Human Milk Glycans Research Articles

2'-Fucosyllactose Inhibits Human Norovirus Replication in Human Intestinal Enteroids.

Human noroviruses (HuNoVs) are the leading cause of acute gastroenteritis worldwide. Currently, there are no targeted antivirals for the treatment of HuNoV infection. Histo-blood group antigens (HBGAs) on the intestinal epithelium are cellular attachment factors for HuNoVs; molecules that block the binding of HuNoVs to HBGAs thus have the potential to be developed as antivirals. Human milk oligosaccharides (HMOs) are glycans in human milk with structures analogous to HBGAs. HMOs have been shown to act as decoy receptors to prevent the attachment of multiple enteric pathogens to host cells. Previous X-ray crystallography studies have demonstrated the binding of HMO 2'-fucosyllactose (2'FL) in the same pocket as HBGAs for some HuNoV strains. We evaluated the effect of 2'FL on the replication of a globally dominant GII.4 Sydney [P16] HuNoV strain using human intestinal enteroids (HIEs) from adults and children. A significant reduction in GII.4 Sydney [P16] replication was seen in duodenal and jejunal HIEs from multiple adult donors, all segments of the small intestine from an adult organ donor and in two pediatric duodenal HIEs. However, 2'FL did not inhibit HuNoV replication in two infant jejunal HIEs that had significantly lower expression of α1-2-fucosylated glycans. 2'FL can be synthesized in large scale, and safety and tolerance have been assessed previously. Our data suggest that 2'FL has the potential to be developed as a therapeutic for HuNoV gastroenteritis.

O-linked glycosylations in human milk casein and major whey proteins during lactation

As glycosylations are difficult to analyze, their roles and effects are poorly understood. Glycosylations in human milk (HM) differ across lactation. Glycosylations can be involved in antimicrobial activities and may serve as food for beneficial microorganisms. This study aimed to identify and analyze O-linked glycans in HM by high-throughput mass spectrometry. 184 longitudinal HM samples from 66 donors from day 3 and months 1, 2, and 3 postpartum were subjected to a post-translational modification specific enrichment-based strategy using TiO2 and ZrO2 beads for O-linked glycopeptide enrichment. β-CN was found to be a major O-linked glycoprotein, additionally, αS1-CN, κ-CN, lactotransferrin, and albumin also contained O-linked glycans. As glycosyltransferases and glycosidases are involved in assembling the glycans including O-linked glycosylations, these were further investigated. Some glycosyltransferases and glycosidases were found to be significantly decreasing through lactation, including two O-linked glycan initiator enzymes (GLNT1 and GLNT2). Despite their decrease, the overall level of O-linked glycans remained stable in HM over lactation. Three different motifs for O-linked glycosylation were enriched in HM proteins: Gly-Xxx-Xxx-Gly-Ser/Thr, Arg-Ser/Thr and Lys-Ser/Thr. Further O-linked glycan motifs on β-CN were observed to differ between intact proteins and endogenous peptides in HM.

Factors Associated with Exclusive Breastfeeding at the Bulili Health Center, South Bureaubuli Village, Palu City

Exclusive breastfeeding is the best nutrient for babies. The impacts of not receiving exclusive breastfeeding are infection, high risk of infectious diseases such as diarrhea and pneumonia, low immune system, and decreased brain development. breastfeeding reduces baby and toddler morbidity and mortality. Human milk glycans, which contain oligosaccharides in both their free and conjugated forms, are demonstrated in studies to be a component of a natural immune process that explains how human milk protects breastfed children from diarrheal illness. This study aims to analyze factors associated with exclusive breastfeeding in the work area of Bulili Public Health Center South Birobuli Subdistrict Palu. This quantitative research was conducted through a cross-sectional approach. This study involved 90 mothers with babies aged 7-12 months, which was carried out from March 2023. The sample was selected using simple random sampling based on inclusion criteria. Direct interviews were used to collect primary data, and secondary data were obtained from Indonesia's health profile, data from the Palu City Health Office 2021, and data on exclusive breastfeeding. The data were processed using the chi-square statistical test with a significance degree of α = 0.05. The results of the study were knowledge (p=0.018), attitude (p=0.001), interest in formula milk promotion (p=0.018), and family support (p=0,002). In Conclusion, there is a significant association between knowledge, attitude, interest in formula milk promotion, and family support with exclusive breastfeeding practice. It is suggested for mothers to increase knowledge, attitude, and family support by gaining information and using social media appropriately.

