Background 2′-Fucosyllactose (2′-FL) is one of the most abundant components among human milk oligosaccharides (HMOs), attracting significant attention for its crucial role in infant immune system development and infection resistance. 2′-FL not only prevents pathogens from binding to intestinal epithelial cells but also exhibits potential protective effects against infections by enteric pathogens such as rotavirus. Rotavirus is a primary pathogen causing acute gastroenteritis in infants and young children, responsible for over 200,000 infant deaths globally each year. Although vaccine use has reduced infection rates, rotavirus remains a significant cause of infant morbidity and mortality worldwide. This paper explores the inhibitory effect of 2′-FL on rotavirus infection and its potential clinical applications. Research Design and Methods This study analysed the effects of 2′-FL on rotavirus infection through in vitro experiments. MA104 African green monkey kidney epithelial cells were employed as the infection model, inoculated with two globally prevalent rotavirus strains: G1P[8] and G2P[4]. Fluorescence-based assays assessed the effects of varying 2′-FL concentrations on viral infection and replication. Experimental protocols were designed to evaluate whether 2′-FL retained inhibitory activity following viral adsorption. Key Findings 1. 2′-FL significantly reduces the infectivity of rotavirus G1P[8] Experimental results demonstrate that 2′-FL treatment markedly diminishes the infectivity of rotavirus G1P[8]. This indicates that 2′-FL competes with cell surface receptors, thereby preventing viral adhesion and reducing opportunities for infection. Figure 1: 2′-FL reduces the infectivity of rotavirus G1P[8] in MA104 cells 2. Inhibitory Effect of 2′-FL on Rotavirus G2P[4] Infection Although infection mechanisms may vary across different viral strains, 2′-FL still exhibits inhibitory effects against G2P[4] infection. At concentrations of 2.5 mg/mL and 5 mg/mL, 2′-FL effectively reduced the infectivity of G2P[4], demonstrating its broad-spectrum activity against multiple rotavirus strains. Figure 2: 2′-FL reduces the infectivity of rotavirus G2P[4] 3. 2′-FL retains its inhibitory effect on infection following viral uptake In experiments where 2′-FL was added after viral adsorption, it was found to significantly reduce the infectivity of GIP[8], with a maximum reduction of 62%. This indicates that 2′-FL not only prevents viral entrycells but may also further diminish infection by interfering with the virus's replication process within the cell. Figure 2: Addition of 2′-FL following viral uptake reduces the infectivity of human rotavirus strain G1P[8] in MA104 cells. Discussion and Outlook This study confirms the efficacy of 2′-FL in inhibiting rotavirus infection, demonstrating particular effectiveness in reducing viral infectivity and lowering viral load. Through its decoy receptor action, 2′-FL effectively prevents viral binding to host cells, thereby reducing opportunities for rotavirus infection. Furthermore, 2′-FL not only acts prior to viral entrycells but also exhibits the capacity to inhibit viral replication following adsorption. The inhibitory effect of 2′-FL was observed across various rotavirus strains, further indicating its broad application prospects as a crucial component in infant health protection. It serves not only as a key element in the development of the infant immune system but also holds promise as an important tool in antiviral therapy. Future research may further explore the antiviral mechanisms of 2′-FL in vivo and validate its potential for application in clinical settings. Reference [1] Laucirica D R, Triantis V, Schoemaker R, et al. Milk oligosaccharides inhibit human rotavirus infectivity in MA104 cells[J]. The Journal of Nutrition, 2017, 147(9): 1709-1714.

