Milk Microbiome Research Articles

A Review Focusing on Microbial Vertical Transmission during Sow Pregnancy

Microorganisms are closely related to the body's physiological activities and growth and development of the body, and participate in many physiological metabolic activities. Analysis of the structure and source of early colonizing bacteria in the intestinal tract of humans and rodents shows that early colonizing bacteria in the intestinal tract of mammals have solid maternal characteristics, and maternal microbes play an essential role in the formation of progeny intestinal flora. The placental microbiome, maternal microbiome and breast milk microbiome are currently hot topics in the field of life science. This paper discusses the vertical transmission and endogenous sources of the mother-to-piglet microbiome through these three pathways, aiming to provide a new research idea for intervention in the intestinal microbiome in young piglets.

Interrogating the role of the milk microbiome in mastitis in the multi-omics era.

There is growing interest in a functional understanding of milk-associated microbiota as there is ample evidence that host-associated microbial communities play an active role in host health and phenotype. Mastitis, characterized by painful inflammation of the mammary gland, is prevalent among lactating humans and agricultural animals and is associated with significant clinical and economic consequences. The etiology of mastitis is complex and polymicrobial and correlative studies have indicated alterations in milk microbial community composition. Recent evidence is beginning to suggest that a causal relationship may exist between the milk microbiota and host phenotype in mastitis. Multi-omic approaches can be leveraged to gain a mechanistic, molecular level understanding of how the milk microbiome might modulate host physiology, thereby informing strategies to prevent and ameliorate mastitis. In this paper, we review existing studies that have utilized omics approaches to investigate the role of the milk microbiome in mastitis. We also summarize the strengths and challenges associated with the different omics techniques including metagenomics, metatranscriptomics, metaproteomics, metabolomics and lipidomics and provide perspective on the integration of multiple omics technologies for a better functional understanding of the milk microbiome.

Characterisation of human milk bacterial DNA profiles in a small cohort of Australian women in relation to infant and maternal factors.

Human milk is composed of complex microbial and non-microbial components that shape the infant gut microbiome. Although several maternal and infant factors have been associated with human milk microbiota, no study has investigated this in an Australian population. Therefore, we aimed to investigate associations between human milk bacterial composition of Australian women and maternal factors (body mass index (BMI), mode of delivery, breast pump use, allergy, parity) and infant factors (sex, mode of feeding, pacifier use, and introduction of solids). Full-length 16S rRNA gene sequencing was used to characterise milk bacterial DNA profiles. Milk from mothers with a normal BMI had a higher relative abundance of Streptococcus australis than that of underweight mothers, while milk from overweight mothers had a higher relative abundance of Streptococcus salivarius compared with underweight and obese mothers. Mothers who delivered vaginally had a higher relative abundance of Streptococcus mitis in their milk compared to those who delivered via emergency caesarean section. Milk of mothers who used a breast pump had a higher relative abundance of Staphylococcus epidermidis and Streptococcus parasanguinis. Milk of mothers whose infants used a pacifier had a higher relative abundance of S. australis and Streptococcus gwangjuense. Maternal BMI, mode of delivery, breast pump use, and infant pacifier use are associated with the bacterial composition of human milk in an Australian cohort. The data from this pilot study suggests that both mother and infant can contribute to the human milk microbiome.

Identification of <i>Lactobacillus</i> and <i>Bifidobacterium</i> 16S rRNA Gene in Breast Milk of Some Healthy Women in Kinshasa (DR Congo)

Breast milk is important for infant health. Some of its benefits are due to the presence of a specific population of bacteria in the microflora. However, the microbiome of breast milk is influenced by many parameters such as maternal diet, breastfeeding and geographic location. Culture and non-culture methods have been used in studies of this bacterial population worldwide. But in the DR Congo, there was no study reporting the use of culture-independent techniques to characterize the bacterial diversity of human milk. The aim of this study was to identify the bacterial 16S rRNA gene from two genera Lactobacillus and Bifidobacterium. The 16S rRNA gene was also identified from four species: Lactobacillus reuteri, Lactobacillus rhamnosus, Bifidobacterium longum and Bifidobacterium lactis. This analytical cross-sectional study was conducted in Kinshasa. Breast milk from some healthy women was collected from February 2 to 28, 2018. A culture-independent protocol using the classical polymerase chain reaction (PCR) was used to identify the bacterial 16S rRNA gene. The 68 samples of breast milk were collected in a sterile condition. The bacteria-specific ribosomal gene 16S rRNA was detected in 91.18% of Lactobacillus and 32.35% of Bifidobacterium at genus level. At of species level, only Lactobacillus reuteri 16S rRNA gene was identified in 89.71%. The 16S rRNA gene from the other species could not be amplified. There was also an association between educational level and the presence of Bifidobacterium and Lactobacillus 16S rRNA genes in the breast milk (p = 0.008*, p Lactobacillus and Bifidobacterium in breast milk at the genus and Lactobacillus reuteri at species level. A further study on the diet, use of antibiotics during pregnancy and lactation practice will provide a better understanding of the microflora of breast milk.

