**The Benefits of Probiotics in Human and Animal Nutrition**

Camila Boaventura, Rafael Azevedo, Ana Uetanabaro, Jacques Nicoli and Luis Gustavo Braga *Universidade Estadual de Santa Cruz Brazil* 

### **1. Introduction**

74 New Advances in the Basic and Clinical Gastroenterology

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Yan, F. & Polk, D.B. (2002). Probiotic bacterium prevents cytokine-induced apoptosis in intestinal epithelial cells. *J Biol Chem*, Vol. 277, No. 52, pp. (50959-65). Yang, H., Liu, A., Zhang, M., Ibrahim, S.A., Pang, Z., Leng, X. & Ren, F. (2009). Oral

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administration of live Bifidobacterium substrains isolated from centenarians enhances intestinal function in mice. *Curr Microbiol,* Vol. 59, No. 4, pp. (439-445). Yason, C.V., Summers, B.A. & Schat, K.A. (1987). Pathogenesis of rotavirus infection in

various age groups of chickens and turkeys: pathology. *American Journal of* 

of yeast (Saccharomyces cerevisiae) cell components on growth performance, meat quality, and ileal mucosa development of broiler chicks. *Poult Sci*, Vol. 84, No. 7,

> At birth, the gastrointestinal tract of any animals is sterile, and it is rapidly colonized by bacteria from the mother and the environment. This colonization by the gut microbiota plays an important role in intestinal tract maturation of newborn (in terms of anatomy, digestive physiology, and immunology) (Hooper 2004). After this colonization, considering healthy human individuals, the gastrointestinal tract harbors 10 or more times as many microbes than there are eukaryotic cells (1014 viable cells for indigenous microbiota/1013 body cells).These microorganisms, altogether weighing approximately 1.5 kg, can be considered as a complementary major organ, responsible for three main functions: colonization resistance, immunomodulation, and nutritional contribution (Hayashi et al., 2002; Zoetendal et al., 2011). Colonization resistance inhibits the installation of exogenous microorganisms as well as the uncontrolled multiplication of microorganisms belonging to the indigenous microbiota. Immunomodulation maintains the immune system under a watchful state, which permits a faster but adequate response in the case of infectious aggression. Nutritional contribution furnishes complementary sources of vitamins, enzymes, and energy substrates (volatile fatty acids).

> Unfortunately, several factors can disturb both the initial colonization and posterior maintenance of the gut microbiota, leading to a microbial ecosystem with beneficial functions transitorily or irreversibly less efficient. As examples, the type of delivery (cesarean or natural) or the reduction of mother-child contacts (premature baby in an incubator or in an intensive care unit) interfere with the supply of microorganisms necessary for post-natal colonization. Additionally, the alimentation (breast- or formula-fed) and the ingestion of antibacterial drugs may be other factors that modify the normal sequence of colonization (Harmsen et al., 2000; Bonnemaison et al., 2003; Westerbeek et al., 2006; Chen et al., 2007). Once installed, the beneficial functions of the microbiota are very powerful but also fragile and can be disturbed by ingestion of drugs (especially antibiotics), drastic changes in diet or stress. In view of what was presented above, the importance of a correct initial colonization and a subsequent preservation of the gut microbiota is evident to obtain optimal functions from this microbial ecosystem. When disturbances of the indigenous microbiota functions are forecasted or installed, you should think about the possibility of compensating failures of these functions. In this sense, probiotics can be considered as

The Benefits of Probiotics in Human and Animal Nutrition 77

(Servin & Coconnier, 2003; Momose et al., 2008); immunomodulation of the host (Ezendam et al, 2006); and inactivation of microbial toxin (Brandão et al., 1998). Other mechanisms by which probiotics may exert protection is through a recuperation of mucosal barrier function when disturbed (Penna et al., 2008), trapping pathogens on their surface (Martins et al, 2010; Martins et al, 2011) and stimulating mucus production (Caballero-Franco et al., 2007).

