Delivery Mode and the Transition of Pioneering Gut-Microbiota Structure, Composition and Predicted Metabolic Function

Abstract

Cesarean (C-section) delivery, recently shown to cause excess weight gain in mice, perturbs human neonatal gut microbiota development due to the lack of natural mother-to-newborn transfer of microbes. Neonates excrete first the in-utero intestinal content (referred to as meconium) hours after birth, followed by intestinal contents reflective of extra-uterine exposure (referred to as transition stool) 2 to 3 days after birth. It is not clear when the effect of C-section on the neonatal gut microbiota emerges. We examined bacterial DNA in carefully-collected meconium, and the subsequent transitional stool, from 59 neonates [13 born by scheduled C-section and 46 born by vaginal delivery] in a private hospital in Brazil. Bacterial DNA was extracted, and the V4 region of the 16S rRNA gene was sequenced using the Illumina MiSeq (San Diego, CA, USA) platform. We found evidence of bacterial DNA in the majority of meconium samples in our study. The bacterial DNA structure (i.e., beta diversity) of meconium differed significantly from that of the transitional stool microbiota. There was a significant reduction in bacterial alpha diversity (e.g., number of observed bacterial species) and change in bacterial composition (e.g., reduced Proteobacteria) in the transition from meconium to stool. However, changes in predicted microbiota metabolic function from meconium to transitional stool were only observed in vaginally-delivered neonates. Within sample comparisons showed that delivery mode was significantly associated with bacterial structure, composition and predicted microbiota metabolic function in transitional-stool samples, but not in meconium samples. Specifically, compared to vaginally delivered neonates, the transitional stool of C-section delivered neonates had lower proportions of the genera Bacteroides, Parabacteroides and Clostridium. These differences led to C-section neonates having lower predicted abundance of microbial genes related to metabolism of amino and nucleotide sugars, and higher abundance of genes related to fatty-acid metabolism, amino-acid degradation and xenobiotics biodegradation. In summary, microbiota diversity was reduced in the transition from meconium to stool, and the association of delivery mode with microbiota structure, composition and predicted metabolic function was not observed until the passing of the transitional stool after meconium.

Keywords: cesarean section; microbial community; microbiome; microbiota; obesity.

Figure 1

Bacterial alpha diversity in the meconium and transitional stool by delivery mode. Samples were rarefied to 1634 reads per sample. The nonparametric p values were calculated using 100,000 Monte Carlo permutation. Different letters indicate significant differences (e.g., ‘a’ is significantly different from ‘b’, but not significantly different from ‘a’); Statistical significance for phylogenetic diversity (PD) whole tree, p < 0.005; Observed species, p < 0.05. +, outlier samples.

Figure 2

Bacterial diversity in the meconium and transitional stool by delivery mode. (A–C) Principal Coordinate Analysis (PCoA) plot of bacterial communities in meconium (A) and transitional stool (B) by delivery mode. Weighted UniFrac distances were used to evaluate diversity between samples. PERMANOVA was used to test dissimilarity. (C) Box plot of intra-group distances. The non-parametric p values were calculated using 100,000 Monte Carlo permutation. Different letters (a, b and c) indicate significant differences; p < 0.001.

Figure 3

Bacterial taxa comparisons in meconium and transitional stool by delivery mode. (A) Each taxonomy (>1% of average relative abundance in any groups) is indicated by a different color at the genus level. ** Indicates overrepresented taxa (using LDA > 3.0) in comparisons of delivery mode within sample type. (B) Histogram of overrepresented taxa (using LDA > 3.0) in each group.