Metagenomic assessment of the bacterial breastfeeding microbiome in mature milk across lactation.

Research has illustrated the presence of a diverse range of microbiota in human milk. The composition of the milk microbiome varies across different stages of lactation, emphasizing the need to consider the lactation stage when studying its composition. Additionally, the transfer of both milk and skin microbiota during breastfeeding is crucial for understanding their collective impact on infant health and development. Further exploration of the complete breastfeeding microbiome is necessary to unravel the role these organisms play in infant development. We aim to longitudinally assess the bacterial breastfeeding microbiome across stages of lactation. This includes all the bacteria that infants are exposed to during breastfeeding, such as bacteria found within human milk and any bacteria found on the breast and nipple. Forty-six human milk samples were collected from 15 women at 1, 4, 7, and 10 months postpartum. Metagenomic analysis of the bacterial microbiome for these samples was performed by CosmosID (Rockville, MD) via deep sequencing. Staphylococcus epidermidis and Propionibacteriaceae species are the most abundant bacterial species from these samples. Samples collected at 10 months showed higher abundances of Proteobacteria, Streptococcaceae, Lactobacillales, Streptococcus, and Neisseria mucosa compared to other timepoints. Alpha diversity varied greatly between participants but did not change significantly over time. As the bacterial breastfeeding microbiome continues to be studied, bacterial contributions could be used to predict and reduce health risks, optimize infant outcomes, and design effective management strategies, such as altering the maternal flora, to mitigate adverse health concerns.

Human milk-associated bacterial communities associate with the infant gut microbiome over the first year of life.

Microbial communities inhabiting the human infant gut are important for immune system development and lifelong health. One critical exposure affecting the bacterial colonization of the infant gut is consumption of human milk, which contains diverse microbial communities and prebiotics. We hypothesized that human milk-associated microbial profiles are associated with those of the infant gut. Maternal-infant dyads enrolled in the New Hampshire Birth Cohort Study (n = 189 dyads) contributed breast milk and infant stool samples collected approximately at 6 weeks, 4 months, 6 months, 9 months, and 12 months postpartum (n = 572 samples). Microbial DNA was extracted from milk and stool and the V4-V5 region of the bacterial 16S rRNA gene was sequenced. Clustering analysis identified three breast milk microbiome types (BMTs), characterized by differences in Streptococcus, Staphylococcus, Pseudomonas, Acinetobacter, and microbial diversity. Four 6-week infant gut microbiome types (6wIGMTs) were identified, differing in abundances of Bifidobacterium, Bacteroides, Clostridium, Streptococcus, and Escherichia/Shigella, while two 12-month IGMTs (12mIGMTs) differed primarily by Bacteroides presence. At 6 weeks, BMT was associated with 6wIGMT (Fisher's exact test value of p = 0.039); this association was strongest among infants delivered by Cesarean section (Fisher's exact test value of p = 0.0028). The strongest correlations between overall breast milk and infant stool microbial community structures were observed when comparing breast milk samples to infant stool samples collected at a subsequent time point, e.g., the 6-week breast milk microbiome associated with the 6-month infant gut microbiome (Mantel test Z-statistic = 0.53, value of p = 0.001). Streptoccous and Veillonella species abundance were correlated in 6-week milk and infant stool, and 4- and 6-month milk Pantoea species were associated with infant stool Lachnospiraceae genera at 9 and 12 months. We identified clusters of human milk and infant stool microbial communities that were associated in maternal-infant dyads at 6 weeks of life and found that milk microbial communities were more strongly associated with infant gut microbial communities in infants delivered operatively and after a lag period. These results suggest that milk microbial communities have a long-term effect on the infant gut microbiome both through sharing of microbes and other molecular mechanisms.

