**Chapter 5**

*Drug Repurposing - Hypothesis, Molecular Aspects and Therapeutic Applications*

[35] Southall NT, Natarajan M, Lau LP, Jonker AH, Deprez B, Guilliams T, et al. The use or generation of biomedical data and existing medicines to

discover and establish new treatments

recommendations of the IRDiRC Data Mining and Repurposing Task Force. Orphanet Journal of Rare Diseases.

[36] Fda.https://wwwaccessdatafdagov/ scripts/opdlisting/oopd/detailedIndexcf m?cfgridkey=632218 [Online]. Available from: https://www.accessdata.fda.gov/ scripts/opdlisting/oopd/detailedIndex. cfm?cfgridkey=632218 [Accessed:

for patients with rare diseases–

2019;**14**(1):1-8

28 February 2020]

[37] Yella JK, Yaddanapudi S, Wang Y, Jegga AG. Changing trends in computational drug repositioning. Pharmaceuticals. 2018;**11**(2):57

**78**


**3.1 Antibiotics/antifungals**

ATB therapy is used for the management of ASD and is routinely prescribed to treat ASD symptoms associated with several underlying disorders like pediatric autoimmune neuropsychiatric disorder associated with streptococcal infections (PANDAS) and pediatric acute-onset neuropsychiatric syndrome (PANS) [72]. Short-term administration of vancomycin was shown to provide significant improvement in both GI and behavioral disturbances in a subset of children with ASD [73]. Unfortunately, this attempt had only partial success since children relapsed, and benefits waned upon treatment termination. As vancomycin is a poorly absorbed ATB known to destroy Gram-positive anaerobes, observed improvement is believed to be the consequence of temporary elimination of toxinproducing clostridia population. Hence, symptomatic relapse was attributed to the spore-forming capacity of these bacteria, and spores resistant to ATB would later germinate into vegetative forms once treatment has stopped [46].

Children with ASD, particularly those with GI disease, are sometimes treated with antifungal agents, as they may have increased incidence of fungal infection. Despite the lack of evidence of fungal overgrowth in children with ASD, parents find that antifungal therapies can often be beneficial and may be a useful adjuvant for the treatments of ASD [74].

Thus, it is possible to speculate that anti-infectives, through modulation of GM, should be able to influence symptoms and expression of psychiatric disorders. However, ATB resistance (and to some extent antifungal resistance) is a major public health concern, making the safety of ATB/antifungal treatments ethically problematic, if they do prove to be beneficial. Therefore, extensive and prolonged use of ATB/antifungal treatments may not be advisable as a long-term therapy for ASD.

#### **3.2 Probiotics**

One of the most nutritional popular approaches selectively modulating the GM is probiotic supplementation due to ease of use, wide availability, and good safety profile. Probiotics are defined as live microorganisms that when administered in adequate amounts confer a health benefit to the host [75]. The main species used are one *Escherichia coli* strain, several lactic acid-producing *Lactobacillus*, a number of bifidobacterial strains, and a yeast *Saccharomyces boulardii*. Research on probiotics has shown efficacy in prevention or treatment of a wide variety of diseases associated with GI difficulties. Recently, some evidence has been accumulated regarding the possible role of probiotics in modulating symptoms of certain psychological diseases such as depression and anxiety as well as ASD [76–78]. Hence, a new class of probiotics, termed as psychobiotics, has emerged and refers to living organisms with beneficial effects on mental health [79]. In spite of the well-documented beneficial effects of probiotics, testing for ASD is still in its infancy, and the exact mechanism of their action has not been thoroughly elucidated to date, though there are several hypotheses. Implicating correction of both composition and/or activities of GM is the first mechanism of action of probiotics through several mechanisms including bacteriocins and metabolites, such as lactic, propionic, and acetic acids.

Hence, because ASD patients present GI dysbiosis, which may exacerbate the disease, these patients could benefit from GM modulation through probiotic supplementation. Conversely, other investigations have shown that probiotic administration could act independently from GM alterations by inducing a stabilization of the

**177**

*Could Gut Modulation through Probiotic Supplementation Be Beneficial in Autism Spectrum…*