Human milk whey glycoprotein N-glycans varied greatly among different maternal secretor status

Human milk glycans are complex carbohydrates, which play a pivotal role in infant health and neonatal development. Maternal secretor status is known to affect free oligosaccharides in milk. Here, the milk N-glycome of secretor (Se+) and nonsecretor (Se−) individuals was qualitatively and quantitatively analyzed by hydrophilic interaction chromatography-electrospray ionization-tandem mass spectrometry. The total glycosylation, fucosylation, and sialylation of N-glycans was three times higher in the Se+ group compared to the Se− group (p < 0.001) per equal volume of milk. Importantly, 52 out of 63 N-glycans—including the eight most abundant ones—differed greatly between Se+ and Se− individuals (p < 0.05). Moreover, nine N-glycans (H5N3F1, H6N3, H3N5F1, H5N5F1, H5N5F1S1, H5N4F3S1, H6N4F2S1, H6N5F4S1, and H8N7S1) were >10 times more abundant in Se+ milk than in Se− milk. These findings lay a glycomics-basis for designing personalized nutrition supplements for infants.

Enzymatic production of a suite of human milk oligosaccharides directly in milk

Human milk oligosaccharides (HMOs) denote specific glycans in human breast milk. They function as prebiotics, immune modulating, and antimicrobial agents in the gut of breastfed infants, and certain HMOs even promote the cognitive development of the baby. HMOs are virtually absent in cow’s milk and hence in infant formula, which provides a huge incentive for identifying ways in which HMOs can be produced to improve infant formulas. Here, we show that different sialylated and fucosylated HMOs can be generated in cow’s milk via different simultaneous enzymatic transglycosylation reactions catalyzed by an engineered sialidase (EC 3.2.1.18, from Trypanosoma rangeli) and an 1,2-α-L-fucosidase (EC 3.2.1.63, from Tannerella forsinthia) acting on the lactose in the milk and on casein glycomacropeptide, two types of commercially available HMOs, i.e. 2′-fucosyllactose and lacto-N-neotetraose, added to the milk. We also outline the details of the individual reactions in aqueous systems, demonstrate that the enzymatic reactions can be accomplished at 5 °C, and validate the products formed by LC-MS and NMR analysis. Enzymatic production of HMOs directly in milk provides opportunities for enriching milk and infant formulas and extends the use of enzymatic transglycosylation reactions to synthesis of HMOs in milk and eventually in other beverages.

Variations in Oligosaccharides and N/O-Glycans in Human Milk through the Eight-Month Lactation Period.

Oligosaccharides and N/O-glycans are abundant in human milk and have numerous biological functions (for instance sialylated glycans provide sialic acid for the growth of infant brains), but their variation trends during lactation need further exploration. Qualitative and quantitative analyses of oligosaccharides and N/O-glycans in human milk at different lactation stages (from 7 days to 8 months) were performed using UHPLC-ESI-MS/MS. Thirty-four oligosaccharides, twenty-three N-glycans, and six O-glycans were identified. Oligosaccharides showed the highest abundance in human colostrum and decreased with the progression of lactation, and the abundance of N/O-glycans fluctuated as lactation progressed, while a high abundance of sialylated oligosaccharides and sialylated N/O-glycans was observed in human colostrum. These findings provide evidence for breastfeeding support and contribute to the development of infant formula supplemented with human milk glycans.

Effective Separation of Human Milk Glycosides using Carbon Dioxide Supercritical Fluid Chromatography.

Carbohydrate purification remains problematic due to the intrinsic diversity of structural isomers present in nature. Although liquid chromatography-based techniques are suitable for analyzing or preparing most glycan structures acquired either from natural sources or through chemical or enzymatic synthesis, the separation of regioisomers or linkage isomers with a clear resolution remains challenging. Herein, a carbon dioxide supercritical fluid chromatography (SFC) method was devised to resolve 18 human milk glycosides: oligomers (disaccharides to hexasaccharides), fucosylated regioisomers (lacto-N-fucopentaose I, III, and V; lacto-N-neofucopentaose V; lacto-N-difucohexaose III; blood group H1 antigen; and TF-LNnT), and connectivity isomers (lacto-N-tetraose/lacto-N-neotetraose and para-lacto-N-hexaose/para-lacto-N-neohexaose/type-1 hexasaccharide). The analysis of these glycosides represents a major limitation associated with conventional carbohydrate analysis. The unprecedented resolution achieved by the SFC method indicates the suitability of this key technology for revealing complex human milk glycomes.

