Anti-pathogenic properties of non-digestible oligosaccharides: The fight against bacterial pathogens and toxins

Despite effective prevention and control efforts over the past few decades, infectious diseases continue to pose a serious threat to global public health, causing millions of deaths annually. Numerous antibiotics can treat bacterial infections. However, practically almost all antimicrobial drugs now have some level of resistance, creating a severe global health issue and the urgent and rapid development of antibiotic alternatives. It has been demonstrated that NDOs are significant inhibitors of the development of pathogenic infections. Several anti-pathogenic effects can be achieved by NDOs, depending on their type and structural characteristics. This thesis suggests that Non-digestible oligosaccharides (NDOs) may serve as an alternative to antibiotics in treating bacterial infections. In our studies we investigated the effects of NDOs (AOS, COS, FOS and GOS) on the defense against several common bacterial pathogens (E. coli, S. agalactia, S. aureus, and C. difficile) in different steps of the pathogenesis: bacterial growth, bacterial adhesion, biofilm formation, inflammation and bacterial toxin activity. Each of these NDOs had some unique potentials against particular pathogens. Likewise, COS can be described as a potent biofilm inhibitor against S. aureus, a strong inhibitor of shiga toxin binding to its receptor (Gb3), a great inhibitor of cytotoxicity and cytopathogenicity of TcdA, an effective anti-inflammatory agent against inflammation caused by E. coli with intestinal barrier-protective properties and an enhancer of human intestinal epithelial cell integrity. Besides, AOS can be defined as a powerful inhibitor of the growth of both GBS and E. coli, an effective inhibitor of the adhesion of E. coli to intestinal epithelial cells, a potent anti-inflammatory agent against inflammation induced by E. coli, and an effective mediator on TcdA's ability to disrupt the intestinal barrier. The knowledge about the antimicrobial strategies of NDOs was used to further investigate in combination therapy by combining NDO with antibiotics to increase bacterial sensitivity to specific antibiotics. Likewise, a combination of COS and clindamycin was successful in preventing the growth of a S. aureus biofilm, as was a combination of AOS and ampicillin in the case of E. coli growth. AOS and trimethoprim were also successful in treating GBS when combined. NDOs are a desirable weapon in the fight against pathogens and antibiotic resistance due to their variety of antibacterial capabilities and minimal reported adverse effects. Moreover, a healthy microbiota plays a vital role in infection prevention by inhibiting pathogenic bacteria and/or coordinating proper immune responses, therefore the effect of NDOs on promoting beneficial bacteria in the gut should not be ignored

Direct Action of Non-Digestible Oligosaccharides against a Leaky Gut

The epithelial monolayer is the primary determinant of mucosal barrier function, and tight junction (TJ) complexes seal the paracellular space between the adjacent epithelial cells and represent the main "gate-keepers" of the paracellular route. Impaired TJ functionality results in increased permeation of the "pro-inflammatory" luminal contents to the circulation that induces local and systemic inflammatory and immune responses, ultimately triggering and/or perpetuating (chronic) systemic inflammatory disorders. Increased gut leakiness is associated with intestinal and systemic disease states such as inflammatory bowel disease and neurodegenerative diseases such as Parkinson's disease. Modulation of TJ dynamics is an appealing strategy aiming at inflammatory conditions associated with compromised intestinal epithelial function. Recently there has been a growing interest in nutraceuticals, particularly in non-digestible oligosaccharides (NDOs). NDOs confer innumerable health benefits via microbiome-shaping and gut microbiota-related immune responses, including enhancement of epithelial barrier integrity. Emerging evidence supports that NDOs also exert health-beneficial effects on microbiota independently via direct interactions with intestinal epithelial and immune cells. Among these valuable features, NDOs promote barrier function by directly regulating TJs via AMPK-, PKC-, MAPK-, and TLR-associated pathways. This review provides a comprehensive overview of the epithelial barrier-protective effects of different NDOs with a special focus on their microbiota-independent modulation of TJs

