**5. Clinical uses of probiotics**

Probiotics are live bacteria meant to inoculate the gut of the host and incorporate into an already diverse microbiota. Probiotics are broadly used in three categories: immunomodulation, normalization of intestinal microbiota, and metabolic effects [45]. In general, the quality of evidence for use in clinical conditions remains low. The literature to support their use has been most clear in necrotizing enterocolitis (NEC) in neonates and pouchitis in ulcerative colitis (UC) patients. However, the use of probiotics well beyond the gastrointestinal tract is ongoing. We will review the studies about the current state of probiotics used in various disease states in this section (**Table 1**).

### **5.1 Antibiotic-associated diarrhea**

Antibiotic-associated diarrhea (AAD) is a common side-effect of the antibiotics that can affect up to a third of patients receiving antibiotics [58]. Broad-spectrum

*Translation of Immunomodulatory Effects of Probiotics into Clinical Practice DOI: http://dx.doi.org/10.5772/intechopen.109864*


**Table 1.**

*Summarization of known effects of probiotics in specific disease states.*

antibiotics with activity against anaerobes are associated with higher rates of the AAD [47]. AAD may last two months after the onset of antibiotic therapy resulting in significant morbidity [47]. Several randomized-controlled trials and meta-analyses, including bacterial strain-specific trials, have shown that the use of Lactobacillus and Saccharomyces has shown potential benefits of probiotics in addressing AAD [59, 60]. It is postulated that probiotics could antagonize the pathogenic microorganisms in the human flora when the host has been exposed to antibiotics [61]. The mechanism of their interference involves interference with cell signaling, direct production of bacteriocins, and augmentation of the systemic immune response of the host [62–66]. Most studies in AAD have focused on inpatients who were on intravenous antibiotics at higher concentrations where concurrent administration of probiotics has conferred a protective effect in some instances [67]. Probiotics have also been shown in meta-analyses to have a protective effect in outpatients receiving antibiotics without adverse side effects [68]. However, there remains a dearth of direct comparisons between specific strains and their effectiveness when used in conjunction with specific antibiotics. In addition, in the studies finding evidence of benefit, there are inconsistent definitions of diarrhea, specific infections treated, and the types of

antibiotics being used [59]. This makes it challenging for clinicians to target probiotic treatment regiments to specific diseases. Thus, clinicians are not able to make specific recommendations to patients despite the strong interest and high prevalence of AAD.

To date, there is no global consensus on the use of the probiotics for AAD. The World Gastroenterology Organization (WGO) has supported their use of AAD in both adults and pediatrics. The use of *L. rhamnosus* GG and *Saccharomyces boulardii* was recommended by the Canadian Agency for Drugs and Technologies (CADTH) [69]. However, this recommendation is not shared by AGA or the IDSA (Infectious Disease Society of America) [49]. While there are certainly benefits to using probiotics in patients with AAD, the recommendations have not been able to clearly define the most appropriate patient or context.

### **5.2 Probiotics in** *Clostridioides difficile* **infections (CDI)**

*C. difficile* is the most prevalent AAD for inpatients and outpatients leading to significant morbidity and mortality [70]. CDI is often associated with exposure to anaerobic coverage and antibiotics such as clindamycin, fluoroquinolones, or cephalosporins. Strategies to prevent *C. difficile* spread have typically involved patient segregation and hygiene measures. However, attempts to alter the host microbiota with fecal transplant or probiotics have become mainstream and shown themselves to be conclusive. In hospitalized patients receiving antibiotics, prophylactic administration of probiotics has been shown to significantly reduce the risk of developing *C. difficile*-associated diarrhea [71–75].

A proposed mechanism of this has been seen in *Saccharomyces boulardii,* which in murine models was shown to make a 54-kDa serine protease that cleaves toxin A and its intestinal receptor [50, 72, 76]. This has also been replicated in humans where toxin A and B cytotoxic effects in the human colon were attenuated when incubated in purified *S. boulardii* protease prior to being placed in the human colon [77]. When used in combination with metronidazole or vancomycin it reduced the number or relapses of diarrhea [78]. Efforts made for targeted primary prevention of CDI have typically focused on a multi-modal approach involving hygiene, antibiotics, and probiotics. A specific formulation of probiotics known as Bio-K+, which includes *L. acidophilus* CL1285, *L. casei* LBC80R, and *L. rhamnosus* CLR2, has been marketed in North America since 1996. Mouse models exposed to Bio-K+ have been found to increase concentrations of *lactobacilli* while decreasing levels of *staphylococci* [1]. The pathology in the human colon arises from toxins A and B of *C. difficile* that affects the colonic epithelium, which results in loss of cellular integrity and disruption of the colon mucosal cell cytoskeleton. Bio-K+ strains have been shown to produce supernatants (extracellular products) that inhibit toxin A/B in human enterocytes Caco-2 and HT-29 cells [1].

