We report an investigation of antibodies to Toxoplasma gondii in 52 wild birds and 20 wild mammals from northern and central areas of Portugal by using the modified agglutination test. The birds comprised 26 common buzzards (Buteo buteo), five tawny owls (Strix aluco), four white storks (Ceconia ceconia), three Eurasian eagle owls (Bubo bubo), three northern goshawks (Accipiter gentilis), two booted eagles (Hieraaetus pennatus), two common barn owls (Tyto alba), two Eurasian sparrowhawks (Accipiter nisus), two short-toed eagles (Circaetus gallicus), one black kite (Milvus migrans), one Griffin vulture (Gyps fulvus), and one peregrine falcon (Falco peregrinus). The mammals were eight wild boars (Sus scrofa), six red foxes (Vulpes vulpes), two common genets (Genetta genetta), two European badgers (Meles meles), one European roe deer (Capreolus capreolus), and one Iberian wolf (Canis lupus signatus). Fifty percent of the wild birds and 90% of the wild mammals were seropositive; the overall seroprevalence of infection was 61.1%. When comparing the prevalence of antibodies in birds and mammals from northern Portugal, a significant difference was found, but the same was not true for birds and mammals from central Portugal. Seroprevalence levels were 30.0% in juvenile and 62.5% in adult birds (p=0.046), 0.0% in juvenile and 94.7% in adult mammals (p=0.100), 80.0% in female and 66.7% in male birds (p=1.000), and 81.8% in female and 100% in male mammals (p=0.479). This is the first study performed on T. gondii in birds of prey, white storks, and wild carnivores in Portugal.