Among the lactic acid bacteria (LAB) present in the oenological microbial ecosystem, Oenococcus oeni, an acidophilic lactic acid bacterium, is essential during winemaking. It outclasses all other bacterial species during malolactic fermentation (MLF). Oenological performances, such as malic acid degradation rate and sensorial impact, vary significantly according to the strain. The genetic diversity of the O. oeni species was evaluated using a multilocus sequence typing (MLST) scheme. Seven housekeeping genes were sequenced for a collection of 258 strains that had been isolated all over the world (particularly Burgundy, Champagne, and Aquitaine, France, Chile, South Africa, and Italy) and in several wine types (red wines, white wines, and champagne) and cider. The allelic diversity was high, with an average of 20.7 alleles per locus, many of them being rare alleles. The collection comprised 127 sequence types, suggesting an important genotypic diversity. The neighbor-joining phylogenetic tree constructed from the concatenated sequence of the seven housekeeping genes showed two major phylogenetic groups, named A and B. One unique strain isolated from cider composed a third group, rooting the phylogenetic tree. However, all other strains isolated from cider were in group B. Eight phylogenetic subgroups were statistically differentiated and could be delineated by the analysis of only 32 mutations instead of the 600 mutations observed in the concatenated sequence of the seven housekeeping genes. Interestingly, in group A, several phylogenetic subgroups were composed mostly of strains coming from a precise geographic origin. Three subgroups were identified, composed of strains from Chile, South Africa, and eastern France.