microbial communities making them less susceptible to perturbations from stressors such as ATBs, poor diet, and psychological stress [80]. Moreover, probiotics may correct the imbalance in the activity of the GM without changing its composition, via their metabolites released in the gut lumen or by potentially correcting the overproduction of harmful and underproduction of beneficial gut bacterial products [81]. The increased intestinal permeability in ASD may also be ameliorated by probiotics which are able to enhance the mucosal barrier. This supports the concept that probiotics can provide protective effect by preventing the metabolites of exogenous toxic substances from leaving the gut and entering the bloodstream to affect the brain [70]. Probiotics may therefore maintain or improve gut barrier integrity and aid in ASD rehabilitation by promoting "leaky gut" healing. Lastly, given the multiple findings of aberrant immune activation and higher levels of gut inflammation in a subset of individuals with ASD, that a major part of the immune system is concentrated in/ around the intestinal mucosa, and that the GM plays an important role in the maturation and the regulation of the immune response, another mechanism of action of probiotics may be on the immune system [82]. Probiotics can downregulate gut and CNS inflammatory pathways in a species- and strain-specific manner [83], by promoting the production of regulatory T cells, diminishing the levels of LPS, providing tolerogenic signals, and boosting the brain-derived neurotrophic factor (BDNF). BDNF is a protein that promotes the survival of existent neurons in the developing brain, fosters the growth and differentiation of new neurons, and regulates the formation and plasticity of synaptic connections, thereby playing an essential role in the normal neurological development [6]. Impairment in BDNF signaling in early developmental phases is thought to be related to CNS abnormalities, the most severe forms of ASD, as well as intellectual disability [84]. Probiotics have been shown also to modulate the intestinal immune system by the production of secreted factors and metabolites that affect growth and function of intestinal epithelial and immune cells [85]. Moreover, probiotic immunomodulation may occur through inhibiting the production of pro-inflammatory cytokines such as IL-12, TNF-α, and INF-α or increase the expression of anti-inflammatory mediators such as IL-10 and transforming growth factor beta (β-TFGF). Thus, by alleviating gut inflammation and suppressing dysregulated immune functions, probiotic supplementation may be effective for improving both gut microbial and behavioral problems in ASD. Lastly, probiotics could act via the vagus nerve-mediated GBA to influence neurotransmission and mood states [86]. They can exert central actions by influencing several neuroactive metabolites such as gamma-aminobutyric acid (GABA) and serotonin which are also associated with neuropsychiatric disorders [87, 88]. Consequently, administration of probiotics regulates the behavior in a way that significantly reduces anxiety, depression, and stress and promotes positive emotional changes. Therefore, probiotic administration might be a useful new therapeutic option to restore normal GM, reduce inflammation, restore epithelial barrier function, and possibly improve some

*DOI: http://dx.doi.org/10.5772/intechopen.89375*

behavioral symptoms associated with ASD.

A role for probiotics has been suggested for children with ASD as well, as preliminary findings from experimental animals studies provide some evidence that administration of selected probiotics may be effective in reducing neurologic signs and symptoms associated with gut dysbiosis. In a summary of studies to date, several probiotic strains, i.e., *Bifidobacterium* sp. (*B. longum***,** *B. breve*, *B. infantis*, and *B. bifidum*), *Lactobacillus* sp. (*L. acidophilus*, *L. helveticus*, *L. rhamnosus*, *L. plantarum*, *L. sporogenes*, *L. bulgaricus*, *L. delbrueckii*, *L. salivarius*, *L. casei*, and *L. paracasei*), *Streptococcus* sp. (*S. thermophilus* and *S. salivarius*), and *Bacteroides fragilis*, are presumed to be effective in ameliorating CNS functions related to mental disorders, as shown through several animal models [18, 89–92]. In a mouse model of ASD induced by maternal immune activation, oral administration of human commensal

### *Could Gut Modulation through Probiotic Supplementation Be Beneficial in Autism Spectrum… DOI: http://dx.doi.org/10.5772/intechopen.89375*

microbial communities making them less susceptible to perturbations from stressors such as ATBs, poor diet, and psychological stress [80]. Moreover, probiotics may correct the imbalance in the activity of the GM without changing its composition, via their metabolites released in the gut lumen or by potentially correcting the overproduction of harmful and underproduction of beneficial gut bacterial products [81]. The increased intestinal permeability in ASD may also be ameliorated by probiotics which are able to enhance the mucosal barrier. This supports the concept that probiotics can provide protective effect by preventing the metabolites of exogenous toxic substances from leaving the gut and entering the bloodstream to affect the brain [70]. Probiotics may therefore maintain or improve gut barrier integrity and aid in ASD rehabilitation by promoting "leaky gut" healing. Lastly, given the multiple findings of aberrant immune activation and higher levels of gut inflammation in a subset of individuals with ASD, that a major part of the immune system is concentrated in/ around the intestinal mucosa, and that the GM plays an important role in the maturation and the regulation of the immune response, another mechanism of action of probiotics may be on the immune system [82]. Probiotics can downregulate gut and CNS inflammatory pathways in a species- and strain-specific manner [83], by promoting the production of regulatory T cells, diminishing the levels of LPS, providing tolerogenic signals, and boosting the brain-derived neurotrophic factor (BDNF). BDNF is a protein that promotes the survival of existent neurons in the developing brain, fosters the growth and differentiation of new neurons, and regulates the formation and plasticity of synaptic connections, thereby playing an essential role in the normal neurological development [6]. Impairment in BDNF signaling in early developmental phases is thought to be related to CNS abnormalities, the most severe forms of ASD, as well as intellectual disability [84]. Probiotics have been shown also to modulate the intestinal immune system by the production of secreted factors and metabolites that affect growth and function of intestinal epithelial and immune cells [85]. Moreover, probiotic immunomodulation may occur through inhibiting the production of pro-inflammatory cytokines such as IL-12, TNF-α, and INF-α or increase the expression of anti-inflammatory mediators such as IL-10 and transforming growth factor beta (β-TFGF). Thus, by alleviating gut inflammation and suppressing dysregulated immune functions, probiotic supplementation may be effective for improving both gut microbial and behavioral problems in ASD. Lastly, probiotics could act via the vagus nerve-mediated GBA to influence neurotransmission and mood states [86]. They can exert central actions by influencing several neuroactive metabolites such as gamma-aminobutyric acid (GABA) and serotonin which are also associated with neuropsychiatric disorders [87, 88]. Consequently, administration of probiotics regulates the behavior in a way that significantly reduces anxiety, depression, and stress and promotes positive emotional changes. Therefore, probiotic administration might be a useful new therapeutic option to restore normal GM, reduce inflammation, restore epithelial barrier function, and possibly improve some behavioral symptoms associated with ASD.