According to De Vrese & Marteau (2007), mechanism and efficiency of probiotic effect depend mainly on the interactions between probiotic microorganisms and microbiota of the

Althought they had not been completely elucidated, the classical mechanisms of action of bacteria used as probiotics are described as: i) competition for bound sites: also known as "competitive exclusion", where bacteria of probiotics are linked to the bound site in the intestinal mucosa, making a physical barrier, impeding the bound by pathogenic bacteria; ii) production of anti-bacterial substances: bacteria of probiotics synthetize compounds as for example bacteriocins, hydrogen peroxide, which has antibacterial action, mainly in relation to pathogenic bacteria, in addition to the production of organic acid which reduced pH in the gastrointestinal tract, preventing growth of many pathogens and development of certain species of *Lactobacillus*; iii) competition for nutrients: shortage of available nutrients which can be used by pathogenic bacteria is a limiting factor for their maintenance; iv) stimulus to the immune system: some bacteria of the probiotics are directly linked to the stimuli of immune response by increasing antibodies production, activation of macrophages, T cells

Action mechanism of yeasts still needs studies for their evidencing. A probable mechanism of action of yeast is related to total (*in vitro*) or partial inhibition of pathogenic microorganisms. Dead yeasts contain in their walls important quantities of polysaccharides and proteins able to act positively in the immune system and on nutrient absorption. Moreover, yeasts produce nutritive metabolites in the digestive tract which increase animal performance, in addition of having minerals (Mn, Co, Zn) and vitamins (A, B12, D3) which

Although some mechanisms had been suggested on the action of probiotics, they are not completely clarified, but it is known that they inhibit growth of pathogenic microorganism by producing antimicrobial compounds; they compete with pathogens for adhesion sites and nutrients; and they model immune system of the host (Oelschlaeger, 2010). In the present, a more complete view on the possible mechanisms of action are been studied, based

The composition of human microbiota has about 10 - 100 trillion members and varies within the gut and among individuals (Zoetendal et al., 2011; Hayashi et al., 2002). These members belong mainly to the dominant bacteria, but there are also representatives from Archaea (Eckburg et al., 2003), Eukarya, and viruses, including bacteriophages (Breitbart et al., 2003). Intestinal microbiota plays a fundamental role in maintaining immune homeostasis which, in other words, involves minimizing the adverse health effects of intestinal microbiota, such as shifts in microbial community structure, changes in the diet of the host or overt

According to Sonnenburg, et al. (2006) some evidences show that by manipulating the microbiota with probiotics could influence the host health and probiotic bacteria could be

host or with imunocompetent cell of the intestinal mucous.

proliferation and interferon production (Fuller, 1992; Jin et al., 1997).

improve action of beneficial microorganisms (Hill et al., 2006).

mainly on the manipulation of normal microbiota.

pathogenic challenge (Hooper & Macpherson, 2010).

biotherapeutics to be used in microbial ecosystems during the installation phase (colonization of the newborn), or with installed (treatment) or forecasted disturbances (prophylaxis).

Probiotics have been defined in a joint meeting of the Food and Agriculture Organization/World Health Organization as "live microorganisms which when administered in adequate amounts confer a benefit to host health" (WHO / FAO, 2002). The objective of its use is to install, enhance or compensate the functions of the indigenous microbiota inhabiting the digestive tract or other body surfaces. The suggestion of using fermented food to obtain some benefits for the health is not new. It was mentioned in the Persian version of the Old Testament (Genesis 18:8) that "Abraham attributed his longevity to the consumption of sour milk." Later in 76 BC Pline, a Roman historian, recommended the use of fermented milk products for the treatment of gastroenteritis (Schrezenmeir & de Vres, 2001). However, a scientific approach, recognizing the beneficial role of certain microorganisms has been only applied in the first decades of the twentieth century with the suggestion of the use of *Lactobacillus* (Elie Metchnikoff attributing the longevity of Bulgarian populations to the yogurt consumption in 1907), *Bifidobacterium* ( Henri Tissier observing a higher presence of bifidobacteria in the feces of healthy breast-fed children in 1906) and *Saccharomyces boulardii* (Henri Boulard noting the use of a tropical fruit colonized by this yeast to treat diarrhea by local populations in Eastern Orient during a cholera outbreak in 1920) to prevent or treat gastrointestinal disorders (Shortt, 1999). The probiotics most often used at the moment are bacteria producing lactic acid (*Lactobacillus, Bifidobacterium*) and yeasts (*Saccharomyces boulardii*).