Infant gut microbiota modulation by human milk disaccharides in humanized microbiome mice

ABSTRACT Human milk glycans present a unique diversity of structures that suggest different mechanisms by which they may affect the infant microbiome development. A humanized mouse model generated by infant fecal transplantation was utilized here to evaluate the impact of fucosyl-α1,3-GlcNAc (3FN), fucosyl-α1,6-GlcNAc, lacto-N-biose (LNB) and galacto-N-biose on the fecal microbiota and host–microbiota interactions. 16S rRNA amplicon sequencing showed that certain bacterial genera significantly increased (Ruminococcus and Oscillospira) or decreased (Eubacterium and Clostridium) in all disaccharide-supplemented groups. Interestingly, cluster analysis differentiates the consumption of fucosyl-oligosaccharides from galactosyl-oligosaccharides, highlighting the disappearance of Akkermansia genus in both fucosyl-oligosaccharides. An increment of the relative abundance of Coprococcus genus was only observed with 3FN. As well, LNB significantly increased the relative abundance of Bifidobacterium, whereas the absolute levels of this genus, as measured by quantitative real-time PCR, did not significantly increase. OTUs corresponding to the species Bifidobacterium longum, Bifidobacterium adolescentis and Ruminococcus gnavus were not present in the control after the 3-week intervention, but were shared among the donor and specific disaccharide groups, indicating that their survival is dependent on disaccharide supplementation. The 3FN-feeding group showed increased levels of butyrate and acetate in the colon, and decreased levels of serum HDL-cholesterol. 3FN also down-regulated the pro-inflammatory cytokine TNF-α and up-regulated the anti-inflammatory cytokines IL-10 and IL-13, and the Toll-like receptor 2 in the large intestine tissue. The present study revealed that the four disaccharides show efficacy in producing beneficial compositional shifts of the gut microbiota and in addition, the 3FN demonstrated physiological and immunomodulatory roles.

Associations between human milk oligosaccharides (HMOs) and eating behaviour in Hispanic infants at 1 and 6 months of age

Human milk oligosaccharides (HMOs) are naturally occurring glycans in human breast milk that act as prebiotics in the infant gut. Prebiotics have been demonstrated to suppress appetite in both adults and children. Therefore, HMOs may affect infant eating behaviour. To determine if HMOs in breast milk are associated with eating behaviour in Hispanic infants. Cross-sectional analysis of a prospective cohort of Hispanic mother-infant dyads (1-month, n = 157; 6-months, n = 69). Breast milk samples were screened for 19 HMOs using high pressure liquid chromatography, and eating behaviour was assessed using the Baby Eating Behaviour Questionnaire (BEBQ). We conducted multiple linear regressions to examine associations between HMOs and BEBQ scores, adjusted for maternal pre-pregnancy BMI, infant sex, birthweight, delivery mode and number of breastfeedings per day. We stratified by HMO secretor status-a genetic determinant of the types of HMOs produced. At 1 month, LNnT (lacto-N-neotetraose; P = .04) was negatively associated with food responsiveness in the total sample, while DFLNT (difucosyllacto-N-tetrose; P = .03) and DSLNT (disialyl-LNT; P = .04) were negatively associated with food responsiveness in secretors only. At 6 months, LSTc (sialyllacto-N-tetraose c; P = .01), FLNH (fucosyllacto-N-hexaose; P = .03), LNH (lacto-N-hexaose; P = .006) and DSLNH (disialyllacto-N-hexaose; P = .05) were positively associated with food responsiveness in both the total sample and secretors only. We found several HMOs that were both positively and negatively associated with infant food responsiveness, which is a measure of drive to eat.

