Strong Multivariate Relations Exist Among Milk, Oral, and Fecal Microbiomes in Mother-Infant Dyads During the First Six Months Postpartum.

BACKGROUND: Neonatal gastrointestinal (GI) bacterial community structure may be related to bacterial communities of the mother, including those of her milk. However, very little is known about the diversity in and relationships among complex bacterial communities in mother-infant dyads. OBJECTIVE: Our primary objective was to assess whether microbiomes of milk are associated with those of oral and fecal samples of healthy lactating women and their infants. METHODS: Samples were collected 9 times from day 2 to 6 mo postpartum from 21 healthy lactating women and their infants. Milk was collected via complete breast expression, oral samples via swabs, and fecal samples from tissue (mothers) and diapers (infants). Microbiomes were characterized using high-throughput sequencing of the 16S ribosomal RNA (rRNA) gene. Alpha and beta diversity indices were used to compare microbiomes across time and sample types. Membership and composition of microbiomes were analyzed using nonmetric multidimensional scaling and canonical correlation analysis (CCA). The contribution of various bacterial communities of the mother-infant dyad to both milk and infant fecal bacterial communities were estimated using SourceTracker2. RESULTS: Bacterial community structures were relatively unique to each sample type. The most abundant genus in milk and maternal and infant oral samples was Streptococcus (47.1% ± 2.3%, 53.9% ± 1.3%, and 69.1% ± 1.8%, respectively), whereas Bacteroides were predominant in maternal and infant fecal microbiomes (22.9% ± 1.3% and 21.4% ± 2.4%, respectively). The milk microbiome was more similar to the infant oral microbiome than the infant fecal microbiome. However, CCA suggested strong associations between the complex microbial communities of milk and those of all other sample types collected. CONCLUSIONS: These findings suggest complex microbial interactions between breastfeeding mothers and their infants and support the hypothesis that variation in the milk microbiome may influence the infant GI microbiome.

Comparison of commercially-available preservatives for maintaining the integrity of bacterial DNA in human milk.

BACKGROUND: Inhibiting changes to bacteria in human milk between sample collection and analysis is necessary for unbiased characterization of the milk microbiome. Although cold storage is considered optimal, alternative preservation is sometimes necessary. RESEARCH AIM/QUESTION: The objective of this study was to compare the effectiveness of several commercially-available preservatives with regard to maintaining bacterial DNA in human milk for delayed microbiome analysis. Specifically, we compared Life Technologies' RNAlater® stabilization solution, Biomatrica's DNAgard® Saliva, Advanced Instruments' Broad Spectrum Microtabs II™, and Norgen Biotek Corporation's Milk DNA Preservation and Isolation Kit. METHODS: Aliquots of 8 pools of human milk were treated with each preservative. DNA was extracted immediately and at 1, 2, 4, and 6wk, during which time milk was held at 37°C. The V1-V3 region of the bacterial 16S rRNA gene was amplified and sequenced. Changes in bacterial community structure and diversity over time were evaluated. RESULTS: Comparable to other studies, the most abundant genera were Streptococcus (33.3%), Staphylococcus (14.0%), Dyella (6.3%), Pseudomonas (3.0%), Veillonella (2.5%), Hafnia (2.0%), Prevotella (1.7%), Rhodococcus (1.6%), and Granulicatella (1.4%). Overall, use of Norgen's Milk DNA Preservation and Isolation Kit best maintained the consistency of the bacterial community structure. Total DNA, diversity, and evenness metrics were also highest in samples preserved with this method. CONCLUSIONS: When collecting human milk for bacterial community analysis in field conditions where cold storage is not available, our results suggest that Norgen's Milk DNA Preservation and Isolation Kit may be a useful method, at least for a period of 2weeks.

Human Milk Microbial Community Structure Is Relatively Stable and Related to Variations in Macronutrient and Micronutrient Intakes in Healthy Lactating Women.