Many health benefits have been related to human and animal intake of probiotic. Several studies have supplied clinical evidences of the benefits generated by probiotics, as for example in diarrhea treatment (Billooet al., 2006; De Vrese & Marteau, 2007), lactose intolerance (He et al., 2008), irritable bowel syndrome (De Vrese et al., 2001; Nagala & Routray, 2011), allergies (Jain et al., 2010), cancer (Chen et al., 2009) and hypercholesterolemia (Baroutkoub, 2010). According to recent meta-analysis based on well conducted clinical trials with probiotics, a clear protective effect was evident, which did not vary significantly between products containing *S. boulardii, Lactobacillus rhamnosus* GG, *L. acidophilus, L. bulgaricus, L. casei, Bifidobacterium longum, B. bifidum* var. *infantis* and *B. animalis* var. *lactis* (Sazawal et al., 2006).

Probiotics have also received special attention by animal nutrition researchers who search for alternatives to the use of traditional growth promoters (antibiotics). Therefore, the use of probiotics is seen more and more as an alternative to the use of antibiotics in animal production, and many scientific works show the beneficial effects of supplementation with probiotic strains in diets fed to chicken, swine, cattle and fish (Veizaj-Delia, 2010; Soleimani et al., 2010; Ignatova, 2009; Aly et al., 2008). Therefore, this chapter will approach action mechanisms and the effects on health of probiotics for human and animal use.

#### **1.1 Mechanisms of action**

The potential mechanisms by which probiotic agents might exert their protective effect include: antagonism by the production of substances that inhibit or kill the pathogen (Servin, 2004); competition with the pathogen for adhesion sites or nutritional sources

biotherapeutics to be used in microbial ecosystems during the installation phase (colonization of the newborn), or with installed (treatment) or forecasted disturbances

Probiotics have been defined in a joint meeting of the Food and Agriculture Organization/World Health Organization as "live microorganisms which when administered in adequate amounts confer a benefit to host health" (WHO / FAO, 2002). The objective of its use is to install, enhance or compensate the functions of the indigenous microbiota inhabiting the digestive tract or other body surfaces. The suggestion of using fermented food to obtain some benefits for the health is not new. It was mentioned in the Persian version of the Old Testament (Genesis 18:8) that "Abraham attributed his longevity to the consumption of sour milk." Later in 76 BC Pline, a Roman historian, recommended the use of fermented milk products for the treatment of gastroenteritis (Schrezenmeir & de Vres, 2001). However, a scientific approach, recognizing the beneficial role of certain microorganisms has been only applied in the first decades of the twentieth century with the suggestion of the use of *Lactobacillus* (Elie Metchnikoff attributing the longevity of Bulgarian populations to the yogurt consumption in 1907), *Bifidobacterium* ( Henri Tissier observing a higher presence of bifidobacteria in the feces of healthy breast-fed children in 1906) and *Saccharomyces boulardii* (Henri Boulard noting the use of a tropical fruit colonized by this yeast to treat diarrhea by local populations in Eastern Orient during a cholera outbreak in 1920) to prevent or treat gastrointestinal disorders (Shortt, 1999). The probiotics most often used at the moment are bacteria producing lactic acid (*Lactobacillus, Bifidobacterium*) and

Many health benefits have been related to human and animal intake of probiotic. Several studies have supplied clinical evidences of the benefits generated by probiotics, as for example in diarrhea treatment (Billooet al., 2006; De Vrese & Marteau, 2007), lactose intolerance (He et al., 2008), irritable bowel syndrome (De Vrese et al., 2001; Nagala & Routray, 2011), allergies (Jain et al., 2010), cancer (Chen et al., 2009) and hypercholesterolemia (Baroutkoub, 2010). According to recent meta-analysis based on well conducted clinical trials with probiotics, a clear protective effect was evident, which did not vary significantly between products containing *S. boulardii, Lactobacillus rhamnosus* GG, *L. acidophilus, L. bulgaricus, L. casei, Bifidobacterium longum, B. bifidum* var. *infantis* and *B.* 

