Given the problem of hybridization with domestic cats, there is a growing need to identify populations of the European wildcat Felis silvestris silvestris in order to protect the genetic integrity of this subspecies. In this paper, we use known locations of observations of wildcats or recovered carcasses to reassess the distribution of the wildcat in France and, in cases where carcasses were collected, we use both phenotypic and molecular genetic analyses to distinguish wildcats from hybrids with domestic cats. Spatially explicit multivariate analysis of wildcat' genotypes was then performed to define genetic units. Our study confirms the presence of wildcats in a large area of c. 155 000 km2, suggestive of a range of expansion, and divided into two clearly distinct and unconnected areas – the Pyrenees and the north-eastern part of France. However, European wildcat populations may be decreasing in the French Pyrenees, whereas the north-eastern part represents the main area (MA) of wildcat presence. This extension does not appear to be primarily due to hybrids, as both wildcats and hybrids were located throughout the MA. In addition, we found that genetic diversity of wildcats in the MA is remarkably high, suggesting that French populations are not threatened by a lack of genetic diversity. Furthermore, wildcats of the MA are structured into two genetically distinct populations that are contiguous and probably extend into Germany to form the largest area of wildcat presence in Europe and an area of major interest for their conservation. Our study calls for localized examination of the feasibility and usefulness of wildlife corridors to enhance connectivity between the different populations, thereby allowing sufficient levels of immigration and gene flow within the regional meta-population to ensure the long-term viability of these populations.