THE HUMAN MICROBIOME - WHERE IS IT HIDING?

Tomasz Gosiewski*, Dominika Salamon, Agnieszka Sroka-Oleksiak, Agnieszka Krawczyk, Monika Brzychczy-Włoch

Department of Molecular Medical Microbiology, Jagiellonian University Collegium Medicum, Department of
Microbiology, Poland
*E-mail: tomasz.gosiewski@uj.edu.pl

Microbiome is a term that defines the genomes of all microorganisms (bacteria, archaeons, fungi or viruses) present in the host organism. The microbiome is therefore a genetic representation of the microbiota, or living cells (viruses escape this classification) interacting with the host. In humans, it has become accepted dogma that the microbiota naturally colonizes the oral cavity, pharynx, gastrointestinal tract (with accumulation in the colon), skin, urethra and vagina. These anatomical locations (ecologists would call them habitats) contain microorganisms that are relatively easy to grow and thus establish their presence, which in taxonomic terms (qualitative as well as quantitative) is relatively constant and typical of a healthy person. What then of other regions of the human body, such as the vascular bed (blood), bronchial tree, uterus with fetus (amniotic fluid), cerebrospinal fluid? Obviously, as a result of the expansion of microorganisms into these regions (invasive infections), an infection occurs with a whole range of clinical picture and the possibility of growing a pathogenic etiological agent.

 

The matter therefore seems simple and for decades was almost an axiom of microbiology and infection medicine. Classical culture-based diagnostic methods continue to lead the way in microbiology, despite the fact that their versatility often prevents the growth of specific species under laboratory conditions. It is this weakness of cultures, which have often failed to yield the expected results in the diagnosis of blood or central nervous system infections, despite all the clinical signs of an ongoing inflammatory process with an infectious basis, that has prompted microbiologists to reach for an arsenal of methods based on the detection of microbial nucleic acids. Methods such as PCR, FISH (Fluorescence in situ hybridization) or next-generation sequencing (NGS) do not depend on whether microbial cells will or will not be able to proliferate in the laboratory, but instead target genetic markers for the presence of microorganisms in a sample. This approach significantly increases the sensitivity of detection, and often reduces the time to wait for test results to a few hours or even tens of minutes. Molecular biology methods have thus made it possible to confirm blood or cerebrospinal fluid (CSF) infection, even if the culture is negative.

Given the diagnostic successes achieved by molecular biology, one began to wonder what would happen if one harnessed these methods to test considered "physiologically sterile" body fluids, such as blood or amniotic fluid and others. It has turned out that PCR or FISH techniques make it possible to detect the presence of bacterial DNA and RNA in test samples. Of course, there were allegations that what was really being detected was microbial contamination of the samples, rather than the actual presence of microbiota due to the significantly higher sensitivity of molecular than culture-based methods. These doubts persisted until the NGS method came into widespread use, enabling high-throughput and comprehensive analysis of microbial taxonomic profiles in samples. This was a revolutionary change in microbiome research. NGS allows the detection of marker 16S rRNA sequences of bacteria and archaeons, or ITS of fungi, and in addition allows their quantification. The obtained genetic library sequences are compared with genetic databases, and based on this, the microbial taxa in the samples are identified. Since NGS is limited only by the detail of genetic databases, almost every detected sequence in a sample can be assigned a specific taxon. This has led to the fact that the previously known and described in microbiology textbooks profiles of cultured bacteria and fungi from the skin, vagina, gastrointestinal tract, oral cavity, urethra, turned out to be just the tip of the iceberg, and that microbiomes are much more biodiverse than thought. In the colon alone, more than 1,000 species of bacteria have been identified, of which perhaps 30% manage to be cultured. By using NGS to study "physiologically sterile" samples, detailed taxonomic profiles of bacteria have been obtained where they were not previously expected (except for symptomatic infections). Literature databases are filling up very quickly with more papers proving that blood, amniotic fluid, bronchial tree and perhaps even PMR are not free from the presence of microorganisms, which, however, do not cause clinical signs of ongoing infection. Data are still being collected and microorganisms will be discovered where we would not have expected them until recently. What's more, scientific evidence is being provided, of strong links between the microbiota and the physiology or pathophysiology of the host. So, do microbes permeate us thoroughly and are there no taboos for them?

 

Funding source: National Science Center - 2017/26/E/NZ5/00266