Probiotics have also received special attention by animal nutrition researchers who search for alternatives to the use of traditional growth promoters (antibiotics). Therefore, the use of probiotics is seen more and more as an alternative to the use of antibiotics in animal production, and many scientific works show the beneficial effects of supplementation with probiotic strains in diets fed to chicken, swine, cattle and fish (Veizaj-Delia, 2010; Soleimani et al., 2010; Ignatova, 2009; Aly et al., 2008). Therefore, this chapter will approach action

The potential mechanisms by which probiotic agents might exert their protective effect include: antagonism by the production of substances that inhibit or kill the pathogen (Servin, 2004); competition with the pathogen for adhesion sites or nutritional sources

mechanisms and the effects on health of probiotics for human and animal use.

(prophylaxis).

yeasts (*Saccharomyces boulardii*).

*animalis* var. *lactis* (Sazawal et al., 2006).

**1.1 Mechanisms of action** 

(Servin & Coconnier, 2003; Momose et al., 2008); immunomodulation of the host (Ezendam et al, 2006); and inactivation of microbial toxin (Brandão et al., 1998). Other mechanisms by which probiotics may exert protection is through a recuperation of mucosal barrier function when disturbed (Penna et al., 2008), trapping pathogens on their surface (Martins et al, 2010; Martins et al, 2011) and stimulating mucus production (Caballero-Franco et al., 2007).

According to De Vrese & Marteau (2007), mechanism and efficiency of probiotic effect depend mainly on the interactions between probiotic microorganisms and microbiota of the host or with imunocompetent cell of the intestinal mucous.

Althought they had not been completely elucidated, the classical mechanisms of action of bacteria used as probiotics are described as: i) competition for bound sites: also known as "competitive exclusion", where bacteria of probiotics are linked to the bound site in the intestinal mucosa, making a physical barrier, impeding the bound by pathogenic bacteria; ii) production of anti-bacterial substances: bacteria of probiotics synthetize compounds as for example bacteriocins, hydrogen peroxide, which has antibacterial action, mainly in relation to pathogenic bacteria, in addition to the production of organic acid which reduced pH in the gastrointestinal tract, preventing growth of many pathogens and development of certain species of *Lactobacillus*; iii) competition for nutrients: shortage of available nutrients which can be used by pathogenic bacteria is a limiting factor for their maintenance; iv) stimulus to the immune system: some bacteria of the probiotics are directly linked to the stimuli of immune response by increasing antibodies production, activation of macrophages, T cells proliferation and interferon production (Fuller, 1992; Jin et al., 1997).

Action mechanism of yeasts still needs studies for their evidencing. A probable mechanism of action of yeast is related to total (*in vitro*) or partial inhibition of pathogenic microorganisms. Dead yeasts contain in their walls important quantities of polysaccharides and proteins able to act positively in the immune system and on nutrient absorption. Moreover, yeasts produce nutritive metabolites in the digestive tract which increase animal performance, in addition of having minerals (Mn, Co, Zn) and vitamins (A, B12, D3) which improve action of beneficial microorganisms (Hill et al., 2006).

Although some mechanisms had been suggested on the action of probiotics, they are not completely clarified, but it is known that they inhibit growth of pathogenic microorganism by producing antimicrobial compounds; they compete with pathogens for adhesion sites and nutrients; and they model immune system of the host (Oelschlaeger, 2010). In the present, a more complete view on the possible mechanisms of action are been studied, based mainly on the manipulation of normal microbiota.

