Composition Of Human Milk Research Articles

Antimicrobial Effect of Zn2+ Ions Governs the Microbial Quality of Donor Human Milk.

Donor human milk (HM) obtained at HM banks is exceptionally crucial for the feeding and treatment of preterm infants. Bacterial contaminations of HM in various stages of its handling are very common and can lead to disqualification of donations or severe infections in worse cases. Hence, HM donations are subject to strict bacteriological evaluations pre- and post-pasteurization. The main contaminating species vary between countries, banks and donors and even exhibit inter-individual variation. We initiated an assessment of the bacteriological composition of HM donated by women hospitalized in a neonatal intensive care unit in Israel. The most common bacterium identified was Staphylococcus epidermidis, found in all but one of the HM samples; the presence of several species of coagulase-negative Staphylococci was also noted. Next, we sought to develop a platform towards antibacterial treatment using Zn2+ ions that have recently been found to be potent against contaminants isolated from bovine milk. Zn2+ efficiently inhibited the growth of viable aerobic population and S. epidermidis in HM. Growth was also inhibited in other Gram-positive bacteria such as Bacillus cereus, a well-known food-borne pathogen. S. epidermidis and B. cereus cells grown in the presence of zinc were taken for microscopic evaluation, aiming to demonstrate zinc’s antimicrobial mode of action morphologically. Images obtained using scanning electron microscopy indicated leakage of cellular content and cell lysis in S. epidermidis. Besides, B. cereus cells showed abnormalities in their cell surface and complete loss of flagella upon treatment with zinc. Along with the above findings, it should be noted that this was a pilot study that tested how high doses of Zn2+ affect breast milk as a product. Further research is likely needed on the safety of consumption of Zn2+-treated HM in infants and older children.

Resolving Human Lactation Heterogeneity Using Single Milk-Derived Cells, a Resource at the Ready.

Single cell RNA sequencing (scRNAseq) of human milk-derived cells (HMDCs) makes highly detailed analyses of the biology of human lactation possible. We explore this powerful application as an exciting tool to inspect the cellular composition of human milk. We point out some important challenges unique to this approach and highlight the importance of collaborations between biologists and well-trained bioinformaticians to utilize these data to their maximum potential. We extend this focus by discussing the first two such studies that describe HMDCs via scRNAseq and a variety of important questions in the field that warrant attention through further research. The stage is set to apply scRNAseq in human lactation biology, potentially leading to new insights regarding the molecular and cellular diversity of human secretory mammary epithelial cells.

The implications of routine milk fortification for the short and long-term health of preterm babies

Fortification refers to the practice of enriching human milk feeds for very preterm babies with macronutrients, minerals and vitamins. Though standard of care in some parts of the world, adoption of fortification is not universal. Fortification entered into use on the assumption that human milk macronutrient content, principally protein, is insufficient to support the growth and development of very preterm babies. However, because of the substantial variability in human milk composition, routine fortification risks exposing some babies to very high protein intakes, which may be dangerous. Some clinicians fear fortification with cow-milk derived products will increase the risk of necrotising enterocolitis, leading them to favour commercial fortifiers made from pooled human milk over cow milk based products, a practice that has additional ethical implications. Randomised controlled trials of multi-nutrient fortification to-date are inadequate. No trial has had power to detect important functional effects; the majority are methodologically weak and focus primarily upon short-term growth. Evidence to guide practice is inadequate. There is an urgent need for collaboration to conduct high-quality research to end these long-standing uncertainties.

Maternal Diet and Infant Feeding Practices Are Associated with Variation in the Human Milk Microbiota at 3 Months Postpartum in a Cohort of Women with High Rates of Gestational Glucose Intolerance

