Relationship Between Calcium and Gut Microbial Composition and Metabolic Pathways in Children with Autism

Background/Objectives: Trace elements may influence autism spectrum disorder (ASD) severity through interactions with the gut microbiota and microbial metabolic functions, but calcium-related evidence remains limited. This cross-sectional study examined associations among hair calcium, gut microbial taxa, metabolic pathways, and behavioral phenotypes in children with ASD. Methods: We analyzed 183 children with ASD who had behavioral assessments, hair calcium measurements, and fecal shotgun metagenomic sequencing data. Participants in the lowest and highest calcium quartiles were first compared to characterize group-level microbiome differences. Full-sample analyses then tested associations among continuous hair calcium, microbial taxa, metabolic pathways, and behavioral measures after covariate adjustment. Benjamini-Hochberg false discovery rate correction was applied for multiple testing. Results: Hair calcium was positively associated with CARS, ATEC-Total, ATEC-1, and ATEC-3 scores, with the strongest associations involving ATEC-1 and ATEC-3. Alpha and beta diversity did not differ significantly between calcium quartile groups, but group-based microbiome analyses identified 63 differential species and 22 differential MetaCyc pathways. Full-sample integrated analyses connected calcium-associated microbial taxa, metabolic pathways, and ASD behavioral measures. Conclusions: Hair calcium was associated with ASD behavioral severity, selected gut microbial species, and microbial metabolic pathways. These findings support an association framework connecting longer-term calcium-related mineral profiles, gut microbial functional potential, and behavioral phenotypes, providing a basis for future longitudinal and multi-omics studies.