Modulation of gene expression in immune-related organs by in ovo stimulation with probiotics and prophybiotics in broiler chickens.

In ovo stimulation has been studied intensively as an alternative to antibiotic use in poultry production. We investigated the potential use of a probiotic in combination with a phytobiotic as a prophybiotic for in ovo stimulation and reported its beneficial effects on the gut microbiome of broiler chickens. The current study further investigates the gene expression in the immune-related organs of these chickens to understand the tissue-specific immunomodulatory effects of the treatments. The selected prophybiotic (Leuconostoc mesenteroides with garlic aqueous extract) and its probiotic component alone were injected into ROSS308 chicken eggs on the 12th day of incubation, and gene expression in cecal tonsils, spleen, and liver at 35 days of age was determined using qPCR method. The relative expression of each treatment was compared to the positive control, chickens injected with physiological saline in ovo. The results displayed a downregulation of pro- and anti-inflammatory cytokines in the cecal tonsils of the probiotic group and the liver of the prophybiotic group. The spleen displayed upregulated AVBD1 in both groups and upregulated IL1-ß in the probiotic group. The probiotic group displayed increased expression of genes related to metabolism of energy (COX16), protein (mTOR), and lipids (CYP46A1) whereas the prophybiotic group displayed reduced expression of genes related to cholesterol synthesis (SREBP1) and glucose transportation (SLC2A2) in the liver. In conclusion, Leuconostoc mesenteroides differentially modulated gene expression in chickens when administered in ovo in combination with garlic aqueous extract. Further in ovo studies with different prophybiotic combinations are required to optimize the benefits in broiler chickens.

In-vitro selection of lactic acid bacteria to combat Salmonella enterica and Campylobacter jejuni in broiler chickens.

Campylobacter and Salmonella are the two most prominent foodborne zoonotic pathogens reported in the European Union. As poultry is one of the major sources of these pathogens, it is imperative to mitigate the colonization of these pathogens in poultry. Many strains of lactic acid bacteria (LAB) have demonstrated anti-Salmonella and anti-Campylobacter characteristics to varying degrees and spectrums which are attributed to the production of various metabolites. However, the production of these compounds and consequent antimicrobial properties are highly strain dependent. Therefore, the current study was performed to select a potent LAB and determine its causal attribute in inhibiting Salmonella enterica and Campylobacter jejuni, in-vitro. Six LAB (Lactiplantibacillus plantarum (LP), Lacticaseibacillus casei (LC), Limosilactobacillus reuteri (LR), Lacticaseibacillus rhamnosus (LRh), Leuconostoc mesenteroides (LM) and Pediococcus pentosaceus (PP)) and three serovars of Salmonella enterica (Typhimurium, Enterica and Braenderup) and Campylobacter jejuni were used in the current study. Spot overlays, well diffusion, co-culture and co-aggregation assays against Salmonella and well diffusion assays against Campylobacter jejuni were performed. Organic acid profiling of culture supernatants was performed using HPLC. The results indicated that LRh, LM and PP had the most significant anti-Salmonella effects while LP, LC, LM and PP displayed the most significant anti-Campylobacter effects. Lactic acid and formic acid detected in the culture supernatants seem the most likely source of the anti-Salmonella and anti-Campylobacter effects exhibited by these LAB. In conclusion, Leuconostoc mesenteroides displayed the most significant overall anti-pathogenic effects when compared to the other LAB strains studied, indicating its potential application in-vivo.

Prophybiotics for in-ovo stimulation; validation of effects on gut health and production of broiler chickens.

