Rett Syndrome: A Focus on Gut Microbiota

Abstract

Rett syndrome (RTT) is an X-linked neurodevelopmental disorder affecting 1 in 10,000 live female births. Changes in microbiota composition, as observed in other neurological disorders such as autism spectrum disorders, may account for several symptoms typically associated with RTT. We studied the relationship between disease phenotypes and microbiome by analyzing diet, gut microbiota, and short-chain fatty acid (SCFA) production. We enrolled eight RTT patients and 10 age- and sex-matched healthy women, all without dietary restrictions. The microbiota was characterized by 16S rRNA gene sequencing, and SCFAs concentration was determined by gas chromatographic analysis. The RTT microbiota showed a lower α diversity, an enrichment in Bacteroidaceae, Clostridium spp., and Sutterella spp., and a slight depletion in Ruminococcaceae. Fecal SCFA concentrations were similar, but RTT samples showed slightly higher concentrations of butyrate and propionate, and significant higher levels in branched-chain fatty acids. Daily caloric intake was similar in the two groups, but macronutrient analysis showed a higher protein content in RTT diets. Microbial function prediction suggested in RTT subjects an increased number of microbial genes encoding for propionate and butyrate, and amino acid metabolism. A full understanding of these critical features could offer new, specific strategies for managing RTT-associated symptoms, such as dietary intervention or pre/probiotic supplementation.

Keywords: Rett syndrome; diet; microbiota; short-chain fatty acids.

Figure 1

α rarefaction curves according to Faith’s phylogenetic diversity index. (A) α-diversity plot of Rett (red) versus control (blue) samples. RTT patients show a reduced biodiversity compared to healthy controls. Differences are not statistically significant (p > 0.05); (B) Data grouped according to disease severity. Severe stage (purple) samples show a reduced biodiversity compared to healthy controls (blue), mild (orange), or intermediate (red) disease stage. Differences are not statistically significant (p > 0.05).

Figure 2

Principal Coordinate Analysis (PCoA) according to unweighted Unifrac distance. The first two components of the variance are represented. (A) PCoA of Rett (red) versus control (blue) samples. The two groups tend to separate according to PCoA 1 component. Differences are not statistically significant (p > 0.05); (B) Samples are colored according to disease severity. Severe stage (purple) significantly (p < 0.05) separates from healthy controls. Other differences are not statistically significant (p > 0.05).

Figure 2

Principal Coordinate Analysis (PCoA) according to unweighted Unifrac distance. The first two components of the variance are represented. (A) PCoA of Rett (red) versus control (blue) samples. The two groups tend to separate according to PCoA 1 component. Differences are not statistically significant (p > 0.05); (B) Samples are colored according to disease severity. Severe stage (purple) significantly (p < 0.05) separates from healthy controls. Other differences are not statistically significant (p > 0.05).

Figure 3

Bar charts representing the average relative abundance of Rett (RTT, n = 8) and control (CTR, n = 10) microbiota, classified using the 16S rRNA gene. (A) Mean relative abundances of fecal bacterial phyla; (B) Mean relative abundances of fecal bacterial families.

Figure 4

Box-and-whisker plots of fecal microbial metabolites in feces of Rett (RTT, magenta) and control (CTR, blue) subjects: (A–D) short-chain fatty acids (SCFAs); (E,F) branched-chain fatty acids (BCFAs). Data refer to mg/g wet weight (mg/g ww) of feces. Significant differences are indicated by ** (p < 0.01), Mann–Whitney test.

Figure 5

Correlations between intestinal family- and genus-level phylogenetic groups and fecal short-chain fatty acids (SCFAs) (heatmap). Right panel: correlations between intestinal bacteria and SCFAs are indicated by colors (magenta: positive; blue: negative). Asterisks (*) indicate statistically significant correlations (p < 0.05); Left panel: mean relative abundances in RTT and CTR are reported. Significant differences are indicated by * p < 0.05, and ** p < 0.01, Mann–Whitney test.

Figure 6

Functional characterization of carbohydrate metabolism, differentially represented in the gut microbiota of RTT and control subjects. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways have been identified by linear discriminant analysis coupled with effect size (LEfSe) (LDA  >  2, p < 0.05). Magenta histograms: pathways enriched in RTT patients; blue histograms: pathways enriched in control subjects.