A role for probiotics has been suggested for children with ASD as well, as preliminary findings from experimental animals studies provide some evidence that administration of selected probiotics may be effective in reducing neurologic signs and symptoms associated with gut dysbiosis. In a summary of studies to date, several probiotic strains, i.e., *Bifidobacterium* sp. (*B. longum***,** *B. breve*, *B. infantis*, and *B. bifidum*), *Lactobacillus* sp. (*L. acidophilus*, *L. helveticus*, *L. rhamnosus*, *L. plantarum*, *L. sporogenes*, *L. bulgaricus*, *L. delbrueckii*, *L. salivarius*, *L. casei*, and *L. paracasei*), *Streptococcus* sp. (*S. thermophilus* and *S. salivarius*), and *Bacteroides fragilis*, are presumed to be effective in ameliorating CNS functions related to mental disorders, as shown through several animal models [18, 89–92]. In a mouse model of ASD induced by maternal immune activation, oral administration of human commensal

*Prebiotics and Probiotics - Potential Benefits in Nutrition and Health*

germinate into vegetative forms once treatment has stopped [46].

**3.1 Antibiotics/antifungals**

for the treatments of ASD [74].

**3.2 Probiotics**

transplantation (FMT) has been shown to improve not only GI disturbances but also behavioral and neurophysiological abnormalities associated with ASD [18, 24].

ATB therapy is used for the management of ASD and is routinely prescribed to treat ASD symptoms associated with several underlying disorders like pediatric autoimmune neuropsychiatric disorder associated with streptococcal infections (PANDAS) and pediatric acute-onset neuropsychiatric syndrome (PANS) [72]. Short-term administration of vancomycin was shown to provide significant improvement in both GI and behavioral disturbances in a subset of children with ASD [73]. Unfortunately, this attempt had only partial success since children relapsed, and benefits waned upon treatment termination. As vancomycin is a poorly absorbed ATB known to destroy Gram-positive anaerobes, observed improvement is believed to be the consequence of temporary elimination of toxinproducing clostridia population. Hence, symptomatic relapse was attributed to the spore-forming capacity of these bacteria, and spores resistant to ATB would later

Children with ASD, particularly those with GI disease, are sometimes treated with antifungal agents, as they may have increased incidence of fungal infection. Despite the lack of evidence of fungal overgrowth in children with ASD, parents find that antifungal therapies can often be beneficial and may be a useful adjuvant

Thus, it is possible to speculate that anti-infectives, through modulation of GM,

One of the most nutritional popular approaches selectively modulating the GM is probiotic supplementation due to ease of use, wide availability, and good safety profile. Probiotics are defined as live microorganisms that when administered in adequate amounts confer a health benefit to the host [75]. The main species used are one *Escherichia coli* strain, several lactic acid-producing *Lactobacillus*, a number of bifidobacterial strains, and a yeast *Saccharomyces boulardii*. Research on probiotics has shown efficacy in prevention or treatment of a wide variety of diseases associated with GI difficulties. Recently, some evidence has been accumulated regarding the possible role of probiotics in modulating symptoms of certain psychological diseases such as depression and anxiety as well as ASD [76–78]. Hence, a new class of probiotics, termed as psychobiotics, has emerged and refers to living organisms with beneficial effects on mental health [79]. In spite of the well-documented beneficial effects of probiotics, testing for ASD is still in its infancy, and the exact mechanism of their action has not been thoroughly elucidated to date, though there are several hypotheses. Implicating correction of both composition and/or activities of GM is the first mechanism of action of probiotics through several mechanisms including bacteriocins and metabolites, such as lactic, propionic, and acetic acids. Hence, because ASD patients present GI dysbiosis, which may exacerbate the disease, these patients could benefit from GM modulation through probiotic supplementation. Conversely, other investigations have shown that probiotic administration could act independently from GM alterations by inducing a stabilization of the

should be able to influence symptoms and expression of psychiatric disorders. However, ATB resistance (and to some extent antifungal resistance) is a major public health concern, making the safety of ATB/antifungal treatments ethically problematic, if they do prove to be beneficial. Therefore, extensive and prolonged use of ATB/antifungal treatments may not be advisable as a long-term therapy for ASD.