Background 2′-Fucosyllactose (2′-FL) is a key component of human milk oligosaccharides (HMOs), which have garnered significant attention in recent years for their role in enhancing infant immunity and maintaining gut health. Research indicates that 2′-FL not only confers significant benefits for gut health but also modulates immune responses during respiratory viral infections. Notably, 2′-FL exhibits pronounced antiviral effects in regulating immunity against respiratory syncytial virus (RSV). RSV is a primary pathogen causing severe respiratory infections in infants and young children, who are particularly vulnerable due to their immature immune systems. Consequently, investigating the immune-enhancing effects of 2′-FL against RSV holds significant clinical application potential. This paper will focus on discussing the immunomodulatory role of 2′-FL in RSV infection and its potential applications. Research Design and Methods This study explored the effects of 2′-FL on RSV infection through in vitro experiments. Research was conducted using 16HBE human bronchial epithelial cells and a polarised Calu-3 epithelial cell model. Following RSV infection of the cells, varying concentrations of 2′-FL were added to assess its impact on viral replication and inflammatory cytokine expression. Secretion levels of pro-inflammatory factors (including IL-6, IL-8, MIP-1α, and TNF-α) were measured via ELISA and qPCR, whilst viral load was evaluated through viral titration assays. Key Findings 1. 2′-FL significantly reduces RSV viral load Experimental results demonstrate that RSV viral load is markedly diminished in 16HBE cells treated with 2′-FL, particularly at concentrations exceeding 50 μg/mL. This finding indicates that 2′-FL inhibits RSV replication even at low concentrations, revealing its potent antiviral potential. Figure 1: Analysis of the effect of 2′-FL on RSV NS1 gene expression in 16HBE epithelial cells via qRT-PCR. *p<0.05 2. 2′-FL Inhibits RSV-Associated Pro-inflammatory Factors Studies indicate that in RSV-infected 16HBE cells, 2′-FL significantly reduced the secretion of pro-inflammatory factors such as IL-6, IL-8, MIP-1α, and TNF-α. This reduction in inflammatory mediators demonstrates that 2′-FL effectively suppresses excessive inflammatory responses triggered by viral infection, thereby mitigating virus-induced tissue damage. Figure 2: 2′-FL reduces inflammatory cytokine markers in 16HBE epithelial cells infected with RSV. *p<0.05, (A) IL-6; (B) IL-8; (C) MIP-1α; (D) MCP-1; (E) TNF-α. 3. Expression of 2′-FL in Calu-3 cells Further studies confirmed that 2′-FL similarly reduced viral load in polarised Calu-3 epithelial cells, demonstrating effective anti-RSV activity and indicating its broad utility across diverse cellular models. Figure 3: Effect of 2′-FL on RSV NS1 gene expression in Calu-3 epithelial cells. *p<0.05 Discussion and Outlook This study has established the immune-enhancing role of 2′-FL in RSV infection. By reducing viral load and decreasing the secretion of inflammatory cytokines, 2′-FL demonstrates significant antiviral and anti-inflammatory effects. Its specific action in RSV infection provides a theoretical basis for the development of future antiviral therapies and prophylactic products, further confirming its potential as a health protection product for infants and young children. Future research may further explore the broad-spectrum efficacy of 2′-FL in other viral infections, particularly its potential application in respiratory health management for infants and young children. Moreover, as a natural antiviral and immune enhancer, 2′-FL holds promise for widespread use in infant nutritional supplementation and the prevention of respiratory diseases in adults, especially among populations with compromised immune systems. Reference [1] Duska-McEwen G, Senft A P, Ruetschilling T L, et al. Human milk oligosaccharides enhance innate immunity to respiratory syncytial virus and influenza in vitro[J]. Food and Nutrition Sciences, 2014, 2014.