Human milk contains a diverse community of bacteria believed to play a role in breast health and inoculation of the infant's gastrointestinal tract. The role of maternal nutrition and infant feeding practices on the human milk microbiota remains poorly understood. Our aim was to explore the associations between maternal diet (delivery to 3 mo postpartum), infant feeding practices, and the microbial composition and predicted function in milk from women with varied metabolic status. This was an exploratory analysis of a previously completed prospective cohort study of women with varying degrees of gestational glucose intolerance (NCT01405547). Milk samples (n=93 mothers) were collected at 3 mo postpartum. Maternal dietary information (validated food-frequency questionnaire) and infant feeding practices (human milk exclusivity, frequency of direct breastfeeding per day) were collected. V4-16S ribosomal RNA gene sequencing (Illumina MiSeq) was conducted to determine microbiota composition. Intake of polyunsaturated fat [β estimate (SE): 0.036 (0.018), P=0.047] and fiber from grains [0.027 (0.013), P=0.048] were positively associated with ɑ-diversity (Shannon index) of human milk. Overall microbial composition of human milk clustered based on human milk exclusivity (weighted UniFrac R2=0.034, P=0.015; Bray-Curtis R2=0.041, P=0.007), frequency of direct breastfeeding per day (Bray-Curtis R2=0.057, P=0.026), and maternal fiber intake from grains (Bray-Curtis R2=0.055, P=0.040). Total fiber, fiber from grains, dietary fat, and infant feeding practices were also associated with a number of differentially abundant taxa. The overall composition of predicted microbial functions was associated with total fiber consumption (Bray-Curtis R2=0.067, P=0.036) and human milk exclusivity (Bray-Curtis R2=0.041, P=0.013). Maternal consumption of fiber and fat, as well as mother's infant feeding practices, are important determinants of the human milk microbiota. Understanding whether these microbial changes impact an infant's overall health and development requires future study.

Preterm human milk composition and dietary intake of breastfeeding mothers in the Indonesian population

Background Human milk consumption is a protective factor against infection. However, the high variability of human milk composition makes it difficult for clinicians and researchers to assess the adequacy of nutritional intake.
 Objective To identify the nutritional profiles and composition of premature human milk and dietary intake of breastfeeding mother.
 Methods Human milk specimens were collected from mothers who gave birth to very premature and/or very low birth weight infants. Infants had been admitted to the Neonatology Unit of Dr. Cipto Mangunkusumo Hospital, Jakarta. The milk specimens were analyzed with a mid-infrared milk analyzerTM (MIRIS) for four weeks, and maternal dietary macronutrient intake was determined with food record questionnaires on food consumption frequency and food recall.
 Results Significant changes in nutritional composition of human milk were observed, with the fat concentration and calories increasing with time, and protein concentration decreasing with time. There were no significant differences observed in carbohydrate concentration of milk over the four weeks (P=0.447). Maternal intake of protein, carbohydrates, fat and calories was lower than the recommended values for breastfeeding women.
 Conclusion The macronutrient (protein and fat) and energy contents of human milk change weekly. Dietary intake of breastfeeding women are lower than recommended values.

Obesogenic Programming Effects during Lactation: A Narrative Review and Conceptual Model Focusing on Underlying Mechanisms and Promising Future Research Avenues.

Animal studies have consistently demonstrated that maternal obesity and a high-fat diet during lactation enhances obesity risk in the offspring. However, less is known about these potential obesogenic programming effects in obese humans. We propose three important pathways that may explain obesogenic programming effects of human breastmilk. First, human milk components and hormones may directly affect child eating and satiety characteristics. Second, human milk constituents can affect child microbiota that, in turn, may influence child eating and weight outcomes. Third, human milk composition may affect child eating and weight outcomes through flavor exposure. We reviewed a few very recent findings from well-powered longitudinal or experimental human research with regard to these three pathways. Moreover, we provide a research agenda for future intervention research with the overarching aim to prevent excessive pediatric weight gain during lactation and beyond. The ideas presented in this paper may represent important “black box” constructs that explain obesogenic programming effects during lactation. It should be noted, however, that given the scarcity of studies, findings should be seen as working hypotheses to further test in future research.

Tandem Breastfeeding: A Descriptive Analysis of the Nutritional Value of Milk When Feeding a Younger and Older Child.

Breastfeeding is a gold standard of feeding of newborns and infants. Tandem breastfeeding (TBF) is feeding two children of different ages at the same time. The knowledge about the composition of human milk in prolonged lactation is still scarce. Milk from tandem breastfeeding women and after weaning was examined. Milk samples were collected from 13 TBF mothers. A 24-h milk collection was done. Analyses of fat, protein, carbohydrate and energy content were performed using MIRIS. Sociodemographic characteristics of TBF mothers was done. Higher fat content, energy value and total protein concentration was found in TBFM milk during tandem breastfeeding, than in milk after weaning the older child. The carbohydrate content remained stable. The composition of breastmilk, in terms of macronutrients, changes after weaning, taking into account the nutritional requirements of the younger child. The milk of nursing mothers in tandem did not show diurnal variability in individual components. These findings suggest an adaptive role of human milk to nutrient requirements of newborn and older children. The results may support the promotion of long breastfeeding, including tandem breastfeeding.