The composition of human microbiota has about 10 - 100 trillion members and varies within the gut and among individuals (Zoetendal et al., 2011; Hayashi et al., 2002). These members belong mainly to the dominant bacteria, but there are also representatives from Archaea (Eckburg et al., 2003), Eukarya, and viruses, including bacteriophages (Breitbart et al., 2003). Intestinal microbiota plays a fundamental role in maintaining immune homeostasis which, in other words, involves minimizing the adverse health effects of intestinal microbiota, such as shifts in microbial community structure, changes in the diet of the host or overt pathogenic challenge (Hooper & Macpherson, 2010).

According to Sonnenburg, et al. (2006) some evidences show that by manipulating the microbiota with probiotics could influence the host health and probiotic bacteria could be

The Benefits of Probiotics in Human and Animal Nutrition 79

Afterwards, a new culture is prepared only with the target colonies for *in vivo* assessment (pathogen inhibition, target species pathogenicity; resistance to host conditions; among others). If there are no restriction to the use of the target species, experiments with in vivo supplementation at big and small scale are carried out to check if there are real benefits to the host. Finally, the probiotic which presented significant satisfactory results can be

One of the main applications of probiotic microorganisms is at preventing or in the treatment of gastrointestinal disturbances. In a clinical trial carried out with children hospitalized for acute rotavirus diarrhea, three treatments were assessed. The first group of children received oral rehydration therapy plus placebo; the second group was submitted to oral rehydration plus *Saccharomyces boulardii* treatment and the third group received oral rehydration and a compound containing *Lactobacillus acidophilus*, *Lactobacillus rhamnosus*, *Bifidobacterium longum* e *Saccharomyces boulardii*. Mean duration of diarrhea was shorter for the children who received the treatment with *Saccharomyces boulardii strain* (58 hours) and for the children who received the compound with for different microorganism strains (60 hours), when compared to the control group (84.5 hours) (Grandy et al., 2010). In Brazil, a double-blind, placebo controlled trial showed that protection against diarrhea (32.2% reduction in diarrhea during the first year of life) was obtained by oral inoculation with a single dose of plasmid-free human *Escherichia coli* EMO soon after birth (Figueiredo et al.,

Treatment with antibiotics can cause an unbalance in the indigenous microbiota, increasing concentration of pathogenic microorganisms and toxin production, promoting diarrhea symptoms (Vasiljevic & Shah, 2008). A significant effect was observed in a study carried out with patients who presented diarrhea caused by antibiotics, in which intake of a probiotic drink containing *L. casei*, *L. bulgaricus* e *S. thermophilus* reduced the incidence of diarrhea (Hickson et al., 2007). In a double-blind, formula controlled trial performed in Brazil, a milk lyophilized formulation supplemented with *B. bifidum* and *S. thermophilus* was compared with another without supplementation for the prevention of antibiotic-associated diarrhea in children 6 to 36 months old. The authors observed a significant reduction of diarrhea frequency in children treated with the probiotic formula (16% of 80 patients) when compared to the control group (31% of 77 patients) (Corrêa et al., 2005). In another doubleblind, placebo controlled trial also performed in Brazil, the treatment with a lyophilized preparation of *S. boulardii* in children with acute diarrhea was evaluated and a reduction in duration of rotavirus diarrhea was observed in the group treated with the probiotic yeast (Corrêa et al., 2011). Other examples of clinical trials related to prevention or shortening of

Irritable bowel syndrome is a diseased characterized by abdominal pain, diarrhea, constipation and mucus secretion along with feces (Vahedi et al., 2010). Although many physiopathology factors had been correlated to the cause of this disease, in the last years, researchers have considered feed intolerance and unbalance of intestinal microbiota as the main factors responsible for symptoms of the irritable bowel syndrome. Probiotics are a

diarrhea symptoms by using probiotics are summarized in Table 1.

**2.3 Irritable bowel syndrome** 

commercially produced and used.

2001).

**2.2 Prevention or reduction of diarrhea symptoms** 

used as a therapeutic strategy to improve human health. The precise mechanisms influencing the crosstalk between the microbe and the host are still unclear but there are evidences suggesting that bacteria in the gut could modulate the functioning of the immune system at systemic and mucosal levels (Ng et al., 2009).