Background Human milk oligosaccharides (HMOs) play a pivotal role in infant gut health and immune system development. Among these, 2′-fucosyllactose (2′-FL) is one of the most prevalent HMOs. 2′-FL not only promotes the growth of beneficial bacteria but also exhibits anti-infective and anti-inflammatory properties, particularly in regulating intestinal inflammatory responses. In intestinal inflammation, CD14 serves as a pivotal receptor that recognises and binds bacterial components such as lipopolysaccharide (LPS), thereby initiating immune responses. IL-8, a pro-inflammatory cytokine, attracts immune cells to the site of infection during the inflammatory process, amplifying the inflammatory response. 2′-FL can mitigate inflammation triggered by intestinal pathogens to some extent by regulating CD14 expression and IL-8 release. Study Design and Methods This study employed human intestinal epithelial cells to simulate the intestinal environment. Cells were first exposed to LPS to induce inflammation, followed by the addition of 2′-FL across different experimental groups to observe its effects on CD14 expression. Gene and protein expression levels of CD14 were assessed via quantitative PCR and Western blot analysis, while IL-8 release was evaluated using ELISA to determine 2′-FL's anti-inflammatory efficacy. Additionally, the study investigated whether 2′-FL influences pathogen adhesion capacity to intestinal epithelial cells. Key Findings 1. Suppressed CD14 Expression Following LPS treatment, CD14 expression in intestinal epithelial cells increased significantly, indicating successful induction of the inflammatory response. However, the cell group treated with 2'-FL exhibited markedly reduced levels of both CD14 gene and protein expression, demonstrating 2'-FL's role in mitigating the inflammatory response by downregulating CD14 expression. Figure 1: 2'-FL alters CD14 expression and distribution in cells T84 cells treated with 2'-FL for 48 hours were subsequently challenged with 100 μg/mL LPS to measure 2'-FL-induced changes in CD14 mRNA levels, protein levels, and intracellular distribution. 2. Reduced IL-8 secretion Following LPS treatment, IL-8 release from intestinal epithelial cells increased significantly, reflecting heightened inflammation. However, in the experimental group treated with 2′-FL, IL-8 release was markedly diminished, suggesting that 2′-FL may mitigate inflammatory responses by inhibiting CD14-mediated signalling pathways. Figure 2: 2'-FL Inhibits LPS-Induced IL-8 Cells were treated with 2'-FL for 48 hours, then challenged with LPS. The degree of inflammatory response was measured by IL-8 levels. 3. Pathogen adhesion is inhibited Research has also revealed that 2′-FL significantly reduces pathogen adhesion to intestinal epithelial cells. This may occur because 2′-FL mimics the sugar chains on the surface of intestinal epithelial cells, competitively preventing pathogens from binding to host cells and thereby lowering the risk of infection. Discussion and Outlook This study elucidates the potential mechanism by which 2′-FL modulates LPS-induced intestinal inflammatory responses, demonstrating that 2′-FL not only plays a crucial role in preventing infection but also alleviates inflammation triggered by intestinal pathogens by regulating immune responses. This holds potential benefits for intestinal health in both infants and adults. Future research may further validate the anti-inflammatory effects of 2′-FL in more complex in vivo models, particularly regarding its application in diseases associated with intestinal inflammation. Moreover, given the diverse array of HMOs present in human milk, the potential of other oligosaccharides in immunomodulation warrants exploration. The clinical application prospects for 2′-FL are extensive, extending beyond infant formula to its potential use as a natural anti-inflammatory agent in adult inflammatory bowel disease (IBD) and other gut disorders. Consequently, future research should prioritise establishing the safety and efficacy of 2′-FL, thereby providing scientific justification for its incorporationfunctional foods or pharmaceutical development. Reference [1] He, YingYing, et al. “The human milk oligosaccharide 2′-fucosyllactose modulates CD14 expression in human enterocytes, thereby attenuating LPS-induced inflammation.” Gut 65.1 (2016): 33-46.