Comparative Lipidomics Analysis of Human Milk and Infant Formulas Using UHPLC-Q-TOF-MS.

The lipidome and fatty acid composition of human milk and different infant formulas with animal- and/or plant-based fat sources are analyzed and compared in this study. The results obtained using positive and negative ionization modes indicate that there are 48 and 71 lipid species, respectively, that are common between the human milk and infant formulas. Moreover, the fatty acid composition in infant formulas varies significantly, depending on the fat source. Human milk is rich in triacylglycerols that contain linoleic acid, α-linolenic acid, arachidonic acid, and docosahexaenoic acid. Meanwhile, the triacylglycerols in IFB comprise long-chain fatty acids at the sn-1,3 position. Compared to human milk, IFC has the same level of sphingomyelin species. Based on univariate and multivariate analyses, there are 37, 34, 31, and 36 lipid species that can be used to distinguish between human milk and infant formulas. Overall, the results reported herein are useful in designing new milk formulas that better mimic human milk.

Medium- and long-chain triacylglycerols composition in preterm and full-term human milk across different lactation stages

Human milk is a natural source of medium-chain fatty acids (MCFAs). It is generally acknowledged that MCFAs exist in medium-chain triacylglycerols (MCTs). However, a previous study revealed that human milk is naturally rich in medium- and long-chain triacylglycerols (MLCTs). Few data are available on the MLCT composition of preterm milk. This study compared the MCFA and MLCT profiles in human milk from mothers after preterm (28–37 weeks) and term (>37 weeks) delivery during three lactational stages. The results showed that almost half of the triacylglycerol species in human milk are MLCT, accounting for approximately 30% of the total TAGs, mainly composed of one MCFA and two long-chain fatty acids, that is, MLL-type MLCT. In addition, MCFAs and MLCTs showed an increase from colostrum to transitional milk, while they were comparable between preterm and full-term milk. This research could help to develop infant formula, which is of great significance for infants.

Changes in macronutrients of human milk after bolus feeding: a simulation study.

Enteral nutrition supply to preterm infants requires feeding through a feeding tube. The aim of this study was to evaluate changes in macronutrient composition of human milk (HM) while passing through a gastric feeding tube. Simulated real-life practice tube feeding was performed by using an infusion pump connected to a feeding tube. A human milk analyzer was used to compare the pre infusion and post infusion macronutrient contents of HM. There was a significant decrease in fat (from 4.06 ± 0.6 g/100 ml to 3.95 ± 0.6 g/100 ml), carbohydrates (from 7.51 ± 0.4 g/100 ml to 7.33 ± 0.5 g/100 ml), and energy content (from 70.77 ± 5.4 kcal/100 ml to 69.72 ± 0.5 kcal/100 ml) after passing through the feeding tube (P < 0.001). A simulated real-life bolus tube feeding model demonstrated small but significant decreases in fat, carbohydrate, and energy content. The biological significance of our results to the very low birth weight infants should be further studied.

Human milk composition promotes optimal infant growth, development and health

Human milk is a living dynamic fluid that promotes optimal nutrition and development of the infant and impacts health across the lifespan. This review reports on the diverse range of nutrients, immune protection factors, hormones, microbes and metabolites in human milk and their impacts on infant nutrition and health. While many of these components are stable across lactation and similar between women, some vary over time, and in response to maternal and infant health status, maternal diet and geographic location. Human milk may be considered as personalized nutrition, with many components working synergistically to stimulate and support the infant's immature immune system, while enhancing appropriate development, growth and body composition.