Probiotics and phytobiotics have demonstrated effective improvement of gut health in broiler chickens when individually administered in-ovo. However, their combined use in-ovo, has not been studied to date. We coined the term "prophybiotic" (probiotic + phytobiotic) for such a combination. The current study therefore, aimed to elucidate the effects of combined use of a selected probiotic and a phytobiotic in-ovo, on broiler gut health and production parameters, as opposed to use of probiotics alone. ROSS 308 hatching eggs were injected with either Leuconostoc mesenteroides (probiotic: PB) or L. mesenteroides with garlic aqueous extract (prophyiotic: PPB) on the 12th day of incubation. Relative abundances of bacteria in feces and cecal content (qPCR), immune related gene expression in cecal mucosa (qPCR) and histomorphology of cecal tissue (PAS staining) were analyzed along with production parameters (hatch quality, body weight, feed efficiency and slaughter and meat quality). PPB treatment increased the abundance of faecalibacteria and bifidobacteria in feces (d 7) and Akkermansia sp. in cecal content. Moreover, it decreased Escherichia coli abundance in both feces (d 34) and cecal content. PB treatment only increased the faecalibacteria in feces (d 7) and Akkermansia sp. in the cecal content. Moreover, PPB treatment resulted in up-regulation of immune related genes (Avian beta defensing 1, Free fatty acid receptor 2 and Mucin 6) and increased the crypt depth in ceca whereas PB treatment demonstrated a higher crypt depth and a tendency to increase Mucin 6 gene expression. Both treatments did not impair the production parameters studied. In conclusion, our results suggest that in-ovo PPB treatment may have enhanced potential in boosting the immune system without compromising broiler production and efficiency, as compared to the use of probiotic alone. Our study, highlights the potential of carefully selected PPB combinations for better results in improving gut health of broiler chickens.

Molecular strategies for the utilisation of human milk oligosaccharides by infant gut-associated bacteria.

A number of bacterial species are found in high abundance in the faeces of healthy breast-fed infants, an occurrence that is understood to be, at least in part, due to the ability of these bacteria to metabolize human milk oligosaccharides (HMOs). HMOs are the third most abundant component of human milk after lactose and lipids, and represent complex sugars which possess unique structural diversity and are resistant to infant gastrointestinal digestion. Thus, these sugars reach the infant distal intestine intact, thereby serving as a fermentable substrate for specific intestinal microbes, including Firmicutes, Proteobacteria, and especially infant-associated Bifidobacterium spp. which help to shape the infant gut microbiome. Bacteria utilising HMOs are equipped with genes associated with their degradation and a number of carbohydrate-active enzymes known as glycoside hydrolase enzymes have been identified in the infant gut, which supports this hypothesis. The resulting degraded HMOs can also be used as growth substrates for other infant gut bacteria present in a microbe-microbe interaction known as 'cross-feeding'. This review describes the current knowledge on HMO metabolism by particular infant gut-associated bacteria, many of which are currently used as commercial probiotics, including the distinct strategies employed by individual species for HMO utilisation.

In-vitro screening of compatible synbiotics and (introducing) "prophybiotics" as a tool to improve gut health.

Synbiotics have been intensively studied recently to improve gut health of humans and animals. The success of synergistic synbiotics depends on the compatibility of the prebiotic and probiotic components. Certain plant extracts possess both antimicrobial and prebiotic properties representing a potential use in combination with probiotics to improve the gut health. Here, we coined the term "prophybiotics" to describe this combined bioactivity. The current study aimed to select prebiotics that are preferred as an energy source and antimicrobial plant extracts which do not inhibit the growth, of six strains of lactic acid bacteria (LAB namely; Lactiplantibacillus plantarum, Lacticaseibacillus casei, Limosilactobacillus reuteri, Lacticaseibacillus rhamnosus, Leuconostoc mesenteroides, and Pediococcus pentosaceus) in-vitro to identify compatible combinations for potential synbiotic/prophybiotic use, respectively. Their growth kinetics were profiled in the presence of prebiotics: Inulin, Raffinose, and Saccharicterpenin with glucose, as the control, using carbohydrate free MRS broth media. Similarly, their growth kinetics in MRS broth supplemented with turmeric, green tea, and garlic extracts at varying concentrations were profiled. The results revealed the most compatible pairs of prebiotics and LAB. Turmeric and garlic had very little inhibitory effect on the growth of the LAB while green tea inhibited the growth of all LAB in a dose-dependent manner. Therefore, we conclude that turmeric and garlic have broad potential for use in prophybiotics, while the prebiotics studied here have limited use in synbiotics, with these LAB.

