Fecal microbiota composition of breast-fed infants is correlated with human milk oligosaccharides consumed.
ABSTRACT: This study tested the hypothesis that the fecal bacterial genera of breast-fed (BF) and formula-fed (FF) infants differ and that human milk oligosaccharides (HMOs) modulate the microbiota of BF infants.Fecal samples were obtained from BF (n = 16) or FF (n = 6) infants at 3-month postpartum. Human milk samples were collected on the same day when feces were collected. The microbiota was assessed by pyrosequencing of bacterial 16S ribosomal RNA genes. HMOs were measured by high-performance liquid chromatography-chip time-of-flight mass spectrometry.The overall microbiota of BF differed from that of FF (P = 0.005). Compared with FF, BF had higher relative abundances of Bacteroides, lower proportions of Clostridium XVIII, Lachnospiraceae incertae sedis, Streptococcus, Enterococcus, and Veillonella (P < 0.05). Bifidobacterium predominated in both BF and FF infants, with no difference in abundance between the 2 groups. The most abundant HMOs were lacto-N-tetraose + lacto-N-neotetraose (LNT + LNnT, 22.6%), followed by 2'-fucosyllactose (2'FL, 14.5%) and lacto-N-fucopentaose I (LNFP I, 9.5%). Partial least squares regression of HMO and microbiota showed several infant fecal bacterial genera could be predicted by their mothers' HMO profiles, and the important HMOs for the prediction of bacterial genera were identified by variable importance in the projection scores.These results strengthen the established relation between HMO and the infant microbiota and identify statistical means whereby infant bacterial genera can be predicted by milk HMO. Future studies are needed to validate these findings and determine whether the supplementation of formula with defined HMO could selectively modify the gut microbiota.
Project description:Several factors affect gut microbiota development in early life, among which breastfeeding plays a key role. We followed 24 mother-infant pairs to investigate the associations between concentrations of selected human milk oligosaccharides (HMOs) in breastmilk, infant faeces, and the faecal microbiota composition in healthy, breastfed infants at two, six and 12 weeks of age. Lactation duration had a significant effect on breastmilk HMO content, which decreased with time, except for 3-fucosyllactose (3FL) and Lacto-N-fucopentaose III (LNFP III). We confirmed that microbiota composition was strongly influenced by infant age and was associated with mode of delivery and breastmilk LNFP III concentration at two weeks, with infant sex, delivery mode, and concentrations of 3'sialyllactose (3'SL) in milk at six weeks, and infant sex and Lacto-N-hexaose (LNH) in milk at 12 weeks of age. Correlations between levels of individual breastmilk HMOs and relative abundance of OTUs found in infant faeces, including the most predominant Bifidobacterium OTUs, were weak and varied with age. The faecal concentration of HMOs decreased with age and were strongly and negatively correlated with relative abundance of OTUs within genera Bifidobacterium, Parabacteroides, Escherichia-Shigella, Bacteroides, Actinomyces, Veillonella, Lachnospiraceae Incertae Sedis, and Erysipelotrichaceae Incertae Sedis, indicating the likely importance of these taxa for HMO metabolism in vivo.
Project description:Human milk oligosaccharides (hMOs) are important bioactive components in mother's milk contributing to infant health by supporting colonization and growth of gut microbes. In particular, Bifidobacterium genus is considered to be supported by hMOs. Approximately 200 different hMOs have been discovered and characterized, but only a few abundant hMOs can be produced in sufficient amounts to be applied in infant formula. These hMOs are usually supplied in infant formula as single molecule, and it is unknown which and how individual hMOs support growth of individual gut bacteria. To investigate how individual hMOs influence growth of several relevant intestinal bacteria species, we studied the effects of three hMOs (2'-fucosyllactose, 3-fucosyllactose, and 6'-sialyllactose) and an hMO acid hydrolysate (lacto-N-triose) on three Bifidobacteria and one Faecalibacterium and introduced a co-culture system of two bacterial strains to study possible cross-feeding in presence and absence of hMOs. We observed that in monoculture, Bifidobacterium longum subsp. infantis could grow well on all hMOs but in a structure-dependent way. Faecalibacterium prausnitzii reached a lower cell density on the hMOs in stationary phase compared to glucose, while B. longum subsp. longum and Bifidobacterium adolescentis were not able to grow on the tested hMOs. In a co-culture of B. longum subsp. infantis with F. prausnitzii, different effects were observed with the different hMOs; 6'-sialyllactose, rather than 2'-fucosyllactose, 3-fucosyllactose, and lacto-N-triose, was able to promote the growth of B. longum subsp. infantis. Our observations demonstrate that effects of hMOs on the tested gut microbiota are hMO-specific and provide new means to support growth of these specific beneficial microorganisms in the intestine.