Anti-pathogenic properties of non-digestible oligosaccharides: The fight against bacterial pathogens and toxins

Despite effective prevention and control efforts over the past few decades, infectious diseases continue to pose a serious threat to global public health, causing millions of deaths annually. Numerous antibiotics can treat bacterial infections. However, practically almost all antimicrobial drugs now have some level of resistance, creating a severe global health issue and the urgent and rapid development of antibiotic alternatives. It has been demonstrated that NDOs are significant inhibitors of the development of pathogenic infections. Several anti-pathogenic effects can be achieved by NDOs, depending on their type and structural characteristics. This thesis suggests that Non-digestible oligosaccharides (NDOs) may serve as an alternative to antibiotics in treating bacterial infections. In our studies we investigated the effects of NDOs (AOS, COS, FOS and GOS) on the defense against several common bacterial pathogens (E. coli, S. agalactia, S. aureus, and C. difficile) in different steps of the pathogenesis: bacterial growth, bacterial adhesion, biofilm formation, inflammation and bacterial toxin activity. Each of these NDOs had some unique potentials against particular pathogens. Likewise, COS can be described as a potent biofilm inhibitor against S. aureus, a strong inhibitor of shiga toxin binding to its receptor (Gb3), a great inhibitor of cytotoxicity and cytopathogenicity of TcdA, an effective anti-inflammatory agent against inflammation caused by E. coli with intestinal barrier-protective properties and an enhancer of human intestinal epithelial cell integrity. Besides, AOS can be defined as a powerful inhibitor of the growth of both GBS and E. coli, an effective inhibitor of the adhesion of E. coli to intestinal epithelial cells, a potent anti-inflammatory agent against inflammation induced by E. coli, and an effective mediator on TcdA's ability to disrupt the intestinal barrier. The knowledge about the antimicrobial strategies of NDOs was used to further investigate in combination therapy by combining NDO with antibiotics to increase bacterial sensitivity to specific antibiotics. Likewise, a combination of COS and clindamycin was successful in preventing the growth of a S. aureus biofilm, as was a combination of AOS and ampicillin in the case of E. coli growth. AOS and trimethoprim were also successful in treating GBS when combined. NDOs are a desirable weapon in the fight against pathogens and antibiotic resistance due to their variety of antibacterial capabilities and minimal reported adverse effects. Moreover, a healthy microbiota plays a vital role in infection prevention by inhibiting pathogenic bacteria and/or coordinating proper immune responses, therefore the effect of NDOs on promoting beneficial bacteria in the gut should not be ignored

Differential effects of oligosaccharides on the effectiveness of ampicillin against Escherichia coli in vitro

Background: The mounting antibiotic resistance emphasizes an urgent need for alternatives. Recent investigations indicate that non-digestible oligosaccharides (NDOs), besides their prebiotic properties, can directly interact with pathogenic bacteria. In this study, the protective effect of alginate-oligosaccharides (AOS), chitosanoligosaccharides (COS), galacto-oligosaccharides (GOS) and fructo-oligosaccharides, against enteropathogenicEscherichia. coli was investigated. Methods: The effect of these NDOs onE. coli growth, adhesion and E. coli-induced inflammatory response (IL-8 release) of HT-29 intestinal epithelial cells were determined in vitro in the presence or absence of ampicillin, using minimum inhibitory concentration (MIC) assay, anti-adhesion assay and ELISA, respectively. Results: At low concentrations 0.5 % and 1%, AOS decreased theE. coli growth, while high GOS concentrations (6%, 8%, 10 %) were effective. Interestingly, the combination of the low concentrations of AOS with ampicillin (2 μg/mL) exerted a 2-fold decrease in the MIC level of ampicillin against E. coli. AOS also concentration dependently reduced the adherence of E. coli to HT-29 cells. The combination of AOS with ampicillin further increased these anti-adhesive properties. Pre-incubation of HT-29 cells with AOS, COS or GOS significantly hampered the E. coli-induced IL-8 release. Conclusion: Current study highlights the direct effects of NDOs onE. coli growth, adhesion and inflammatory responses of HT-29 cells in vitro

