**1. The Gut Microbiota**

Our digestive system, particularly our intestines, harbors a vast amount of microorganisms including bacteria, archaea, bacteriophages, eukaryotic viruses, and fungi called the microbiota [1], whose genetic makeup is referred to as the microbiome.

The number of these microorganisms inhabiting the human gastrointestinal tract is extremely high, reaching, according to some estimates, a ratio of equality (about 1:1) vis-àvis all cells in the human body, while the genetic material of these microorganisms appears to have at least 100 times more genetic diversity than that of the entire human genome [2,3].

A "superorganism" is defined as the set of the host and all the microorganisms that colonize it [2].

In terms of their composition, most of these bacteria belong to the *Firmicutes* phyla (64%), followed by *Bacteroidetes* (23%) and *Proteobacteria* (8%) to Gram-negative bacteria such as *E. coli* and *H. pylori* [4] (Figure 1).

The various gastrointestinal regions, because of their different characteristics, represent different microenvironments, in which specific microorganisms grow [5].

The stomach has special characteristics, such as its acidic ph, so most of the microorganisms that colonize it are acid resistant.

The most important microorganism residing in the gastric lumen is Helicobacter pylori, which influences the growth of other secondary species, which may play a mutualistic or pathogenic role [6].

**Citation:** Piccioni, A.; Rosa, F.; Manca, F.; Pignataro, G.; Zanza, C.; Savioli, G.; Covino, M.; Ojetti, V.; Gasbarrini, A.; Franceschi, F.; et al. Gut Microbiota and *Clostridium difficile*: What We Know and the New Frontiers. *Int. J. Mol. Sci.* **2022**, *23*, 13323. https://doi.org/10.3390/ ijms232113323

Academic Editor: Maria Teresa Mascellino

Received: 29 September 2022 Accepted: 29 October 2022 Published: 1 November 2022

**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

**Figure 1.** Composition of the gut microbiota. The most represented phyla are Firmicutes (64%) and Bacteroidetes (23%), followed by Proteobacteria (8%), Fusobacteria, verrucomicrobia, and actinobacteria (3%).

Regarding the small intestine, it is characterized by the presence of oxygen, rapid luminal flow, and bactericidal secretions such as bile acids.

In the duodenum, the predominant phyla are Firmicutes and Actinobacteria [7].

The colon, on the other hand, is in a condition of anaerobiosis, where the slower passage of food, absorption of water, and fermentation of undigested food take place.

For these reasons, the most common microorganisms are *Bacteroides*, *Bifidobacterium*, *Streptococcus*, *Enterobacteriaceae*, *Enterococcus*, *Clostridium*, *Lactobacillus*, and *Ruminococcus* [8] (Table 1).

**Table 1.** Summary of the predominant microorganisms in the various tracts of the digestive system with their basic characteristics.


This type of complex mutualistic interaction between these microorganisms and their host appears to have evolved over thousands of years [2].

In fact, the ingestion of germs that will later go on to contribute to the formation of human intestinal flora has been known since ancestral times, traces of which have been found in cave paintings, dating as far back as the Neolithic age [3].

The interaction between diet, microbiota, and the human host has fascinated researchers since the turn of the last century when Metchnikoff tried to link this complex interaction and senescence [4].

One of the controversial aspects that characterize this topic, is that in the early days, it was not very clear whether it was the microbiota that predisposed toward certain pathological conditions or whether the opposite was the case [9], while currently, most researchers argue that it is an alteration of the gut microbiota that predisposes toward the onset of certain diseases.

Most of these microorganisms play a commensal role with their host [9].

In fact, this complex host–individual interaction could become so long-lived only by bringing, most of the time, benefits to both parties.

In recent years, more and more efforts are being made to investigate certain aspects that characterize the microbiome, and one of these is to understand its interindividual diversity.

The mechanism by which each individual develops his or her own microbiome appears to be multifactorial in origin, with several factors being called into play, some genetic and others environmental such as those related to childbirth and early life, followed by the type of infant feeding, medications taken, and lifestyle [9].

We have seen how the gut microbiota is greatly affected by environmental influences. An interesting line of research is on strategies to maintain good health through the homeostasis of the gut microbiota.

Some authors suggest practicing a vegetarian diet, while wheat gluten, red meat, and alcohol are related to dysbiosis that can trigger a chronic inflammatory response [10].

The very fact that so many factors come into play therefore makes it very arduous to know about each one.

Another fascinating field of study in recent years is trying to identify and understand the many functions performed by the microbiome.

The roles played by the gut microbiota are multiple, going on to act on food digestion, drug metabolism, and regulation of intestinal endocrine function, and going as far as being involved in host immune defense mechanisms (Table 2) [1].

**Table 2.** Gut microbiota functions.


The roles played by the microbiota are indeed multifaceted; in fact, it is not surprising to learn that some researchers are focusing on the possible role of microorganisms in fighting cancer, a new frontier with little evidence at the moment but very promising [11].

Again, the real challenge is to try to disentangle all these different functions that are performed in different individuals.

In our case, we will see how crucial its protective function is in the healthy individual and how disastrous its lack is in patients who will develop *Clostridium difficile* infection.
