A bacterial strain designated M1MS02(T) was isolated from a surface-sterilized nodule of Medicago sativa in Zamora (Spain). The 16S rRNA gene sequence of this strain showed 96.5 and 96.2 % similarity, respectively, with respect to Gluconacetobacter liquefaciens IFO 12388(T) and Granulibacter bethesdensis CGDNIH1(T) from the family Acetobacteraceae. The novel isolate was a Gram-stain-negative, non-sporulating, aerobic coccoid to rod-shaped bacterium that was motile by a subpolar flagellum. The major fatty acid was C18 : 1ω7c and the major ubiquinone was Q-10. The lipid profile consisted of diphosphatidylglycerol, phosphatidylethanolamine, two aminophospholipids, three aminolipids, four glycolipids, two phospholipids and one lipid. Strain M1MS02(T) was catalase-positive and oxidase- and urease-negative. Acetate and lactate were not oxidized. Acetic acid was produced from ethanol in culture media supplemented with 2 % CaCO3. Ammonium sulphate was assimilated in glucose medium. The strain produced dihydroxyacetone from glycerol. Phylogenetic and phenotypic analyses commonly used to differentiate genera within the family Acetobacteraceae showed that strain M1MS02(T) should be classified as representing a novel species of a new genus within this family, for which the name Endobacter medicaginis gen. nov., sp. nov. is proposed. The type strain of the type species is M1MS02(T) ( = LMG 26838(T) = CECT 8088(T)). To our knowledge, this is the first report of a member of the Acetobacteraceae occurring as a legume nodule endophyte.