Macronutrient content of pasteurised donor human milk: Variability between batches from single-donor pools at an Australian milk bank.

AIM: This study aimed to characterise the between-batch variability of pasteurised donor human milk (PDHM) produced from single-donor pools at Australian Red Cross Lifeblood's milk bank and identify key donor characteristics that predict macronutrient content. METHODS: Macronutrient content from 200 batches of PDHM was measured using a mid-infrared human milk analyser (Miris, Uppsala, Sweden). Linear mixed models were used to study the impact of stage of lactation and gestational age on macronutrient content. Coefficients of determination (R2 ) were calculated to estimate the impact of the individual donor on overall variability. RESULTS: Macronutrient content of PDHM varied considerably, with between-batch variations of 2.8 and 6.4-fold for protein and fat content, respectively. Mean crude protein content was 1.16 g/100 mL, ranging from 0.7 to 1.96 g/100 mL. Mean fat content was 3.85 g/100 mL, ranging from 1.46 to 9.39 g/100 mL. Stage of lactation was identified as a predictor for protein content and gestational age at birth for fat content. Individual donor effect explained 55 and 35% of the variance for fat and protein content, respectively. CONCLUSIONS: This study highlights the variation in macronutrient content in PDHM at an Australian milk bank. Variability could be reduced through the implementation of targeted multiple-donor pooling using the key donor characteristics identified in this study along with the measurement of macronutrient content of individual donors at the time of first donation. However, the clinical benefit of a reduction in between-batch variation, achieved through multiple-donor pooling, would need to be assessed to justify additional efforts associated with PDHM processing changes.

Evaluating human milk as a drug delivery vehicle for clofazimine to premature infants.

Human milk is proposed as a drug delivery vehicle suitable for use in neonatal patients. Clofazimine, traditionally used for the treatment of leprosy and tuberculosis, is emerging as a treatment for cryptosporidiosis in infants, however its poor aqueous solubility has led to its commercial formulation as a waxy lipid formulation in a capsule, a format that is not suitable for infants. In this study, the evaluation of pasteurised human milk for the delivery of clofazimine was investigated using an in vitro lipolysis model to simulate gastric and intestinal digestion. The total lipid composition of the human milk was characterised alongside the liberated fatty acid species following digestion for comparison to alternative lipid-based delivery systems. Small-angle X-ray scattering was used to measure the presence of crystalline clofazimine during digestion and hence the extent of drug solubilisation. High-performance liquid chromatography was used to quantify the mass of clofazimine solubilised per gram of human milk fat (drug-to-fat ratio) in digested and undigested human milk. The digestion process was essential for the solubilisation of clofazimine, with digested human milk solubilising a sufficient dose of clofazimine for treatment of a premature infant. Human milk solubilised the clofazimine to a greater extent than bovine milk and infant formula during digestion, most likely as a result of differing lipid composition and increased long-chain fatty acid concentrations. These findings show that human milk enhances the solubility of clofazimine as a model drug and may be a suitable drug delivery vehicle for infant populations requiring therapeutic treatment.

Emotional factors, medical interventions and mode of birth among low-risk primiparous women in Poland.

Background and objectives: Birth is a critical event in women's lives. Since humans have evolved to give birth in the context of social support, not having it in modern settings might lead to more complications during birth. Our aim was to model how emotional factors and medical interventions related to birth outcomes in hospital settings in Poland, where c-section rates have doubled in the last decade. Methodology: We analysed data from 2363 low-risk primiparous women who went into labor with the intention of giving birth vaginally. We used a model comparison approach to examine the relationship between emotional and medical variables and birth outcome (vaginal or c-section), including sociodemographic control variables in all models. Results: A model with emotional factors better explained the data than a control model (ΔAIC = 470.8); women with continuous personal support during labor had lower odds of a c-section compared to those attended by hospital staff only (OR = 0.12, 95% CI = 0.09 - 0.16). A model that included medical interventions also better explained the data than a control model (ΔAIC = 133.6); women given epidurals, in particular, had increased odds of a c-section over those who were not (OR = 3.55, 95% CI = 2.95 - 4.27). The best model included variables for both the level of personal support and the use of epidural (ΔAIC = 598.0). Conclusions and implications: Continuous personal support during childbirth may be an evolutionarily informed strategy for reducing complications, including one of the most common obstetrical complications in modern hospital settings, the c-section.