The dynamic communities of oral microbiome in neonates.

The oral microbiome, associated with both oral disease and systemic disease, is in dynamic status along the whole life, and many factors including maternal microbiomes could impact the oral microbiome. While fewer studies have been conducted to study the characteristics of the oral microbiome in neonates and the associated maternal factors. Hence, we collected the microbiome of 15 mother-infant pairs across multiple body sites from birth up to 4 days postpartum and used high-throughput sequencing to characterize the microbiomes in mothers and their neonates. The oral microbiome in the neonates changed obviously during the 4 days after birth. Many bacteria originating from the vagina, skin, and environment disappeared in oral cavity over time, such as Prevotella bivia and Prevotella jejuni. Meanwhile, Staphylococcus epidermidis RP62A phage SP-beta, predominate bacterium in maternal skin microbiome and Streptococcus unclassified, main bacterium in vaginal microbiome, obviously increased in neonatal oral microbiome as time went on. Interestingly, as time progressed, the composition of the oral microbiome in the neonates was more similar to that of the milk microbiome in their mothers. Moreover, we found that the changes in the predominant bacteria in the neonates were in line with those in the neonates exposed to the environment. Together, these data described the sharp dynamics of the oral microbiome in neonates and the importance of maternal efforts in the development of the neonatal microbiome.

Influence of dietary n-3 long-chain fatty acids on microbial diversity and composition of sows' feces, colostrum, milk, and suckling piglets' feces.

Very little is known about the impact of n-3 long-chain fatty acids (n-3 LCFAs) on the microbiota of sows and their piglets. The aim of this study was to evaluate the effect of n-3 LCFA in sow diets on the microbiota composition of sows' feces, colostrum, and milk as well as that of piglets' feces. Twenty-two sows were randomly assigned to either a control or an n-3 LCFA diet from service to weaning. Sows' and piglets' performance was monitored. The gestating and lactating sows' microbiomes in feces, colostrum, and milk were characterized by 16s ribosomal RNA gene sequencing. The fecal microbiome from the two lowest (>800 g) and the two highest birth weight piglets per litter was also characterized, and the LPS levels in plasma were analyzed at weaning. n-3 LCFA increased microbiota alpha diversity in suckling piglets' and gestating sows' feces. However, no effects were observed in colostrum, milk, or lactating sows' feces. Dietary n-3 LCFA modified the microbiota composition of gestating sows' feces, milk, and suckling piglets' feces, without affecting lactating sows' feces or colostrum. In gestating sows' feces and milk, the decrease in genus Succinivibrio and the increase of Proteobacteria phylum, due to the increased genera Brenneria and Escherichia, respectively, stand out. In the feces of suckling piglets, the higher abundance of the beneficial genus Akkermansia and Bacteroides, and different species of Lactobacillus are highlighted. In addition, positive correlations for families and genera were found between lactating sows' feces and milk, milk and suckling piglets' feces, and lactating sows' feces and suckling piglets' feces. To conclude, dietary n-3 LCFA had a positive impact on the microbiome of suckling piglet's feces by increasing microbial diversity and some beneficial bacteria populations, had a few minor modifications on the microbiome of milk and gestating sows' feces and did not change the microbiome in lactating sows' feces or colostrum. Therefore, this study shows the effect of dietary n-3 LCFA on the microbiota of sows, colostrum, milk, and suckling piglets during the lactation period providing crucial information on the microbiota status at the early stages of life, which have an impact on the post-weaning.

Human Milk Microbiome and Microbiome-Related Products: Potential Modulators of Infant Growth

Infant growth trajectory may influence later-life obesity. Human milk provides a wide range of nutritional and bioactive components that are vital for infant growth. Compared to formula-fed infants, breastfed infants are less likely to develop later-onset obesity, highlighting the potential role of bioactive components present in human milk. Components of particular interest are the human milk microbiota, human milk oligosaccharides (HMOs), short-chain fatty acids (SCFAs), and antimicrobial proteins, each of which influence the infant gut microbiome, which in turn has been associated with infant body composition. SCFAs and antimicrobial proteins from human milk may also systemically influence infant metabolism. Although inconsistent, multiple studies have reported associations between HMOs and infant growth, while studies on other bioactive components in relation to infant growth are sparse. Moreover, these microbiome-related components may interact with each other within the mammary gland. Here, we review the evidence around the impact of human milk microbes, HMOs, SCFAs, and antimicrobial proteins on infant growth. Breastfeeding is a unique window of opportunity to promote optimal infant growth, with aberrant growth trajectories potentially creating short- and long-term public health burdens. Therefore, it is important to understand how bioactive components of human milk influence infant growth.