Project description:The infant's gut microbiome is generally rich in the Bifidobacterium genus. The mother's milk contains natural prebiotics, called human milk oligosaccharides (HMOs), as the third most abundant solid component after lactose and lipids, and of the different gut microbes, infant gut-associated bifidobacteria are the most efficient in assimilating HMOs. Indeed, the fecal concentration of HMOs was found to be negatively correlated with the fecal abundance of Bifidobacterium in infants. Given these results, two HMO molecules, 2'-fucosyllactose and lacto-N-neotetraose, have recently been industrialized to fortify formula milk. As of now, however, our knowledge about the HMO consumption pathways in infant gut-associated bifidobacteria is still incomplete. The recent studies indicate that HMO assimilation abilities significantly vary among different Bifidobacterium species and strains. Therefore, to truly maximize the effects of prebiotic and probiotic supplementation in commercialized formula, we need to understand HMO consumption behaviors of bifidobacteria in more detail. In this review, we summarized how different Bifidobacterium species/strains are equipped with varied gene sets required for HMO assimilation. We then examined the correlation between the abundance of the HMO-related genes and bifidobacteria-rich microbiota formation in the infant gut through data mining analysis of a deposited fecal microbiome shotgun sequencing dataset. Finally, we shortly described future perspectives on HMO-related studies.
Project description:Numerous benefits of breastfeeding over infant formula are fully established. The superiority of human milk over bovine milk-based formula is partly due to human milk oligosaccharides (HMOs), a family of over 100 molecules present specifically and substantially in human milk that resemble mucosal glycans. To uncover novel physiological functions and pathways of HMOs, we screened a panel of 165 G-protein coupled receptors (GPCRs) using a blend of 6 HMOs (3'-O-sialyllactose (3'SL), 6'-O-sialyllactose (6'SL), lacto-N-tetraose (LNT), lacto-N-neo-tetraose (LNnT), 2-O-fucosyllactose (2'FL), and difucosyllactose (diFL)), and followed up positive hits with standard receptor assays. The HMO blend specifically activated GPR35. LNT and 6'SL individually activated GPR35, and they showed synergy when used together. In addition, in vitro fermentation of infant stool samples showed that 2'FL upregulates the production of the GPR35 agonist kynurenic acid (KYNA) by the microbiota. LNT?+?6'SL and KYNA showed additive activation of GPR35. Activation by 6'SL and LNT of GPR35, a receptor mediating attenuation of pain and colitis, is to our knowledge the first demonstration of GPCR activation by any HMO. In addition, we demonstrated a remarkable cooperation between nutrition and microbiota towards activation of a host receptor highlighting the close interplay between environment and host-microbe interactions.
Project description:SCOPE:Understanding biological functions of different free human milk oligosaccharides (HMOs) in shaping gastrointestinal tract microbiota during infancy is of great interest. We examined a link between HMOs in maternal milk and infant faecal microbiota composition and investigated the role of microbiota in degrading HMOs within the GI tract of healthy, breastfed, one-month old infants. METHODS AND RESULTS:Maternal breast milk and corresponding infant faeces originated from the KOALA Birth Cohort. HMOs were quantified in milk and infant faecal samples using PGC-UPLC-MS and HPAEC-PAD. Faecal microbiota composition was characterised using Illumina HiSeq amplicon 16S rRNA sequencing. The composition associated with gender, mode of delivery, and milk HMOs: Lacto-N-fucopentaose I and 2'-Fucosyllactose. Overall, Bifidobacterium, Bacteroides, Escherichia-Shigella and Parabacteroides were predominating genera. We detected three different patterns in the infant faecal microbiota structure. Gastrointestinal degradation of HMOs was strongly associated with faecal microbiota composition, and there was a link between utilisation of specific HMOs and abundance of various phylotypes (OTUs). CONCLUSIONS:HMOs in maternal milk are among important factors shaping GI tract microbiota composition in one-month old breastfed infants. Infant's ability to metabolise different HMOs strongly correlate with faecal microbiota composition, and with phylotypes within genera Bifidobacterium, Bacteroides and Lactobacillus. This article is protected by copyright. All rights reserved.
