Commensal Pathogen - HMO+ HMO Glycan receptor HMO - HMO+ HMO Effect of fucosyl- and sialyllactoses on bacterial adhesion to intestinal epithelial cells Maria L. Skalkam 1, Maria Frank 2 and Dennis S. Nielsen 1 1 Department of Food Sciences, Food Microbiology, University of Copenhagen, Rolighedsvej 26, 1958 Frederiksberg C, Denmark; 2 Department of Food Science, Food Chemistry and technology, University of Aarhus, Blichers Allé 20, 8830 Tjele, Denmark FACULTY OF SCIENCE UNIVERSITY OF COPENHAGEN Human milk is considered the golden standard for infant feeding, and is shown to confer several health benefits to the breastfed infant. Human milk oligosaccharides (HMOs) are believed to contribute significantly to the observed health effects associated with breastfeeding. Health benefits associated with breastfeeding includes fewer infections, reduced incidence of asthma and allergies, reduced risk of developing lifestyle diseases such as diabetes and cardiovascular diseases, but also neurological development are believed to be positively influenced by breastfeeding. HMOs, which especially stimulate the growth of bifidobacteria, are believed to play a role in several of the observed health benefits of breast milk and are gaining increasingly more attention as a possible functional food supplement, especially in infant formulas, but also possible roles as an anti-adhesive agent is being investigated. Caco-2 cells were cultured for 21 days to obtain a fully functional differentiated monolayer. Cronobacter sakazakii (CCUG28860) was added to the Caco-2 monolayer at an approximate inoculum size of 1*10 8 cells/well, together with single HMO at a final concentration of 10 mg/ml, and incubated for 1½ hour in a humidified atmosphere at 37⁰C with 5% CO 2. Following incubation, cells were washed 3 times with PBS to remove non-adherent bacteria and lysed with 1% Triton X-100. Bacterial enumaration was done by serial dilutions and plating on brain-heart infusion agar. HMOs can act as receptor analogues, binding pathogenic bacteria in the lumen and thus reduce adherence to intestinal cells. Preliminary results show a varied effect on adhesion, which seems to be structure and species specific. A probiotic, Lactobacillus acidophilus NCFM, showed increased adhesion when added to Caco-2 cells together with 6-sialyllactose (6-SL). Pathogenic bacteria showed different responses with both increased and decreased adhesion following incubation with different HMOs onto Caco-2 cells. Cronobacter sakazakii, which is a major concern in reconstituted infant formula, show reduced adherence in the presence of 2-fucosyllactose (2-FL) and, although non-significant, 3-fucosyllactose (3-FL) show the same tendency. 3-sialyllactose (3- SL) does not seem to affect adhesion properties of either probitic or pathogenic bacteria. Schematic illustration of prebiotic and anti-adhesive effects of HMOs. A. HMOs stimulate growth of commensal bacteria, causing a reduction in pathogenic bacteria through exclusion and nutrient competition. B. HMOs act as receptor analogues and prevent binding of pathogenic bacteria to intestinal cells. Improved infant formula Our results show a potential for the use of some simple HMOs as a functional food ingredient, which aside from presumed prebiotic effects, have the potential to decrease bacterial infections by acting as decoy receptors. The addition of HMOs to powdered infant formula could have several positive effects, however further studies are needed. HMOs can affect adhesion of different bacteria to Caco-2 cells Adherence of Listeria monocytogenes LO28Adherence of Cronobacter sakazakiiAdherence of Lb. acidophilus NCFM 2-FL3-FL3-SL6-SL2-FL3-FL3-SL6-SL2-FL3-FL3-SL6-SL * * * Results Introduction Hypothesis Method Perspectives B A