**176**

*B. fragilis* corrected intestinal permeability, restored GM, improved GI physiology, and abolished ASD-like behavioral disruptions [18]. Interestingly, the probiotic also corrected altered expression of the tight junction proteins in the colon and restored the increase in the pro-inflammatory cytokine IL-6. Moreover, *B. fragilis* treatment mitigated elevations in several maternal immune activation-induced serum metabolites associated with ASD. One metabolite of particular interest was 4-ethylphenylsulfate (4EPS) which is chemically similar to p-cresol and is thought to be a possible putative urinary biomarker for ASD [59]. Hence, the administration of *B. fragilis* in a mouse model for ASD was able to reverse autistic symptoms and metabolic derangement. Similarly, reconstituting GM with a human breast milk and gut commensal *Lactobacillus reuteri* completely corrected social deficits and reversed aberrant neurotransmission in maternal high-fat diet offspring [89]. This is intriguing in light of reports that risperidone, an FDA-approved treatment for ASD, does not correct social abnormalities [93]. Treatment with *L. reuteri* resulted also in reduction of stress-induced corticosterone, and stimulation of the production of oxytocin, a key regulator of social behaviors, involved in the mesolimbic dopamine reward system, which is thought to be dysregulated in ASD [94]. On another hand, a recent study performed on hamsters in which autistic-like behaviors were induced by propionic acid and clindamycin administration studied the therapeutic effect of a 3-week oral treatment with a mixture of bifidobacteria and lactobacilli strains (Protexin®) [91]. Protexin® administration was effective in rebalancing GM, ameliorating oxidative stress, and counteracting behavioral deficits. Finally, in maternal separation-induced early-life stress mice model, oral administration of *L. plantarum* as a psychobiotic strain significantly reduced anxiety while improving locomotor activities and exploratory behavior [92]. According to various animal studies, *L. plantarum* can modulate the levels of neurotransmitters in the brain by influencing gene expression in the CNS and increasing dopamine level in the prefrontal cortex [92]. Summing up, by conducting behavioral tests in animal models, research provided convincing evidence for the efficacy of psychobiotic strains in improving ASD-like behaviors. However, the experimental evidence for the positive behavioral changes observed in animal models after administration of probiotics raises the interesting question whether the same result also applies to humans.

To date only few studies explored the effects of probiotics on ASD clinical features in humans. **Table 1** synthesizes available evidence on the efficacy and safety of probiotic supplementation as an adjunctive treatment for ASD [49, 51, 73, 95–106]. In these studies, probiotic interventions varied across all of the trials. Concentrations of the probiotic strains administered ranged from 107 to 1010 colony-forming units (CFU)/dose, and their usage by recipients differed. Strains were administered alone or as mixed strains with or without other additives (immunomodulators [99] and ATBs [73]). In summary, despite the variability in species, strains, dosages, and duration utilized, all studies pointed toward a similar trend of improvement in both caregiver-reported ASD and GI symptoms after probiotic therapies. Accordingly, a recent survey found that almost 20% of physicians treating ASD patients encourage probiotic use, and almost 60% accept their use [107]. In a recent study, the positive effect of probiotic treatment was represented not only by the ability to consistently normalize the *Bacteroidetes*/*Firmicutes* ratio and restore the amounts of *Desulfovibrio* and *Bifidobacterium* in children with ASD [51] but also by the tendency to reduce intestinal inflammation and permeability. Similarly, another study associated changes in the GM composition to improvements in GI symptoms and functioning [102]. After probiotic supplementation in the latter study, children with ASD experienced a significant increase in *Bifidobacterium* and *Lactobacillus* in their fecal stool samples with a simultaneous reduction of *Clostridium* species. Furthermore, in a third study, probiotic feeding of children

**179**

*Could Gut Modulation through Probiotic Supplementation Be Beneficial in Autism Spectrum…*

Not reported Strain and dosage not provided

CARS *L. acidophilus*, *L. bulgaricus*, and *B. bifidum*, 40 × 109

QD for 4 wk

4.5 × 1010 CFU QD for 3 wk

ATEC *L. rhamnosus*, *B. bifidum*, *L. acidophilus*,

and *L. sporogenes* Dosage not provided BID for 21 d

ATEC Strain and dosage not provided QD

DSM-IV *L. acidophilus* (strain Rosell-11) 5 × 109

ICD-10 *L. rhamnosus* GG

 CFU/g BID for 2 mo

Not reported *L. acidophilus*, *L. casei*, *L. delbrueckii*, *B. longum*, and *B. bifidum* 1 × 1010 CFU/capsule One capsule TID for 6 mo

> 1 × 1010 CFU QD for 6 mo

ICD-10 3 strains of *Lactobacillus* (60%, one is

DSM-V + ADOS-2 *B. breve*, *B. longum*, *B. infantis*, *L.*

*L. casei*), 2 strains of *Bifidobacterium* (25%, one is *B. longum*), and 1 strain of *Streptococcus* (15%, exact strain information not provided) Dosage not provided One capsule TID for 4 mo

*acidophilus*, *L. plantarum*, *L. paracasei*, *L. bulgaricus*, *L. delbrueckii*, *S. thermophilus*, and *S. salivarius* 9 × 1010 CFU/g *Bifidobacteria* 8 × 1010 CFU/g *Lactobacilli*, and 20 × 1010 CFU/g *Streptococci* QD for 4 months

Not reported *L. plantarum* WCFS1

Administration for 2 mo

CFU/mL

*B. infantis*, *B. longum*, *S. thermophilus*, *L. plantarum*, *L. salivarius*, *L. reuteri*, *L. casei*, *L. bulgaricus*, *L. acidophilus* DDS-1,

**Authors Study design Diagnosis Probiotic therapy**

*DOI: http://dx.doi.org/10.5772/intechopen.89375*

ASD

Case report of a 6 y with

Open-label with selfcontrol study 11 ASD subjects (10 M, 3.5–7 y)

Randomized, doubleblind, PBO-controlled, cross overdesigned feeding study 17 ASD subjects (14 M, 4–16 y)

Open-label with selfcontrol study 10 ASD subjects (Gender not reported,

Retrospective case cohort study 57 ASD subjects: 19 PS vs. 38 no PS (50 M, 3–9 y)

Open-label with selfcontrol study 22 ASD subjects (20 M, 4–10 y)

Open-label with selfcontrol study 33 ASD subjects (Gender not reported,

Randomized, doubleblind, PBO-controlled prospective follow-up study 75 ASD subjects 40 PS (24 M, 13 y) vs.