Structural basis for broad substrate specificity of UDP-glucose 4-epimerase in the human milk oligosaccharide catabolic pathway of Bifidobacterium longum

Infant gut-associated bifidobacteria has a metabolic pathway that specifically utilizes lacto-N-biose I (Gal-β1,3-GlcNAc) and galacto-N-biose (Gal-β1,3-GalNAc) from human milk and mucin glycans. UDP-glucose 4-epimerase (GalE) from Bifidobacterium longum (bGalE) catalyzes epimerization reactions of UDP-Gal into UDP-Glc and UDP-GalNAc into UDP-GlcNAc with the same level of activity that is required to send galacto-hexoses into glycolysis. Here, we determined the crystal structures of bGalE in three ternary complex forms: NAD+/UDP, NAD+/UDP-GlcNAc, and NAD+/UDP-Glc. The broad specificity of bGalE was explained by structural features of the binding pocket for the N-acetyl or C2 hydroxy group of the substrate. Asn200 is located in a pocket of the C2 group, and its side chain adopts different conformations in the complex structures with UDP-Glc and UDP-GlcNAc. On the other side, Cys299 forms a large pocket for the C5 sugar ring atom. The flexible C2 pocket and the large C5 pocket of bGalE are suitable for accommodating both the hydroxy and N-acetyl groups of the substrate during sugar ring rotation in the catalytic cycle. The substrate specificity and active site structure of bGalE were distinct from those of Esherichia coli GalE but similar to those of human GalE.

Fucosylated Human Milk Oligosaccharides and N-Glycans in the Milk of Chinese Mothers Regulate the Gut Microbiome of Their Breast-Fed Infants during Different Lactation Stages.

The milk glycobiome has a significant impact on the gut microbiota of infants, which plays a pivotal role in health and development. Fucosylated human milk oligosaccharides (HMOs) and N-glycans on milk proteins are beneficial for the development of healthy gut microbiota, and the fucosylation levels of these glycans can be affected by the maternal fucosyltransferase 2 gene (FUT2). Here, we present results of longitudinal research on paired milk and stool samples from 56 Chinese mothers (CMs) and their breast-fed children. Changes of HMOs and fucosylated N-glycans in milk of CMs at different lactation stages were detected, which allowed characterization of the major differences in milk glycans and consequential effects on the gut microbiome of infants according to maternal FUT2 status. Significant differences in the abundance of total and fucosylated HMOs between secretor and nonsecretor CMs were noted, especially during early lactation. Despite a tendency toward decreasing milk protein concentrations, the fucosylation levels of milk N-glycans increased during late lactation. The changes in the levels of fucosylated HMOs and milk N-glycans were highly correlated with the growth of Bifidobacterium spp. and Lactobacillus spp. in the gut of infants during early and later lactation, respectively. Enriched expression of genes encoding glycoside hydrolases, glycosyl transferases, ATP-binding cassette (ABC) transporters, and permeases in infants fed by secretor CMs contributed to the promotion of these bacteria in infants. Our data highlight the important role of fucosylated milk glycans in shaping the gut microbiome of infants and provide a solid foundation for development of "personalized" nutrition for Chinese infants. IMPORTANCE Human milk glycans provide a broad range of carbon sources for gut microbes in infants. Levels of protein glycosylation in human milk vary during lactation and may also be affected by the stages of gestation and lactation and by the secretor status of the mother. This was the first study to evaluate systematically dynamic changes in human milk oligosaccharides and fucosylated N-glycans in the milk of Chinese mothers with different secretor statuses during 6 months of lactation. Given the unique single nucleotide polymorphism site (rs1047781, A385T) on the fucosyltransferase 2 gene among Chinese populations, our report provides a specific insight into the milk glycobiome of Chinese mothers, which may exert effects on the gut microbiota of infants that differ from findings from other study cohorts.

Glycosaminoglycanomic profiling of human milk in different stages of lactation by liquid chromatography-tandem mass spectrometry

Glycans in human milk serve several important biological functions that promote infant health. As kind of important glycans, glycosaminoglycans (GAGs) are a complex family of polyanionic carbohydrate, participating in a variety of critical physiological and pathological processes. In this study, the content and the detailed composition of human milk GAGs from Chinese mothers in different stages of lactation, based on a liquid chromatography-tandem mass spectrometry approach was investigated. The results showed that the GAG fraction in the human milk samples was very complex as it was composed of heparan sulfate, chondroitin sulfate, and hyaluronic acid. With lactation extending, the total amount of GAGs in human milk decreased. This study provided an important guide for the demands of GAGs during different stages of lactation. The results were also beneficial for studies on the composition and functional properties of infant formula.

Persistence of Supplemented Bifidobacterium longum subsp. infantis EVC001 in Breastfed Infants.

