The Microbiome and Pediatric Health

Measuring the Microbiome

Prior knowledge of microbes on and around the human body was based on specific methods to cultivate organisms. Molecular technologies have revolutionized the identification of poorly cultivatable microbes, rare microbes, and microbes in complex communities such as those associated with the human body ( Fig. 196.1 ). The development of the polymerase chain reaction (PCR) and the availability of modern nucleic acid sequencing have improved the sensitivity of detection of many organisms and resulted in the discovery of new organisms. Modern sequencing technologies, called next-generation sequencing platforms, allow sequencing in high volume and depth, with millions of sequences obtained from a single biologic sample. Three major approaches utilize next-generation sequencing to understand the composition, diversity, and activity of the microbiome: (1) sequencing species-specific regions of genomes such as ribosomal RNA–encoding tracks and intergenic regions termed metagenomics , (2) total DNA sequencing from a sample (e.g., feces, saliva) and assembly of the sequence fragments into large genome pieces termed shotgun metagenomics , and (3) RNA transcript sequencing to decipher the composition and, as a surrogate for functional activity, the transcriptional activity of a microbiome termed metatranscriptomics . Massive computational power and new bioinformatics tools have allowed the analysis and comparison of the large datasets arising from these methods.

Two additional approaches to measure the microbiome phenotype have rapidly developed as well. First, large-scale measurements of the peptide composition of the microbiota, called proteomics , have been increasingly used to describe the activity of a microbiome sample, as peptides provide information about the composition and function of a microbiome. Second, in a complementary approach called metabolomics , microbiome-derived metabolites are measured using advanced gas chromatography and mass spectrometry techniques. Together, proteomics and metabolomics better describe the activity of a microbiome than the nucleotide-sequencing approaches; however, at this point, they provide less depth of resolution and specificity relative to the composition and phenotype of a microbiome.

Despite the power of these methodologies to interrogate the microbiome, they do not yet replace cultivation of microbes in many clinical circumstances. Cultivation of organisms still represents the most practical means to differentiate potential pathogenic species from more benign species and to provide clinically actionable information such as susceptibility to a range of antimicrobials.

Early Childhood Development of the Microbiome

Emerging studies suggest that the placenta and fetus are exposed to microbes in utero, but the effect of such an exposure remains to be fully appreciated. Prematurity as a complication of an infection of the fetal membranes and either subclinical or clinical chorioamnionitis may alter the in utero exposure to microbes. The rupture of the fetal membranes and subsequent delivery provide substantial exposure to new maternal and environmental microbes that will assume common places in the developing microbiota. Mode of delivery has a major influence on the early life microbiome, with vaginally delivered infants becoming acutely colonized with intestinal organisms that reflect the mother's vaginal tract and infants delivered by cesarean section becoming colonized with organisms reflective of maternal skin and oral cavity, including staphylococci and streptococci, as well as the surrounding environment.

In the term infant delivered vaginally, the first intestinal microbes, so-called pioneering organisms, include Escherichia and other Enterobacteriaceae , Bacteroides, and Parabacteroides . Exclusive breastfeeding has been reported to result in high levels of bifidobacteria and Lactobacillus in the week following the start of feeding. These probiotic organisms have unique capacities to exclude would-be pathogens from colonization by sequestering nutrients and producing antimicrobial factors while stimulating the intestinal epithelium to tighten cellular junctions and express antimicrobial peptides. However, these genera have been notably deficient from some breastfed infant cohorts, particularly in the United States.

The premature infant is more likely to be delivered by cesarean section and thus is more abundantly colonized with skin-related organisms such as coagulase-negative staphylococci, similar to the term infant delivered by cesarean section. However, the premature infant may fail to progress through the same stages of expansion and diversification of the microbiome over the 1st week to month of life as the term infant. The factors related to the delayed maturation are not fully clear but are predictably related to delayed or limited enteral feeding, normal environmental exposure to the household environment, and exposure to medical interventions such as antimicrobial therapy.