The American College of Gastroenterology (ACG), ESCMID (European Society of Clinical Microbiology and Infectious Diseases), and IDSA recommend probiotics for prevention or treatment of primary and recurrent *C. difficile* infections. However, the AGA is in favor of the use of *S. boulardii*, *L. acidophilus* CL1285, and *L. casei* for adults and children who are being treated with antibiotics except in situations of severe illness [79]. The difference among professional organizations comes from lack of clear evidence on the safety profiles and whether there is a true benefit [49]. Given the conclusive evidence on fecal microbiota transplants as a definitive treatment for recurrent CDI, there is no question of the significance that microbiota plays in the development of CDI and the potential manipulating it has in the

prophylaxis and treatment of CDI. Furthermore, optimizing probiotic supplementation may have a meaningful role in CDI treatment.

### **5.3 Inflammatory bowel disease and probiotics**

IBD pathophysiology involves a complex interplay between genetics, the host microbiome, environmental conditions, and the individual's immune response [80, 81]. Changes in the intestinal mucosa and microbiota may disrupt homeostasis between the human immune system and the flora [82]. These changes may then trigger a reaction of the human immune system playing a role in development of the IBD. Indeed, specific intestinal microbiota profiles have been associated with active disease [83]. CD and UC patients have been found to have less *Firmicutes* and *Bacteroidetes* and more *Proteobacteria* and *Actinobacteria* when compared to healthy controls [84]. In addition, CD patients have been found to have reduced levels of *Bacteroides*, *Eubacterium*, *Faecalibacterium*, and *Ruminococcus* possibly leading to increased gut permeability [84]. A technology to help distinguish commensal from quiescent pathologic bacteria has been developed known as IgA-SEQ, which combines cell sorting with 16srRNA gene sequencing to quantify the amount of IgA on various taxa of bacteria found in the gastrointestinal tract. By measuring the amount of IgA coating, immunostimulatory and immunoregulatory taxa of the microbiota can be measured more accurately. This can then be used to confer susceptibility to IBD. While IgA itself does not contribute to the inflammatory response in IBD, this technology revealed three potential bacteria, which were associated with disease progression in IBD and three protective taxa. Taxa with relatively low abundance (based on 16S rRNA) were *Erysipelotrichaceae* sp. and *Faecalibacterium prausnitzii*, as well as low IgA coating of *Oscillospira* was associated with less progression to surgery [85]. These studies identify disease-modifying taxa and biomarkers for disease severity and progression. By identifying bacteria taxa as so-called "bad actors" in the human microbiome there may be a framework for the development of more refined biomarkers impacting disease courses and the possibility of microbiome-based therapeutics.

There has also been an association between CD and the colonization of adherentinvasive *Escherichia coli* (AIEC). AIEC is thought to impair mitochondrial function in epithelial cells of the gastrointestinal mucosa by invading the Peyer's patches and the lamina propria *via* M cells [86]. It is thought that AIEC incorporates into macrophages and possibly increases the proinflammatory cytokine TNF-α. Patients with highly expressed CEACAM6 and CHI3L1 receptors, which are often expressed during times of inflammation, have been shown to promote the adhesion of AIEC and consequently bacteria invasion at the ileum [87]. Monocyte-derived macrophages (MDM) taken from patients with CD are unable to restrict AIEC as compared to healthy controls MDM *in vitro* models leading to pathologic immune response [88]. The overall prevalence of AIEC in healthy individuals is about 0–16% in the colon and 6–19% in ileal samples compared to 21–63% in CD patients suggesting that AIEC may be an additive factor in the pathogenesis of CD [51, 89, 90].

The increasing interest in the immune response to the gut microbiome in IBD has been met with interest in probiotic supplementation in this condition for induction and maintenance of remission. Specifically, it is thought that probiotics might be able to impact IBD pathophysiology by improving epithelium integrity, downregulating inflammatory bacterial byproducts, and reducing mucin production [91, 92]. Certain probiotic strains such as *lactobacilli* and *bifidobacteria* produce bacteriocins that act as antimicrobial peptides [54, 93]. *Lactobacillus paracasei* L74 CBA often found

in fermented milk products like Kefir has been shown to inhibit pro-inflammatory cytokines such as NF-Kb potentially providing anti-inflammatory properties to the host. In addition, Duary et al. found that TNF-α was reduced by *Lactiplantibacillus plantarum* Lp91. While these findings have not been shown in human-based clinic trials, they provide a potential mechanism by which probiotics may have some clinical value in IBD [94].