HMO-primed bifidobacteria exhibit enhanced ability to adhere to intestinal epithelial cells.

The ability of gut commensals to adhere to the intestinal epithelium can play a key role in influencing the composition of the gut microbiota. Bifidobacteria are associated with a multitude of health benefits and are one of the most widely used probiotics for humans. Enhanced bifidobacterial adhesion may increase host-microbe, microbe-nutrient, and/or microbe-microbe interactions, thereby enabling consolidated health benefits to the host. The objective of this study was to determine the ability of human milk oligosaccharides (HMOs) to enhance bifidobacterial intestinal adhesion in vitro. This study assessed the colonisation-promoting effects of HMOs on four commercial infant-associated Bifidobacterium strains (two B. longum subsp. infantis strains, B. breve and B. bifidum). HT29-MTX cells were used as an in vitro intestinal model for bacterial adhesion. Short-term exposure of four commercial infant-associated Bifidobacterium strains to HMOs derived from breastmilk substantially increased the adherence (up to 47%) of these probiotic strains. Interestingly, when strains were incubated with HMOs as a four-strain combination, the number of viable bacteria adhering to intestinal cells increased by >90%. Proteomic analysis of this multi-strain bifidobacterial mixture revealed that the increased adherence resulting from exposure to HMOs was associated with notable increases in the abundance of sortase-dependent pili and glycosyl hydrolases matched to Bifidobacterium bifidum. This study suggests that HMOs may prime infant gut-associated Bifidobacterium for colonisation to intestinal epithelial cells by influencing the expression of various colonization factors.

Human milk oligosaccharide-sharing by a consortium of infant derived Bifidobacterium species.

Bifidobacteria are associated with a host of health benefits and are typically dominant in the gut microbiota of healthy, breast-fed infants. A key adaptation, facilitating the establishment of these species, is their ability to consume particular sugars, known as human milk oligosaccharides (HMO), which are abundantly found in breastmilk. In the current study, we aimed to characterise the co-operative metabolism of four commercial infant-derived bifidobacteria (Bifidobacterium bifidum R0071, Bifidobacterium breve M-16V, Bifidobacterium infantis R0033, and Bifidobacterium infantis M-63) when grown on HMO. Three different HMO substrates (2'-fucosyllactose alone and oligosaccharides isolated from human milk representing non-secretor and secretor status) were employed. The four-strain combination resulted in increased bifidobacterial numbers (> 21%) in comparison to single strain cultivation. The relative abundance of B. breve increased by > 30% during co-cultivation with the other strains despite demonstrating limited ability to assimilate HMO in mono-culture. HPLC analysis revealed strain-level variations in HMO consumption. Metabolomics confirmed the production of formate, acetate, 1,2-propanediol, and lactate with an overall increase in such metabolites during co-cultivation. These results support the concept of positive co-operation between multiple bifidobacterial strains during HMO utilisation which may result in higher cell numbers and a potentially healthier balance of metabolites.

Characterization and Analysis of Food-Sourced Carbohydrates.

Food carbohydrates are macronutrients that are found in fruits, grains, vegetables, and milk products. These organic compounds are present in foods in the form of sugars, starches, and fibers and are composed of carbon, hydrogen, and oxygen. These wide ranging macromolecules can be classified according to their chemical structure into three major groups: low molecular weight mono- and disaccharides, intermediate molecular weight oligosaccharides, and high molecular weight polysaccharides. Notably, the digestibility of specific carbohydrate components differ and nondigestible carbohydrates can reach the large intestine intact where they act as food sources for beneficial bacteria. In this review, we give an overview of advances made in food carbohydrate analysis. Overall, this review indicates the importance of carbohydrate analytical techniques in the quest to identify and isolate health-promoting carbohydrates to be used as additives in the functional foods industry.