The most significant shift in the intestinal microbiota appears to occur after weaning and the introduction of solid foods. As the infant transitions from breast milk to a solid-food diet containing complex plant-derived polysaccharides, the microbiota begins to reshape progressively into a more mature composition beginning to resemble the adult microbiota. At the same time, the metabolic potential of the microbiome shifts to accommodate the changing diet, with the newborn microbiome enriched with phosphotransferase system (PTS) genes and then shifting to increasing abundance of lactose transporter genes by 4 mo of age, reflecting milk intake, and further shifting to a high abundance of genes such as β-glucoside transporters and enzymes necessary to break down complex carbohydrates by age 12 mo. The maturation of the childhood microbiome after the early years to adulthood is less well understood, and more studies are required in large numbers of children to understand fully the developmental stages of maturation and similarity to the mature, healthy adult state.

The oral microbiota of the newborn is of maternal origin, with vaginally born infants having predominantly Lactobacillus, Prevotella, and Sneathia, whereas cesarean-born infants have more maternal skin organisms, including Staphylococcus, Corynebacterium, and Propionobacterium .Within the 1st day of life, Firmucutes dominate the oral cavity, including Streptococcus and Staphylococcus. Formula-fed babies acquire more Bacteroidetes, whereas breast-fed babies have more bacteria of the phyla Proteobacteria and Actinobacteria. With the eruption of first teeth, new environmental niches are formed to foster microbial communities. Although cariogenic bacteria such as Streptococcus mutans were thought to be acquired after dentition, recent data demonstrate the presence of these organisms prior to tooth eruption in a soft tissue reservoir, highlighting the importance of good infant oral care even before primary dentition.

By age 3 yr, the childhood oral and salivary microbiome is complex but less diverse than the adult microbiome. The composition of the microbiota within the oral cavity in the presence of full adult dentition has an estimated 1,000 bacterial species. Even with oral health, the diversity in the gingiva of different types of teeth ( geodiversity ) is substantial, and the diversity changes dramatically with the development of oral disease such as periodontitis. How the microbiota evolves between preschool and adulthood remains a topic for future study. Furthermore, the placement and removal of oral hardware for orthodontics is common in childhood and may produce significant alterations in the microbiome of the oral cavity.

During the 1st yr of life, the infant skin microbiome increases in diversity, including species richness and evenness. The skin of the younger infant is relatively undifferentiated between body sites, with more shared species among different body sites such as arms, forehead, and buttocks than the older infant when the microbial communities at each site undergo differentiation. As with the early infancy oral microbiota, skin of the young infant skin is predominantly colonized by Firmicutes, including Streptococcaceae and Staphylococcaceae, with the inclusion of bacteria from other phyla such as Actinobacteria, Proteobacteria, and Bacteroidetes as the skin matures. The adult skin microbiome displays a high degree of geodiversity—differences in composition depending on site and local physiology, with major differences in dry and wet skin sites. However, the linkage between skin development in childhood and maturation of the skin microbiome remains a subject of ongoing studies.

Social structure and family interactions likely play a significant role in the development of the early life microbiome. Breast milk feeding provides a microbiologic link between mothers and infants, including transmission of probiotic-like organisms such as lactobacilli and bifidobacteria, each of which may have some protective effects, including protection against diarrheal diseases and atopy. Pediatricians have long been aware of the infectious disease risks and benefits of daycare attendance, with examples of shared pneumococcal strains producing otitis media and outbreaks of respiratory syncytial virus infection and associations with reduced atopy, allergy, and possibly asthma. Family contacts are risks for acquisition of methicillin-resistant Staphylococcus aureus and subsequent disease. Studies also demonstrate transmission of parts of the human microbiome between household individuals and domesticated pets such as dogs and cats. For example, family members share the same strains of Escherichia coli known to produce urinary tract infections in one of the household members. There may be differences in the oral microbiota among infants for whom the parents did and did not use the practice of pacifier sucking for cleaning. In rural settings, microbiome sharing extends to livestock, household surfaces, and household members. Thus, development of the microbiome during childhood with environmental interactions is a complicated process that continues to be explored.

The Microbiome and Physiologic Development

Increasingly complex roles are being identified for the microbiome in the development of mammalian physiology ( Fig. 196.2 ). These roles include the development of the enteral tract, respiratory tract, immune system, hematologic system, metabolic-endocrine system, and neurologic system. The details of how the microbiome contributes to these developmental processes in humans are still under intense investigation; however, modeling in other mammalian systems predicts that the microbiome will have a critical role across species.