# Exploring gut microbiome and nutritional status among children with Autism Spectrum Disorder (MY-ASD Microbiome): A study protocol

## TL;DR

This study explores gut and oral microbiota differences in children with autism and how these relate to nutrition, aiming to improve early intervention strategies.

## Contribution

The study introduces a protocol to investigate gut microbiota and nutritional status in ASD children in Malaysia, linking microbial profiles to dietary behaviors.

## Key findings

- The study will identify oral and gut microbiota differences between children with ASD and typically developing controls.
- It will explore associations between microbiota profiles and nutritional status in children with ASD.
- Findings may inform personalized nutrition and healthcare strategies for ASD children.

## Abstract

Autism Spectrum Disorder (ASD) is a neurodevelopmental disorder characterised by persistent deficits in social communication and the presence of restricted, repetitive behaviours or interests. Previous literature has identified a link between the gut and ASD; however, the underlying mechanisms remain unclear. Gut microbiota dysbiosis has been extensively reported in cohort studies of ASD, and specific microbial metabolites or by-products may serve as potential biomarkers for ASD. Additionally, children with ASD often exhibit food refusal, have a limited food repertoire and display a tendency to consume the same foods frequently; thus, these behaviours increase their risk of malnutrition (over-nutrition or under-nutrition) compared to typically developing (TD) healthy children. This study primarily aims to identify oral and gut microbiota among children with ASD and TD healthy children. The secondary aim is to determine the associations between oral and gut microbiota with nutritional status among children with ASD. The findings will enhance understanding of the aetiology of ASD and inform early intervention strategies to mitigate disease severity and early identification of malnutrition in genetically at-risk children.

This observational, age-matched, case-control study is conducted in Malaysia among 40 male children with ASD and age-matched with 40 TD healthy controls aged 4–10 years. The dependent variables include the microbiota profile, identified through metagenomic sequencing analysis of saliva and faecal samples, and autism severity, assessed through validated questionnaires. Independent variables include nutritional status, determined through Subjective Global Nutrition Assessment (SGNA), anthropometry and dietary measurements, gastrointestinal symptoms, eating behaviour, behavioural profile, and sleep quality. Data collection is expected to be completed by June 2026. The study nature may limit causality establishment. Analyses will use chi-square/ANOVA for group comparisons, SparCC for microbiota correlations, and mixed-effects logistic regression to model associations.

This study advances understanding of ASD-related microbiota, guiding personalised nutrition and precision healthcare in Malaysia.