Project description:Human milk oligosaccharides (HMOs) play an important role in the health of an infant as substrate for beneficial gut bacteria. Little is known about the effects of HMO composition and its changes on the morbidity and growth outcomes of infants living in areas with high infection rates. Mother's HMO composition and infant gut microbiota from 33 Gambian mother/infant pairs at 4, 16, and 20 weeks postpartum were analyzed for relationships between HMOs, microbiota, and infant morbidity and growth. The data indicate that lacto-N-fucopentaose I was associated with decreased infant morbidity, and 3'-sialyllactose was found to be a good indicator of infant weight-for-age. Because HMOs, gut microbiota, and infant health are interrelated, the relationship between infant health and their microbiome were analyzed. While bifidobacteria were the dominant genus in the infant gut overall, Dialister and Prevotella were negatively correlated with morbidity, and Bacteroides was increased in infants with abnormal calprotectin. Mothers nursing in the wet season (July to October) produced significantly less oligosaccharides compared to those nursing in the dry season (November to June). These results suggest that specific types and structures of HMOs are sensitive to environmental conditions, protective of morbidity, predictive of growth, and correlated with specific microbiota.
Project description:Newborns are colonized with an intestinal microbiota shortly after birth, but the factors governing the retention and abundance of specific microbial lineages are unknown. Nursing infants consume human milk oligosaccharides (HMOs) that pass undigested to the distal gut, where they may be digested by microbes. We determined that the prominent neonate gut residents, Bacteroides thetaiotaomicron and Bacteroides fragilis, induce the same genes during HMO consumption that are used to harvest host mucus glycans, which are structurally similar to HMOs. Lacto-N-neotetraose, a specific HMO component, selects for HMO-adapted species such as Bifidobacterium infantis, which cannot use mucus, and provides a selective advantage to B. infantis in vivo when biassociated with B. thetaiotaomicron in the gnotobiotic mouse gut. This indicates that the complex oligosaccharide mixture within HMOs attracts both mutualistic mucus-adapted species and HMO-adapted bifidobacteria to the infant intestine that likely facilitate both milk and future solid food digestion.
Project description:Human milk oligosaccharides (HMOs) may provide health benefits to infants partly by shaping the development of the early-life intestinal microbiota. In a randomized double-blinded controlled multicentric clinical trial, healthy term infants received either infant formula (control) or the same formula with two HMOs (2'-fucosyllactose and lacto-N-neotetraose; test) from enrollment (0 to 14?days) to 6?months. Then, all infants received the same follow-up formula without HMOs until 12?months of age. Breastfed infants (BF) served as a reference group. Stool microbiota at 3 and 12?months, analyzed by 16S rRNA gene sequencing, clustered into seven fecal community types (FCTs) with marked differences in total microbial abundances. Three of the four 12-month FCTs were likely precursors of the adult enterotypes. At 3?months, microbiota composition in the test group (n?=?58) appeared closer to that of BF (n?=?35) than control (n?=?63) by microbiota alpha (within group) and beta (between groups) diversity analyses and distribution of FCTs. While bifidobacteriaceae dominated two FCTs, its abundance was significantly higher in one (FCT BiH for Bifidobacteriaceae at high abundance) than in the other (FCT Bi for Bifidobacteriaceae). HMO supplementation increased the number of infants with FCT BiH (predominant in BF) at the expense of FCT Bi (predominant in control). We explored the association of the FCTs with reported morbidities and medication use up to 12?months. Formula-fed infants with FCT BiH at 3?months were significantly less likely to require antibiotics during the first year than those with FCT Bi. Previously reported lower rates of infection-related medication use with HMOs may therefore be linked to gut microbiota community types. (This study has been registered at ClinicalTrials.gov under registration number NCT01715246.)IMPORTANCE Human milk is the sole and recommended nutrition for the newborn infant and contains one of the largest constituents of diverse oligosaccharides, dubbed human milk oligosaccharides (HMOs). Preclinical and clinical association studies indicate that HMOs have multiple physiological functions largely mediated through the establishment of the gut microbiome. Until recently, HMOs were not available to investigate their role in randomized controlled intervention trials. To our knowledge, this is the first report on the effects of 2 HMOs on establishing microbiota in newborn infants. We provide a detailed description of the microbiota changes observed upon feeding a formula with 2 HMOs in comparison to breastfed reference infants' microbiota. Then, we associate the microbiota to long-term health as assessed by prescribed antibiotic use.