Direct Action of Non-Digestible Oligosaccharides against a Leaky Gut

The epithelial monolayer is the primary determinant of mucosal barrier function, and tight junction (TJ) complexes seal the paracellular space between the adjacent epithelial cells and represent the main “gate-keepers” of the paracellular route. Impaired TJ functionality results in increased permeation of the “pro-inflammatory” luminal contents to the circulation that induces local and systemic inflammatory and immune responses, ultimately triggering and/or perpetuating (chronic) systemic inflammatory disorders. Increased gut leakiness is associated with intestinal and systemic disease states such as inflammatory bowel disease and neurodegenerative diseases such as Parkinson’s disease. Modulation of TJ dynamics is an appealing strategy aiming at inflammatory conditions associated with compromised intestinal epithelial function. Recently there has been a growing interest in nutraceuticals, particularly in non-digestible oligosaccharides (NDOs). NDOs confer innumerable health benefits via microbiome-shaping and gut microbiota-related immune responses, including enhancement of epithelial barrier integrity. Emerging evidence supports that NDOs also exert health-beneficial effects on microbiota independently via direct interactions with intestinal epithelial and immune cells. Among these valuable features, NDOs promote barrier function by directly regulating TJs via AMPK-, PKC-, MAPK-, and TLR-associated pathways. This review provides a comprehensive overview of the epithelial barrier-protective effects of different NDOs with a special focus on their microbiota-independent modulation of TJs

Anti-pathogenic functions of non-digestible oligosaccharides in vitro

Non-digestible oligosaccharides (NDOs), complex carbohydrates that resist hydrolysis by salivary and intestinal digestive enzymes, fulfill a diversity of important biological roles. A lot of NDOs are known for their prebiotic properties by stimulating beneficial bacteria in the intestinal microbiota. Human milk oligosaccharides (HMOs) represent the first prebiotics that humans encounter in life. Inspired by these HMO structures, chemically-produced NDO structures (e.g., galacto-oligosaccharides and chito-oligosaccharides) have been recognized as valuable food additives and exert promising health effects. Besides their apparent ability to stimulate beneficial microbial species, oligosaccharides have shown to be important inhibitors of the development of pathogenic infections. Depending on the type and structural characteristics, oligosaccharides can exert a number of anti-pathogenic effects. The most described effect is their ability to act as a decoy receptor, thereby inhibiting adhesion of pathogens. Other ways of pathogenic inhibition, such as interference with pathogenic cell membrane and biofilm integrity and DNA transcription, are less investigated, but could be equally impactful. In this review, a comprehensive overview of In vitro anti-pathogenic properties of different NDOs and associated pathways are discussed. A framework is created categorizing all anti-pathogenic effects and providing insight into structural necessities for an oligosaccharide to exert one of these effects

Non-Digestible Oligosaccharides and Short Chain Fatty Acids as Therapeutic Targets against Enterotoxin-Producing Bacteria and Their Toxins

Enterotoxin-producing bacteria (EPB) have developed multiple mechanisms to disrupt gut homeostasis, and provoke various pathologies. A major part of bacterial cytotoxicity is attributed to the secretion of virulence factors, including enterotoxins. Depending on their structure and mode of action, enterotoxins intrude the intestinal epithelium causing long-term consequences such as hemorrhagic colitis. Multiple non-digestible oligosaccharides (NDOs), and short chain fatty acids (SCFA), as their metabolites produced by the gut microbiota, interact with enteropathogens and their toxins, which may result in the inhibition of the bacterial pathogenicity. NDOs characterized by diverse structural characteristics, block the pathogenicity of EPB either directly, by inhibiting bacterial adherence and growth, or biofilm formation or indirectly, by promoting gut microbiota. Apart from these abilities, NDOs and SCFA can interact with enterotoxins and reduce their cytotoxicity. These anti-virulent effects mostly rely on their ability to mimic the structure of toxin receptors and thus inhibiting toxin adherence to host cells. This review focuses on the strategies of EPB and related enterotoxins to impair host cell immunity, discusses the anti-pathogenic properties of NDOs and SCFA on EPB functions and provides insight into the potential use of NDOs and SCFA as effective agents to fight against enterotoxins

