**2. The gap between healthy and septic gut microbiomes**

Sepsis is a multifaceted host response to an infecting pathogen that may be significantly amplified by endogenous factors [6]. The broader perspective also emphasizes the significant biological and clinical heterogeneity in affected individuals such as their age, underlying comorbidities, concurrent injuries (including surgery) and medications, and source of infection adding to further complexity [7]. The success of antibiotic treatment depends on rapid and accurate identification of relevant pathogens and is complicated by the increasing rate of antimicrobial resistance conditioned by the dynamic changes in the bacterial population in which aerobic and facultative anaerobic bacteria predominant at the onset of sepsis are replaced by anaerobic species as the oxygen levels deplete. Broad-spectrum therapy is administered in the absence of bacterial identification, but this may not accurately reflect causative pathogens [8].

For a better understanding of how to treat, we probably should change the paradigm from "anthropocentrism" to "microbiocentrism," as we think.

The presence of over hundreds of species in the gut of a healthy adult host is a way to survive in an ever-changing world and the ability to receive energy from different sources of food. In critical condition, the advantage is obtained by those species that are capable of surviving in more extreme conditions with less oxygen and a lack of nutrients and trace elements. For example, *Enterococcus* is one of the few microorganisms capable of surviving and thriving in the presence of bile acids, an increased concentration (6.5%) of NaCl, hydrogen peroxide, and changes in the pH level [9]. The most frequent cause of abdominal sepsis is a leakage of fecal material from the intestinal lumen into the peritoneal cavity [10]. The leakage introduces gut bacteria, including *Enterobacteriaceae*, *Enterococcus* spp., *Streptococcus* spp., and *Staphylococcus* spp., into the sterile peritoneal environment. Another prospective study of 32 patients admitted to the ICU after the trauma and acute care surgery service similarly found a replacement of intestinal *Faecalibacterium* and *Ruminococcus* with the more pathogenic *Enterococcus* [11]. The site of infection is not usually in the gut, but the metabolic influence of the pathogens on the gut microbiota and host tends to be persistently overlooked. For example, microbes that flourished in the guts of elite athletes boosted the time that lab mice ran on a treadmill. These particular microbes seem to take lactate, pumped out by muscles during exercise, and turn it into a compound that may contribute to endurance [12].

rRNA gene encodes highly specific RNA of bacterial ribosomes and is present in genomes of all known microorganisms. Its structure is quite conservative, but variable-specific regions allow identifying microorganisms of different species and strains. The study pattern is quite simple but rather laborious: at the first stage, DNA is isolated from a sample, and then a so-called genome library containing copies of gene 16S rRNA belonging to different bacteria is obtained. The library is "read" using high-performance sequenators providing reception of several thousand nucleotide sequences of gene 16S rRNA for each sample. The next stage deals with analysis of a huge body of received data using bioinformatic techniques. Results are represented in a way most suitable in each particular case. The introduction of latest technologies, for example, nanopore sequencing, allows fast identification of bacteria in samples and finding markers of resistance to antimicrobial drugs within 5–10 minutes with the portable real-time device for DNA and RNA sequencing "MinION" that weighs less than 100 grams. This method is currently undergoing clinical testing [4]. However, in a typical microbiome experiment, several aspects of microbial communities still remain inaccessible. These include low-abundance but potentially crucial taxa whose genetic material is not sampled by sequencing techniques due to being present below the level of detection [5]. The real value of all this novel knowledge to understand the pathogenesis of sepsis has yet to be established. In this chapter, we are discussing the important role of bacterial metabolites in comparison with

taxonomic structure of the septic gut microbiota.

"anthropocentrism" to "microbiocentrism," as we think.

pathogens [8].

26 Infectious Process and Sepsis

**2. The gap between healthy and septic gut microbiomes**

Sepsis is a multifaceted host response to an infecting pathogen that may be significantly amplified by endogenous factors [6]. The broader perspective also emphasizes the significant biological and clinical heterogeneity in affected individuals such as their age, underlying comorbidities, concurrent injuries (including surgery) and medications, and source of infection adding to further complexity [7]. The success of antibiotic treatment depends on rapid and accurate identification of relevant pathogens and is complicated by the increasing rate of antimicrobial resistance conditioned by the dynamic changes in the bacterial population in which aerobic and facultative anaerobic bacteria predominant at the onset of sepsis are replaced by anaerobic species as the oxygen levels deplete. Broad-spectrum therapy is administered in the absence of bacterial identification, but this may not accurately reflect causative

For a better understanding of how to treat, we probably should change the paradigm from

The presence of over hundreds of species in the gut of a healthy adult host is a way to survive in an ever-changing world and the ability to receive energy from different sources of food. In critical condition, the advantage is obtained by those species that are capable of surviving in more extreme conditions with less oxygen and a lack of nutrients and trace elements. For example, *Enterococcus* is one of the few microorganisms capable of surviving and thriving in the presence of bile acids, an increased concentration (6.5%) of NaCl, hydrogen peroxide, and changes in the pH level [9]. The most frequent cause of abdominal sepsis is a leakage of fecal In our preliminary study, we used gas chromatography-mass spectrometry (GC-MS) analysis of blood serum and feces simultaneously and at the same time analyzed the taxonomic composition of the gut microbiota using 16S rRNA gene-based metagenomic analysis in groups of patients with sepsis, n = 9, and healthy, n = 5. The sepsis was diagnosed according to the Sepsis 3 definition [7].

