Metagenomic analysis of the intestinal archeobiom and mycobiom of children with Crohn's disease compared to healthy subjects
Project manager: Agnieszka Krawczyk, M.Sc.
Implementation period: 2020 - 2023
Project type: Prelude 17
The human gastrointestinal tract harbors a huge number of microorganisms, the total mass of which accounts for up to 2 kilograms. The microbiota of the gastrointestinal tract consists primarily of bacteria, as well as somewhat less numerous populations of fungi, viruses, protozoa or archaeons. Their composition and abundance varies significantly in different sections of the digestive tract. The greatest activity, diversity and number of microorganisms is observed in the large intestine. It is estimated that this part of the digestive tract is home to up to 1,000 different species of microorganisms, belonging to 45 genera and 17 families of microorganisms. Over the past few years, thanks to the development of methods based on molecular biology techniques, the complex interactions that occur between microorganisms and the human body are becoming better understood. As a result, there is an increase in knowledge of the impact of the gut microbiota on the course of many diseases, such as inflammatory bowel disease, systemic connective tissue inflammation, anorexia, obesity, diabetes, or conditions associated with mental disorders such as depression, autism and Parkinson's disease.
Recently, much attention has been paid to the role of intestinal dysbiosis in the induction of Crohn's Disease (CD). CD is a chronic autoimmune inflammatory disease of the gastrointestinal tract, with periods of remission and exacerbation. Although the disease has been known for more than a century, both the causes of its recurrence and its etiopathogenesis remain unexplained. As numerous studies show an abnormal composition of the gastrointestinal microbiota among CD patients, it has begun to be hypothesized that the disease is the result of genetic and immunological abnormalities, which, due to intestinal dysbiosis, result in an overreactive immune system against microorganisms. Consequently, this leads to permanent, non-specific damage to the mucosa of the gastrointestinal tract. The involvement of microorganisms in the pathogenesis is supported by a number of clinical observations, including the improvement of patients' health after the use of antibiotics and intestinal cleansing procedures, the alleviation of disease symptoms or the achievement of remission due to the modification of the gastrointestinal microbiota with probiotic drugs, the recurrence of the disease due to food poisoning and gastrointestinal infections, the absence of intestinal inflammation among experimental animals raised in sterile conditions of the so-called "germ free". "germ free", or achieving remission among patients after fecal transplantation from healthy individuals.
To date, the analyses that have been conducted of the gut microbiota of CD patients have focused primarily on abnormalities within the bacteriome. However, there is a lack of more comprehensive studies involving the archaeobiom or mycobiom, which is also a component of the natural gut microbiome. This gap can be filled by the presented research project, which aims to determine the taxonomic composition of fungi and archaeons colonizing the gut of children with newly diagnosed CD in comparison to healthy subjects. Using modern molecular biology techniques, such as Next Generation Sequencing (NGS) and Real-Time PCR, it will be possible to accurately and comprehensively assess qualitatively and quantitatively the fungi and archaeons of the gastrointestinal tract, even those that cannot be cultured on artificial media. These techniques, are based on the analysis of DNA isolated directly from microorganisms found in a given environment, using a molecular marker such as the rDNA sequence. The obtained nucleotide sequences will be subjected to bioinformatics analysis, which in turn will allow accurate assignment of the tested fungi and archaeons to a specific taxonomic group. There is no doubt that the composition of the microbiome significantly affects the maintenance of internal balance and health, and the quantitative and/or qualitative change of microorganisms is an important factor inducing pathological processes.Given that both fungi and archaeons are an important component of the intestinal microbiota, it is not impossible that a change in their abundance or composition could affect the occurrence of certain diseases, especially CD, which is the research hypothesis of this project.
If a difference in the composition and/or quantity of these microorganisms is found between a group of patients with CD and a healthy population, this would be a premise confirming the involvement of these microorganisms in the development of the disease. The finding of such a disparity will perhaps allow in the future to implement preventive measures to modify the gastrointestinal microbiota with probiotics, targeted antibiotic therapy, antifungal agents or fecal transplantation, which could alleviate the symptoms of CD or even prevent the onset of the disease.