Preterm infants at low risk for early-onset sepsis differ in early fecal microbiome assembly

Abstract

Antibiotics are administered near-universally to very low birth weight (VLBW) infants after birth for suspected early-onset sepsis (EOS). We previously identified a phenotypic group of VLBW infants, referred to as low-risk for EOS (LRE), whose risk of EOS is low enough to avoid routine antibiotic initiation. In this cohort study, we compared 18 such infants with 30 infants categorized as non-LRE to determine if the lower risk of pathogen transmission at birth is accompanied by differences in microbiome acquisition and development. We did shotgun metagenomic sequencing of 361 fecal samples obtained serially. LRE infants had a higher human-to-bacterial DNA ratio than non-LRE infants in fecal samples on days 1-3 after birth, confirming lower bacterial acquisition among LRE infants. The microbial diversity and composition in samples from days 4-7 differed between the groups with a predominance of Staphylococcus epidermidis in LRE infants and Enterobacteriaceae sp. in non-LRE infants. Compositional differences were congruent with the distribution of virulence factors and antibiotic resistant genes. After the first week, the overall composition was similar, but changes in relative abundance for several taxa with increasing age differed between groups. Of the nine late-onset bacteremia episodes, eight occurred in non-LRE infants. Species isolated from the blood culture was detected in the pre-antibiotic fecal samples of the infant for all episodes, though these species were also found in infants without bacteremia. In conclusion, LRE infants present a distinct pattern of microbiome development that is aligned with their low risk for EOS. Further investigation to determine the impact of these differences on later outcomes such as late-onset bacteremia is warranted.

Keywords: Preterm; low-risk; microbiome; sepsis; vertical transmission.

Figure 1.

Clinical characteristics of the study cohort. (a) Depiction of study design and comparison groups. (b) Timeline of diet, clinical culture results, antibiotic exposure, and fecal sample collection. Each square of the grid represents one day in the life of an infant enrolled in the study. Infant study IDs are followed by codes to indicate sex (F = female; M = male) and delivery mode (C = cesarean delivery; V = vaginal delivery). A filled black dot (•) denotes a fecal sample, a large open circle (O)denotes a negative blood culture, a diagonal cross (✕) denotes a positive blood culture, and a line (―) denotes antibiotics.

Figure 2.

Microbiome of LRE and non-LRE infants in the first week of life. Fecal samples of infants are shown as open circles when obtained on day 1–3 (0–72 h) after birth and closed circles when obtained on day 4–7 (72–168 h) after birth. (a) The proportion of human reads obtained in shotgun metagenomic sequencing was higher in LRE infants on days 1–3 but not different on days 4–7. (b) The Shannon diversity of bacterial species was not different on days 1–3 but was lower in LRE infants on days 4–7. (c) Comparison of bacterial community composition based on Bray-Curtis distance between samples, and taxon relative abundance in two high-density sample clusters. (d) Relative abundance of taxa (logarithmic scale) found to be different between LRE and non-LRE infants on days 4–7.

Figure 3.

Microbiome of LRE and non-LRE infants after the first week of life. (a) Shannon diversity of samples after the first week of life. (b) Comparison of bacterial community composition by Bray-Curtis distance between samples. The species with highest abundance in each sample is indicated by color. (c) Time course of the most abundant species in infant fecal samples after the first week of life. Stacked bars at each time point indicate the most abundant species in each sample. Curves connect samples from the same infant that were collected at consecutive time points.

Figure 4.

Virulence factors and antibiotic resistance genes in LRE and non-LRE infants. (a) Selected virulence factors and (b) antibiotic resistance genes found to be different between LRE and non-LRE infants (complete set in Figure S4 and Figure S5, respectively). In each plot, the x-axis has units of reads per kilobase gene length per million reads (RPKM). Lines extend from the first to the third quartile, and the median is indicated by a point. An asterisk indicates a difference in gene abundance between LRE and non-LRE infants after controlling for a 5% false discovery rate. (c) Partial correlation analysis of virulence factor and (d) antibiotic resistance gene abundance and species abundance. Grey squares represent genes not found in the species based on a database search. An asterisk indicates a statistically significant correlation after controlling for a 5% false discovery rate.

Figure 5.

Fecal microbiome during episodes of bacteremia. (a) Relative abundance of bacterial species in pre-antibiotic fecal samples obtained prior to clinical evaluation resulting in the diagnosis of bacteremia. (b) Time course of relative abundance for the bacterial species matching those obtained from blood culture in each episode. Dashed lines indicate the days that antibiotics started and ended during each episode. Episodes E5 and E8 did not have post-antibiotic samples. The final sample in episode E7 was obtained prior to completion of therapy, as the infant was transferred to a different facility.