The Mediterranean diet (MD) has beneficial effects on the intestinal microbiota by the promotion of bacteria associated with a healthy gut. However, its impact on intestinal fungi, among others, is still unknown, and how it affects digestive symptoms and different biomarkers in patients with gastrointestinal (GI) disorders has hardly been explored. The present study evaluated the effect of the MD on gut microbial diversity and structure and intestinal symptoms and biomarkers after 6 weeks of dietary intervention in 46 patients with GI disorders. Dysbiosis in fungal composition and diversity was observed, with a significantly lower abundance of Sordariomycetes, Leotiomycetes, and Orbiliomycetes; a significantly higher abundance of Saccharomycetes; the Chytridiomycota and Mucoromycota phyla were significantly reduced; and the bacterial microbiota remained unchanged. In addition, various GI disorders decreased and associations between stool consistency and intestinal permeability were found with the bacterial genera