Centering human milk composition as normal human biological variation

AbstractObjectivesHuman biological variation in the phenotype is the cornerstone of modern human biology, evolutionary anthropology, and related studies of human evolution. Minimal dialogue, however, has considered human milk to be part of this phenotypic variation. This may reflect researcher bias‐mental models oriented around commercial infant formula and homogenized cow's milk, both of which present milk composition as static. A general lack of research outside primarily Western, well‐nourished populations has also contributed to this underestimation of biological variation.MethodsThis review analyzes published research on breast milk composition, developmental metabolic programming, and maternal body composition to articulate the ways in which population‐based studies of human milk outside the United Sates are necessary to better understanding biological variation in human milk phenotypes.ResultsThis review discusses some of the common issues in current research on the biological variation in human milk composition and argues that anthropological inquiries that frame milk as part of an adaptive phenotype are necessary to better understand the biological significance of human milk composition in the production of human biological variation.ConclusionsBiological anthropology is uniquely positioned to investigate biological variation in human milk, using evolutionary theory, cutting edge biology, and anthropologically informed perspectives that challenge the biomedical framing of lactation and often act to privilege well nourished, primarily western populations and formula feeding as normatives for infant feeding research.

MACRONUTRIENTS OF MOTHERS' MILK OF VERY LOW BIRTH WEIGHT INFANTS: ANALYSIS ACCORDING TO GESTATIONAL AGE AND MATERNAL VARIABLES.

ABSTRACTObjective: To analyze the composition of macronutrients present in the milk of mothers of preterm newborn infants (PTNB) - protein, fat, carbohydrate, and calories - by gestational age (GA), chronological age (CA) and maternal variables.Methods: Longitudinal study that analyzed 215 milk samples from the 51 mothers of PTNB admitted in three Neonatal Intensive Care Units of Rio de Janeiro from May/2013-January/2014. Milk collection was performed by pickup pump, on a fixed day of each week until discharge. The spectrophotometric technique with Infrared Analysis (MilkoScan Minor 104) was used for the quantitative analysis. A sample of 7 mL of human milk was taken from the total volume of milk extracted by the mother. The data was grouped by GA (25-27, 28-31, 32-36, 37-40 weeks) and by CA (zero to 4, 5-8, 9-12, 13-16 weeks).Results: Protein, carbohydrate, fat and calories did not show any pattern of change, with no difference among groups of GA. When the macronutrients were analyzed by groups of CA, protein decreased, with significant difference between the first two groups of CA. Carbohydrates, fat and calories presented increasing values in all groups, without significant differences. Weight gain during pregnancy, maternal hypertension and maternal age were associated with changes in fat and calories in the first moment of the analysis of milk.Conclusions: There was a significant decrease in the levels of protein during the first eight weeks after birth. CA may be an important factor in the composition of human milk.

Customized Human Milk Fortification Based on Measured Human Milk Composition to Improve the Quality of Growth in Very Preterm Infants: A Mixed-Cohort Study Protocol.

Adequate nutrition of very preterm infants comprises fortification of human milk (HM), which helps to improve their nutrition and health. Standard HM fortification involves a fixed dose of a multi-nutrient HM fortifier, regardless of the composition of HM. This fortification method requires regular measurements of HM composition and has been suggested to be a more accurate fortification method. This observational study protocol is designed to assess whether the target HM fortification method (contemporary cohort) improves the energy and macronutrient intakes and the quality of growth of very preterm infants, compared with the previously used standard HM fortification (historical cohorts). In the contemporary cohort, a HM multi-nutrient fortifier and modular supplements of protein and fat are used for HM fortification, and the enteral nutrition recommendations of the European Society for Paediatric Gastroenterology Hepatology and Nutrition for preterm infants will be considered. For both cohorts, the composition of HM is assessed using the Miris Human Milk analyzer (Uppsala, Sweden). The quality of growth will be assessed by in-hospital weight, length, and head circumference growth velocities and a single measurement of adiposity (fat mass percentage and fat mass index) performed just after discharge, using the air displacement plethysmography method (Pea Pod, Cosmed, Italy). ClinicalTrials.gov registration number: NCT04400396.

Analysis of dietary patterns and nutritional adequacy in lactating women: a multicentre European cohort (ATLAS study).

Eating habits of lactating women can influence the nutrient composition of human milk, which in turn influences nutrient intake of breastfed infants. The aim of the present study was to identify food patterns and nutritional adequacy among lactating women in Europe. Data from a multicentre European longitudinal cohort (ATLAS study) were analysed to identify dietary patterns using cluster analysis. Dietary information from 180 lactating women was obtained using 3-d food diaries over the first 4 months of lactation. Four dietary patterns were identified: 'vege-oils', 'fish-poultry', 'confectionery-salads' and 'mixed dishes'. Nutrition adequacy was not significantly different between clusters, but the 'vege-oils' cluster tended to yield the highest nutrition adequacy measured by Mean Adequacy Ratio. Compared with European dietary reference values (DRVs) for lactating women, women in all clusters had inadequate intakes of energy, pantothenic acid, folate, vitamin C, vitamin A, vitamin D, zinc, iodine, potassium and linoleic acid. Adequate intake for fibre and α-linolenic acid was only achieved in the 'vege-oils' cluster. Overall, fat intake was above DRVs. The present study showed that various dietary patterns do not adequately supply all nutrients, indicating a need to promote overall healthy dietary habits for European lactating women.