Understanding Mothers' Experiences of Being Ineligible to Donate Their Milk to a Not-for-Profit Milk Bank.

Background: Donor milk banks have strict donor screening criteria to ensure that donor milk is safe for premature or hospitalized babies. Yet little evidence is available to understand how potential donors, who are often breastfeeding their own infants, experience being ineligible ("deferred") to donate their milk to a milk bank. Materials and Methods: Interviews were conducted with 10 mothers who were permanently or temporarily deferred from donating to a large, not-for-profit milk bank in Australia. Interviews focused on becoming a donor and being deferred, meanings of deferral, impact of deferral on feeding own infant, and improving the deferral process. Results: Thematic analysis of interviews identified nine themes: (1) donation as a solution to wasting milk; (2) eligibility questions were acceptable and understandable; (3) more information early on allows self-deferral; (4) deferral is not always clear; (5) deferral is disappointing but does not prevent future donation; (6) deferral did not prevent feeding own infant; (7) early information enables preparation for donation; (8) slow communication disrupts perfect timing to donate; and (9) alternatives to wasting milk. Conclusions: Milk banks have a duty of care to both milk recipients and donors. While mothers who want to donate milk are disappointed by deferrals, clear communication protects their breastfeeding relationships with their own infants. Milk banks can improve their screening processes by providing information up-front and ensuring timely contact with mothers. Mothers can then make informed decisions about donating and not feel as if their milk and resources are "wasted."

Donor and recipient safety in human milk banking.

AIM: Australian Red Cross Lifeblood supplies pasteurised donor human milk (PDHM) to more than 30 partner hospitals across Australia. Preterm infants who receive PDHM are a highly vulnerable population but formal biovigilance programs are rare in human milk banking. Lifeblood Milk performs ongoing surveillance for both donor and recipient adverse events. This study aimed to formally review adverse events reported to Lifeblood Milk since 2018. METHODS: Milk donor infectious diseases testing outcomes and donor adverse events (DAEs) are prospectively recorded at Lifeblood. Infant recipient adverse events are contractually reported back to Lifeblood Milk by hospitals and assessed according to severity and likelihood of relationship to PDHM administration. Donor and recipient adverse events over a 3.5-year period (July 2018 to December 2021) were reviewed. RESULTS: There were three DAEs (3/976 = 0.31%) related to phlebotomy; these included two vasovagal reactions and one phlebotomy site haematoma. Eight (8/976 = 0.81%) additional donors had biological false reactive (BFR) infectious diseases serology results. There were 10 reported suspected adverse events in recipients. Six were infection-related; other events included milk curd obstruction, high urinary iodine levels, sudden cardiac death and nasogastric tube obstruction. All reported suspected adverse events in recipients were classified as unlikely to be related, or definitely not related, to PDHM administration. CONCLUSIONS: Milk donor adverse events were rare but biological false reactive serology results were not uncommon. There were no recipient adverse events considered causally related to pasteurised donor human milk, which is generally a low-risk biological product. Ongoing biovigilance remains essential.

Human Milk Oligosaccharide Compositions Illustrate Global Variations in Early Nutrition.

