community surveys will address questions of disease pathogenicity, variations in population structure based on the degree/type of medical occurrence, and characterization of the gallbladder ecology. Although the GallBiome project is still on-going, the clinical & research team has impacted the greater scientific society by defining SOPs for human gallbladder microbiome sampling, collection, and processing. The contingency plan enacted in pigs provided a unique opportunity to investigate diet as a link to gallbladder disease, and facilitated collection of additional data (plasma, heart ventricle size, liver weight) not possible in humans. This work will lead to establishment of the pig as a model organism for studying cardiological effects of westernized diets, and will provide an excellent comparative dataset when more human gallbladder samples are obtained.

Ultimately, new knowledge relevant to pathophysiology, microbiology, and taxonomy will be generated through these projects. This research will highlight new models of gallbladder infection, including the bacterial participants and novel insights into pathogen recognition, and will inform the development of next generation diagnostic tools for the prevention and treatment of cholelithiasis. Cholecystectomies may affect normal gastrointestinal functions, therefore, results of this proposal could aid in the identification of consequences for gallbladder removal not once realized. It is expected that this action will reveal the culprits of disease, a result that could alter the treatment procedures currently in place or offer alternatives to surgery. As such, these findings are necessary for the evaluation of better patient management practices and contribute to the attainment of EU strategic goals.
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