Human milk oligosaccharides: Shaping the infant gut microbiota and supporting health.

Human milk oligosaccharides (HMO) are complex sugars which are found in breast milk at significant concentrations and with unique structural diversity. These sugars are the fourth most abundant component of human milk after water, lipids, and lactose and yet provide no direct nutritional value to the infant. Recent research has highlighted that HMOs have various functional roles to play in infant development. These sugars act as prebiotics by promoting growth of beneficial intestinal bacteria thereby generating short-chain fatty acids which are critical for gut health. HMOs also directly modulate host-epithelial immune responses and can selectively reduce binding of pathogenic bacteria and viruses to the gut epithelium preventing the emergence of a disease. This review covers current knowledge related to the functional biology of HMOs and their associated impact on infant gut health.

A Whey Fraction Rich in Immunoglobulin G Combined with Bifidobacterium longum subsp. infantis ATCC 15697 Exhibits Synergistic Effects against Campylobacter jejuni.

Evidence that whey proteins and peptides have health benefits beyond basic infant nutrition has increased dramatically in recent years. Previously, we demonstrated that a whey-derived immunoglobulin G-enriched powder (IGEP) enhanced adhesion of Bifidobacterium longum subsp. infantis ATCC 15697 (B. infantis) to HT-29 cells. In this study, we investigated the synergistic effect of IGEP-treated B. infantis on preventing the attachment of highly invasive Campylobacter jejuni 81-176 (C. jejuni) to intestinal HT-29 cells. The combination decreased the adherence of C. jejuni to the HT-29 cells by an average of 48% compared to the control (non-IGEP-treated B. infantis). We also confirmed that treatment of IGEP with sodium metaperiodate, which disables the biological recognition of the conjugated oligosaccharides, reduced adhesion of B. infantis to the intestinal cells. Thus, glycosylation of the IGEP components may be important in enhancing B. infantis adhesion. Interestingly, an increased adhesion phenotype was not observed when B. infantis was treated with bovine serum-derived IgG, suggesting that bioactivity was unique to milk-derived immunoglobulin-rich powders. Notably, IGEP did not induce growth of B. infantis within a 24 hours incubation period, as demonstrated by growth curves and metabolite analysis. The current study provides insight into the functionality of bovine whey components and highlights their potential in positively impacting the development of a healthy microbiota.

Immunoglobulin G from bovine milk primes intestinal epithelial cells for increased colonization of bifidobacteria.

A bovine colostrum fraction (BCF) was recently shown to enhance the adherence of several commensal organisms to intestinal epithelial cells through modulating the epithelial cell surface. In this study, the main components of the BCF were examined to investigate the active component/s responsible for driving the changes in the intestinal cells. The adherence of various bifidobacteria to HT-29 cells was increased when the intestinal cells were pre-incubated with immunoglobulin G (IgG). Modulation of the intestinal cells by IgG was concentration dependent with 16 mg/mL IgG resulting in a 43-fold increase in the adhesion of Bifidobacterium longum NCIMB 8809 to HT-29 cells. Periodate treatment of colostral IgG prior to performing the colonization studies resulted in a reduction in the adhesion of the strain to the intestinal cells demonstrating that the glycans of IgG may be important in modulating the intestinal cells for enhanced commensal adhesion. IgG isolated from mature milk also resulted in significant increases in adhesion of the Bifidobacterium strains tested albeit at reduced levels (3.9-fold). The impact of IgG on the HT-29 cells was also visualised via scanning electron microscopy. This study builds a strong case for the inclusion of IgG ingredients sourced from cow's milk in functional foods aimed at increasing numbers of health promoting bacteria in the human gut.

Bifidobacterium longum subsp. infantis ATCC 15697 and Goat Milk Oligosaccharides Show Synergism In Vitro as Anti-Infectives against Campylobacter jejuni.