Case control cohort

9 non-ASD siblings (7 M, 5–17 y) 10 non-ASD controls (10 M; 2–11 y)

10 ASD subjects (9 M,

Case report of a 12 y boy

3–16 y)

35 PBO (16 M, 13 y)

study

2–9 y)

with ASD

4–15 y)

Blades M., 2000, UK [95]

Sandler et al., 2000, USA [73]

Parracho et al., 2010, UK [96]

Ray et al., 2010, USA [97]

Adams et al., 2011 USA [49]

Kałzuna-Czaplinska et al., 2012, Poland [98]

West et al., 2013, USA [99]

Partty et al., 2015, Finland [100]

Tomova et al., 2015, Slovakia [51]

Grossi et al., 2016, Italy [101]

*Could Gut Modulation through Probiotic Supplementation Be Beneficial in Autism Spectrum… DOI: http://dx.doi.org/10.5772/intechopen.89375*

*Prebiotics and Probiotics - Potential Benefits in Nutrition and Health*

interesting question whether the same result also applies to humans.

To date only few studies explored the effects of probiotics on ASD clinical features in humans. **Table 1** synthesizes available evidence on the efficacy and safety of probiotic supplementation as an adjunctive treatment for ASD [49, 51, 73, 95–106]. In these studies, probiotic interventions varied across all of the trials. Concentrations of the probiotic strains administered ranged from 107

colony-forming units (CFU)/dose, and their usage by recipients differed. Strains were administered alone or as mixed strains with or without other additives (immunomodulators [99] and ATBs [73]). In summary, despite the variability in species, strains, dosages, and duration utilized, all studies pointed toward a similar trend of improvement in both caregiver-reported ASD and GI symptoms after probiotic therapies. Accordingly, a recent survey found that almost 20% of physicians treating ASD patients encourage probiotic use, and almost 60% accept their use [107]. In a recent study, the positive effect of probiotic treatment was represented not only by the ability to consistently normalize the *Bacteroidetes*/*Firmicutes* ratio and restore the amounts of *Desulfovibrio* and *Bifidobacterium* in children with ASD [51] but also by the tendency to reduce intestinal inflammation and permeability. Similarly, another study associated changes in the GM composition to improvements in GI symptoms and functioning [102]. After probiotic supplementation in the latter study, children with ASD experienced a significant increase in *Bifidobacterium* and *Lactobacillus* in their fecal stool samples with a simultaneous reduction of *Clostridium* species. Furthermore, in a third study, probiotic feeding of children

to 1010

*B. fragilis* corrected intestinal permeability, restored GM, improved GI physiology, and abolished ASD-like behavioral disruptions [18]. Interestingly, the probiotic also corrected altered expression of the tight junction proteins in the colon and restored the increase in the pro-inflammatory cytokine IL-6. Moreover, *B. fragilis* treatment mitigated elevations in several maternal immune activation-induced serum metabolites associated with ASD. One metabolite of particular interest was 4-ethylphenylsulfate (4EPS) which is chemically similar to p-cresol and is thought to be a possible putative urinary biomarker for ASD [59]. Hence, the administration of *B. fragilis* in a mouse model for ASD was able to reverse autistic symptoms and metabolic derangement. Similarly, reconstituting GM with a human breast milk and gut commensal *Lactobacillus reuteri* completely corrected social deficits and reversed aberrant neurotransmission in maternal high-fat diet offspring [89]. This is intriguing in light of reports that risperidone, an FDA-approved treatment for ASD, does not correct social abnormalities [93]. Treatment with *L. reuteri* resulted also in reduction of stress-induced corticosterone, and stimulation of the production of oxytocin, a key regulator of social behaviors, involved in the mesolimbic dopamine reward system, which is thought to be dysregulated in ASD [94]. On another hand, a recent study performed on hamsters in which autistic-like behaviors were induced by propionic acid and clindamycin administration studied the therapeutic effect of a 3-week oral treatment with a mixture of bifidobacteria and lactobacilli strains (Protexin®) [91]. Protexin® administration was effective in rebalancing GM, ameliorating oxidative stress, and counteracting behavioral deficits. Finally, in maternal separation-induced early-life stress mice model, oral administration of *L. plantarum* as a psychobiotic strain significantly reduced anxiety while improving locomotor activities and exploratory behavior [92]. According to various animal studies, *L. plantarum* can modulate the levels of neurotransmitters in the brain by influencing gene expression in the CNS and increasing dopamine level in the prefrontal cortex [92]. Summing up, by conducting behavioral tests in animal models, research provided convincing evidence for the efficacy of psychobiotic strains in improving ASD-like behaviors. However, the experimental evidence for the positive behavioral changes observed in animal models after administration of probiotics raises the