Attempts to alter intestinal dysbiosis via administration of probiotics have consistently shown that colonization with the administered microbes is transient. This study sought to determine whether provision of an initial course of Bifidobacterium longum subsp. infantis (B.infantis) would lead to persistent colonization of the probiotic organism in breastfed infants. Mothers intending to breastfeed were recruited and provided with lactation support. One group of mothers fed B.infantis EVC001 to their infants from day 7 to day 28 of life (n = 34), and the second group did not administer any probiotic (n = 32). Fecal samples were collected during the first 60postnatal days in both groups. Fecal samples were assessed by 16S rRNA gene sequencing, quantitative PCR, mass spectrometry, and endotoxin measurement. B.infantis-fed infants had significantly higher populations of fecal Bifidobacteriaceae, in particular B.infantis, while EVC001 was fed, and this difference persisted more than 30days after EVC001 supplementation ceased. Fecal milk oligosaccharides were significantly lower in B.infantis EVC001-fed infants, demonstrating higher consumption of human milk oligosaccharides by B. infantis EVC001. Concentrations of acetate and lactate were significantly higher and fecal pH was significantly lower in infants fed EVC001, demonstrating alterations in intestinal fermentation. Infants colonized by Bifidobacteriaceae at high levels had 4-fold-lower fecal endotoxin levels, consistent with observed lower levels of Gram-negative Proteobacteria and Bacteroidetes. IMPORTANCE The gut microbiome in early life plays an important role for long-term health and is shaped in large part by diet. Probiotics may contribute to improvements in health, but they have not been shown to alter the community composition of the gut microbiome. Here, we found that breastfed infants could be stably colonized at high levels by provision of B.infantis EVC001, with significant changes to the overall microbiome composition persisting more than a month later, whether the infants were born vaginally or by caesarean section. This observation is consistent with previous studies demonstrating the capacity of this subspecies to utilize human milk glycans as a nutrient and underscores the importance of pairing a probiotic organism with a specific substrate. Colonization by B.infantis EVC001 resulted in significant changes to fecal microbiome composition and was associated with improvements in fecal biochemistry. The combination of human milk and an infant-associated Bifidobacterium sp. shows, for the first time, that durable changes to the human gut microbiome are possible and are associated with improved gut function.

Maternal inheritance of bifidobacterial communities and bifidophages in infants through vertical transmission

BackgroundThe correct establishment of the human gut microbiota represents a crucial development that commences at birth. Different hypotheses propose that the infant gut microbiota is derived from, among other sources, the mother’s fecal/vaginal microbiota and human milk.ResultsThe composition of bifidobacterial communities of 25 mother-infant pairs was investigated based on an internal transcribed spacer (ITS) approach, combined with cultivation-mediated and genomic analyses. We identified bifidobacterial strains/communities that are shared between mothers and their corresponding newborns. Notably, genomic analyses together with growth profiling assays revealed that bifidobacterial strains that had been isolated from human milk are genetically adapted to utilize human milk glycans. In addition, we identified particular bacteriophages specific of bifidobacterial species that are common in the viromes of mother and corresponding child.ConclusionsThis study highlights the transmission of bifidobacterial communities from the mother to her child and implies human milk as a potential vehicle to facilitate this acquisition. Furthermore, these data represent the first example of maternal inheritance of bifidobacterial phages, also known as bifidophages in infants following a vertical transmission route.

Human Milk Oligosaccharide Specificities of Human Galectins. Comparison of Electrospray Ionization Mass Spectrometry and Glycan Microarray Screening Results.

The affinities of thirty-two free human milk oligosaccharides (HMOs) for four human galectin proteins, a stable mutant of hGal1 (hGal-1), a C-terminal fragment of hGal-3 (hGal-3C), hGal-7, and an N-terminal fragment of hGal-9 (hGal-9N), were measured using electrospray ionization mass spectrometry (ESI-MS). The binding data show that each of the four galectins recognize the majority of the HMOs tested (hGal-1 binds thirty-two HMOs, hGal-3C binds twenty-six, hGal-7 binds thirty-one, and hGal-9N binds twenty-six). Twenty-five of the HMOs tested bind all four galectins, with affinities ranging from 103 to 105 M-1. The reliability of the ESI-MS assay for quantifying the affinities of HMOs for lectins was established from the agreement found between the ESI-MS data and affinities of a small number of HMOs for hGal-1, hGal-3C, and hGal-7 measured by isothermal titration calorimetry (ITC). Comparison of the relative affinities (of 14 HMOs) measured by ESI-MS with the reported specificities of hGal-1, hGal-3, hGal-7, and hGal-9 for these same HMOs established using the shotgun human milk glycan microarray (HM-SGM-v2) showed fair-to-poor correlation, with evidence of false positives and false negatives in the microarray data. The results of this study suggest that HMO specificities of lectins established using microarrays may not accurately reflect their true HMO-binding properties and that the use of "in solution" assays such as ESI-MS and ITC is to be preferred.