  Background Research indicates that 2′-fucosyllactose (2′-FL) specifically stimulates the proliferation of Bifidobacterium species, thereby establishing a healthy gut microbial environment. Escherichia coli O157 is a common pathogen capable of causing severe haemorrhagic colitis and haemolytic uraemic syndrome (HUS), particularly in children, the elderly, and immunocompromised individuals. Given its potent virulence and high antibiotic resistance, identifying effective preventive and therapeutic strategies is paramount. Against this backdrop, 2′-FL, as a functional food ingredient, may play a significant role in preventing and mitigating E. coli O157 infections. Study Design and Methods This study employed a mouse infection model to analyse the inhibitory effect of 2′-FL on intestinal E. coli O157 infection following dietary administration. Experiments included extracting bacterial DNA from mouse caecal contents, performing PCR amplification and gene analysis, and utilising 16S rDNA gene sequencing to analyse the gut microbiome. Inflammation levels were assessed by measuring IL-6, TNF-α, and IL-1β concentrations in blood, ileum, and colon. Additionally, qPCR methods were employed to detect expression levels of mucin-2 (MUC2) and tight junction protein-related genes. Key Findings 1. 2′-FL inhibits Escherichia coli O157 colonisation in the gut In both the ileum and colon, the colonisation rate of Escherichia coli O157 in the 2′-FL group was significantly lower than that in the control group (MC group), at 8.63% ± 6.87% and 6.16% ± 2.25% respectively. This indicates that 2′-FL possesses a significant anti-pathogenic colonisation effect, contributing to a reduction in the intestinal pathogen burden. Figure 1: Colonisation of Escherichia coli O157 in the ileum and pathological sections of ileal tissue. (A) Colonisation in the ileum; (B) Colonisation in the colon; (C) Representative image of ileal tissue section from the control group (CK) (HE staining, ×100); (D) Representative image of ileal tissue section from the FL group (HE staining, ×100); (E) Representative image of ileal tissue section from the model control group (MCG) (HE staining, ×100). 2. Effects on Gut Microbiota Composition and Community Structure At the phylum level, prolonged supplementation with 2′-FL significantly reduced the abundance of Firmicutes and Patescibacteria, while increasing that of Bacteroidetes and Proteobacteria. Specifically, at the genus level, Parabacteroides, Eubacterium, Mucispirillum, Anaerotruncus, and Alistipes were markedly diminished. These alterations reflect 2′-FL's substantial influence on gut microbial ecological equilibrium. Furthermore, the 2′-FL group exhibited heightened diversity and stability. Such changes not only aid in resisting pathogenic colonisation but may also enhance overall host health by promoting the growth of beneficial bacteria. Figure 2: Relative abundance, diversity, and functional prediction of gut microbiota in different mouse groups: (A) Phylum; (B) Genus; (C) Dilution curve; (D) PCoA analysis based on operational taxonomic unit (OTU) results. 3. Effects on Inflammation Levels, Gene Expression, and Host Immune Responses 2′-FL significantly reduced levels of inflammatory cytokines (IL-6, TNF-α, and IL-1β) in the ileum and colon of mice, indicating its efficacy in suppressing gut inflammatory responses triggered by pathogenic bacterial infection. Concurrently, 2′-FL upregulated the expression of mucin-2 (MUC2) and tight junction protein genes in the colon. Enhanced expression of these genes contributes to strengthened intestinal barrier function, reducing pathogen invasion and colonisation. Moreover, following 2′-FL supplementation, the host immune system demonstrated improved regulatory capacity, manifested as a balanced profile of pro-inflammatory and anti-inflammatory cytokines. This immunomodulatory effect may represent an indirect action of 2′-FL mediated through the gut microbiota. Figure 3: Relative mRNA levels of inflammatory cytokines and mucins, normalised against β-actin mRNA expression: (A) inflammatory cytokines in the ileum; (B) inflammatory cytokines in the colon; (C) mucin expression in the colon. Different letters (a–c) denote significant differences in the same factor between groups (p < 0.05). Discussion and Outlook Research indicates that 2′-FL exerts significant effects in regulating the gut microbiota and reducing pathogenic bacterial colonisation, particularly demonstrating inhibitory activity against Escherichia coli O157. Furthermore, its efficacy in ameliorating intestinal inflammation and enhancing barrier function has been validated. Nevertheless, further investigations are required to elucidate its precise mechanisms and explore its potential applications against other pathogenic bacterial infections. Future investigations may elucidate its comprehensive role in gut health by exploring synergistic interactions between 2′-FL and other prebiotics and probiotics. Reference [1] Wang Y , Zou Y , Wang J ,et al.The Protective Effects of 2′-Fucosyllactose against E. Coli O157 Infection Are Mediated by the Regulation of Gut Microbiota and the Inhibition of Pathogen Adhesion[J].Nutrients, 2020, 12(5):1284.DOI:10.3390/nu12051284.