Bifidobacteria are known to inhibit, compete with and displace the adhesion of pathogens to human intestinal cells. Previously, we demonstrated that goat milk oligosaccharides (GMO) increased the attachment of Bifidobacterium longum subsp. infantis ATCC 15697 to intestinal cells in vitro. In this study, we aimed to exploit this effect as a mechanism for inhibiting pathogen association with intestinal cells. We examined the synergistic effect of GMO-treated B. infantis on preventing the attachment of a highly invasive strain of Campylobacter jejuni to intestinal HT-29 cells. The combination decreased the adherence of C. jejuni to the HT-29 cells by an average of 42% compared to the control (non-GMO treated B. infantis). Increasing the incubation time of the GMO with the Bifidobacterium strain resulted in the strain metabolizing the GMO, correlating with a subsequent 104% increase in growth over a 24 h period when compared to the control. Metabolite analysis in the 24 h period also revealed increased production of acetate, lactate, formate and ethanol by GMO-treated B. infantis. Statistically significant changes in the GMO profile were also demonstrated over the 24 h period, indicating that the strain was digesting certain structures within the pool such as lactose, lacto-N-neotetraose, lacto-N-neohexaose 3'-sialyllactose, 6'-sialyllactose, sialyllacto-N-neotetraose c and disialyllactose. It may be that early exposure to GMO modulates the adhesion of B. infantis while carbohydrate utilisation becomes more important after the bacteria have transiently colonised the host cells in adequate numbers. This study builds a strong case for the use of synbiotics that incorporate oligosaccharides sourced from goat's milk and probiotic bifidobacteria in functional foods, particularly considering the growing popularity of formulas based on goat milk.

Symposium review: Dairy-derived oligosaccharides-Their influence on host-microbe interactions in the gastrointestinal tract of infants.

Oligosaccharides are the third most abundant component in human milk. It is widely accepted that they play several important protective, physiological, and biological roles, including selective growth stimulation of beneficial gut microbiota, inhibition of pathogen adhesion, and immune modulation. However, until recently, very few commercial products on the market have capitalized on these functions. This is mainly because the quantities of human milk oligosaccharides required for clinical trials have been unavailable. Recently, clinical studies have tested the potential beneficial effects of feeding infants formula containing 2'-fucosyllactose, which is the most abundant oligosaccharide in human milk. These studies have opened this field for further well-designed studies, which are required to fully understand the role of human milk oligosaccharides. However, one of the most striking features of human milk is its diversity of oligosaccharides, with over 200 identified to date. It may be that a mixture of oligosaccharides is even more beneficial to infants than a single structure. For this reason, the milk of domestic animals has become a focal point in recent years as an alternative source of complex oligosaccharides with associated biological activity. This review will focus specifically on free oligosaccharides found in bovine and caprine milk and the biological roles associated with such structures. These dairy streams are ideal sources of oligosaccharides, given their wide availability and use in so many regularly consumed dairy products. The aim of this review was to provide an overview of research into the functional role of bovine and caprine milk oligosaccharides in host-microbial interactions in the gut and provide current knowledge related to the isolation of oligosaccharides as ingredients for incorporation in functional or medical foods.

New insights on the colonization of the human gut by health-promoting bacteria.

We are beginning to see how the microbiota of the human gastrointestinal tract (GIT) can drive the development of new products to benefit human health and wellbeing. Despite the growing market for prebiotics and probiotics, there are currently no commercial products available that aid or increase the attachment of health-promoting bacteria to the gut mucosal surface. Components in milk have the potential to increase commensal adherence in the gut by priming the bacteria or the mucosal surface for colonization. Such compositions have potential for supplementation in many products aimed at individuals at different life stages or those suffering from various disease states where lower numbers of health-promoting bacteria such as bifidobacteria are evident. This review will explore how milk ingredients may lead to the attachment of larger numbers of bacteria with health-promoting properties in the gut.

