Microbiome assembly across multiple body sites in low-birthweight infants

Abstract

The purpose of this study was to evaluate the composition and richness of bacterial communities associated with low-birthweight (LBW) infants in relation to host body site, individual, and age. Bacterial 16S rRNA genes from saliva samples, skin swabs, and stool samples collected on postnatal days 8, 10, 12, 15, 18, and 21 from six LBW (five premature) infants were amplified, pyrosequenced, and analyzed within a comparative framework that included analogous data from normal-birthweight (NBW) infants and healthy adults. We found that body site was the primary determinant of bacterial community composition in the LBW infants. However, site specificity depended on postnatal age: saliva and stool compositions diverged over time but were not significantly different until the babies were 15 days old. This divergence was primarily driven by progressive temporal turnover in the distal gut, which proceeded at a rate similar to that of age-matched NBW infants. Neonatal skin was the most adult-like in microbiota composition, while saliva and stool remained the least so. Compositional variation among infants was marked and depended on body site and age. Only the smallest, most premature infant received antibiotics during the study period; this heralded a coexpansion of Pseudomonas aeruginosa and a novel Mycoplasma sp. in the oral cavity of this vaginally delivered, intubated patient. We conclude that concurrent molecular surveillance of multiple body sites in LBW neonates reveals a delayed compositional differentiation of the oral cavity and distal gut microbiota and, in the case of one infant, an abundant, uncultivated oral Mycoplasma sp., recently detected in human vaginal samples.

Importance: Complications of premature birth are the most common cause of neonatal mortality. Colonization by the indigenous microbiota, which begins at delivery, may predispose some high-risk newborns to invasive infection or necrotizing enterocolitis (NEC), and protect others, yet neonatal microbiome dynamics are poorly understood. Here, we present the first cultivation-independent time series tracking microbiota assembly across multiple body sites in a synchronous cohort of hospitalized low-birthweight (LBW) neonates. We take advantage of archived samples and publically available sequence data and compare our LBW infant findings to those from normal-birthweight (NBW) infants and healthy adults. Our results suggest potential windows of opportunity for the dispersal of microbes within and between hosts and support recent findings of substantial baseline spatiotemporal variation in microbiota composition among high-risk newborns.

Figures

FIG  1

Stacked bar plots depicting the relative abundances of the 30 most abundant genus-level taxa in the LBW infants. Taxa were ranked according to their mean abundance across all samples (percentages at right). Ten taxa had mean abundances of >1.00% (percentages in bold type within parentheses). Cs, Cesarean section delivery; V, vaginal delivery.

FIG  2

Unweighted UniFrac-based principal coordinate analysis (PCoA) of LBW infant-associated bacterial communities. Each symbol represents the value for a sample, with the shape of the symbol indicating the infant (infants 1 to 6) and the color indicating the body site. The percentages of variation explained by the plotted principal coordinates (PCo1 and PCo2) are indicated on the axes.

FIG  3

Average unweighted UniFrac distances between LBW infants (present study) and healthy adults (references , and ; see Materials and Methods) for oral, skin surface, and stool microbiota (250 sequences per sample). Values that are significantly different by Tukey’s posthoc tests are indicated by bars and 4 asterisks (P < 0.0001). Error bars represent 95% confidence intervals.

FIG 4

Relationship between neonatal stool microbiota composition and time. (a) Average (95% CI) within-subject, unweighted UniFrac distance plotted against the age difference (lag) in days for LBW (R2 = 0.5) and NBW (R2 = 0.5) infants. Lines indicate best fit linear regressions. The NBW infants were not sampled on day 18; thus, for this analysis, the corresponding age was excluded from the LBW data set. (b) For LBW infants at various ages, average (95% CI) unweighted UniFrac distance compared to a healthy reference cohort (the 21-day-old NBW infants).

FIG  5

Relative abundances of three OTUs belonging to the Mycoplasmataceae from oral samples from extremely LBW baby 3. OTU 15, a novel, uncultivated Mycoplasma sp., is plotted against the upper y axis. OTUs 53 and 144, which are closely related to OTUs from cultivated strains, are plotted against the lower y axis. Expansion of OTU 15 coincided with antibiotic treatment from DOL 13 to 19 (see Table 2 for details). Feedings were delivered via naso- or orogastric tube. For antibiotics, the date range is indicated. The baby’s diet (breast milk or formula) is given for each sample date.