The taxonomic composition of the gut microbiota in a group at the phylum level as determined by the metagenomic analysis of feces is shown in **Figure 1**. The major four phyla of the human gut microbiota, *Bacteroidetes*, *Firmicutes*, *Proteobacteria*, *Actinobacteria*, were the predominant phyla in most patients. The composition of the gut microbiota was not stable in any of the patients, and dynamic changes were observed in all nine patients. At the same time, the absolute percentage of *Proteobacteria* in septic patients was several times higher than in healthy volunteers. This was confirmed at the family level. The *Enterobacteriaceae* family, which is a part of the *Proteobacteria*, was shown to represent the leading species among the top 10 in sepsis. However, clear understanding cannot be reached using only taxonomy since it allows to observe only a handful of processes taking place in the development of any infection (**Figure 1**).

As we have shown earlier, high levels of some aromatic microbial metabolites (AMMs) in serum are related to the severity and mortality of critically ill patients [13]. The sum of the level of eight most relevant metabolites, benzoic (BA), phenylpropionic (PhPA), phenyllactic (PhLA), p-hydroxyphenylbenzoic (p-HBA), p-hydroxyphenylacetic (p-HPhAA), p-hydroxyphenylpropionic (p-HPhPA), homovanillic (HVA), and p-hydroxyphenyllactic acids (p-HPhLA), in serum samples from septic patients was higher than in healthy people 3.7 (1.2–8.0) μM and 1.3 (1.0–1.6) μM, respectively (p < 0.05). In the septic group, the maximum values of the sum of these metabolites were more than 10 μM which is higher than in patients with lethal outcome. The differences in the AMM quality profiles of simultaneously serum and fecal samples (SFS) of patients with sepsis and healthy are presented in **Figure 2**. The results showed that the feces of healthy people abound with such metabolites, p-PhAA, p-HPhPA, and p-HPhLA, supporting data obtained by Jenner et al. [14]. At the same time, we observed prevalence of BA, PhLA, and p-PhAA in sepsis with a higher level of BA in the gut of non-survivors. Differences in the proportion of AMM in the blood compared to the intestine can be explained by the fact that most hydrophilic (p-HPhAA, p-HPhLA, and PhLA) metabolites are excreted by the kidneys, while lipophilic metabolites (BA, PhAA, and PhPA) are absorbed by cells of tissue barriers (intestinal wall, lymphoid tissue, liver, vascular endothelium, etc.).

Experimental study of the proximal part of the gastrointestinal tract showed that BA had a bacteriostatic and bactericidal dose-dependent effect on coliform and lactic acid bacteria [15]. On the one hand, it is important to emphasize that the dysfunction of the microbiota is manifested by excessive production of certain microbial metabolites as a reflection of the high microbial load with pathological colonization by bacteria involved in the development of sepsis. On the other hand, microbiota function, which is very important for host homeostasis, such as microbial biodegradation of an excess of endogenous biologically active compounds, due to a decrease in biodiversity in the intestine, primarily a deficiency of indigenous anaerobes, is disturbed [16]. The altered profile of aromatic metabolites in the blood may be an integral indicator reflecting these dramatic disturbances and possibly other functions of the "invisible organ."

Microbiota-Oriented Diagnostics and Therapy in Sepsis: Utopia or Necessity?

http://dx.doi.org/10.5772/intechopen.89187

29

**3. The gut microbial metabolites in the pathogenesis of sepsis**

ecules (**Figure 3**).

in comparison.

It was shown that in vitro some sepsis-associated AMM in clinically significant concentrations can inhibit the phagocytic activity of neutrophils [17]; cause mitochondrial dysfunction [18]; influence on platelet aggregation [19]; reduce tyrosine hydroxylase activity, thus limiting the synthesis of catecholamines; and participate in the pathogenesis of septic shock [20]. Numerous data obtained in vitro allow us to hypothesize that AMM acts as signaling mol-

It is impossible to exclude the presence of common signaling pathways, cell receptors, transmembrane transporters, and other mechanisms of humans and bacteria, as well as the direct participation of microbial metabolites in the pathogenesis of sepsis. Thus, today, we should not confine ourselves to studying eukaryotic cells while searching for new molecular mechanisms of sepsis-associated organ failure and septic shock [20]. We should consider

**Figure 3.** Schematic representation of levels of some biochemical parameters, metabolites, and hormones in blood serum

**Figure 1.** Taxonomic composition of the gut microbiota by metagenomic analysis. Comparison the taxonomic composition of the gut microbiota: (a) at the major phylum and (b) by top 10 families.

**Figure 2.** Metabolic profile of aromatic metabolites in: (A) the gut and (B) the blood serum. The data are presented by median of the proportion of each acid among all AMMs.

In particular, serum samples of healthy people are characterized by a predominance of BA and PhPA, while hydrophilic AMMs are detected in sepsis with the appearance of high levels of HVA in the serum of non-survivors. BA is a product of the synthesis of bacteria, plants, and fungi, but a significant content is formed as a result of biodegradation of phenylalanine. Experimental study of the proximal part of the gastrointestinal tract showed that BA had a bacteriostatic and bactericidal dose-dependent effect on coliform and lactic acid bacteria [15].

On the one hand, it is important to emphasize that the dysfunction of the microbiota is manifested by excessive production of certain microbial metabolites as a reflection of the high microbial load with pathological colonization by bacteria involved in the development of sepsis. On the other hand, microbiota function, which is very important for host homeostasis, such as microbial biodegradation of an excess of endogenous biologically active compounds, due to a decrease in biodiversity in the intestine, primarily a deficiency of indigenous anaerobes, is disturbed [16]. The altered profile of aromatic metabolites in the blood may be an integral indicator reflecting these dramatic disturbances and possibly other functions of the "invisible organ."