Infant Gut Microbiome is Enriched with Bifidobacterium longum ssp. infantis in Old Order Mennonites With Traditional Farming Lifestyle

Growing up on traditional, single-family farms is associated with fewer early manifestations of asthma and possibly other atopic conditions. This study examined gut microbiome and metabolome composition in rural Old Order Mennonite (OOM) infants at low risk and Rochester, NY infants at high risk of atopic conditions.This cohort included OOM mother-infant pairs from the Finger Lakes region of New York State (N = 65) and urban or suburban mother-infant pairs from Rochester, NY (N = 39); most infants were exclusively breastfed.Microbiome composition was evaluated by 16S rRNA gene sequencing of 101 infant stool and 61 human milk samples. Bifidobacterium spp. and B. longum ssp. infantis (B. infantis), a consumer of human milk oligosaccharides, were quantified using quanititative polymerase chain reation. Fatty acids (FAs) were analyzed in 34 stool and 24 human milk samples. Symptoms and diagnoses of atopic conditions were assessed by telephone.At a median age of 2 months, stool was enriched with Bifidobacteriaceae, Clostridiaceae, and Aerococcaceae in OOM compared with Rochester infants. B. infantis was more abundant (P < .001) and prevalent (P < .001) in OOM (70%) compared with Rochester (21%) infants. Stool colonized with B. infantis had higher levels of lactate (P < .05) and several medium-to long or odd-chain FAs (P < .05). Human milk α diversity was similar between the groups. Paired human milk was enriched with a distinct pattern of FAs, including butyrate. At age 3 years, fewer atopic conditions, particularly eczema and food allergy, were reported in OOM children (6.5%) compared with Rochester children (35%, P < .001). Atopic conditions were not associated with B. infantis presence (P = .08), or enrichment of gut or human milk microbiome α diversity.In this small, cross-sectional, pilot study, OOM infants had a high rate of B. infantis colonization, similar to infants in developing countries, and were at lower risk of developing early atopic conditions.OOM infants have a distinct gut microbiome and metabolome composition and fewer early-onset atopic conditions. Prospective examination of the associations in the gut and human milk microbiome and presence of B. infantis in a larger cohort of OOM infants may help to further elucidate the mechanisms of these associations.URL: www.pediatrics.org/cgi/doi/10.1542/peds.10.1111/all.14877

Human milk microbial species are associated with mild growth deficits during early infancy among Guatemalan mother–infant dyads

Growth faltering is common in Guatemalan indigenous communities, but the possibility that it may be related to milk microbial composition has not been explored. For this cross-sectional study, unrelated mother–infant dyads (n = 64) from eight communities in the remote Western Highlands of Guatemala were recruited. Milk samples and infant length-for-age and weight-for-age Z-scores were collected at two stages of lactation: early (6–46 days postpartum, n = 29) or late (109–184 days postpartum, n = 35). Within each stage of lactation, infants were subdivided into mildly underweight [weight-for-age Z-score (WAZ) &amp;lt; –1 SD] or normal weight (WAZ ≥ –1 SD) and mildly stunted [length-for-age Z-score (LAZ) &amp;lt; –1.5 SD] or non-stunted (LAZ ≥ –1.5 SD). 16S ribosomal RNA gene sequencing was used to identify milk microbial communities, and DESeq2 was used to compare the differential abundance (DA) of human milk microbiota at the species level for WAZ and LAZ subgroups at each stage of lactation. A total of 503 ESVs annotated as 256 putative species across the 64 human milk samples were identified. Alpha diversity did not differ, but beta-diversity redundancy analysis identified four distinct clusters among the four WAZ (p = 0.004) and LAZ subgroups (p = 0.001). DA identified 15 different taxa in the WAZ and 11 in the LAZ groups in early lactation and 8 in the WAZ and 19 in the LAZ groups in late lactation. Mothers’ milk had more DA taxa of oropharyngeal and environmental bacteria with opportunistic activities in the LAZ &amp;lt; –1.5 SD infants, whereas the LAZ ≥ –1.5 SD had DA taxa with potential probiotic and antimicrobial inhibitory activity against pathogens. In particular, milk microbial communities of infants not classified as underweight or stunted had more beneficial species including Lactococcus_lactis. These findings suggest the potential associations between the milk microbiome at the species level with infant growth prior to 6 months of age. These data provide important evidence of the associations between the human milk microbiome and the growth of breastfed infants.