Background 2′-Fucosyllactose (2′-FL) is a human milk oligosaccharide (HMO) abundant in breast milk. Human milk oligosaccharides are recognised for their prebiotic and immunomodulatory effects, influencing infant gut microbiota and enhancing immune system function. This study aims to investigate the immunomodulatory and prebiotic effects of 2′-FL in lactating rats, elucidating its potential health benefits by analysing its impact on microbial communities, the immune system, and metabolic products. Study Design and Methods This study employed a randomised controlled trial design, with newborn Wistar rats randomly assigned to a control group and an experimental group. During the lactation period, the control group received standard feed while the experimental group was fed a diet supplemented with 2′-FL. Throughout the study, changes in gut microbiota, immunoglobulin levels, and metabolite concentrations were monitored via faecal sampling and blood analysis. Microbial communities in faecal samples were analysed using 16S rRNA gene sequencing, while serum immunoglobulin content was measured by enzyme-linked immunosorbent assay (ELISA). Key Findings 1. No significant differences in growth and faecal characteristics The study results indicate that supplementation with 2′-FL had no significant effect on body weight gain or faecal characteristics in lactating rats. This suggests that 2′-FL is safe and well-tolerated at the doses used, without inducing adverse physiological responses. 2. Elevated immunoglobulin and cytokine levels Rats supplemented with 2′-FL exhibited reduced levels of highly inflammatory-associated factors such as IL-1β, IL-6, and IFN-γ, demonstrating 2′-FL's anti-inflammatory potential. Furthermore, animals supplemented with 2′-FL exhibited elevated levels of circulating immunoglobulin G (IgG) and immunoglobulin A (IgA), alongside enhanced proliferation of B cells, T cells, and macrophages within the spleen. This further indicates 2′-FL's positive role in immune system development. Figure 1: Effects of 2′-FL on Gut Gene Expression and Cytokines (A) Day 8: Quantitative assessment via real-time PCR of IgA secretion, Toll-like receptors, cytokines, epithelial barrier integrity, and maturation (B) Day 8: Pooled quantification of cytokines (IL-1β, IL-4, IL-6, IL-10, IL-12, IFN-γ, TNF-α) in intestinal lavage fluid from animals (C) Day 16: Quantification of cytokine (IL-1β, IL-4, IL-6, IL-10, IL-12, IFN-γ, TNF-α) levels in pooled intestinal lavage fluid from animals Figure 2: Effect of 2′-FL supplementation on plasma immunoglobulin levels 3. Improved intestinal tissue morphology Administration of 2′-FL demonstrated enteral nutritional effects in the small intestine, such as increased villus height and surface area, indicating that 2′-FL contributes to maintaining intestinal health. Figure 3: Effects of 2′-FL on intestinal tissue morphology at day 8 (A) Representative images of distal jejunal sections stained with haematoxylin and eosin, 100× (B) Villus height, villus width, villus area, crypt depth, villus height/crypt depth ratio, and jejunal circumference in suckling mice 4. Significant alterations in faecal microbiota composition Rats in the 2′-FL group exhibited marked changes in faecal microbiota composition, particularly at the phylum, family and genus levels. This demonstrated increased abundance of beneficial bacteria such as Bifidobacterium and Lactobacillus, alongside a reduction in potential pathogens. This indicates that 2′-FL exerts a positive effect on maintaining intestinal health. Figure 4: Effects of 2′-FL on faecal microbiota composition on day 8 (A) Relative proportions of bacteria at phylum, family, genus and species levels (B) Principal component analysis (PCA) of bacterial abundance at family level (C) Venn diagram showing the number of families and genera in the REF and 2′-FL groups 5. Elevated levels of short-chain fatty acids in urinary metabolism Urine metabolomics analysis revealed that rats in the 2′-FL group exhibited higher concentrations of short-chain fatty acids—such as acetate, propionate, and butyrate—metabolic products closely associated with gut health. Discussion and Outlook This study demonstrates that 2′-FL exhibits significant immunomodulatory and prebiotic effects in lactating rats. By regulating the gut microbiota, increasing immunoglobulin levels and short-chain fatty acid concentrations, 2′-FL improves intestinal health and enhances immune function. Future research may further explore the mechanisms of 2′-FL in other animal models and humans, particularly its potential in preventing and treating intestinal disorders. Moreover, as a functional food ingredient, 2′-FL holds broad application prospects and warrants in-depth development within the food and health supplement sectors. Reference [1] Ignasi Azagra-Boronat, M. J. et al. (2019). Immunomodulatory and Prebiotic Effects of 2′-Fucosyllactose in Suckling Rats. Frontiers in Immunology, 10, 1773. doi:10.3389/fimmu.2019.01773

