Accumulation of Black Carbon Particles in Placenta, Cord Blood, and Childhood Urine in Association with the Intestinal Microbiome Diversity and Composition in Four- to Six-Year-Old Children in the ENVIR ON AGE Birth Cohort

Abstract

Background: The gut microbiome plays an essential role in human health. Despite the link between air pollution exposure and various diseases, its association with the gut microbiome during susceptible life periods remains scarce.

Objectives: In this study, we examined the association between black carbon particles quantified in prenatal and postnatal biological matrices and bacterial richness and diversity measures, and bacterial families.

Methods: A total of 85 stool samples were collected from 4- to 6-y-old children enrolled in the ENVIRonmental influence ON early AGEing birth cohort. We performed 16S rRNA gene sequencing to calculate bacterial richness and diversity indices (Chao1 richness, Shannon diversity, and Simpson diversity) and the relative abundance of bacterial families. Black carbon particles were quantified via white light generation under femtosecond pulsed laser illumination in placental tissue and cord blood, employed as prenatal exposure biomarkers, and in urine, used as a post-natal exposure biomarker. We used robust multivariable-adjusted linear models to examine the associations between quantified black carbon loads and measures of richness (Chao1 index) and diversity (Shannon and Simpson indices), adjusting for parity, season of delivery, sequencing batch, age, sex, weight and height of the child, and maternal education. Additionally, we performed a differential relative abundance analysis of bacterial families with a correction for sampling fraction bias. Results are expressed as percentage difference for a doubling in black carbon loads with 95% confidence interval (CI).

Results: Two diversity indices were negatively associated with placental black carbon [Shannon: $-4.38\%$ (95% CI: $-8.31\%$, $-0.28\%$); Simpson: $-0.90\%$ (95% CI: $-1.76\%$, $-0.04\%$)], cord blood black carbon [Shannon: $-3.38\%$ (95% CI: $-5.66\%$, $-0.84\%$); Simpson: $-0.91$ (95% CI: $-1.66\%$, $-0.16\%$)], and urinary black carbon [Shannon: $-3.39\%$ (95% CI: $-5.77\%$, $-0.94\%$); Simpson: $-0.89\%$ (95% CI: $-1.37\%$, $-0.40\%$)]. The explained variance of black carbon on the above indices varied from 6.1% to 16.6%. No statistically significant associations were found between black carbon load and the Chao1 richness index. After multiple testing correction, placental black carbon was negatively associated with relative abundance of the bacterial families Defluviitaleaceae and Marinifilaceae, and urinary black carbon with Christensenellaceae and Coriobacteriaceae; associations with cord blood black carbon were not statistically significant after correction.

Conclusion: Black carbon particles quantified in prenatal and postnatal biological matrices were associated with the composition and diversity of the childhood intestinal microbiome. These findings address the influential role of exposure to air pollution during pregnancy and early life in human health. https://doi.org/10.1289/EHP11257.

Figure 1.

Participant flow chart depicting the selection of participants enrolled in the ENVIRONAGE birth cohort for arriving at the final study sample size. Note: Only mother–child pairs who already participated in the 4-y follow-up study up to 1 y before the house visit or who were going to participate within 1 y after the house visit, mother–child pairs who did not (plan to) move between the house visit study and the 4-y follow-up study and who had no major renovations planned during the house visit study were eligible for inclusion in this study. ASV, amplicon sequence variant; ENVIRONAGE, ENVIRonmental influence ON early AGEing birth cohort.

Figure 2.

Evidence of black carbon particles in (A) placental tissue, (B) cord blood, and (C) urine. White light generation originating from black carbon particles (yellow, indicated with a white arrow) under femtosecond pulsed laser illumination (excitation $810\text{\hspace{0.17em}nm}$, 120 fs, 80 MHz) was observed. Images represent the overlap of channel 1 (green, emission $450–650\text{\hspace{0.17em}nm}$) and channel 2 (red, emission $400–410\text{\hspace{0.17em}nm}$). All samples were collected in the ENVIRONAGE birth cohort and images were randomly selected from different participants. Scale bar: $50\mathrm{\mu }\mathrm{m}$. Note: ENVIRONAGE, ENVIRonmental influence ON early AGEing birth cohort.

Figure 3.

Correlation graphs between (A) placental black carbon and residential black carbon exposure averaged over the entire pregnancy ($n=63$), (B) cord blood black carbon and residential black carbon exposure averaged over the entire pregnancy ($n=80$), and (C), (D), and (E) urinary black carbon normalized for osmolality ($n=80$) and residential black carbon exposure averaged over the (C) preceding month, (D) preceding 6 months, and (E) preceding year of the house visit. See Table S5 for corresponding numeric data. Participants are enrolled in the ENVIRONAGE birth cohort. Note: ENVIRONAGE, ENVIRonmental influence ON early AGEing birth cohort.

Figure 4.

Overview of the relative abundance (percentage) of the 10 most abundant bacterial families in relation to all other taxa. Height of bars represents the relative abundance. Families are ranked in increasing order from bottom to top. Participants were enrolled in the ENVIRONAGE birth cohort. $n=85$. ENVIRONAGE, ENVIRonmental influence ON early AGEing birth cohort.

Figure 5.

(Partial) Spearman correlation coefficients with 95% CI between Chao1 richness index, Shannon diversity index, and Simpson diversity index and placental black carbon load, cord blood black carbon, or urinary black carbon normalized for osmolality, based on corrected (partial/adjusted) and uncorrected models. Partial Spearman and adjusted coefficient of determination models were adjusted for parity, season of delivery, batch, age, sex, weight, height, and maternal education per categories included in Table 1. See Table S7 for corresponding numeric data. Participants were enrolled in the ENVIRONAGE birth cohort. * indicates $p$ $\le 0.05$. $p$-Values were calculated using pairwise (partial) Spearman correlation. Placenta $n=63$, cord blood $n=80$, and urine $n=80$. Note: BC, black carbon; CI, confidence interval; ENVIRONAGE, ENVIRonmental influence ON early AGEing birth cohort.

Figure 6.

Percentage of variance ($R^2$) in Shannon or Simpson diversity explained by different covariables. Covariables were categorized as depicted in Table 1. See Table S8 for corresponding numeric data. Participants were enrolled in the ENVIRONAGE birth cohort. Note: ENVIRONAGE: ENVIRonmental influence ON early AGEing birth cohort.