Oligosaccharides Isolated from MGO™ Manuka Honey Inhibit the Adhesion of Pseudomonas aeruginosa, Escherichia Coli O157:H7 and Staphylococcus Aureus to Human HT-29 cells.

Historically, honey is known for its anti-bacterial and anti-fungal activities and its use for treatment of wound infections. Although this practice has been in place for millennia, little information exists regarding which manuka honey components contribute to the protective nature of this product. Given that sugar accounts for over 80% of honey and up to 25% of this sugar is composed of oligosaccharides, we have investigated the anti-infective activity of manuka honey oligosaccharides against a range of pathogens. Initially, oligosaccharides were extracted from a commercially-available New Zealand manuka honey-MGO™ Manuka Honey (Manuka Health New Zealand Ltd)-and characterized by High pH anion exchange chromatography coupled with pulsed amperiometric detection. The adhesion of specific pathogens to the human colonic adenocarcinoma cell line, HT-29, was then assessed in the presence and absence of these oligosaccharides. Manuka honey oligosaccharides significantly reduced the adhesion of Escherichia coli O157:H7 (by 40%), Staphylococcus aureus (by 30%), and Pseudomonas aeruginosa (by 52%) to HT-29 cells. This activity was then proven to be concentration dependent and independent of bacterial killing. This study identifies MGO™ Manuka Honey as a source of anti-infective oligosaccharides for applications in functional foods aimed at lowering the incidence of infectious diseases.

Lectin microarray profiling and monosaccharide analysis of bovine milk immunoglobulin G oligosaccharides during the first 10 days of lactation.

Immunoglobulin G (IgG) in bovine milk is credited with ensuring efficient passive immunity for newborn calves. Bovine milk IgG glycosylation may also have positive impacts on the health of nonbovine consumers of cow's milk. Milk IgG's glycosylation contributes to effector function and may also protect it from protease digestion, allowing IgG to reach the intestine for absorption. However, relatively little is known about changes in milk IgG oligosaccharide presentation and composition over early lactation. In this work, IgG was isolated from milk pooled from three cows at four time points over the first 10 days of lactation postparturition. Purified IgG was labeled with a fluorescent dye and interrogated with a microarray consisting of 48 carbohydrate-binding proteins (lectins) from plant, fungal, and bacterial sources. Lectin microarray profiles suggested that only subtle changes in the glycosylation of IgG occurred during days 2 and 3 of lactation, but by day 10, the lectin profile diverged from the other three time points. Monosaccharide analysis carried out after hydrolysis confirmed that the ratios of oligosaccharide components remained relatively stable through day 3 and also that sialylation was substantially reduced by day 10. The differences that were observed for glycosylation suggest that different functionalities associated with IgG glycosylation may be required in the first days of life.

Interrogation of Milk-Driven Changes to the Proteome of Intestinal Epithelial Cells by Integrated Proteomics and Glycomics.

Bovine colostrum is a rich source of bioactive components which are important in the development of the intestine, in stimulating gut structure and function and in preparing the gut surface for subsequent colonization of microbes. What is not clear, however, is how colostrum may affect the repertoire of receptors and membrane proteins of the intestinal surface and the post-translational modifications associated with them. In the present work, we aimed to characterize the surface receptor and glycan profile of human HT-29 intestinal cells after exposure to a bovine colostrum fraction (BCF) by means of proteomic and glycomic analyses. Integration of label-free quantitative proteomic analysis and lectin array profiles confirmed that BCF exposure results in changes in the levels of glycoproteins present at the cell surface and also changes to their glycosylation pattern. This study contributes to our understanding of how milk components may regulate intestinal cells and prime them for bacterial interaction.

Bovine colostrum-driven modulation of intestinal epithelial cells for increased commensal colonisation.