**178**

*ADOS, Autism Diagnostic Observation Schedule; ATEC, Autism Treatment Evaluation Checklist; CARS, Childhood Autism Rating Scale; DSM-IV, Diagnostic and Statistical Manual-4th Edition; ICD-10, International Classification of Diseases-10th Edition; d, days; wk, weeks; mo, months; CFU, colony-forming unit; g, gram; QD, once a day; BID, two times a day; TID, three times a day; QID, four times a day.*

#### **Table 1.**

*Main clinical trials investigating the effectiveness of probiotic supplementation in autism spectrum disorders.*

with ASD significantly increased levels of the beneficial bacteria, specifically, the amount of lactobacilli and enterococci, and reduced their fecal *Clostridium* counts [96]. Additionally, in the same study, the efficacy of probiotic supplementation seemed to be age-dependent, with better effects noticed on younger children, underscoring the importance of early interventions. Although all aforementioned studies showed changes in GM after the implementation diet with probiotics, only some of them analyzed the correlations with GI symptoms and reported improvement of GI function indices [96, 98, 101, 102, 105]. In fact, a survey conducted on caregivers showed that daily administration of a specific five-strain probiotic decreased the severity of comorbid GI problems [99]. Concurrent with increases in the proportion of normal stools, there was an increased appetite and willingness to consume novel foods that may be due to reduced abdominal pain or improved stool evacuation [105]. Besides, in one case study report, a multi-strain mixture of 10 probiotics not only relieved the GI disturbances but also improved ASD core symptoms [101]. However, benefits reversed after the term of the therapy. In addition to the alteration in GI dysfunction and GM, more than half of the studies also included assessment of change in behavior after probiotic therapy. Most of these studies recorded a reduction in the severity of ASD symptoms, although not all reached significance. Noted positive effects on mood and general behavior were (i) statistically significant differences in behavioral scores for disruptive/ antisocial behavior, anxiety problems, communication disturbances, and selfabsorbed behavior compared to baseline [96, 104]; (ii) decrease in the severity of

**181**

**3.3 Prebiotics**

*Could Gut Modulation through Probiotic Supplementation Be Beneficial in Autism Spectrum…*

speech/language/communication performance [97, 101]; (iii) progress in sociability, sensory and cognitive awareness, physical health, and behavior [99, 102]; and (iv) changes in adaptive, repetitive, and aberrant behaviors including irritability, lethargy, stereotypy, and hyperactivity [105]. Conversely, in other investigations, no statistically significant differences in behavioral scores were detected between probiotics and placebo control groups [73, 103]. Lastly, probiotic treatment reversed the metabolic disruptions and improved behavioral performance. In one of the studies, probiotics significantly lowered the concentration of myeloperoxidase (a marker of inflammation and oxidation) in ASD patients compared to other autistic individuals not taking probiotics [106]. Metabolic changes were also observed in another former study in which probiotics substantially diminished fecal levels of propionic acid in ASD individuals [49]. In a third study, probiotic supplementation significantly reduced metabolic products of the pathogenic *Candida* yeast species [98], which are remarkably elevated in the urines of individuals with ASD [108]. This biological improvement also corresponded to behavioral outcomes, where children showed significant ameliorations in their ability to concentrate and fulfill orders. In 2015, an intriguing randomized trial demonstrated that early postnatal probiotic administration has a preventive effect on ASD and may reduce the risk of developing further neuropsychiatric disorder [100]. Results showed that from children randomly assigned to probiotic or placebo groups during the first 6 months of life, 17% in the placebo group had a diagnosis of ASD at the age of 13, while none

At last, though probiotics are considered a relatively risk-free option for individuals with ASD, the current literature cannot confidently state their safety as there is a paucity of systematic reporting of adverse events. However, among the studies that monitored side effects, the reported ones (bloating, skin rash, worsening constipation or diarrhea, and weight loss) appear to be mild, transient, and infrequent [96, 99, 102, 105]. Conclusively, due to the large heterogeneity between trials, studies provide only suggestive but not conclusive evidence regarding the efficacy of probiotics on GI and behavioral symptoms among ASD patients. Thus, future probiotic research holds hopes for discovering the optimal species, strains, strength, and length of probiotic therapy for the particular comorbidity profile of different individuals with ASD.

The International Scientific Association for Probiotics and Prebiotics (ISAPP) defines prebiotics as substrates "selectively utilized by host microorganisms, conferring a health benefit" [75]. Such benefits are not limited to gut homeostasis but can extend elsewhere in the organism, leading to improvements of the immune, metabolic, endocrine, or nervous functions. Fructans, comprising fructooligosaccharides (FOS) and inulin, and galactans (galactooligosaccharides (GOS)) are the most recognized prebiotics. Differently from most dietary fibers, which promote growth of a wide variety of microorganisms, prebiotics display a selective effect, being a substrate for beneficial strains only, while excluding metabolism by pathogenic bacteria [6]. Thus, the main reason for a potential influence of prebiotics on the treatment of ASD concerns the selective enrichment of *Lactobacillus* and/or *Bifidobacterium* spp. Besides, various other mechanisms have been identified through which prebiotics can act, including generation of SCFA that have an influence on gut energy metabolism, barrier function, water fluxes, motility [109], elongation of microvilli, increase in mucus layer thickness, and consequent protection of gut epithelium. However, as with probiotics, prebiotic studies regarding their impact in neurological problems are few and not conclusive. In rats, oral administration of GOS elevates BDNF levels [110], normalizes LPS-induced anxiety, and modulates cortical IL-1β levels [111],

*DOI: http://dx.doi.org/10.5772/intechopen.89375*

of the children in the probiotic group did.