BACKGROUND: Human milk oligosaccharides (HMOs) are an abundant class of compounds found in human milk and have been linked to the development of the infant, and specifically the brain, immune system, and gut microbiome. OBJECTIVES: Advanced analytical methods were used to obtain relative quantitation of many structures in approximately 2000 samples from over 1000 mothers in urban, semirural, and rural sites across geographically diverse countries. METHODS: LC-MS-based analytical methods were used to profile the compounds with broad structural coverage and quantitative information. The profiles revealed their structural heterogeneity and their potential biological roles. Comparisons of HMO compositions were made between mothers of different age groups, lactation periods, infant sexes, and residing geographical locations. RESULTS: A common behavior found among all sites was a decrease in HMO abundances during lactation until approximately postnatal month 6, where they remained relatively constant. The greatest variations in structural abundances were associated with the presence of α(1,2)-fucosylated species. Genomic analyses of the mothers were not performed; instead, milk was phenotyped according to the abundances of α(1,2)-fucosylated structures. Mothers from the South American sites tended to have higher proportions of phenotypic secretors [mothers with relatively high concentrations of α(1,2)-fucosylated structures] in their populations compared to the rest of the globe, with Bolivia at ∼100% secretors, Peru at ∼97%, Brazil at ∼90%, and Argentina at ∼85%. Conversely, the cohort sampled in Africa manifested the lowest proportion of secretors (South Africa ∼ 63%, the Gambia ∼ 64%, and Malawi ∼ 75%). Furthermore, we compared total abundances of HMOs in secretors compared with nonsecretors and found that nonsecretors have lower abundances of HMOs compared to secretors, regardless of geographical location. We also observed compositional differences of the 50+ most abundant HMOs between milk types and geographical locations. CONCLUSIONS: This study represents the largest structural HMO study to date and reveals the general behavior of HMOs during lactation among different populations.

Tailoring Human Milk Oligosaccharides to Prevent Necrotising Enterocolitis Among Preterm Infants.

Necrotising enterocolitis (NEC) is a devastating disease affecting preterm infants, with little improvement in mortality rates and treatment strategies in the last 30 years. Human milk oligosaccharides (HMOs) are emerging as a potential preventive therapy, with multiple protective functions postulated. Our aim is to summarise the evidence concerning the role of HMOs in NEC development and emerging strategies to tailor the delivery of HMOs to preterm infants. Most research efforts to date have focused on supplementing preterm infants with simple oligosaccharides, which are structurally different to HMOs and derived mainly from plants. Clinical trials demonstrate limited benefits for NEC prevention arising from the use of these supplements. Alternative strategies under investigation include optimising HMOs for infants receiving donor human milk, concentrating oligosaccharides from donor human milk and from animal milks, as well as more sophisticated synthetic oligosaccharide production strategies. Critically, high quality evidence to support implementation of any of these approaches in the neonatal unit is lacking. Whether it is a specific HMO alone or a combination of HMOs that exert protective effects remains to be elucidated. Further challenges include how best to manufacture and administer oligosaccharides whilst retaining bioactivity and safety, including evaluation of the long-term effects of altering the balance of HMOs and gut microbiota in preterm infants. While several human clinical trials are underway, further research is needed to understand whether a tailored approach to oligosaccharide supplementation is beneficial for preterm infants.

What are Optimal Bacteriological Screening Test Cut-Offs for Pasteurized Donor Human Milk Intended for Feeding Preterm Infants?

BACKGROUND: Definitive criteria for microbial screening of pasteurized donor human milk are not well established and international recommendations vary. AIMS: (1) To review pasteurized donor human milk batch discard due to failed microbial screening criteria at our milk bank (following United Kingdom National Institute of Clinical Excellence guidelines), and (2) to compare our known milk discard proportion with estimated milk discard proportions that would be required by other international milk bank guidelines. METHODS: We reviewed our microbial screening results (N = 783) over 18-months (July 2018-December 2019) and compared our known milk discard proportion with estimated milk discard proportions using other international milk bank guidelines. RESULTS: Of samples, n = 50 (6.4%) failed pre-pasteurization screening, most commonly due to the presence of >104 CFU/mL Enterobacterales in the pre-pasteurization sample (n = 30; 3.8%). Two (0.3%) samples failed post-pasteurization screening, with Bacillus cereus identified in both cases, resulting in total discard proportion of 6.7% (n = 52) of batches. Applying European Milk Bank Association recommended bacterial screening criteria, approximately 23.3% (n = 183) of milk batches would have been discarded. CONCLUSIONS: Further research is required to justify the stringent European Milk Bank Association recommendations for pre-pasteurization discard criteria, although we believe that a post-pasteurization acceptance criterion of <1 CFU/mL is appropriate and aligns with international guidance. Further work is needed to understand pasteurized donor human milk microbiological safety risks, to better integrate screening criteria within current food standards regulation, and to consider risk-based assessment including the impact on availability and affordability.