Nutritional intake may influence the intestinal epithelial glycome and in turn the available attachment sites for bacteria. In this study, we tested the hypothesis that bovine colostrum may influence the intestinal cell surface and in turn the attachment of commensal organisms. Human HT-29 intestinal cells were exposed to a bovine colostrum fraction (BCF) rich in free oligosaccharides. The adherence of several commensal bacteria, comprising mainly bifidobacteria, to the intestinal cells was significantly enhanced (up to 52-fold) for all strains tested which spanned species that are found across the human lifespan. Importantly, the changes to the HT-29 cell surface did not support enhanced adhesion of the enteric pathogens tested. The gene expression profile of the HT-29 cells following treatment with the BCF was evaluated by microarray analysis. Many so called "glyco-genes" (glycosyltransferases and genes involved in the complex biosynthetic pathways of glycans) were found to be differentially regulated suggesting modulation of the enzymatic addition of sugars to glycoconjugate proteins. The microarray data was further validated by means of real-time PCR. The current findings provide an insight into how commensal microorganisms colonise the human gut and highlight the potential of colostrum and milk components as functional ingredients that can potentially increase commensal numbers in individuals with lower counts of health-promoting bacteria.

Mining Milk for Factors which Increase the Adherence of Bifidobacterium longum subsp. infantis to Intestinal Cells.

Bifidobacteria play a vital role in human nutrition and health by shaping and maintaining the gut ecosystem. In order to exert a beneficial effect, a sufficient population of bifidobacteria must colonise the host. In this study, we developed a miniaturised high-throughput in vitro assay for assessing the colonising ability of bacterial strains in human cells. We also investigated a variety of components isolated from different milk sources for their ability to increase the adherence of Bifidobacterium longum subsp. infantis ATCC 15697, a common member of the gastrointestinal microbiota of breastfed infants, to HT-29 cells. Both conventional and miniaturised colonisation assays were employed to examine the effect of 13 different milk-derived powders on bacterial adherence, including positive controls which had previously resulted in increased bifidobacterial adherence (human milk oligosaccharides and a combination of 3'- and 6'-sialylactose) to intestinal cells. Immunoglobulin G enriched from bovine whey and goat milk oligosaccharides resulted in increased adhesion (3.3- and 8.3-fold, respectively) of B. infantis to the intestinal cells and the miniaturised and conventional assays were found to yield comparable and reproducible results. This study highlights the potential of certain milk components to favourably modulate adhesion of bifidobacteria to human intestinal cells.

Glycomacropeptide Reduces Intestinal Epithelial Cell Barrier Dysfunction and Adhesion of Entero-Hemorrhagic and Entero-Pathogenic Escherichia coli in Vitro.

In recent years, the potential of glycosylated food components to positively influence health has received considerable attention. Milk is a rich source of biologically active glycoconjugates which are associated with antimicrobial, immunomodulatory, anti-adhesion, anti-inflammatory and prebiotic properties. Glycomacropeptide (GMP) is the C-terminal portion of kappa-casein that is released from whey during cheese-making by the action of chymosin. Many of the biological properties associated with GMP, such as anti-adhesion, have been linked with the carbohydrate portion of the protein. In this study, we investigated the ability of GMP to inhibit the adhesion of a variety of pathogenic Escherichia coli strains to HT-29 and Caco-2 intestinal cell lines, given the importance of E. coli in causing bacterial gastroenteritis. GMP significantly reduced pathogen adhesion, albeit with a high degree of species specificity toward enteropathogenic E. coli (EPEC) strains O125:H32 and O111:H2 and enterohemorrhagic E. coli (EHEC) strain 12900 O157:H7. The anti-adhesive effect resulted from the interaction of GMP with the E. coli cells and was also dependent on GMP concentration. Pre-incubation of intestinal Caco-2 cells with GMP reduced pathogen translocation as represented by a decrease in transepithelial electrical resistance (TEER). Thus, GMP is an effective in-vitro inhibitor of adhesion and epithelial injury caused by E. coli and may have potential as a biofunctional ingredient in foods to improve gastrointestinal health.