*Could Gut Modulation through Probiotic Supplementation Be Beneficial in Autism Spectrum… DOI: http://dx.doi.org/10.5772/intechopen.89375*

speech/language/communication performance [97, 101]; (iii) progress in sociability, sensory and cognitive awareness, physical health, and behavior [99, 102]; and (iv) changes in adaptive, repetitive, and aberrant behaviors including irritability, lethargy, stereotypy, and hyperactivity [105]. Conversely, in other investigations, no statistically significant differences in behavioral scores were detected between probiotics and placebo control groups [73, 103]. Lastly, probiotic treatment reversed the metabolic disruptions and improved behavioral performance. In one of the studies, probiotics significantly lowered the concentration of myeloperoxidase (a marker of inflammation and oxidation) in ASD patients compared to other autistic individuals not taking probiotics [106]. Metabolic changes were also observed in another former study in which probiotics substantially diminished fecal levels of propionic acid in ASD individuals [49]. In a third study, probiotic supplementation significantly reduced metabolic products of the pathogenic *Candida* yeast species [98], which are remarkably elevated in the urines of individuals with ASD [108]. This biological improvement also corresponded to behavioral outcomes, where children showed significant ameliorations in their ability to concentrate and fulfill orders. In 2015, an intriguing randomized trial demonstrated that early postnatal probiotic administration has a preventive effect on ASD and may reduce the risk of developing further neuropsychiatric disorder [100]. Results showed that from children randomly assigned to probiotic or placebo groups during the first 6 months of life, 17% in the placebo group had a diagnosis of ASD at the age of 13, while none of the children in the probiotic group did.

At last, though probiotics are considered a relatively risk-free option for individuals with ASD, the current literature cannot confidently state their safety as there is a paucity of systematic reporting of adverse events. However, among the studies that monitored side effects, the reported ones (bloating, skin rash, worsening constipation or diarrhea, and weight loss) appear to be mild, transient, and infrequent [96, 99, 102, 105]. Conclusively, due to the large heterogeneity between trials, studies provide only suggestive but not conclusive evidence regarding the efficacy of probiotics on GI and behavioral symptoms among ASD patients. Thus, future probiotic research holds hopes for discovering the optimal species, strains, strength, and length of probiotic therapy for the particular comorbidity profile of different individuals with ASD.

#### **3.3 Prebiotics**

*Prebiotics and Probiotics - Potential Benefits in Nutrition and Health*

Prospective, open-label

30 ASD subjects (19 M,

30 age/sex-matched non-ASD siblings

Case report of a 16 y child with ASD

Double-blind, crossover, randomized clinical trial 8 ASD subjects (7 M, 4–11 y)

*two times a day; TID, three times a day; QID, four times a day.*

A double-blind, randomized, PBOcontrolled, parallel feeding study 71 ASD subjects: 36 PS (36 M, 7–15 y) vs.

cohort study

5–9 y)

35 PBO (35 M, 7–15 y)

Shaaban et al., 2017, Egypt [102]

Liu et al., 2019, Taiwan [103]

Kobliner et al., 2019, USA [104]

Sanctuary et al., 2019, USA [105]

**Table 1.**

**Authors Study design Diagnosis Probiotic therapy**

DSM-V + ADOS +

*L. acidophilus*, *L. rhamnosus*, and *B.*

 CFU/g 5 g QD for 3 mo

3 × 1010 CFU/capsule 1 capsule QD for 4 wk

 CFU/capsule Week 1: 6 capsules QD. Weekly increases reaching a final dose of 12 capsules BID. After 3 mo, weaning

down to 3 capsules BID

*longum* 10 × 107

3 × 109

ADOS *B. longum* subsp. *infantis*

2 × 1010 CFU QD for 5 wk

DSM-V + ADI-R *L. plantarum* PS128

Not reported *S. boulardii*

ADI-R

with ASD significantly increased levels of the beneficial bacteria, specifically, the amount of lactobacilli and enterococci, and reduced their fecal *Clostridium* counts [96]. Additionally, in the same study, the efficacy of probiotic supplementation seemed to be age-dependent, with better effects noticed on younger children, underscoring the importance of early interventions. Although all aforementioned studies showed changes in GM after the implementation diet with probiotics, only some of them analyzed the correlations with GI symptoms and reported improvement of GI function indices [96, 98, 101, 102, 105]. In fact, a survey conducted on caregivers showed that daily administration of a specific five-strain probiotic decreased the severity of comorbid GI problems [99]. Concurrent with increases in the proportion of normal stools, there was an increased appetite and willingness to consume novel foods that may be due to reduced abdominal pain or improved stool evacuation [105]. Besides, in one case study report, a multi-strain mixture of 10 probiotics not only relieved the GI disturbances but also improved ASD core symptoms [101]. However, benefits reversed after the term of the therapy. In addition to the alteration in GI dysfunction and GM, more than half of the studies also included assessment of change in behavior after probiotic therapy. Most of these studies recorded a reduction in the severity of ASD symptoms, although not all reached significance. Noted positive effects on mood and general behavior were (i) statistically significant differences in behavioral scores for disruptive/ antisocial behavior, anxiety problems, communication disturbances, and selfabsorbed behavior compared to baseline [96, 104]; (ii) decrease in the severity of