A longitudinal census of the bacterial community in raw milk correlated with Staphylococcus aureus clinical mastitis infections in dairy cattle

BackgroundStaphylococcus aureus is a common cause of clinical mastitis (CM) in dairy cattle. Optimizing the bovine mammary gland microbiota to resist S. aureus colonization is a growing area of research. However, the details of the interbacterial interactions between S. aureus and commensal bacteria, which would be required to manipulate the microbiome to resist infection, are still unknown. This study aims to characterize changes in the bovine milk bacterial community before, during, and after S. aureus CM and to compare bacterial communities present in milk between infected and healthy quarters.MethodsWe collected quarter-level milk samples from 698 Holstein dairy cows over an entire lactation. A total of 11 quarters from 10 cows were affected by S. aureus CM and milk samples from these 10 cows (n = 583) regardless of health status were analyzed by performing 16S rRNA gene amplicon sequencing.ResultsThe milk microbiota of healthy quarters was distinguishable from that of S. aureus CM quarters two weeks before CM diagnosis via visual inspection. Microbial network analysis showed that 11 OTUs had negative associations with OTU0001 (Staphylococcus). A low diversity or dysbiotic milk microbiome did not necessarily correlate with increased inflammation. Specifically, Staphylococcus xylosus, Staphylococcus epidermidis, and Aerococcus urinaeequi were each abundant in milk from the quarters with low levels of inflammation.ConclusionOur results show that the udder microbiome is highly dynamic, yet a change in the abundance in certain bacteria can be a potential indicator of future S. aureus CM. This study has identified potential prophylactic bacterial species that could act as a barrier against S. aureus colonization and prevent future instances of S. aureus CM.

The Human Milk Microbiota Produces Potential Therapeutic Biomolecules and Shapes the Intestinal Microbiota of Infants.

Human milk not only provides a perfect balance of nutrients to meet all the needs of the infant in the first months of life but also contains a variety of bacteria that play a key role in tailoring the neonatal faecal microbiome. Microbiome analysis of human milk and infant faeces from mother-breastfed infant pairs was performed by sequencing the V1-V3 region of the 16S rRNA gene using the Illumina MiSeq platform. According to the results, there is a connection in the composition of the microbiome in each mother-breastfed infant pair, supporting the hypothesis that the infant's gut is colonised with bacteria from human milk. MiSeq sequencing also revealed high biodiversity of the human milk microbiome and the infant faecal microbiome, whose composition changes during lactation and infant development, respectively. A total of 28 genetically distinct strains were selected by hierarchical cluster analysis of RAPD-PCR (Random Amplified Polymorphic DNA-Polymerase Chain Reaction) electrophoresis profiles of 100 strains isolated from human milk and identified by 16S RNA sequencing. Since certain cellular molecules may support their use as probiotics, the next focus was to detect (S)-layer proteins, bacteriocins and exopolysaccharides (EPSs) that have potential as therapeutic biomolecules. SDS-PAGE (Sodium Dodecyl-Sulfate Polyacrylamide Gel Electrophoresis) coupled with LC-MS (liquid chromatography-mass spectrometry) analysis revealed that four Levilactobacillus brevis strains expressed S-layer proteins, which were identified for the first time in strains isolated from human milk. The potential biosynthesis of plantaricin was detected in six Lactiplantibacillus plantarum strains by PCR analysis and in vitro antibacterial studies. 1H NMR (Proton Nuclear Magnetic Resonance) analysis confirmed EPS production in only one strain, Limosilactobacillus fermentum MC1. The overall microbiome analysis suggests that human milk contributes to the establishment of the intestinal microbiota of infants. In addition, it is a promising source of novel Lactobacillus strains expressing specific functional biomolecules.