Breastfeeding and the origins of health: Interdisciplinary perspectives and priorities.

Breastfeeding and human milk (HM) are critically important to maternal, infant and population health. This paper summarizes the proceedings of a workshop that convened a multidisciplinary panel of researchers to identify key priorities and anticipated breakthroughs in breastfeeding and HM research, discuss perceived barriers and challenges to achieving these breakthroughs and propose a constructive action plan to maximize the impact of future research in this field. Priority research areas identified were as follows: (1) addressing low breastfeeding rates and inequities using mixed methods, community partnerships and implementation science approaches; (2) improving awareness of evidence‐based benefits, challenges and complexities of breastfeeding and HM among health practitioners and the public; (3) identifying differential impacts of alternative modes of HM feeding including expressed/pumped milk, donor milk and shared milk; and (4) developing a mechanistic understanding of the health effects of breastfeeding and the contributors to HM composition and variability. Key barriers and challenges included (1) overcoming methodological limitations of epidemiological breastfeeding research and mechanistic HM research; (2) counteracting ‘breastfeeding denialism’ arising from negative personal breastfeeding experiences; (3) distinguishing and aligning research and advocacy efforts; and (4) managing real and perceived conflicts of interest. To advance research on breastfeeding and HM and maximize the reach and impact of this research, larger investments are needed, interdisciplinary collaboration is essential, and the scientific community must engage families and other stakeholders in research planning and knowledge translation.

Metabolomic and Metataxonomic Fingerprinting of Human Milk Suggests Compositional Stability over a Natural Term of Breastfeeding to 24 Months.

Sparse data exist regarding the normal range of composition of maternal milk beyond the first postnatal weeks. This single timepoint, observational study in collaboration with the ‘Parenting Science Gang’ citizen science group evaluated the metabolite and bacterial composition of human milk from 62 participants (infants aged 3–48 months), nearly 3 years longer than previous studies. We utilised rapid evaporative ionisation mass spectrometry (REIMS) for metabolic fingerprinting and 16S rRNA gene metataxonomics for microbiome composition analysis. Milk expression volumes were significantly lower beyond 24 months of lactation, but there were no corresponding changes in bacterial load, composition, or whole-scale metabolomic fingerprint. Some individual metabolite features (~14%) showed altered abundances in nursling age groups above 24 months. Neither milk expression method nor nursling sex affected metabolite and metataxonomic fingerprints. Self-reported lifestyle factors, including diet and physical traits, had minimal impact on metabolite and metataxonomic fingerprints. Our findings suggest remarkable consistency in human milk composition over natural-term lactation. The results add to previous studies suggesting that milk donation can continue up to 24 months postnatally. Future longitudinal studies will confirm the inter-individual and temporal nature of compositional variations and the use of donor milk as a personalised therapeutic.

Maternal adiposity alters the human milk metabolome: associations between nonglucose monosaccharides and infant adiposity