*Main clinical trials investigating the effectiveness of probiotic supplementation in autism spectrum disorders.*

*y, years; M, male; PBO, placebo; PS, probiotic supplementation; ADI-R, Autism Diagnostic Interview-Revised; ADOS, Autism Diagnostic Observation Schedule; ATEC, Autism Treatment Evaluation Checklist; CARS, Childhood Autism Rating Scale; DSM-IV, Diagnostic and Statistical Manual-4th Edition; ICD-10, International Classification of Diseases-10th Edition; d, days; wk, weeks; mo, months; CFU, colony-forming unit; g, gram; QD, once a day; BID,*

**180**

The International Scientific Association for Probiotics and Prebiotics (ISAPP) defines prebiotics as substrates "selectively utilized by host microorganisms, conferring a health benefit" [75]. Such benefits are not limited to gut homeostasis but can extend elsewhere in the organism, leading to improvements of the immune, metabolic, endocrine, or nervous functions. Fructans, comprising fructooligosaccharides (FOS) and inulin, and galactans (galactooligosaccharides (GOS)) are the most recognized prebiotics. Differently from most dietary fibers, which promote growth of a wide variety of microorganisms, prebiotics display a selective effect, being a substrate for beneficial strains only, while excluding metabolism by pathogenic bacteria [6]. Thus, the main reason for a potential influence of prebiotics on the treatment of ASD concerns the selective enrichment of *Lactobacillus* and/or *Bifidobacterium* spp. Besides, various other mechanisms have been identified through which prebiotics can act, including generation of SCFA that have an influence on gut energy metabolism, barrier function, water fluxes, motility [109], elongation of microvilli, increase in mucus layer thickness, and consequent protection of gut epithelium. However, as with probiotics, prebiotic studies regarding their impact in neurological problems are few and not conclusive. In rats, oral administration of GOS elevates BDNF levels [110], normalizes LPS-induced anxiety, and modulates cortical IL-1β levels [111],

thus confirming the potential role for prebiotics in ASD where anxiety and neuroinflammation are prominent clinical features. Lastly, only two clinical trials examined the use of prebiotics in children with ASD [105, 112]. In the first study, GOS alone did not have a significant effect on GI symptoms, while when combined with bovine colostrum, GI symptoms improved as well as irritability scores and stereotypy [105]. Similarly, in the second trial, GOS intervention did not show a significant impact when provided alone. Nevertheless, when associating GOS treatment with an exclusion diet, notable increases occurred in beneficial bacteria supporting improvement in antisocial behavior [112]. GOS also affected SCFA production decreasing propionic acid as a result of normalized GM composition. Thus, it is sensible to postulate that combined intervention therapies might have a better impact on ASD individuals than single dietary approach and prebiotics could be a useful option for ASD children treatment in early life.


**9. Importance of project teams**

Projects are executed in teams. In general, projects are managed using teams in a work environment that is complex for two reasons: first, each project is unique, and second, conditions for team selection and motivation are often far from ideal [3]. In addition to uniqueness and complexity, unfamiliarity is often described as one of the characteristics of projects and as a result, projects are often associated with change. Consequently, successful project performance requires strong leadership, which provides vision and ability to cope with change.

Projects, by definition, are unique and are often associated with uncertainties and unknowns. It is reasonable to assume that in project management, it is not if the plans will change, it is when, what will change, and by how much. If anything is constant in a project, it is the change. When changes are significant in a project, which is often the case, leadership role assumes greater importance. Leadership has its efforts directed towards convincing people about the need to change, aligning them to a new direction, and motivating people to work together to achieve project objectives under difficult and demanding work environments [12].

Motivating the project team involves getting the team members to do the tasks that need to be done, not because they have to do them but because they understand the value of their work to the overall success of the project and they want to do them. If people want to do what we need to have them do, their commitment is bound to be greater, and the job is likely to be accomplished much better than if they were being forced to do the work.

Project managers must understand the personal aspirations of project team members and support them. The project manager's leadership skills play an important role in motivating and guiding the project team to grow as professionals while accomplishing project goals at the same time. In essence, project managers should ensure that both personal and project goals are accomplished and that the conflict between these two goals is minimized [7].

The project manager's job is to get others to do what the project manager needs them to do, not because they have to but because they want to. Empowerment means providing freedom to people (not control) so that they can successfully do what they want to do. This is very different from making them do what you want them to do. Empowerment makes sense on projects. Projects typically employ a multidisciplinary approach, requiring subject matter experts from different

functional areas. Each person brings unique expertise, and experience to the project team. By letting project team members make decisions and set goals pertaining to their roles and functions, the project manager can empower the team.


**5.5. Category or concept structure**

People are sensitive to the value of both labeled and unlabeled stimuli, and this depends on the structure of the concept being learned [24]. Markant and Gureckis [31] showed that the effectiveness of AML might interact with the particular structure of the target categories. Two types of category structures were used in that study: rule-based (RB), in which the decision rule is defined as a criterion along a single dimension, and information integration, in which the decision rule is a function of at least two dimensions.