Human milk microbial species are associated with infant head-circumference during early and late lactation in Guatemalan mother-infant dyads.

Human milk contains abundant commensal bacteria that colonize and establish the infant's gut microbiome but the association between the milk microbiome and head circumference during infancy has not been explored. For this cross-sectional study, head-circumference-for-age-z-scores (HCAZ) of vaginally delivered breastfed infants were collected from 62 unrelated Mam-Mayan mothers living in eight remote rural communities in the Western Highlands of Guatemala during two stages of lactation, 'early' (6-46 days postpartum, n = 29) or 'late' (109-184 days postpartum, n = 33). At each stage of lactation, infants were divided into HCAZ ≥ -1 SD (early: n = 18; late: n = 14) and HCAZ < -1 SD (early: n = 11; late: n = 19). Milk microbiome communities were assessed using 16S ribosomal RNA gene sequencing and DESeq2 was used to compare the differential abundance (DA) of human milk microbiota with infant HCAZ subgroups at both stages of lactations. A total of 503 ESVs annotated 256 putative species across the 64 human milk samples. Alpha-diversity using Chao index uncovered a difference in microbial community richness between HCAZ ≥ -1 SD and HCAZ < -1 SD groups at late lactation (p = 0.045) but not at early lactation. In contrast, Canonical Analysis of Principal Coordinates identified significant differences between HCAZ ≥ -1 SD and HCAZ < -1 SD at both stages of lactation (p = 0.003); moreover, 26 milk microbial taxa differed in relative abundance (FDR < 0.05) between HCAZ ≥ -1 SD and HCAZ < -1 SD, with 13 differentially abundant at each lactation stage. Most species in the HCAZ ≥ -1 SD group were Streptococcus species from the Firmicutes phylum which are considered human colonizers associated with human milk whereas the HCAZ < -1 SD group at late lactation had more differentially abundant taxa associated with environmentally and 'potentially opportunistic' species belonging to the Actinobacteria genus. These findings suggest possible associations between brain growth of breastfed infants and the milk microbiome during lactation. Importantly, these data provide the first evidence of cross talk between the human milk microbiome and the infant brain that requires further investigation.

Fatty acid profile driven by maternal diet is associated with the composition of human milk microbiota

Little is known regarding the impact of diet on the breast milk microbiome. We hypothesized that vegan, vegetarian, and omnivore diets would impact the human milk microbiota. We also aimed to explore associations between human milk fatty acid concentrations and microbial composition. A cross-sectional microbiome diversity analysis of human milk samples (N = 72) was performed using 16S rRNA amplicon sequencing. Human milk microbial diversity was not associated with diet type. However, analysis of microbiome in relation to fatty acid profiles revealed significant differences in the overall composition of the human milk microbiota between high (&amp;gt; 0.7% of total fat) and low (&amp;lt; 0.7%) trans-fatty acid groups (TF) (p = 0.039, pairwise PERMANOVA p = 0.035), high (&amp;gt; 40%) versus low (&amp;lt; 40%) saturated fatty acids (UniFrac p = 0.083, PERMANOVA p = 0.094), and high (&amp;gt;60%) versus low (&amp;lt;60%) unsaturated fatty acids (UF) (UniFrac p = 0.094, PERMANOVA p = 0.093). 84% of samples from omnivore mothers were in the high TF group compared to only 12% of samples from vegans. Gut-associated species (Faecalibacterium, Blautia, Roseburia and Subdoligranulum) and Lactobacillus were characteristic of both high UF and TF groups, but not the low-fat groups. Functional analysis revealed 2,4-dichlorophenol 6-monooxygenase was differentially abundant in the high UF group. Although microbiome diversity did not differ by diet type, TF breast milk content differed by diet group, highlighting the relationship between maternal diet and the microbial profile of human milk.

