Folic acid, folate receptor autoantibodies, glutathione and vitamin D3 as biomarkers of cerebral folate deficiency in children with autism spectrum disorder: A case‑control study from Iraq

Abstract

Objective: To assess serum levels of vitamin D3, folic acid and reduced glutathione in children with autism spectrum disorder (ASD) in relation to the presence of folate receptor alpha (FOLR1) autoantibodies, and to explore associations with ASD severity.

Methods: A case‑control study was conducted between October 2025 and January 2026 at the QAR Autism Rehabilitation Centre and Alshatra General Hospital in Thi Qar, Iraq. The study included 60 children with ASD diagnosed according to DSM‑5 criteria and 60 age‑ and sex‑matched healthy controls. Serum anti‑FOLR1 IgG, vitamin D3, folic acid and reduced glutathione were measured by ELISA and automated chemistry. ASD severity was categorised using the Childhood Autism Rating Scale (CARS). Comparisons between seropositive and seronegative ASD children were performed with Mann–Whitney U and chi‑square tests.

Results: Among the 60 ASD children, 46 (76.7%) were positive for anti‑FOLR1 IgG. Compared with seronegative ASD children, seropositive children had significantly lower levels of vitamin D3 (median 13.74 vs 29.20 ng/mL, p=0.002), folic acid (median 9.94 vs 21.50 ng/mL, p=0.002) and reduced glutathione (median 0.83 vs 2.15 μmol/L, p<0.001). Anti‑FOLR1 positivity was strongly associated with greater ASD severity (severe: 63.0% in seropositive vs 14.3% in seronegative, p=0.002).

Conclusion: The presence of FOLR1 autoantibodies in children with ASD is associated with marked depletion of vitamin D3, folic acid and glutathione, and with more severe clinical presentation. These findings support the existence of a distinct metabolic‑immunological subtype of ASD that may benefit from targeted biomarker‑guided interventions.