SARS-CoV-2 in human milk is inactivated by Holder pasteurisation but not cold storage.

AIM: As the COVID-19 pandemic evolves, human milk banks world-wide continue to provide donor human milk to vulnerable infants who lack access to mother's own milk. Under these circumstances, ensuring the safety of donor human milk is paramount, as the risk of vertical transmission of SARS-CoV-2 is not fully understood. Here, we investigate the inactivation of SARS-CoV-2 in human milk by pasteurisation and the stability of SARS-CoV-2 in human milk under cold storage. METHODS: SARS-CoV-2 was experimentally inoculated into human milk samples from healthy donors or into a control medium. Triplicates of each sample were layered onto uninfected cells after Holder pasteurisation (63°C for 30 min), heating to 56°C for 30 min, or after 48 h of storage at 4°C or -30°C. Infectious titres of virus were determined at 72 h post-infection by endpoint titration. RESULTS: Following heating to 63°C or 56°C for 30 min, replication competent (i.e. live) SARS-CoV-2 was undetected in both human milk and the control medium. Cold storage of SARS-CoV-2 in human milk (either at 4°C or -30°C) did not significantly impact infectious viral load over a 48 h period. CONCLUSION: SARS-CoV-2 is effectively inactivated by Holder pasteurisation, suggesting that existing milk bank processes will effectively mitigate the risk of transmission of SARS-COV-2 to vulnerable infants through pasteurised donor human milk. The demonstrated stability of SARS-CoV-2 in refrigerated or frozen human milk may assist in the development of guidelines around safe expressing and storing of milk from COVID-19 infected mothers.

Declining ages at menarche in an agrarian rural region of Poland.

OBJECTIVE: Age at menarche in Poland has varied with political and socioeconomic changes. An increase in age at menarche corresponded to a period of economic crisis and food rationing between 1976 and 1989. Experiencing food shortages in utero or during childhood development can affect menarcheal timing, but this national effect may be buffered in local agrarian regions growing their own food. Here we examine patterns of age at menarche over time in the rural, agrarian Beskid Wyspowy region of southern Poland. METHODS: This study examined menarcheal timing using data collected from Polish women (n = 1326) recruited at the Mogielica Human Ecology Study Site between 2003 and 2018. Simple linear regressions were used to assess changing ages at menarche over time. Comparisons between ages at menarche for women born before and after the fall of communism in 1989 were assessed via one-way analysis of variance. RESULTS: Age at menarche has declined over time in the Beskid Wyspowy region of southern Poland from 1920 to 2000 (R2 = .08, P < .0001). There was not a statistically significant increase or decrease in age at menarche for women born and growing up during the period of food rationing. CONCLUSIONS: The declining age at menarche is likely reflective of a transitioning environment, suggesting that major socioeconomic changes affect life history traits like pubertal timing. Living in agricultural regions may have helped buffer the increasing ages at menarche seen in other areas of Poland during times of food rationing.

Variation among populations in the immune protein composition of mother's milk reflects subsistence pattern.