Background: The human milk microbiome has been somewhat characterized, but little is known about changes over time and relations with maternal factors such as nutrient intake.Objective: We sought to characterize the human milk microbiome and described associations with maternal nutrient intake, time postpartum, delivery mode, and body mass index (BMI; in kg/m2).Methods: Milk samples (n = 104) and 24-h diet recalls were collected 9 times from 21 healthy lactating women from day 2 to 6 mo postpartum. Women were classified by BMI as healthy weight (<25) or overweight or obese (≥25). Bacterial taxa were characterized with the use of the high-throughput sequencing of the 16S ribosomal RNA gene.Results: The milk microbiome was relatively constant over time, although there were small changes in some of the lesser-abundant genera. Relative abundances of several taxa were associated with BMI, delivery mode, and infant sex. For instance, overweight and obese mothers produced milk with a higher relative abundance of Granulicatella than did healthy-weight women (1.8% ± 0.6% compared with 0.4% ± 0.2%, respectively; P < 0.05). Relative abundances of several bacterial taxa were also associated with variations in maternal dietary intake. For example, intakes of saturated fatty acids (rs = -0.59; P = 0.005) and monounsaturated fatty acids (rs = -0.46; P = 0.036) were inversely associated with the relative abundance of Corynebacterium; total carbohydrates (rs = -0.54; P = 0.011), disaccharides (rs = -0.47; P = 0.031), and lactose (rs = -0.51; P = 0.018) were negatively associated with Firmicutes; and protein consumption was positively correlated with the relative abundance of Gemella (rs = 0.46; P = 0.037).Conclusions: Factors associated with variations in the human milk microbiome are complex and may include maternal nutrient intake, maternal BMI, delivery mode, and infant sex. Future studies designed to investigate the relation between maternal nutrient intake and the milk microbiome should strive to also evaluate dietary supplement usage and analyze the collected milk for its nutrient content.

Glyphosate and aminomethylphosphonic acid are not detectable in human milk.

BACKGROUND: Although animal studies have shown that exposure to glyphosate (a commonly used herbicide) does not result in glyphosate bioaccumulation in tissues, to our knowledge there are no published data on whether it is detectable in human milk and therefore consumed by breastfed infants. OBJECTIVE: We sought to determine whether glyphosate and its metabolite aminomethylphosphonic acid (AMPA) could be detected in milk and urine produced by lactating women and, if so, to quantify typical consumption by breastfed infants. DESIGN: We collected milk (n = 41) and urine (n = 40) samples from healthy lactating women living in and around Moscow, Idaho and Pullman, Washington. Milk and urine samples were analyzed for glyphosate and AMPA with the use of highly sensitive liquid chromatography-tandem mass spectrometry methods validated for and optimized to each sample matrix. RESULTS: Our milk assay, which was sensitive down to 1 µg/L for both analytes, detected neither glyphosate nor AMPA in any milk sample. Mean ± SD glyphosate and AMPA concentrations in urine were 0.28 ± 0.38 and 0.30 ± 0.33 µg/L, respectively. Because of the complex nature of milk matrixes, these samples required more dilution before analysis than did urine, thus decreasing the sensitivity of the assay in milk compared with urine. No difference was found in urine glyphosate and AMPA concentrations between subjects consuming organic compared with conventionally grown foods or between women living on or near a farm/ranch and those living in an urban or suburban nonfarming area. CONCLUSIONS: Our data provide evidence that glyphosate and AMPA are not detectable in milk produced by women living in this region of the US Pacific Northwest. By extension, our results therefore suggest that dietary glyphosate exposure is not a health concern for breastfed infants. This study was registered at clinicaltrials.gov as NCT02670278.

Fecal Microbial Community Structure Is Stable over Time and Related to Variation in Macronutrient and Micronutrient Intakes in Lactating Women.

BACKGROUND: The fecal microbiota has been characterized in some adult populations, but little is known about its community structure during lactation. OBJECTIVES: We characterized the maternal fecal microbiome during lactation and explored possible mediating factors such as nutrition. METHODS: Fecal samples were collected from 20 lactating women from 2 d to 6 mo postpartum, and bacterial taxa were characterized with the use of high-throughput sequencing. Bacterial community structure (at each taxonomic level) and relations between bacterial taxa and environmental and dietary variables were visualized and analyzed with the use of stacked bar charts, principal component analysis, and multivariate analyses such as nonmetric multidimensional scaling and canonical correlation analysis. RESULTS: Complex bacterial community structure was somewhat similar to those previously published for other adult populations (although there were some notable differences), and there were no clear associations with time postpartum or anthropometric or environmental variables. However, Spearman rank correlations suggested that increased intake of pantothenic acid, riboflavin, vitamin B-6, and vitamin B-12 were related to increased relative abundance of Prevotella (r = 0.45, 0.39, 0.34, and 0.24, respectively; P ≤ 0.01) and decreased relative abundance of Bacteroides (r = -0.55, -0.46, -0.32, and -0.35, respectively; P ≤ 0.01). Intakes of copper, magnesium, manganese, and molybdenum were positively associated with Firmicutes (r = 0.33, 0.38, 0.44, and 0.51, respectively; P ≤ 0.01) and negatively associated with Bacteroidetes (r = -0.38, -0.44, -0.48, and -0.53, respectively; P ≤ 0.01). Overall, data consistently suggest that increased consumption of a more nutrient- and calorie-rich diet was positively associated with relative abundance of Firmicutes. CONCLUSIONS: The fecal microbiome of lactating women is relatively stable in the postpartum period and somewhat similar to that of other adult populations. Variation in dietary constituents may be related to that of relative abundance of individual bacterial taxa. Controlled dietary intervention studies will be required to determine whether these associations are causal in nature.