The pampas cat (Leopardus colocolo) is a poorly known small felid that occurs in open habitats of South America. Previous studies have suggested that the disjunct pampas cat population occurring in the critically endangered Uruguayan Savannah ecoregion are a distinct ‘Evolutionarily Significant Unit’ (ESU), and as such should be the target of separate conservation assessment. Here, we developed spatial distribution models for this ESU, using them to estimate population size and then assess its conservation status based on IUCN criteria. We constructed two models (with and without anthropogenic variables), each of them built separately with two different algorithms (Maxent and Maxlike). All models were very similar, with strongly correlated results (rs>0.88; P<0.01), consistently indicating higher suitability (or probability of occurrence [PO]) in grassland areas from sea level to 400 m of altitude. The two models incorporating anthropogenic variables provided the best fit to the data, and yielded a more restricted distribution in higher suitability and PO areas, supporting the conclusion that this pampas cat population is directly affected by human disturbance and enabling us to identify priority areas for its conservation. Using available density estimates, the population size within high suitability and PO areas would be <250 and 1,800–9,000 individuals, respectively, suggesting that this ESU may be Critically Endangered or Endangered. Very little of the high-suitability landscape is presently protected, highlighting the urgent need to expand the few existing protected areas in this region, as well as to create new ones to ensure the long-term survival of this elusive felid.