Metagenomic analysis of the bacteriobiom and mycobiom of the gastrointestinal tract of children and adolescents with celiac disease and their healthy siblings, as well as healthy children carrying the HLA DQ2 and/or DQ8 antigen

Project leader: Tomasz Gosiewski, Ph.D., Prof. UJ
Implementation period: 2018 - 2023
Project type: Sonata Bis 7

The set of microorganisms that populate the human body is referred to as the microbiota. The main component of this collection is bacteria. It is estimated that the human microbiota has about 10 times more bacteria than there are human cells. The most numerous group is the microflora of the gastrointestinal tract, especially the terminal segment - the large intestine. It is considered a specific ecosystem that has a significant impact on the functioning of the human body. Meanwhile, the term microbiome refers to all the genomes of microorganisms living in and on the human body. However, the term microbiome is often used in the sense of microbiota. The importance and role of the gut microbiota is increasingly understood and helps explain the course and causes of many diseases, such as inflammatory bowel disease, diabetes and obesity. There have also been hypotheses that the composition of the microbiota is disturbed in celiac disease (CD) patients compared to healthy individuals. Celiac disease (other terms for the disease are: gluten-dependent visceral disease, gluten-sensitive enteropathy, or non-tropical sprue) is the most common type of food intolerance, and also characterized by the most severe course. The disease has an immunological basis, which means that it is the result of a strong response of the gastrointestinal tract's immune system to a plant protein - gluten - introduced into the body. It is mainly contained in cereal products (wheat, rye and barley) and when it is found in the diet of a person who has a certain genetic predisposition, it causes an excessive immune response, which in the long run leads to atrophy of the intestinal villi and finally to a reduction in the intestinal absorption surface. CD can manifest at any age, but is most often diagnosed in childhood.

Currently, the only effective treatment for this disease is strict adherence to a gluten-free diet. Consumption of gluten generates an immune response, which must affect the state of the gastrointestinal flora. On the other hand, the composition of the gastrointestinal microflora stimulates the intestinal immune system GALT, which under physiological conditions should lead to a balance between the host organism and the microbiota. To date, in the analysis of the intestinal microbiota in celiac patients, only a few studies focusing on selected groups of bacteria have been conducted. Among other things, it has been shown that in the course of CD and adherence to a gluten-free diet, the abundance of bacteria belonging to the Lactobacillus and Bifidobacterium species, considered a reservoir of strains with probiotic potential, decreases. However, more comprehensive studies covering the entire human gut microbiome, which consists of more than 1,000 bacterial species, are lacking. This gap can be filled by the presented research project. By analyzing samples from both the upper gastrointestinal tract (duodenal mucosal fragments collected during gastroscopy) and the lower gastrointestinal tract (fecal samples) and using modern molecular diagnostic methods, it is planned to conduct a detailed study of the intestinal microbiota in terms of the qualitative and quantitative composition of bacteria and selected fungi in children with newly diagnosed celiac disease compared to healthy children. The plan is also to evaluate the gut microbiome in relation to the degree of adherence to a gluten-free diet by children with celiac disease. Such a comprehensive analysis will be possible through the use of state-of-the-art molecular diagnostic methods. Next-generation metagenomic sequencing (NSG), i.e. analysis of DNA isolated directly from microorganisms found in a given environment (without the need for prior culture on artificial media), using the molecular marker that is the rDNA sequence (encodes the rRNA gene - a component of the minor subunit of the ribosome of prokaryotes and eukaryotes), will be used to determine the type and number of bacteria.

Finally, the obtained nucleotide sequences will be subjected to bioinformatics analysis allowing to classify each bacterium and fungus into a given taxonomic group. Obtaining scientific data proving possible links between celiac disease and the microbiome will perhaps allow preventive measures (in the form of appropriate antibiotics or probiotics) to modify the gastrointestinal flora. Such action could prevent the development of the disease in genetically susceptible individuals or alleviate symptoms in those already ill.