Background Staphylococcus aureus (SA) and Pseudomonas aeruginosa (PA) are two common pathogenic bacteria. Staphylococcus aureus can cause various infections, including skin infections, food poisoning, and more severe systemic infections, while Pseudomonas aeruginosa frequently causes respiratory tract infections, wound infections, urinary tract infections, and sepsis in immunocompromised patients. Pseudomonas aeruginosa's high resistance to antibiotics renders treating infections more complex and challenging. This study aims to investigate the effects of 2′-fucosyllactose (2′-FL) on the growth of both pathogens and its influence on phagocytosis by polymorphonuclear neutrophils (PMNs), a crucial type of immune cell. Research Design and Methods In this study, PMNs were isolated from the peripheral blood of healthy volunteers and incubated with 2′-FL (0.5–2.5%) before co-incubation with green fluorescent protein (GFP)-labelled SA or PA for 60 hours. Bacterial growth was assessed by measuring GFP relative fluorescence units (GFP-RFU), and the inhibitory effect of 2′-FL on bacterial growth was evaluated by calculating the lag phase duration. Concurrently, the number of viable bacterial colonies was observed to evaluate growth status. Key Findings 1. Effects of 2′-FL on SA and PA Growth At certain concentrations, 2′-FL significantly inhibited the lag phase duration of SA and PA. Notably, high concentrations of 2′-FL (2.5%) markedly reduced the lag phase duration of both SA and PA (P<0.05). 2′-FL extended the lag phase duration of SA and PA by approximately 2 hours and 1.5 hours respectively, indicating that 2′-FL effectively suppresses the early growth stages of these pathogens. Figure 1: Effect of 2′-FL on SA growth Figure 2: Effect of 2′-FL on PA growth 2. Effect of 2′-FL on CFU Counts of Two Pathogenic Bacteria 2′-FL demonstrated significant efficacy in reducing CFU counts of both pathogenic bacteria. Compared to the control group, the 2′-FL-treated groups exhibited markedly reduced CFU counts for both SA and PA, indicating that 2′-FL not only delays bacterial growth but also diminishes their active reproductive capacity. Specifically, a 2.5% concentration of 2′-FL reduced the CFU count of SA by approximately 60% and that of PA by approximately 50%. Figure 3: CFU after 60 hours of incubation with bacteria, PMNs and 2′-FL 3. Enhancement of PMN Phagocytic Capacity by 2′-FL In the presence of PMNs, 2′-FL further enhanced the phagocytic capacity of PMNs towards bacteria. Experiments demonstrated that 2′-FL-treated PMNs exhibited significantly increased phagocytic activity towards SA and PA. This enhancing effect may be attributable to 2′-FL's capacity to stimulate PMN activation, thereby increasing their recognition and phagocytic efficiency towards pathogens. Figure 4: Effect of 2′-FL on the phagocytosis of bacterial GFP-SA by PMNs Figure 5: Effect of 2′-FL on the phagocytosis of bacterial GFP-PA by PMNs Discussion and Outlook Research indicates that 2′-FL can inhibit the growth of pathogenic bacteria SA and PA while enhancing the phagocytic capacity of PMNs, a crucial type of immune cell. These findings provide a foundation for further investigationthe application of human milk oligosaccharides in preventing and treating bacterial infections. Moreover, human milk oligosaccharides may hold potential clinical value in enhancing immune function and preventing pathogen infection. Future research should further investigate its role and mechanisms across different infection models to validate the feasibility and efficacy of its clinical application. Reference [1] Mortaz, E., Nomani, M., Adcock, I., Folkerts, G., & Garssen, J. (2022). Galactooligosaccharides and 2′-Fucosyllactose can directly suppress growth of specific pathogenic microbes and affect phagocytosis of neutrophils. Nutrition, 96, 111601. https://doi.org/10.1016/j.nut.2022.111601