Effects of Probiotic Supplementation on Symptoms and Microbiome Characteristics in Patients With Non‐Celiac Gluten/Wheat Sensitivity: A Randomized Controlled Double‐Blind Trial

Non-celiac gluten/wheat sensitivity (NCGWS) remains a controversial condition lacking a clear pathophysiological mechanism and specific biomarkers. This randomized, double-blind, placebo-controlled trial aimed to explore whether probiotic supplemen- tation may allow the reintroduction of gluten/wheat in NCGWS individuals. Thirty NCGWS participants were randomized to receive either a probiotic formulation ( n = 15) ( Lactiplantibacillus plantarum , Lacticaseibacillus paracasei , and Ligilactobacillus salivarius ) or placebo ( n = 15) for 6 weeks. The intervention included a 4-week gluten-free diet low in fermentable oligosaccharides, disaccharides, monosaccharides, and polyols (T1), followed by a 2-week gluten reintroduction (T2). At baseline (T0), T1, and T2, participants completed a modified Gastrointestinal Symptom Rating Scale questionnaire and provided fecal samples. Gut microbiome was analyzed using shotgun sequencing. Volatilomic profiling was performed via comprehensive two-dimensional gas chromatography. In probiotic-treated group, 47% of participants exhibited improved tolerance to gluten reintroduction, whereas no improvement was observed in the placebo arm ( p = 0.003). At T2, probiotic-treated participants showed a shift in gut microbiome composition and displayed higher relative abundance of beneficial bacteria (such as L. plantarum , Bifidobacterium adolescentis , and Coprococcus catus ) and lower species correlated to gut inflammation (such as Bacteroides vulgatus and Bacteroides dorei ). Changes in metagenomic functions related to bacteriocin transport and biosynthesis, carbohydrates metabolism, and protein degradation occurred in probiotic-treated individuals. Furthermore, individuals with improved short-term gluten tolerance exhibited higher abundance of genes involved in gliadin hydrolysis and increased propanoic acid levels. Our results suggest that probiotics treatment may improve gluten tolerance in individuals with NCGWS. The beneficial effect might be related to the increased abundance of microbial genes involved in gluten digestion.