# Targeting the gut-brain axis in autism: machine learning analysis of gut microbiome shows association with autism spectrum disorder

D Rigters, L N Peralta-Marzal, N Prince, P Perez-Pardo, A Kraneveld, Alejandro Lopez-Rincon

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## ► To cite this version:

D Rigters, L N Peralta-Marzal, N Prince, P Perez-Pardo, A Kraneveld, et al.. Targeting the gut-brain axis in autism: machine learning analysis of gut microbiome shows association with autism spectrum disorder. FIGON/EUFEPS European Medicines Days, FIGON; EUFEPS, Jun 2022, Leiden, Netherlands. hal-05139817

HAL Id: hal-05139817

https://hal.science/hal-05139817v1

Submitted on 2 Jul 2025

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# Targeting the gut-brain axis in autism: machine learning analysis of gut microbiome shows association with autism spectrum disorder

D. Rigters¹, L.N. Peralta-Marzal¹, N. Prince¹, P. Perez-Pardo¹, A. Kraneveld¹, A. Lopez-Rincon¹

¹Dept. of Pharmacology, Utrecht University

## Aims

Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by reduced social functioning and repetitive behavior. The gut microbiome has been considered as a potential target for treatments, as ASD individuals suffer from intestinal problems and show a differentiated gut microbiome. However, studies from different cohorts rarely agree on the specific bacterial taxa that are involved, and a recent study concluded that the gut microbiome is not related at all¹. Despite this, fecal transplantation has been tentatively shown to alleviate both behavioral and intestinal symptoms². Our aim is to find bacterial taxa, that are relevant to the ASD phenotype.

## Methods

We processed 16S amplicon sequencing data of the gut microbiome of children with ASD and neurotypical children using DADA2. This was done for data from a sibling-controlled study³, for feature selection, and two age-matched studies⁴,⁵ to validate our findings. The data from the sibling-controlled study was analyzed using recursive ensemble feature selection (REFS)⁶ with 8 classifiers. As the gut microbiome is sensitive to environmental factors and siblings share most of these factors, the resulting features are more likely to be associated to ASD.

## Results

REFS, applied to the sibling-controlled data resulted in 26 bacterial features that predict ASD. The reduced feature-set was evaluated with 5 classifiers not part of the ensemble, to avoid over fitting, in a nested 10-cross-validation scheme. The 26 bacterial features were applied to the validation data of 2 different cohort studies and the reduced datasets were evaluated with the same method. The results are summarized in the table below, with the best ROC curves for each dataset:

<table><thead><tr><th></th><th>Sibling-controlled<sup>3</sup></th><th>PRJNA578223<sup>4</sup></th><th>PRJNA589343<sup>5</sup></th></tr></thead><tbody><tr><th>Subjects</th><td>60 ASD, 57 control</td><td>48 ASD, 48 control</td><td>77 ASD, 50 control</td></tr><tr><th>Total features</th><td>2040</td><td>18758</td><td>2030</td></tr><tr><th>REFS features</th><td>26</td><td>20</td><td>22</td></tr><tr><th>AUC</th><td>0.93</td><td>0.85</td><td>0.84</td></tr></tbody></table>

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## Conclusion

Analysis of the gut microbiome has resulted in a set of bacterial taxa that can be used to predict the ASD status of children in 3 distinct cohorts with an AUC over 0.80. Bacteria from the class of *Clostridia* were enriched in ASD subjects, indicating that this class might play a role in this disorder. Our results indicate that the gut microbiome has a robust association with ASD and should not be disregarded as a potential target for treatment.

## References:

1-Yap, C. X. et al. Autism-related dietary preferences mediate autism-gut microbiome associations. Cell 184, 5916–5931 (2021).
2-Kang, D.-W. et al. Long-term benefit of microbiota transfer therapy on autism symptoms and gut microbiota. Sci. Reports **9**, 1–9 (2019).

3-David, Maude M., et al. Children with Autism and Their Typically Developing Siblings Differ in Amplicon Sequence Variants and Predicted Functions of Stool-Associated Microbes. *Msystems* **6.2** (2021): e00193-20.

4-Zou, R. et al. Changes in the gut microbiota of children with autism spectrum disorder. *Autism Res.* **13**, 1614–1625 (2020).

5-Ding, X. et al. Gut microbiota changes in patients with autism spectrum disorders. *J. Psychiatr. Res.* **129**, 149–159 (2020).

6-Lopez-Rincon, et al. Automatic discovery of 100-mirna signature for cancer classification using ensemble feature selection. *BMC bioinformatics* **20**, 1–17 (2019).