Human DC-SIGN binds specific human milk glycans.

Human milk glycans (HMGs) are prebiotics, pathogen receptor decoys and regulators of host physiology and immune responses. Mechanistically, human lectins (glycan-binding proteins, hGBP) expressed by dendritic cells (DCs) are of major interest, as these cells directly contact HMGs. To explore such interactions, we screened many C-type lectins and sialic acid-binding immunoglobulin-like lectins (Siglecs) expressed by DCs for glycan binding on microarrays presenting over 200 HMGs. Unexpectedly, DC-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) showed robust binding to many HMGs, whereas other C-type lectins failed to bind, and Siglec-5 and Siglec-9 showed weak binding to a few glycans. By contrast, most hGBP bound to multiple glycans on other microarrays lacking HMGs. An α-linked fucose residue was characteristic of HMGs bound by DC-SIGN. Binding of DC-SIGN to the simple HMGs 2'-fucosyl-lactose (2'-FL) and 3-fucosyl-lactose (3-FL) was confirmed by flow cytometry to beads conjugated with 2'-FL or 3-FL, as well as the ability of the free glycans to inhibit DC-SIGN binding. 2'-FL had an IC50 of ∼1mM for DC-SIGN, which is within the physiological concentration of 2'-FL in human milk. These results demonstrate that DC-SIGN among the many hGBP expressed by DCs binds to α-fucosylated HMGs, and suggest that such interactions may be important in influencing immune responses in the developing infant.

Galectins are human milk glycan receptors.

The biological recognition of human milk glycans (HMGs) is poorly understood. Because HMGs are rich in galactose we explored whether they might interact with human galectins, which bind galactose-containing glycans and are highly expressed in epithelial cells and other cell types. We screened a number of human galectins for their binding to HMGs on a shotgun glycan microarray consisting of 247 HMGs derived from human milk, as well as to a defined HMG microarray. Recombinant human galectins (hGal)-1, -3, -4, -7, -8 and -9 bound selectively to glycans, with each galectin recognizing a relatively unique binding motif; by contrast hGal-2 did not recognize HMGs, but did bind to the human blood group A Type 2 determinants on other microarrays. Unlike other galectins, hGal-7 preferentially bound to glycans expressing a terminal Type 1 (Galβ1-3GlcNAc) sequence, a motif that had eluded detection on non-HMG glycan microarrays. Interactions with HMGs were confirmed in a solution setting by isothermal titration microcalorimetry and hapten inhibition experiments. These results demonstrate that galectins selectively bind to HMGs and suggest the possibility that galectin-HMG interactions may play a role in infant immunity.

Should Infants Cry Over Spilled Milk? Fecal Glycomics as an Indicator of a Healthy Infant Gut Microbiome