The most used formation of probiotics in IBD patients is known as Visbiome®/ VSL #3® (Italian form), which was developed by Sigma-Tau Healthscience/ Alfasigma. The original formulation was changed in 2016 and there is now a U.S. version known as Visbiome® and an Italian version known as (VSL3®). In CD, the data has remained mixed on the efficacy of probiotics to induce or retain remission as an adjuvant or stand-alone therapy. The mechanism of action possibly includes improving tight junction protein function, positive composition of the intestinal microbiota, and regulating immune-related cytokine expression. In regard to CD, there was one randomized control (RCT) that evaluated the ability of VSL#3 to prevent human recurrence after surgery. This study looked at early and the late administration of VSL#3 and found that early VSL#3 administration was associated with later recurrence after surgery. While there have been no statistical differences in endoscopic recurrence rates at day 90 between patients who received VSL#3 and patients who received placebo. Levels of inflammatory cytokines and recurrence rates leading to repeat surgery were lower among patients who received early VSL#3 (for the entire 365 days). This indicated that this probiotic should be further investigated for prevention of Crohn's disease recurrence [94, 95].

While it is understood that there may be a potential for probiotics in UC, there is still no convincing data to constitute a recommendation. In a small cohort of pediatric patients with UC, *Lactobacillus reuteri* was shown to improve clinical and endoscopic disease activity [96, 97]. This has not been replicated in adults. However, in patients with UC who have undergone total proctocolectomy and ileal pouch-anal anastomosis for UC, a definitive connection to gut microbiota has been made. There has been a potential benefit in VSL#3 containing *Lactobacillus*, *Bifidobacterium*, and *Streptococcus* for prevention of the initial episode of acute pouchitis. To date, there have been four clinical trials showing VSL#3 could prevent or maintain remission in patients with chronic pouchitis [98–101]. A potential mechanism suggested is the improvement of the intestinal barrier function (IBF). While VSL#3 has shown efficacy in chronic pouchitis, an open-label trial showed that most patients on chronic antibiotics for pouchitis were not able to use VSL#3 for long-term therapy largely due to disease recurrence [102]. This formulation has been demonstrated in preventing future episodes and improving inflammation [96]. However, according to the AGA, this only constitutes a weak recommendation due to the small size of the patient population in which these studies were done.

The next generation of probiotics in IBD may involve the use of genetically engineered bacteria that could release therapeutically operative molecules in the intestine. This will involve organisms that could sense and respond to intestinal inflammatory cytokines or topically produce molecules to treat the inflammation. Harnessing the power of the biotherapeutics with synthetic biology could provide a future of personalized medicine in the diverse IBD patient population [103].

### **5.4 Necrotizing enterocolitis**

Preterm birth impacts about 10% of newborns born in the US and 15 million pregnancies worldwide. A preterm infant's gut is exposed to colonization of commensal

### *Translation of Immunomodulatory Effects of Probiotics into Clinical Practice DOI: http://dx.doi.org/10.5772/intechopen.109864*

and pathological bacteria. During this time, their innate immune system is sorting through a constant excess of peptidoglycans and liposaccharides [104]. In this delicate time, NEC inflammation can be driven by Toll-like receptor 4. By influencing the innate and adaptive immune systems, probiotics are thought to aid in the balance of these two systems and prevent the pathogenesis of NEC [104, 105]. NEC is associated with bowel necrosis leading to short bowel syndrome and impaired development, and can be fatal in up to 30% of patients [55]. There have been case-control studies identifying an overpopulation or so-called "bloom" of Gammaproteobacteria tending to precede NEC in many preterm infants [56, 106]. In contrast, commensal bacteria such as bifidobacterial are found to be protective of NEC and plentiful in breastfed infants likely due to the breast milk-specific oligosaccharides that this preferentially consumes [107].

A Cochrane review article found probiotics were superior to placebo in reducing the risk of severe necrotizing enterocolitis (RR = 0.43; 95% CI, 0.33–0.56; 20 studies with 5529 infants) and mortality (RR = 0.65; 95% CI, 0.52–0.81; 17 studies with 5112 infants) [108]. Combinations of certain probiotics containing *Bifidobacterium infantis* and *L. acidophilus* have shown strong association with preventing NEC and reducing need for abdominal surgery and all-cause mortality [109, 110].

There are numerous hypotheses on the mechanism of how they might protect against NEC in infants. One such proposition involves the production of butyrate and other short-chain fatty acids that could supply nutrition to the colonocytes thereby lowering the pH and decreasing the oxygen tension within the intestinal lumen. This ultimately is thought to suppress the growth of *Enterobacteriaceae* (phylum *Proteobacteria*), which is well known to be pathologic in NEC [111, 112]. Other proposed mechanisms include supporting the maturation and functions of the infants' bowels by regulating the Th1:Th2 balance [57, 113]. Specifically, it is known that an imbalance of Th2 levels greater than Th1 levels can predispose to autoimmune disease and gut inflammation by lack of regulation of the gut immune response [57]. According to the AGA, in babies less than 37 weeks of gestational age and low-birthweight infants, it is recommended to use a combination of probiotics containing *Lactobacillus* spp. and *Bifidobacterium* spp. over no probiotics to prevent the development of NEC in this population. This constitutes a conditional recommendation with a moderate to high level of evidence in this population.
