Objectives: The aim of this study was to establish a mother and child cohort in the Chinese population and to investigate human milk (HM) composition and its relationship with infant growth and development during the first three months of life.
Methods: 110 Chinese mother and infant pairs were included in this prospective cohort. Changes of total energy, total fat, total protein, true protein, carbohydrate and osteopontin (OPN) in milk of Chinese mothers at one (T1), two (T2), three (T3) months lactation were analyzed. HM fatty acid (FA) profiles were measured by GC and HM proteomic profiling was conducted by matrix-assisted laser desorption ionization (MALDI), time-of-flight (TOF) mass spectrometry (MS). BMI of the mothers and infant growth indicators, such as weight, length, BMI and head circumference were also recorded at three time points.
Results: Total energy, total fat, total protein, true protein and OPN levels significantly decreased during the first three month of lactation (P < 0.05). Similarly, medium and long-chain saturated FA, including C13:0, C16:0, C20:0, C22:0 and C24:0, and n-6 polyunsaturated (PU) FA including C20:3n6 and C20:4n6, and n-3 PUFA, including C18:3n3, C20:3n3 and n6/n3 ration all significantly decreased over time (P < 0.05). Conversely, short-chain fatty acids, such as butyrate and C6:0, increased during the first three months (P <0.05). HM proteomic analyses distinguished protein composition over time (P = 0.001). Personalized analyses demonstrated that the HM of high-BMI (BMI>25) mothers presented increased total fat, total protein and total energy at T1 and/or T3, and increased OPN at T3 when compared with the normal-BMI (18<BMI<20) mothers (p < 0.05). Similarly, the content of n6 PUFA including C18:3n6 at T1, C20:3n6 at T1-T3 and n6/n3 ratio at T3 were significantly higher in high-BMI mother’s milk. However, the content of MUFA, mainly C18:1 was significantly higher in low-BMI mother’s milk. In addition, BMI of the mothers was positively correlated with the specific FA C20:3n6 (P <0.05,r = 0.27, 0.34, 0.36 respectively) as well as the head circumference (HC)Â of infants (P <0.05,r = 0.31, 0.33, 0.20 respectively) over the three time points.
Conclusion: This study showed that HM changes over time and many of the studied components decreased in concentration during the first three months of lactation. It also concluded that maternal postpartum BMI can influence the FA profile of HM and HC of the infants. This study provides more evidence to the Chinese breast-milk database and further knowledge of HM FA function to support future strategies for the health growth and development of Chinese infants.
Feitong Liu, Xuyi Peng, Jie Li, Juchun Chen, Shuyuan Yan, Jonathan Lane, Patrice Malard
H&H Group Global Research and Technology Center, Guangzhou, China; School of Food Science, South China Agriculture University; Division of Laboratory Medicine, Zhujiang Hospital, Southern Medical University; Child Health Care Center, Changsha Hospital for Maternal and Child Care, Changsha, China
Objectives and Study: Bifidobacteria are associated with positive effects on human health. The influence of human milk oligosaccharides (HMO) on the growth of bifidobacteria is of increasing interest; however, the mechanisms involved in carbohydrate metabolism in a bifidobacterial community are not clearly understood. The aim of this study was to characterize essential gene and protein data sets involved in carbohydrate utilization in a community of four commercial infant Bifidobacterium strains.
Methods: PacBio SMRT sequencing was used to determine the whole genome sequences of all commercial probiotic strains including Bifidobacterium longum subsp. infantis (x2), Bifidobacterium bifidum and Bifidobacterium breve. A subsystem-based comparative genomics approach was used to reconstruct carbohydrate utilization pathways and identify enzymes, transporters and transcriptional regulators related to assimilation of HMO. In order to validate the HMO degradation pathways identified, Bifidobacterium strains were grown in combination in the presence of HMO isolated from human milk. Proteins expressed during growth on HMO were quantified using label-free proteomic analysis via a Q-Exactive mass spectrometer.
Results: Whole genome sequencing of the four Bifidobacterium strains revealed diverse genomic architecture enriched in carbohydrate metabolism genes, which was distinct between species, with some notable differences even amongst the B. infantis strains. One of the B. infantis strains contained the largest proportion of carbohydrate metabolism features (16.4%), while the B. bifidum strain had the lowest proportion of features dedicated to carbohydrate metabolism (11.3%). All strains contained genes encoding for enzymes involved in the GNB/LNB pathway. The B. breve strain, unlike the other strains, lacked genes encoding glycosyl hydrolases (GHs) necessary for hydrolysis of complex HMO structures. Proteome changes during exposure of these strains to HMO were quantified and 235 proteins were detected at higher abundance versus a lactose control. The largest proportions of differentially abundant proteins were involved in carbohydrate metabolic pathways (20%) or transport/localization processes (13%). In B. bifidum, GH proteins involved in assimilation of HMO were found in higher abundances including fucosidases, sialidase, β-galactosidase, and β-N-acetylhexosaminidase. Increased expression of proteins involved in the B. infantis ‘HMO Island’ was also observed, including higher abundances of solute-binding proteins, transporters, a fucosidase, and a β-galactosidase. A >10 fold increase in abundance of lacto-N-phosphorylase in B. breve was observed during growth on HMO, signifying preferential consumption of HMO-derivatives released by other strains within the community.
Conclusion: The combined genomic-proteomic approaches described in this study allows for an in-depth understanding of carbohydrate resource sharing relationships that exist between dominant members of the infant gut microbiota. This study provides new insights on diverse carbohydrate utilization networks in Bifidobacteria that can be employed in metabolic modelling, phenotype prediction and the targeted development of novel symbiotic formulations.