The gut microbiome is rapidly assembled after birth with profound implications for human health and development. Recent research suggests the infant gut microbiome is strongly shaped by microbial exposures at birth followed by a range of other environmental factors, most notably mother’s milk, the one food shared by all humans (1–4). Human milk is an incredibly complex assemblage of nutrients and bioactive factors that evolved under diverse selective pressures to both nourish and protect the infant while simultaneously shaping the infant gut microbiome. Amazingly, the more that is revealed as to the nature of the tripartite relationship between mother’s milk, the infant, and the infant gut microbiome, the more complex the story becomes. For example, one of the most abundant components of milk, the complex assembly of sugars called glycans, are completely indigestible to the infant itself (5). Numerous in vitro studies have shown that these glycan structures are consumed by a relatively small group of early colonizers in the infant gut, primarily of the genera Bifidobacterium (6). However it was unclear if glycan consumption truly takes place inside the infant gut. Is a more saccharolytic microbiota (i.e. high in bifidobacteria) associated with a reduction in milk glycans in feces? Since infants colonized by bifidobacteria show a wealth of favorable health outcomes (7), is there a correlation between their presence and the disappearance of glycans in the feces? Moreover when infants are not colonized by bifidobacteria they often possess a host of other taxa, some of which are less than desirable for infant health (1, 8). Do these latter assemblages consume less milk glycans? In this issue of JGPN, Wang et al (1) helps resolve this issue by showing associations between the milk glycans emerging in the infant feces and various microbial populations in the infant gut. They show that a host of these milk glycans are negatively correlated in abundance with the presence of a suite of microbial taxa such as Bifidobacterium and Bacteroides, known to consume these glycans in vitro. In contrast, microbial populations with less favorable relationships with infant health such as Escherichia and Klebsiella, are not negatively correlated with these human milk glycans, suggesting that they are unable to consume these sugars. However, Wang et al suggest potential cross-feeding among the glycan degraders and these taxa that are not known to consume milk glycans in vitro, pointing to associations between keystone taxa that may be contributing to these populations (1). This work is timely, considering a prior study by De Leoz et al (8) which showed that Bifidobacterium colonization was associated with a concomitant decrease in the presence of milk glycans in the feces of a breast-fed infant while in another infant (not colonized by Bifidobacterium), these structures do not decrease in abundance. These two studies together provide a powerful example of the potential utility of fecal glycomics for assessing the “health” of the developing breast fed infant gut microbiome. If beneficial microbial populations are linked with the presence of specific breast milk glycans and health outcomes, then measurements of these structures could provide a diagnostic readout on infant gut health. Thus the relative absence of these complex milk sugars in the feces would be a measure of a healthy gut microbiome. In contrast, an abundance of ‘spilled milk’ glycans could be a measure of a dysbiotic system. Future work investigating these relationships in larger infant populations is necessary to confirm these trends, but these complementary studies provide great promise in understanding how milk glycans shape the gut microbiome.

A Milk‐Oriented Microbiota (MOM) in Infants: Lessons from Your Other MOM

Human milk contains numerous components that shape the microbial content of the developing infant gastrointestinal tract. A prominent feature of milk is an array of complex free glycans and glycoconjugates that serve a passive immune function by deflecting pathogens while simultaneously enriching a protective microbiota often dominated by bifidobacteria. Human milk glycan structures are also host‐specific and these structural variations differentially influence the composition and function of the microbiota populations enriched. Infant‐borne bifidobacteria utilize human milk glycans as a carbon source for growth however the specific deconstruction mechanisms vary by glycan‐type. These bifidobacteria contain glycosidases and transport systems required to utilize milk glycans either by internalizing large glycans or degrading them extracellularly followed by import and catabolism of specific components. Growth on milk glycans also enhances bifidobacterial interaction with the host through direct and indirect routes—increasing bifidobacterial binding to epithelial cells and producing short chain fatty acids that beneficially modulate host immunity and barrier function. In aggregate, these studies suggest a co‐evolutionary relationship between mammalian milk glycans, infant‐borne bifidobacteria and the infant host that has enabled a programmed enrichment of a protective microbiota during a critical stage of infant development. Importantly, analysis of this natural system serves as a key model for design of probiotic and prebiotic‐based manipulation of the gut microbiota in a range of health settings.

Human Milk Blocks DC-SIGN-Pathogen Interaction via MUC1.

Beneficial effects of breastfeeding are well-recognized and include both immediate neonatal protection against pathogens and long-term protection against allergies and autoimmune diseases. Although several proteins have been identified to have anti-viral or anti-bacterial effects like secretory IgA or lactoferrin, the mechanisms of immune modulation are not fully understood. Recent studies identified important beneficial effects of glycans in human milk, such as those expressed in oligosaccharides or on glycoproteins. Glycans are recognized by the carbohydrate receptors C-type lectins on dendritic cell (DC) and specific tissue macrophages, which exert important functions in immune modulation and immune homeostasis. A well-characterized C-type lectin is dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN), which binds terminal fucose. The present study shows that in human milk, MUC1 is the major milk glycoprotein that binds to the lectin domain of DC-SIGN and prevents pathogen interaction through the presence of Lewis x-type oligosaccharides. Surprisingly, this was specific for human milk, as formula, bovine or camel milk did not show any presence of proteins that interacted with DC-SIGN. The expression of DC-SIGN is found in young infants along the entire gastrointestinal tract. Our data thus suggest the importance of human milk glycoproteins for blocking pathogen interaction to DC in young children. Moreover, a potential benefit of human milk later in life in shaping the infants immune system through DC-SIGN cannot be ruled out.