Project description:Bifidobacterial carbohydrate metabolism has been studied in considerable detail for a variety of both plant- and human-derived glycans, particularly involving the bifidobacterial prototype strain Bifidobacterium breve UCC2003. We recently elucidated the metabolic pathways by which the human milk oligosaccharide (HMO) constituents lacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT) and lacto-N-biose (LNB) are utilized by B. breve UCC2003. However, to date, no work has been carried out on the regulatory mechanisms that control the expression of the genetic loci involved in these HMO metabolic pathways. In this study, we describe the characterization of three transcriptional regulators and the corresponding operator and associated (inducible) promoter sequences, with the latter governing the transcription of the genetic elements involved in LN(n)T/LNB metabolism. The activity of these regulators is dependent on the release of specific monosaccharides, which are believed to act as allosteric effectors and which are derived from the corresponding HMOs targeted by the particular locus.IMPORTANCE Human milk oligosaccharides (HMOs) are a key factor in the development of the breastfed-infant microbiota. They function as prebiotics, selecting for a specific range of microbes, including a number of infant-associated species of bifidobacteria, which are thought to provide a range of health benefits to the infant host. While much research has been carried out on elucidating the mechanisms of HMO metabolism in infant-associated bifidobacteria, to date there is very little understanding of the transcriptional regulation of these pathways. This study reveals a multicomponent transcriptional regulation system that controls the recently identified pathways of HMO metabolism in the infant-associated Bifidobacterium breve prototype strain UCC2003. This not only provides insight into the regulatory mechanisms present in other infant-associated bifidobacteria but also provides an example of a network of sequential steps regulating microbial carbohydrate metabolism.
Project description:Human milk oligosaccharides (HMOs) are bioactive molecules playing a critical role in infant health. We aimed to quantify the composition of HMOs of women with normal weight (18.5-24.9 kg/m<sup>2</sup>), overweight (25.0-29.9 kg/m<sup>2</sup>), or obesity (30.0-60.0 kg/m<sup>2</sup>) and determine the effect of HMO intake on infant growth. Human milk (HM) samples collected at 2 months (2 M; <i>n</i> = 194) postpartum were analyzed for HMO concentrations via high-performance liquid chromatography. Infant HM intake, anthropometrics and body composition were assessed at 2 M and 6 M postpartum. Linear regressions and linear mixed-effects models were conducted examining the relationships between maternal BMI and HMO composition and HMO intake and infant growth over the first 6 M, respectively. Maternal obesity was associated with lower concentrations of several fucosylated and sialylated HMOs and infants born to women with obesity had lower intakes of these HMOs. Maternal BMI was positively associated with lacto-N-neotetraose, 3-fucosyllactose, 3-sialyllactose and 6-sialyllactose and negatively associated with disialyllacto-N-tetraose, disialyllacto-N-hexaose, fucodisialyllacto-N-hexaose and total acidic HMOs concentrations at 2 M. Infant intakes of 3-fucosyllactose, 3-sialyllactose, 6-sialyllactose, disialyllacto-N-tetraose, disialyllacto-N-hexaose, and total acidic HMOs were positively associated with infant growth over the first 6 M of life. Maternal obesity is associated with changes in HMO concentrations that are associated with infant adiposity.