Non-Digestible Oligosaccharides and Short Chain Fatty Acids as Therapeutic Targets against Enterotoxin-Producing Bacteria and Their Toxins

Enterotoxin-producing bacteria (EPB) have developed multiple mechanisms to disrupt gut homeostasis, and provoke various pathologies. A major part of bacterial cytotoxicity is attributed to the secretion of virulence factors, including enterotoxins. Depending on their structure and mode of action, enterotoxins intrude the intestinal epithelium causing long-term consequences such as hemorrhagic colitis. Multiple non-digestible oligosaccharides (NDOs), and short chain fatty acids (SCFA), as their metabolites produced by the gut microbiota, interact with enteropathogens and their toxins, which may result in the inhibition of the bacterial pathogenicity. NDOs characterized by diverse structural characteristics, block the pathogenicity of EPB either directly, by inhibiting bacterial adherence and growth, or biofilm formation or indirectly, by promoting gut microbiota. Apart from these abilities, NDOs and SCFA can interact with enterotoxins and reduce their cytotoxicity. These anti-virulent effects mostly rely on their ability to mimic the structure of toxin receptors and thus inhibiting toxin adherence to host cells. This review focuses on the strategies of EPB and related enterotoxins to impair host cell immunity, discusses the anti-pathogenic properties of NDOs and SCFA on EPB functions and provides insight into the potential use of NDOs and SCFA as effective agents to fight against enterotoxins

Fighting Shigella by Blocking Its Disease-Causing Toxin

Shiga toxin is an AB5 toxin produced by Shigella species, while related toxins are produced by Shiga toxin-producing Escherichia coli (STEC). Infection by Shigella can lead to bloody diarrhea followed by the often fatal hemolytic uremic syndrome (HUS). In the present paper, we aimed for a simple and effective toxin inhibitor by comparing three classes of carbohydrate-based inhibitors: glycodendrimers, glycopolymers, and oligosaccharides. We observed a clear enhancement in potency for multivalent inhibitors, with the divalent and tetravalent compounds inhibiting in the millimolar and micromolar range, respectively. However, the polymeric inhibitor based on galabiose was the most potent in the series exhibiting nanomolar inhibition. Alginate and chitosan oligosaccharides also inhibit Shiga toxin and may be used as a prophylactic drug during shigella outbreaks

Antimicrobial Activities of Alginate and Chitosan Oligosaccharides Against Staphylococcus aureus and Group B Streptococcus

The bacterial pathogens Streptococcus agalactiae (GBS) and Staphylococcus aureus (S. aureus) cause serious infections in humans and animals. The emergence of antibiotic-resistant isolates and bacterial biofilm formation entails the urge of novel treatment strategies. Recently, there is a profound scientific interest in the capabilities of non-digestible oligosaccharides as antimicrobial and anti-biofilm agents as well as adjuvants in antibiotic combination therapies. In this study, we investigated the potential of alginate oligosaccharides (AOS) and chitosan oligosaccharides (COS) as alternative for, or in combination with antibiotic treatment. AOS (2-16%) significantly decreased GBS V growth by determining the minimum inhibitory concentration. Both AOS (8 and 16%) and COS (2-16%) were able to prevent biofilm formation by S. aureus wood 46. A checkerboard biofilm formation assay demonstrated a synergistic effect of COS and clindamycin on the S. aureus biofilm formation, while AOS (2 and 4%) were found to sensitize GBS V to trimethoprim. In conclusion, AOS and COS affect the growth of GBS V and S. aureus wood 46 and can function as anti-biofilm agents. The promising effects of AOS and COS in combination with different antibiotics may offer new opportunities to combat antimicrobial resistance