LAY SUMMARY: Adaptive immune proteins in mothers' milk are more variable than innate immune proteins across populations and subsistence strategies. These results suggest that the immune defenses in milk are shaped by a mother's environment throughout her life. BACKGROUND AND OBJECTIVES: Mother's milk contains immune proteins that play critical roles in protecting the infant from infection and priming the infant's developing immune system during early life. The composition of these molecules in milk, particularly the acquired immune proteins, is thought to reflect a mother's immunological exposures throughout her life. In this study, we examine the composition of innate and acquired immune proteins in milk across seven populations with diverse disease and cultural ecologies. METHODOLOGY: Milk samples (n = 164) were collected in Argentina, Bolivia, Nepal, Namibia, Philippines, Poland and the USA. Populations were classified as having one of four subsistence patterns: urban-industrialism, rural-shop, horticulturalist-forager or agro-pastoralism. Milk innate (lactalbumin, lactoferrin and lysozyme) and acquired (Secretory IgA, IgG and IgM) protein concentrations were determined using triple-quadrupole mass spectrometry. RESULTS: Both innate and acquired immune protein composition in milk varied among populations, though the acquired immune protein composition of milk differed more among populations. Populations living in closer geographic proximity or having similar subsistence strategies (e.g. agro-pastoralists from Nepal and Namibia) had more similar milk immune protein compositions. Agro-pastoralists had different milk innate immune protein composition from horticulturalist-foragers and urban-industrialists. Acquired immune protein composition differed among all subsistence strategies except horticulturist-foragers and rural-shop. CONCLUSIONS AND IMPLICATIONS: Our results reveal fundamental variation in milk composition that has not been previously explored in human milk research. Further study is needed to understand what specific aspects of the local environment influence milk composition and the effects this variation may have on infant health outcomes.

Concentrations of trace elements in human milk: Comparisons among women in Argentina, Namibia, Poland, and the United States.

Human milk contains essential micronutrients for growth and development during early life. Environmental pollutants, such as potentially toxic metals, can also be transferred to the infant through human milk. These elements have been well-studied, but changing diets and environments and advances in laboratory technology require re-examining these elements in a variety of settings. The aim of this study was to characterize the concentrations of essential and toxic metals in human milk from four diverse populations. Human milk samples (n = 70) were collected in Argentina (n = 21), Namibia (n = 6), Poland (n = 23), and the United States (n = 20) using a standardized mid-feed collection procedure. Milk concentrations of calcium, zinc, iron, copper, manganese, lead, arsenic, and cadmium were determined using inductively coupled plasma mass spectrometry (ICP-MS). We used standard multiple linear regression models to evaluate differences among populations, while including infant age, infant sex, and maternal parity status (multiparous or primiparous) as covariates. Concentrations of all elements, except zinc, varied across populations after controlling for infant age, infant sex, and maternal parity. Calcium and magnesium showed more differences across populations than iron or copper. There were no significant differences among population in zinc concentrations. Mean concentrations of lead, but not arsenic, were low compared to recently published values from other populations. The concentrations of trace elements in human milk are variable among populations. Limitations due to small sample sizes and environmental contamination of some samples prevent us from drawing robust conclusions about the causes of these differences.

Relationships between biomarkers of inflammation, ovarian steroids, and age at menarche in a rural Polish sample.

OBJECTIVES: To test the hypothesis that life history trade-offs between maintenance and reproductive effort would be evident through inverse associations between levels of a biomarker of inflammation [C-reactive protein (CRP)], and ovarian hormones. Associations between CRP and age at menarche were also explored. METHODS: Urinary CRP, salivary progesterone, and estradiol were measured over one menstrual cycle from rural Polish women (n = 25), representing a natural fertility sample. Age of menarche was assessed through interview recall methods. We used minimum second-order Akaike Information Criteria as a means of multiple regression model selection, and repeated measures ANOVA to test cycle-dependent hypotheses. RESULTS: Comparisons of individuals in high and low CRP tertiles revealed that those with high CRP had significantly lower progesterone (luteal P = 0.03, mid luteal P = 0.007) but not estradiol (follicular P = 0.21, luteal P = 0.15) concentrations through the menstrual cycle. However, when the age at menarche was included in the analysis, both age at menarche and urinary CRP were negatively associated with estradiol (R(2) = 0.44, P = 0.0007). Age at menarche and estradiol were the strongest negative predictors of CRP (R(2) = 0.52, P = 0.0001). CONCLUSIONS: Inflammation itself may suppress ovarian function, or indicate immune challenges that lead to ovarian suppression. The timing of menarche may also influence adult inflammatory sensitivity and ovarian hormone concentrations. This lends support to existing models of trade-offs between maintenance and reproduction in women.