Clodagh Walsh, Rebecca A. Owens, Jonathan A. Lane, Francesca Bottacini, Wiley Barton, Paul D. Cotter, Douwe van Sinderen, Rita M. Hickey
Teagasc Food Research Centre, Moorepark, Food Bioscience, Cork, Ireland, National University of Ireland, Maynooth, Human Health Institute, Co. Kildare, Ireland, Health and Happiness Group, Global Research and Technology Center, Cork, Ireland, APC Microbiome Institute, University College Cork, School of Microbiology, Cork, Ireland
Objectives and Study: Bifidobacteria are associated with a host of health benefits and are the most widely used probiotics in humans. These bacteria are highly abundant in the intestinal microbiota of healthy breastfed infants. Infant-associated Bifidobacteria spp. include Bifidobacterium longum subsp. infantis, Bifidobacterium bifidum and Bifidobacterium breve. A key adaptation, facilitating the establishment and dominance of these species in the breastfed infant gut microbiota, is an ability to consume and metabolise specific sugars only found in breastmilk which are indigestible by the infant, known as Human Milk Oligosaccharides (HMO). In this study, we aimed to characterise the metabolic capability and co-operative metabolism of a community of four commercial infant Bifidobacterium strains when grown on HMO.
Methods: Phenotypic profiling was performed on the four-strain combination (Bifidobacterium longum subsp. infantis x 2, Bifidobacterium bifidum and Bifidobacterium breve) when culture on HMO to understand the physiology and mechanisms underlying HMO utilization in a community environment. Metabolism was measured using growth curves, HMO-High Performance Liquid Chromatography (HPLC) depletion and organic acid analysis (combination of Gas Chromatography Mass Spectrometry and HPLC). Each analysis was performed using three different HMO substrates (2′-fucosyllactose alone and oligosaccharides isolated from human milk representing non-secretor and secretor status). Media with no carbohydrate source was used as a control.
Results: The four-strain combination resulted in total Bifidobacterium numbers increasing (>16%, pâ‰¤0.05) in comparison to single strain cultivation. Over a 24 hour period, the relative abundance of B. breve increased by >50% during co-cultivation with the other strains despite demonstrating limited ability to consume HMO in pure culture. Growth of the strains on secretor HMO resulted in an increase in growth of 14% and 64% when compared to non-secretor HMO and 2′-FL, respectively. HPLC analysis revealed that an increasing number of HMO were consumed by the bacteria over time. Metabolomics confirmed the production of formate, acetate, 1,2-propanediol and lactate. An overall increase in health-promoting organic acids was noted during co-cultivation of all four strains, thus, demonstrating that syntrophic relationships between bifidobacterial strains, increases flux along central metabolic pathways.
Conclusion: This data highlights the cooperative nature of bifidobacterial strains which may reflect their behaviour within an infant ecosystem and how sharing HMO resources maximises nutrient consumption from the diet. The synergistic effect of multiple HMO structures and multiple Bifidobacterium strains results in higher cell numbers and a healthier balance of metabolites, which may have potential health implications for the host. We propose that this social behaviour adds to the diversity and dominance of Bifidobacterium in early life and highlights opportunities for development of new diet and microbiota based therapies to promote infant health.
Clodagh Walsh, Jonathan A. Lane, Helen Slattery, Douwe van Sinderen, Rita M. Hickey
Teagasc Food Research Centre, Moorepark, Food Bioscience, Cork, Ireland, H&H Group, Global Research and Technology Center, Cork, Ireland, APC Microbiome Institute, University College Cork, School of Microbiology, Cork, Ireland
Objectives and Study: Atopic dermatitis (AD) is a common chronic inflammatory skin disease that is often associated with skin barrier dysfunction leading to a higher frequency of bacterial and viral skin infections. The protective effect of breastfeeding against the development of AD and other allergic diseases is well accepted but not fully understood. In this study, the impact of osteopontin (OPN), a multi-functional glycoprotein found in mothers’ milk, and 2´-fucosyllactose (2′-FL), the predominant human milk oligosaccharide, alone or in combination, was determined on AD-like symptoms.
Methods: Repeated epicutaneous application of 2,4-dinitrochlorobenzene (DNCB) was performed on the ear and dorsal skin of BALB/c mice to induce AD-like symptoms and skin lesions. Oral administrations of OPN and/or 2′-FL at concentrations commonly associated with mature mothers milk were performed and AD-like symptoms, behavior, water loss, immune cell activation, immunoglobulin and cytokine expression was recorded.
Results: OPN and/or 2′-FL decreased serum IgE levels and limited the infiltration of eosinophils and mast cells to the dermal tissues in the DNCB-induced AD mice. Furthermore, OPN and 2′-FL reduced Th2 and Th17 responses, leading to an attenuated cutaneous inflammatory response. Interesting, combinations of OPN and 2-FL had a more pronounced effect on IgE expression, eosinophil and Th cell response when compared to the use of OPN or 2-FL alone.
Conclusion: It could be concluded that OPN and 2′-FL synergistically attenuate DNCB-induced AD-like skin lesion in a murine model through modulating T cell-elicited immune responses and CD4+ T cell polarization.
Xi Chen, Jiaqi Zeng, Jonathan A. Lane, Rush Deng, Patrice Malard, Daming Zuo
Southern Medical University, Department of Immunology School of Basic Medical Sciences, Guangzhou, China, 2H&H Group, Global Research and Technology Center, Cork, Ireland, H&H Group, Global Research and Technology Center, Guangzhou, China