Human milk composition is altered by maternal obesity. The association between milk metabolites and infant outcomes has not been thoroughly investigated. This study aimed to quantify maternal adiposity-related differences in the human milk metabolome and to identify metabolites associated with infant adiposity during the first 6 mo postpartum using untargeted metabolomics. Maternal anthropometrics were assessed≤14 weeks of gestation. Human milk samples were collected at 0.5 mo (n=159), 2 mo (n=131), and 6 mo (n=94) postpartum from normal weight (NW, BMI=18.5-24.9 kg/m2) and obese (OB, BMI>30 kg/m2) mothers. GC-time-of-flight-MS was used to identify metabolic signatures that discriminate NW and OB women. Partial least squared (PLS)-discriminant analysis, and PLS-regression models were assessed to examine relations between metabolites and maternal BMI and fat mass. Metabolites altered by maternal obesity were used in linear mixed effect models to predict infant adiposity. Multivariate modeling identified 23, 17, and 10 metabolites that described maternal adiposity indices at 0.5 mo, 2 mo, and 6 mo postpartum, respectively. Monosaccharides and sugar alcohols were the most representative annotated metabolite classes that were increased in milk from OB women and included: mannose, ribose, lyxose, lyxitol (0.5 mo); mannose, ribitol, glycerol, isothreonic acid, lyxitol (2 mo); lyxitol and isothreonic acid (6 mo). Other discriminant metabolites included: 1-monostearin, xylonolactone, shikimic acid, pseudo uridine, and dodecanol (0.5 mo); N-acetyl-D-hexosamine and fumaric acid (2 mo); uric acid and tyrosine (6 mo). Mannose, lyxitol, and shikimic acid predicted higher infant adiposity over the first 6 mo of life. This study reports on 1 of the largest cohorts to date examining the metabolic profiles in human milk comparing NW and OB women. Maternal adiposity was associated with increased amounts of milk nonglucose monosaccharides. Human milk metabolomics may be useful in predicting infant adiposity. These trials were registered at www.clinicaltrials.gov as NCT01131117 and NCT02125149.

Do Thyroid Diseases during Pregnancy and Lactation Affect the Nutritional Composition of Human Milk?

To identify whether the effects of thyroid disease during pregnancy and lactation affect the nutritional composition of human milk. Systematic review of the scientific literature using the Medical Literature Analysis and Retrieval System Online/MedLine databases to evaluate the association of thyroid diseases during pregnancy and lactation with the nutritional composition of human milk. There was no delimitation by period or by language, and the searches were completed in March 2019. The following descriptors were applied: human milk AND thyroid AND composition, using the preferred reporting items for systematic reviews and meta-analyses (PRISMA) protocol for data search, selection, and extraction. The flowchart proposed for bibliographic search resulted in 12 articles and, of these, four were selected. The articles elected for this review were published between 1976 and 2018. Two studies found significant differences in the nutritional composition of mothers' milk with hypothyroidism or overweight compared with the milk of those without hypothyroidism. Studies have shown that the presence of the disease led to changes in the nutritional composition of human milk, especially a higher concentration of human milk fat. It is extremely important that these women have continuous nutritional follow-up to minimize the impact of these morbidities on the nutritional composition of human milk.

Macronutrient, immunoglobulin a and total antioxidant capacity profiles of human milk from 1 to 24\u2009months: a cross-sectional study in Thailand

BackgroundAn extended duration of breastfeeding of up to two years is encouraged by many health authorities, but information regarding the composition of milk after one year postpartum is limited. The goal of this study was to determine the association between the duration of lactation and macronutrient contents, immunoglobulin A (IgA) levels and total antioxidant capacity (TAC) in human milk (HM), from 1 to 24 months postpartum.MethodsCross-sectional milk samples were collected between January and April 2019 from mothers with healthy full-term children who had been lactating for 1 to 24 months. The HM was biochemically analyzed for protein and carbohydrate contents by colorimetric assays. The fat content was determined by capillary centrifugation, and the energy content was calculated from the results of centrifugation assays. IgA levels and TAC were determined by ELISA and a Trolox equivalent antioxidant capacity (TEAC) assay, respectively. Pearson’s correlation coefficient and Spearman’s rank correlation coefficient were used to determine associations between months of lactation and milk composition, and multiple regression analysis was used to assess associations between months of lactation and milk composition adjusted for relevant covariates. Differences were considered significant at p < 0.05.ResultsOne hundred eighty-four milk samples were analyzed. The month of lactation was positively associated with the fat concentration (B = 0.31, SE = 0.09, p = 0.001), energy content (B = 3.11, SE = 0.92, p = 0.001), and IgA (B = 4.17, SE = 1.08, p < 0.001) but negatively associated with the carbohydrate concentration (B = − 0.22, SE = 0.01, p = 0.04). No association was observed between the month of lactation and the protein concentration or TAC after adjustment for maternal age, maternal BMI, birth order, and breastfeeding frequency.ConclusionThe duration of lactation was found to be positively associated with the fat, energy, and IgA content in HM for up to two years postpartum, and negatively associated with carbohydrate concentration. More prospective cohort studies are needed to obtain evidence-based knowledge regarding the changes in HM composition throughout the course of lactation.