Diversity and temporal dynamics of breast milk microbiome and its influencing factors in Chinese women during the first 6 months postpartum

Human breast milk (HBM) plays an important role in providing nutrients, beneficial microorganisms and bioactive components for infants, helping maturation of their immune system and gastrointestinal development. Here, we present a study aiming to investigate the diversity and temporal dynamics of the milk microbiome across the first 6 month postpartum in Chinese healthy breastfeeding women, and to investigate to what extent other variables (e.g., sampling location, infant sex, and mode of delivery) might also be related to variations in the human milk microbiome, and the association with maternal diet and nutrients. Fifty-three healthy pregnant women from four cities were recruited from a China Maternal and Infant Health Cohort Study and breast milk samples were collected and analyzed using 16S rRNA metagenomic sequencing. We illustrated the diversity and temporal dynamics during lactation (Adonis p-value = 3e-04). Firmicutes and Proteobacteria were the most abundant phyla, and Streptococcus, Staphylococcus, Serratia, and Corynebacterium were the core genera. Partitioning around medoids clustering identified two major internal clusters of breast milk microbiota. Cluster 1 was dominated by Acinetobacter and Pseudomonas, while Cluster 2 was dominated by Streptococcus and Staphylococcus. Among other environmental variables, sampling location showed significant influence on breast milk microbiome (Adonis p-value = 4e-04), while infant sex (Adonis p-value = 0.33) and mode of delivery (Adonis p-value = 0.19) were less related to variations in the human milk microbiome. Maternal diet such as tuber was significantly correlated with the relative abundance of Neisseria (rho = 0.34, adjusted p-value = 0.01) and Cutibacterium (rho = -0.35, adjusted p-value = 0.01), and nutrients such as carbohydrates were significantly correlated with the relative abundance of Aquabacterium (rho = -0.39, adjusted p-value = 0.0027), and vitamin B12 was significantly correlated with the relative abundance of Coprococcus (rho = 0.40, adjusted p-value = 0.0018), etc. These results illustrated the dynamic changes of composition and diversity during the lactation phases of the Chinese breast milk microbiome and addressed the importance of geographic location on milk microbiota, and associations with maternal diet consumption, which have potential benefits on the establishment and future health of breastfeeding infants.

Microbial Community Structure of Colostrum in Women with Antibiotic Exposure Immediately After Delivery.

<b><i>Background:</i></b> The microbial community in human milk is associated with many maternal and neonatal factors. This study aimed to investigate the effect of antibiotic exposure on the microbial community structure of colostrum. <b><i>Methods:</i></b> Twenty women with antibiotic treatment immediately after delivery and 10 age-matched control women were enrolled at the Guangdong Women and Children Hospital. Colostrum samples were collected within postpartum 30 hours. The V4 variable region of the bacterial 16S rRNA gene was sequenced to characterize the microbial profile using Illumina MiSeq platform. <b><i>Results:</i></b> Phyla <i>Proteobacteria</i> and <i>Firmicutes</i> were the predominant bacteria in colostrum samples. The core and abundant genera in colostrum included <i>Streptococcus</i>, <i>Staphylococcus</i>, and <i>Pseudomonas</i>. Compared with the control group, principal coordinate analysis based on the Bray-Curtis distance showed a significant difference in milk microbial community in women with antibiotic exposure, accompanied by a significantly lower alpha diversity and a different microbial ecological network. Furthermore, the relative abundances of genera <i>Actinomyces</i>, <i>Anaerobacter</i>, and <i>Clostridium_</i>sensu<i>_stricto</i> significantly decreased after antibiotic treatment. <b><i>Conclusions:</i></b> This study provided evidence of alterations in the colostrum microbial community with antibiotic exposure, improving our understanding of the effects of antibiotic treatment on the milk microbiome.

22 Young Scholar Talk PHD: Investigating Ovine Mastitis: Microbial Sources and Management Methods to Reduce the Prevalence

