A balance between pro- and anti-inflammatory mechanisms at mucosal interfaces, which are sites of constitutive exposure to microbes and non-microbial foreign substances, allows for efficient protection against pathogens yet prevents adverse inflammatory responses associated with allergy, asthma and intestinal inflammation. Regulatory T (T_reg) cells prevent systemic and tissue-specific autoimmunity and inflammatory lesions at mucosal interfaces. These cells are generated in the thymus (tT_reg cells) and in the periphery (induced (i)T_reg cells), and their dual origin implies a division of labour between tT_reg and iT_reg cells in immune homeostasis. Here we show that a highly selective blockage in differentiation of iT_reg cells in mice did not lead to unprovoked multi-organ autoimmunity, exacerbation of induced tissue-specific autoimmune pathology, or increased pro-inflammatory responses of T helper 1 (T_H1) and T_H17 cells. However, mice deficient in iT_reg cells spontaneously developed pronounced T_H2-type pathologies at mucosal sites—in the gastrointestinal tract and lungs—with hallmarks of allergic inflammation and asthma. Furthermore, iT_reg-cell deficiency altered gut microbial communities. These results suggest that whereas T_reg cells generated in the thymus appear sufficient for control of systemic and tissue-specific autoimmunity, extrathymic differentiation of T_reg cells affects commensal microbiota composition and serves a distinct, essential function in restraint of allergic-type inflammation at mucosal interfaces.