Impact of Ex Vivo Bisphenol A Exposure on Gut Microbiota Dysbiosis and Its Association with Childhood Obesity

Abstract

Dietary exposure to the plasticiser bisphenol A (BPA), an obesogenic and endocrine disruptor from plastic and epoxy resin industries, remains prevalent despite regulatory restriction and food safety efforts. BPA can be accumulated in humans and animals, potentially exerting differential health effects based on individual metabolic capacity. This pilot study examines the impact of direct ex vivo BPA exposure on the gut microbiota of obese and normal-weight children, using 16S rRNA amplicon sequencing and anaerobic culturing combined methods. Results showed that direct xenobiotic exposure induced modifications in microbial taxa relative abundance, community structure, and diversity. Specifically, BPA reduced the abundance of bacteria belonging to the phylum Bacteroidota, while taxa from the phylum Actinomycetota were promoted. Consistently, Bacteroides species were classified as sensitive to BPA, whereas bacteria belonging to the class Clostridia were identified as resistant to BPA in our culturomics analysis. Some of the altered bacterial abundance patterns were common for both the BPA-exposed groups and the obese non-exposed group in our pilot study. These findings were also corroborated in a larger cohort of children. Future research will be essential to evaluate these microbial taxa as potential biomarkers for biomonitoring the effect of BPA and its role as an obesogenic substance in children.

Keywords: dysbiosis; ex vivo BPA exposure; microbiota taxa; obesity.

Figure A1

Stool sampling and ex vivo BPA exposure procedure.

Figure A2

Boxplots showing the relative abundance distribution of significantly different bacterial genera and phyla and their specific ratios between study groups (a) Bifidobacterium; (b) Bacteroides; (c) Clostridium; (d) Ratio Bacteroides/Bifidobacterium; (e) Ratio Bacteroides/Clostridium; (f) Actinomycetota; (g) Bacteroidota; (h) Bacillota; (i) Ratio Bacteroidota/Actinomycetota; (j) Ratio Bacteroidota/Bacillota. NW, green; OB, blue; NW10, pink; OB10, orange.

Figure 1

Description of children’s gut microbiota before and after ex vivo BPA exposure according to the study groups, results based on 16S rRNA gene amplicon sequencing. (a) Box plots of the alpha diversity indices; (b) beta diversity: nMDS plot based on Bray–Curtis distance, with samples as points and ellipses coloured by study groups. PERMANOVA test results (R², F, p-value) are indicated in the plot. NW, green; OB, blue; NW10, pink; OB10, orange. Mean relative abundance of indicated phyla (c) and ASVs (d).

Figure 2

Differentially abundant ASVs between exposed and non-exposed groups. To identify taxa that were differentially abundant due to BPA exposure within each BMI group, ANCOM-BC2 analysis was performed: (a) green bars indicate taxa that were significantly more abundant in the NW group, while pink represents taxa that were significantly more abundant in the NW10 group; (b) blue bars indicate taxa that were significantly more abundant in the OB group, while red represents taxa that were significantly more abundant in the OB10 group. * features not sensitive to pseudo-count addition. ASVs with a log fold change between—1.99 and 0 are not shown here to simplify plot size.

Figure 3

A heatmap displaying the top 50 ASVs with significant changes in relative abundance between the study groups and NW group (used as the reference), detected by MaAsLin2s with default parameters. An increase in relative abundance is shown in red (+) and a decrease in this is shown in blue (−).

Figure 4

Box plot representing CFU/g from each study group after 1, 15, and 30 days of anaerobic culture in recovery enrichment media. NW, green; OB, blue; NW10, pink; OB10, orange.

Figure 5

Heatmap of the relative abundance of identified isolates in each group, accompanied by a phylogenetic tree. Stars highlight species identified by sequencing after MALDI TOF failed to identify them. Circles indicate species sensitive to BPA, cultured in control samples (NW, OB) but absent after being exposed to BPA (NW10, OB10). Triangles indicate species showing resistance to BPA, only cultured after exposure to BPA or those whose relative abundance was at least double that of non-exposed samples.

Figure 6

Comparison of 16S rRNA sequencing and culturing results. (a) Venn diagram depicting overlap between genera identified by 16S rRNA sequencing (blue) and culturing (yellow); (b,c) heatmaps representing the relative abundance of the common genera identified by 16S rRNA sequencing and culturing. “g_Clostridium” represents the relative abundance of “g_Clostridium sensu stricto 1”.

Figure 7

Box plots showing the relative abundance distribution of significantly different bacterial genera and phyla and their specific ratios between study groups: (a) Bifidobacterium; (b) Bacteroides; (c) Clostridium; (d) Bacteroides/Bifidobacterium ratio; (e) Bacteroides/Clostridium ratio; (f) Actinomycetota; (g) Bacteroidota; (h) Bacillota; (i) Bacteroidota/Actinomycetota ratio; (j) Bacteroidota/Bacillota ratio. NW, green; OB, blue.