Abstract Subclinical mastitis, a common intramammary disease in sheep production, can be costly to producers due to compromised performance and decreased longevity. Our research focused on preventative and therapeutic measures on mammary health where the objectives were to identify (1) the impact of Zn supplementation on the milk microbiome and (2) the impact of management practices at lambing and weaning on prevalence and cure rate of subclinical mastitis, the milk microbiome, and levels of antimicrobial resistance in milk bacteria. In study 1, the milk microbiome was characterized during the first 3 wk of lactation and at weaning in ewes divergent in dietary Zn supplementation (1× = CON or 3× recommendations = ZnTRT). Mammary health was assessed using somatic cell count (SCC) class. Amplicon sequencing revealed the presence of Staphylococcus, Mannheimia, Corynebacterium, and Pseudomonas, all major mastitis pathogens. Effects of breed, dietary Zn concentration, and SCC class on diversity were limited to early lactation, where CON milk had greater composition stability than ZnTRT milk (P = 0.02). In study 2, milk bacteria were characterized by culture; Staphylococcus spp. and Streptococcus spp. were tested for antimicrobial susceptibility. Subclinical mastitis prevalence ranged from 22–66% where coagulase-negative staphylococci (CoNS; 59%), bacilli (35%), Mannheimia haemolytica (10%), S. aureus (8%), streptococci (5%), and Corynebacterium spp. (5%) were common. Milk samples were compared by jug bedding treatment: jugs were cleaned and dried with barn lime before adding fresh straw (CLEAN) or fresh straw was added atop soiled bedding (SOILED). Bedding had no effect on subclinical mastitis (P = 0.57), but CoNS had greater sulfadimethoxine resistance in SOILED isolates (P = 0.03). Additional mastitis control methods at weaning were evaluated: penicillin injection (PENN), restricted feed access (FAST), or their combination (COMBO). Weaning treatment did not affect cure rate from weaning to 3-d post- weaning (P = 0.21). Further characterization of these samples via microbial sequencing are underway with source tracking analyses to investigate an exogenous route via retrograde transfer and environmental bacteria or endogenous route via an entero-mammary pathway. Results support that breed, animal husbandry, and mammary health variably affect the milk microbiome and provide novel insight to the impact of management practices in a milk microbiome longitudinal study.

Maternal weight status and the composition of the human milk microbiome: A scoping review.

The human milk microbiome is thought to partly contribute to the assembly of the infant gut microbiome, a microbial community with important implications for infant health and development. While obesity has well-established links with the adult gut microbiome, less is known about how it affects the human milk microbiome. In this scoping review, we synthesize the current literature on the microbial composition of human milk by maternal weight status, defined broadly as BMI (prepregnancy and postpartum) and gestational weight gain (GWG). This study followed the a priori protocol published in Prospero (registration #: CRD42020165633). We searched the following databases for studies reporting maternal weight status and a characterization of milk microbiota through culture-dependent and culture-independent methods: MEDLINE, Embase, Web of Science, CINAHL, and Scopus. After screening 6,365 studies, we found 20 longitudinal and cross-sectional studies investigating associations between maternal weight status and the composition of the milk microbiome. While some studies reported no associations, many others reported that women with a pre-pregnancy or postpartum BMI characterized as overweight or obese, or with excessive GWG, had higher abundances of the genus Staphylococcus, lower Bifidobacterium abundance, and lower alpha diversity (within-sample diversity). This review suggests that maternal weight status is minorly associated with the composition of the milk microbiome in various ways. We offer potential explanations for these findings, as well as suggestions for future research.

Characterization of rumen, fecal, and milk microbiota in lactating dairy cows.

Targeting the gastrointestinal microbiome for improvement of feed efficiency and reduction of production costs is a potential promising strategy. However little progress has been made in manipulation of the gut microbiomes in dairy cattle to improve milk yield and milk quality. Even less understood is the milk microbiome. Understanding the milk microbiome may provide insight into how the microbiota correlate with milk yield and milk quality. The objective of this study was to characterize similarities between rumen, fecal, and milk microbiota simultaneously, and to investigate associations between microbiota, milk somatic cell count (SCC), and milk yield. A total of 51 mid-lactation, multiparous Holstein dairy cattle were chosen for sampling of ruminal, fecal, and milk contents that were processed for microbial DNA extraction and sequencing. Cows were categorized based on low, medium, and high SCC; as well as low, medium, and high milk yield. Beta diversity indicated that ruminal, fecal, and milk populations were distinct (p < 0.001). Additionally, the Shannon index demonstrated that ruminal microbial populations were more diverse (p < 0.05) than were fecal and milk populations, and milk microbiota was the least diverse of all sample types (p < 0.001). While diversity indices were not linked (p > 0.1) with milk yield, milk microbial populations from cows with low SCC demonstrated a more evenly distributed microbiome in comparison to cows with high SCC values (p = 0.053). These data demonstrate the complexity of host microbiomes both in the gut and mammary gland. Further, we conclude that there is a significant relationship between mammary health (i.e., SCC) and the milk microbiome. Whether this microbiome could be utilized in efforts to protect the mammary gland remains unclear, but should be explored in future studies.